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" W*_A 8,D@D@D H5q=]znt9n6{ Ʋ5"W<ˁss5z$r@++qcPD@DTps$ɸv ZD@D@D$?@KkiנE@D@D@O@2s$ɸv ZD@D@D$?@KkiנE@D@D@O@?ulR_}ƍ7o9sԨQWЫW_|ŪUbûp˗/q˗'NLj"3gά_޳K.]z5kVXxjN۷/yEg| v޽hѢ9=و@9(Wy(D@{ƍ1Ǐ߳gOwuuXFyӜ9snϞ=[z'?\|AqÇ{_^nۻwox":n Iy*Ep)ۿW_}Uf՚@d~" -Jڵk/fH1cXOoڴi<&H? /=zN<ِ!CS2sua0;ƋY/1Bz鷽~]3f{T}>yd .ꌏŋ1CzӬ.К>=M;,Kd\@! _|9$&dbϱ4hPhd3tPwʕ+ Yь3]K\_~~V x…л cۉ'$wWRlRر4qC"O@2Ο,E@CYtiT rJ]&M]dvy3vӧO*RӧOV 3zȵQ2$"[ N>u' J" %!7a+CʐS,]j5Váհ1cwM2H`Vҩlt1_Ҭ{!DfΜ$p,X0ydZ ҋ" $ h9ݳ18,eDRnݺaÆmOn"-cЂػwo4'oHΆ2cfM^ ܱc/f.z Xd~" -GH?P۠a]WR4Uc`Zx7t0 *[Ե޿?TuBZuӯBD aS_-8-Meb)XU|N9NqɭkB'hi eXmş,$ WOj[DY ߪrl߾=z|;w8ӔjvZhlg3te˘ɭk)3ɸX'Xsڈ8:}ĉǚyU3w^hQ T$r@+" "P-wWz駟~:l0Ν;5k :4ڵkaÆs4bŊ9s減H1?ݾ}{edN/!C^;zj =ͪk'nٲE'VQ&9)"P*׮][x1CڳgϘ1cLM6 <̟(G ^'O 4>uT{{{Pؙ鷳^*K':u/cǎ,A/'XM;?_oceYqGBD08 aC? sbZ ƍ \2p%j… Y1'|ү_hdrvj?s.O~:{ۉ'"bR ;&1fz$8kYDҥKF+W,4iR6ҬfH߿XeG/IۊZ2ַYV?= ڟOݎ^B+4I`Io5jTʐS,]j5V!԰15e}:wWi)8M&1Mq>9/"P0 #@F֢Uka0`շo_N-c3S͛7]o>fYnۯ[/Ų({qqr@D XhiՀSBMԊ)9>CrO=* Uvizer7ƳqѴ.Ο?ߧ;v`/:G-/+لj8" MMIՔ*Ll^WJߤ-l׮];44dQ?P~X H;S~I3QqP2(%UK6@Sp;5,AOzFƉ-[ #b.ߊU{UӬ'!JܹLqU}fN"$$d"@Kp\[ʀ=zdOU|~U{ʩ|1uׯ_,o3aZ}ً@skyW" $f CEpˤ!!&mDzT;ݣՐ~c[,D{Wï*S2B+4I!ߗTu2z,e?>C'ڦ Ygr=;4!F'B/t-ڔd\!IN ۣgs6oGF:ٳg=teLgf>K gljȹ#149g!u4@GGSQ я~#_/~ӺΝ;ևc&9K6b[oaF?XYs7"O4a0&__v.ٛ7o>=gtʋMb94 f*5@ZM7f)%Bx5Gl?A[[H2l,}F9ĉvTѣGGKQWW " ["@@2N? ($+iO@K`xY 7}t4o5 F5"" E">e mٷSM@MHF IENDB`[(   N:kashi@techunix.technion.ac.il>*racheli@ibexperts.com0ugophna@dal.ca/ 00DArialNew Rot$t\ )0tY 0"DComic Sans MS$t\ )0tY 0B DMS Sans Serif$t\ )0tY 00DTimes New Romant\ )0tY 0c .  @n?" dd@  @@`` 5( ~ n ($%&'  #! " _b$ ov飼kf#A2$%Gg_gDj#A2$nI8i v o&A2$0h,m5A @,7Ab$L-s$֠n:A 0AA ff33@@K2ʚ;n8ʚ;g4BdBd )0hppp@ <42d2d 0$t<4BdBd 0$t( <4!d!dy 0$0___PPT10 ppR___PPT94,&@ &@L@@4@@?  %QPathogenomics in Israel$ff"    Metagenomics LGT = lateral gene transfer Typing of bacterial strains and drug resistance Identification of virulence factors Whole genome analysis 33"   )%#Facilities for pathogenomic studies"$ $G$ff.    Good facilities for genomics, transcriptomics and proteomics in several universities and research institutes High level bioinformatics All facilities are also available as service E33.    Metagenomics ( $ff("   zMetagenomics of culturable and non-culturable microorganism populations present in biofilms of Acute Otitis Media (AOM, Middle Ear Infection) This infection involves a variety of bacterial species, found in the form of biofilms, which are inherently resistant to antibiotic treatment Detection of microbial biodiversity of AOM biofilm is limited due to current cultivation methods The group of Fauzi Silbaq (ArabQual) and Racheli Kreisberg-Zakarin (IBEX) racheli@ibexperts.com in involved in a study the biodiversity of AOM using a metagenomics approach, including microbiology, functional genomics and bioinformatics methodologies v P P33l33 33v3333         &     q  e  <               8   X  0Metagenomics $ff MMetagenomics of intestinal microflora in health and disease and the effect of the TLR mutations Toll-Like Receptors (TLRs) recognize pathogen specific patterns of microorganisms. Mutations in TLRs are associated with inflammatory bowel diseases (Crohn's disease and ulcerative colitis). Patients with those diseases have altered intestinal microflora Uri Gophna, currently with Ford Doolitle in Halifax ugophna@dal.ca performs culture-independent profiling of intestinal microflora of TLR knockout mice under normal conditions and after a challenge which models inflammatory bowel diseaseza Z Z ZaG33G33 G33G33     Q  G  2  W          6   l  0pLGT  lateral gene transfer in relation to pathogenesis (97$ff$8 Uri Gophna studies the role of lateral gene transfer (LGT) in the evolution of pathogens Uses bioinformatics for the identification of laterally acquired genes and pathways in pathogens \ 5E33E33~E33E336    pLGT  lateral gene transfer in relation to pathogenesis (97G$ff(8 lYair Aharonowitz, Ilya Borovok and Gerald Cohen from Tel Aviv University study gluthathione synthesis Identified GshF orthologs, consisting of a -glutamylcysteine ligase (GshA) domain fused to an ATP-grasp domain, in 20 gram-positive and gram-negative bacteria. Remarkably, 95% of these bacteria are mammalian pathogens. Presumably, this fusion gene, once formed, spread between mammalian hosts most likely by horizontal gene transfer Rf ZP8 Z2 ZG33C(        1                  q    1 Typing of bacterial strains and drug resistance 62(/$ff(1 -The group of Chezi Kashi kashi@techunix.technion.ac.il uses novel methods for molecular typing ofVibrio cholera in order to study emerging new pathogenic strains The group of Sima Yaron (Technion) is involved in typing the serovars of Salmonella enterica in respect to drug resistance and virulence l. aG33G33~G33G33.G33       )  J          -  06  #Identification of virulence factors$$G$ff# The group of Sima Yaron has developed a rapid, simple screen for real-time quantification of promoter - activity in S. enterica using a library of plasmids with GFP as a reporter The group of Nir Asherov (Tel Aviv U.) studies Aspergillus fumigatus, which causes serious disease (around 60% mortality) in immunocompromised people. They concentrate on studying novel cell wall proteins and identified 68 such cwp s. They are studying these CWPs in vitro and in infected animals by the use of deletion mutants. They are currently in the process of preparing a Cwp-specific microarrays to study Cwp gene expression patternsl ZtG33 G33G33G33RG33G33uG33z     `   0          9  V    s           Whole genome analysis$ff Two genomes of bacterial pathogens are being sequenced and analyzed in Israel Bacillus anthracis (Shaefferman et al, Biological Institute, Nes Ziona) Septicemic Escherichia coli  strain of serotype O78 (Ron, Tel Aviv U.) N  NE33G 33AG 33G 33G 33G 33 G  33G 33G 33W             *  Virulent E. coli strainsF G$ffG$ff G$ff Most of the E. coli strains are commensal, but a small number are pathogenic Pathogenic E. coli strains are divided into two groups: Intestinal strains. These produce enterotoxins and constitute a major problem, especially in young children and travellers (Montesumu s revenge) Extraintestinal strains  ExPEC (Extraintestinal Pathogenic E. coli)  Z Z Z G33G33FG33G33&G33E33E33E33G33G33       B               Septicemia (colibacillosis)G$ff.    Colisepticemia is the major causes of mortality from community and hospital-acquired infections (more than 80%) Main cause of mortality in immuno-supressed patients (HIV, chemotherapy, old age) Colisepticemia is an emerging disease  83% increase 1980  1992, over 40% of the bacteremia cases in community acquired infectionsG E33e33e33e33e33e33 ~     '  D   ' Goals:6G$ffg ff Define virulence-essential ExPEC-specific genes Profile strains involved in UTI, NBM and sepsis using these ExPEC-specific genes Use the data to define potential targets for development of vaccines and/or antibacterial drugs.| /E33E33QE33_e33 e 33 E 33v    <  o  CIdentification of virulence related sequences in septicemic strainsDDG$ff.1    3Whole genome sequencing Subtractive hybridization >3  3E33E33 4 Subtractive hybridization  G$ff  Obtain pathogen specific sequences, absent from non-pathogenic K12 strain Excellent chance of  hitting pathogenicity islands which are pathogen specific and very large Faster (and much cheaper) than whole genome sequencing E33  Subtractive hybridization2 G$f3G$f3  QA way to study comparative genomics with organisms which have not been sequenced .R PE33E33 R PSearch for unique  septicemic sequences))G f32      PUsing suppression subtractive hybridization (SSH) we identified sequences unique to strain O78-9 and absent from the non-pathogenic strain K-12 Oover 80 O78-specific open reading frames were found (91 to 1473 bp in length) The same experiment was repeated with another septicemic strain O2-1772 117 unique O2 sequences were identified Q g6GgIg'ggZ  <  ;   9 ,Comparison of unique sequences of O2 and O78--G$f3,  ^Although the two strains cause the same disease, there is a high diversity between the SSH libraries of O2 and O78 strains, with only a few shared genes coding for virulence factors. Is this diversity serotype specific? To determine this we profiled additional septicemic strains of the same serotypes the presence of each of the unique sequences l_ Z|e9eeEe:   L   EComparison of unique sequences of two septicemic strains - O2 and O78FFG f32&     high level of genome plasticity there is a high diversity between the septicemic strainsunexpected for strains causing the same disease Septicemic strains of serogroups O2 and O78 contain a large pool of virulence genes which are used in a  mix and match fashionD  Ye0e33e##tG     &       ` #Current-future researchG$ff $Molecular-physiological studies of the newly-identified virulence genes Bioinformatic studies to determine the evolution of these virulence genes (many in PAIs with evidence of LGT) Whole genome sequencing  joint project with Prof. Joerg Hacker and Prof. Gerhard Gottschalk: E3333E33bH   F  J  $ ($ Thank you!!"  G$ff  TAU group Uri Gophna Diana Ideses Daphna Mokady Dr. Dvora Biran Collaborations Wuerzburg University (Prof. Joerg Hacker) Greifswald University (Prof. Michael Hecker) Goetingen University (Prof. Gerhard Gottschalk)   Z7 Z Z Z G337G33G33sG3333*                          ( "Metagenomics ( Uri Gophna, currently with Ford Doolitle in Halifax studies the effect of TLR mutations on micrflora in health and disease by culture-independent profiling of intestinal microflora of TLR knockout mice under normal conditions and after a challenge which models inflammatory bowel diseaseH  0@|    3   F   k MLST of O78 strains" G$f3  Multi Locus Sequence Typing 450  500 bp of 7  housekeeping genes Criteria for chosing genes: 97-98% homology to E. coli K-12 (from blast data) appear in pathogenic and non pathogenic strains map at considerable distance from each other several allels in the populationf_ Z Z_EEEEZ&  (       ! [The MLST results are compatible with the results from subtractive hybridization and sequencing The profile of virulence factors in ExPEC strains is independent of the host and independent of the serotype There is a high diversity of virulence genes in the various E. coli septicemic strains and each strain has its own profile of virulence genesP\ AE##=AAKA@     @ !"#  Both libraries contain many sequences associated with genomic plasticity - evolution by horizontal gene transfer Many sequences of O2 and O78 are homologous to virulence related sequences of human ExPEC strains ns  c  tecEA&      /$%&'  ` 33` Sf3f` 33g` f` www3PP` ZXdbmo` \ғ3y`Ӣ` 3f3ff` 3f3FKf` hk]wwwfܹ` ff>>\`Y{ff` R>&- {p_/̴>?" dd@,|?" dd@   " @ ` n?" dd@   @@``PR    @ ` ` p>> (    6,W  `}  X Click to edit Master title style!!   0Y  `  RClick to edit Master text styles Second level Third level Fourth level Fifth level!    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S  < 6: _   : ^*   < 6F: _Q : `*  H < 0޽h ? 3380___PPT10.\[0  0 0(   x  c $@ `}   x  c $ `  H  0޽h ? 3380___PPT10.\@ $  $(  r  S D `}   r  S  `  H  0޽h ? 3380___PPT10.^pv0  0(  x  c $ `}   x  c $j<  H  0޽h ? 3380___PPT10.\ۇ0   0(   x  c $` `   x  c $<i  H  0޽h ? 3380___PPT10.\ۇ0  0(  x  c $:'g  : x  c $:P L : H  0޽h ? 3380___PPT10.\Y   ((  (r ( S T :q< :  ( s :0e0e #" 0e `}  : H ( 0޽h ? 3380___PPT10.\`WQK? b>Z>@}},=(  ,L , c $X99? , 6  j   lEscherichia coli,  B  4B ,@ # 7D `r  , 6 +  Salmonella typhimurium,  B&     4B ,@ # 7D 3r 4B ,@ # 7 rD  , 6 9AZ  Vibrio cholerae,  B4    4B  ,@ # 7 TH 4B  ,@ # 7 $   , 6Xp t\  Pasteurella multocida0  3F34      4B  ,@ # 7 h{   , 6% T  Haemophilus influenzae,  B4      4B ,@ # 7 h 4B ,@ # 7 h   , 6+ k  Actinobacillus actinomycetemco0  3F34    4B ,@ # 7  4B ,@ # 7   , 62 m  Haemophilus somnus0  3F34     4B , # 7P  4B ,@ # 7    , 649 Actinobacillus pleuropneumonia0  3F34    4B , # 7  4B ,@ # 7 $ 4B ,@ # 7 $ 9  , 6?Tm Pseudomonas aeruginosa,  B&     4B , # 7x9  4B ,@ # 79 q   , 6EM Rhodospirillum rubrum,  B4    4B , # 7Jq 4B ,@ # 7Wq    , 6 Lw3] Anabaena cylindrica,  B&     4B !, # 7b  ", 6\R;S Chlamydia trachomatis,  B&     4B #, # 7 4B $, # 7  W4B %,@ # 7R > W &, 6Wa  Clostridium perfringens0  3F3&     4B ', # 74 R  (, 6]  Clostridium acetobutylicum,  B&    4B ), # 70 R 4B *, # 7R  +, 6c  Enterococcus faecalis0  3F34     4B ,, # 7<  X -, 6,j_@ @ Enterococcus faecium0  3F34     4B ., # 7W  X4B /, # 7X 2c 0, 6poy N Lactobacillus plantarum0  3F3&    4B 1, # 7 -D 2, 6u6  Lactobacillus delbrueckii,  B&    4B 3, # 7-D4B 4, # 7D-2c4B 5, # 7c2D 6, 6{=W n Bacillus subtilis,  B  4B 7, # 7p 8, 6́  Staphylococcus aureus,  B&   4B 9, # 76=4B :, # 7 ;, 6D&v  Listeria monocytogenes0  3F3  4B <, # 7w =, 68H  Listeria innocua0  3F3  4B >, # 7 w4B ?, # 7w4B @, # 7D4B A, # 7?D B, 6@p  Streptococcus uberus0  3F3&   4B C, # 7 D, 6(]X  Streptococcus pyogenes,  B&   4B E, # 74B F, # 7v G, 6  Streptococcus agalactiae0  3F3&    4B H, # 7v 4B I, # 7mv J, 6?  Streptococcus suis0  3F3&   4B K, # 7m4B L, # 7dm M, 6h4U  Streptococcus gordonii0  3F3&   4B N, # 79] O, 6z   Streptococcus pneumoniae,  B&    4B P, # 79x4B Q, # 7d94B R, # 7*d S, 6ܸ-9  Streptococcus sobrinus0  3F3&   4B T, # 75< U, 6|JEa  Streptococcus mutans0  3F3&   4B V, # 75<<4B W, # 7*<54B X, # 7K*  Y, 6T3@   Lactococcus lactis,  BB      4B Z, # 7K!4B [, # 7?K4B \, # 7 ?4B ], # 7 ! ^, 6 s N   Mycoplasma pneumoniae,  BB       4B _, # 7W   `, 6Do/  Borrelia japonica,  B(    4B a, # 7 g b, 6(   Leptospira interrogans,  BB       4B c, # 7 | 4B d, # 7W 4B e, # 7! W4B f, # 7!> R4B g, # 7R> ? [ h, 6 3 Q   Streptomyces coelicolor,  BB       4B i, # 7  j, 6<Z    Corinebacterium glutamicum,  BB      4B k, # 7 " d  l, 6  % vMycobacterium tuberculosis,  B  4B m,@ # 7 4B n, # 7 4B o, # 7>  p, 6d h Hb Thermotoga maritima,  B4     4B q,@ # 7 I N r, 6fA  Thermus aqaticus,  B4    4B s,@ # 7 e 44B t,@ # 7  u, 6  Chloroflexus aurantiacus,  B4      4B v,@ # 7 4B w,@ # 7 > 4B x, # 7[> ? 4B y, # O4B z, # 4B {, # OP |, 6h[ f0.052 gB  } }, 0tw  Distribution of fused glutathione biosynthetic genes mapped onto a universal tree of bacterial 16S rRNA (Minimum Evolution, ME)T( 2"Dff Fff:DffFff&c   H , 0޽h ? 33___PPT10i.#X =+D=' = @B +   0(  x  c $m: Y  : x  c $dn:` : H  0޽h ? 33___PPT10i.\ +D=' = @B +}  04$(  4r 4 S $H: `  : r 4 S H:D : H 4 0޽h ? 33___PPT10i.\@+D=' = @B +  P0(  x  c $d6: `}  : x  c $: ` : H  0޽h ? 33___PPT10i.\iZ+D=' = @B +v   `L\(  Lx L c $({: `}  :  L c $|: `<$ 0 : H L 0޽h ? 33R J ___PPT10* .+} <D' = @B D' = @BA?%,( < +O%,( < +DA' =%(D' =%(D' =A@BBBB0B%(D' =1:Bvisible*o3>+B#style.visibility<*LN%(Ds' =%(D' =%(D' =A@BBBB0B%(D' =1:Bvisible*o3>+B#style.visibility<*LN%(D' =A@BBBB0B%(D' =1:Bvisible*o3>+B#style.visibility<*L%(D' =A@BBBB0B%(D' =1:Bvisible*o3>+B#style.visibility<*L\%(+8+0+L, + PP(  P P s *` *Extraintestinal diseases caused by E. coliH+#G ffG ffG ff"  5 P s *46 / Urinary tract infections (UTI) (pyeolonephritis, kidney failure, productivity loss) UTIs are responsible for > seven million patient visits and one million hospital admissions (due to complications) per year in the United States only. 80 - 90% of the cases are caused by E. coli Neonatal meningitis: bacterial meningitis 0.25 per 1000 live births in industrialized countries (2.66 per 1000 in developing countries). ~30% caused by E. coli , ~10% mortality Intra-abdominal infections, Respiratory tract infections, Wound and surgical infections SepticemiaHU( 2( 2,( 2( 2d( 2ETG33CG##G##C)G33CnG##G##G##YG33 G##@!  &   H P 0޽h ? 33___PPT10i.N+D=' = @B +   pX\(  Xx X c $: `}  :  X c $:0 `P<$ 0 : H X 0޽h ? 33ia___PPT10A.+} <D' = @B D' = @BA?%,( < +O%,( < +DA' =%(D' =%(D' =A@BBBB0B%(D' =1:Bvisible*o3>+B#style.visibility<*Xp%(DA' =%(D' =%(D' =A@BBBB0B%(D' =1:Bvisible*o3>+B#style.visibility<*Xp%(DA' =%(D' =%(D' =A@BBBB0B%(D' =1:Bvisible*o3>+B#style.visibility<*XG%(+8+0+X, +  `0(  `x ` c $< l  l x ` c $ l0 l H ` 0޽h ? 33 `dC(  dR d s *3@  d 61Xd, Welch et al. 2002, PNASjGGGGG  d c bA>comparison cft073_k12_o157 figR H d 0޽h ? 33___PPT10i.-3+D=' = @B +D  l\(  lx l c $: `}  :  l c $: `<$ 0 : H l 0޽h ? 33ff33 ___PPT10.+} <D' = @B DS' = @BA?%,( < +O%,( < +DA' =%(D' =%(D' =A@BBBB0B%(D' =1:Bvisible*o3>+B#style.visibility<*l%(DA' =%(D' =%(D' =A@BBBB0B%(D' =1:Bvisible*o3>+B#style.visibility<*l3%(+8+0+l, +  p6(  p~ p s *l `}  l x p c $l l H p 0޽h ? ̙336  x'(  xx x c $$l\| l  x  0p-: <$ 0 :  z   x ,$D 0B x ZgֳgֳD)?  B x  `gֳgֳD)?  B x ZgֳgֳD)?  B x ZgֳgֳD)?L L B  x  `gֳgֳD)? B  x ZgֳgֳD)? @  @ B  x  `gֳgֳD)?  B  x  `gֳgֳD)? ((B  x  `gֳgֳD)?  B x ZgֳgֳD)?( +  x Tgֳgֳ ? J  ^Pathogen& 0(*2 C   x TDgֳgֳ ? b  b Non-Pathogen& 0(*2 C   x Tgֳgֳ ?  gPathogen Specific&0(*2C  B x ZgֳgֳD)?  B x  `gֳgֳDf)?  B x  `gֳgֳDf)?   x Zgֳgֳ ? ,$D 0B x ZgֳgֳDԔ?P wP ,$D 0: x T lgֳgֳ ?H ,$ 0 "Library of pathogen specific genes(#0(*2#C" B x s *޽h ? ̙33ME___PPT10%.+UDU' = @B D' = @BA?%,( < +O%,( < +DA' =%(D' =%(D' =A@BBBB0B%(D' =1:Bvisible*o3>+B#style.visibility<*xR%(D4' =%(D' =%(D' =4@BBBB%(D' =1:Bvisible*o3>+B#style.visibility<*x%(Dt' =%(D' =%(D' =4@BBBB%())))?D' =1:Bvisible*o3>+B#style.visibility<*x%(Dt' =%(D' =%(D' =4@BBBB%())))?D' =1:Bvisible*o3>+B#style.visibility<*x%(D' =%(D)' =%(D' =A@BBBB0B%())))?D' =1:Bvisible*o3>+B#style.visibility<*x%(DA' =%(D' =%(D' =A@BBBB0B%(D' =1:Bvisible*o3>+B#style.visibility<*xR%(++0+x ++0+x} ++0+x +   \(  x  c $|8lU  l   c $\9l <$ 0 l H  0޽h ? ̙33  ___PPT10 .+} <D* ' = @B D ' = @BA?%,( < +O%,( < +DA' =%(D' =%(D' =A@BBBB0B%(D' =1:Bvisible*o3>+B#style.visibility<*%(DA' =%(D' =%(D' =A@BBBB0B%(D' =1:Bvisible*o3>+B#style.visibility<*%(DA' =%(D' =%(D' =A@BBBB0B%(D' =1:Bvisible*o3>+B#style.visibility<*(%(DA' =%(D' =%(D' =A@BBBB0B%(D' =1:Bvisible*o3>+B#style.visibility<*(Q%(+8+0+, +  0(  x  c $Hl``  l x  c $pIl l Z  Bd޽h @ ? ̙33p  p D(  D8  wY D:HSN  wY D  wY` D 0A? wY D <@ Z`  OO78   D <xD  c  NO2  <N M   D M   D <HbG M  U*G   D <dLF   U*G   D <OF   U*G   D <NbM U*G  D <V U*G  D <PZ U*G H D 0޽h ? 33___PPT10i._;x+D=' = @B +   \(  x  c $DVl``  l   c $$Wl'<$ 0 l H  0޽h ? ̙33ia___PPT10A.+[W_D' = @B D' = @BA?%,( < +O%,( < +DA' =%(D' =%(D' =A@BBBB0B%(D' =1:Bvisible*o3>+B#style.visibility<*!%(DA' =%(D' =%(D' =A@BBBB0B%(D' =1:Bvisible*o3>+B#style.visibility<*!%(DA' =%(D' =%(D' =A@BBBB0B%(D' =1:Bvisible*o3>+B#style.visibility<* %(+8+0+ +$  $(  r  S bl `}  l r  S Pcl ` l H  0޽h ? 3380___PPT10.^mg$   $(  r  S `F `}   r  S G `  H  0޽h ? 3380___PPT10.^0\.0 0(  H  0޽h ? 3380___PPT10.^5[$  $$(  $r $ S }l `}  l r $ S }l ` l H $ 0޽h ? 3380___PPT10.\0K  0(  x  c $lp  l x  c $dlD l Z  Bd޽h @ ? ffD3fy___PPT10Y+D=' = @B +  E=(  =  0ti  P(,$ 0 There is a positive correlation between virulence, invasiveness and clonal origin Clonal division in E. coli O78 strains is host independent - closely related clones reside in different hostsp8 ZeG G G G 4G @D    h X  0A?z H  0޽h ? 33ff33 ___PPT10+JD' = @B DS' = @BA?%,( < +O%,( < +DA' =%(D' =%(D' =A@BBBB0B%(D' =1:Bvisible*o3>+B#style.visibility<*R%(DA' =%(D' =%(D' =A@BBBB0B%(D' =1:Bvisible*o3>+B#style.visibility<*R%(+8+0+5 +   F> (    c $ l<$ 0 l B  s *޽h ? yq___PPT10Q..M@:+JD' = @B D' = @BA?%,( < +O%,( < +DA' =%(D' =%(D' =A@BBBB0B%(D' =1:Bvisible*o3>+B#style.visibility<*`%(DA' =%(D' =%(D' =A@BBBB0B%(D' =1:Bvisible*o3>+B#style.visibility<*`%(DA' =%(D' =%(D' =A@BBBB0B%(D' =1:Bvisible*o3>+B#style.visibility<*\%(+8+0+5 +F me(  R  s *z<  3 A?0 -  6 /p W80 sequences specific to the pathogenic strain and absent from the driver strain K-12. 8X0(2Vgf3cW B  s *޽h ? f333f33fMMMy___PPT10Y+D=' = @B +O vn (  p  6T^VN___PPT90(@@  b00*g g C  <  3 A?     0 P X117 sequences specific to the pathogenic strain and absent from the driver strain K-12. (Y0(2Ygf3X B  s *޽h ? ̙33y___PPT10Y+D=' = @B +  0(  x  c $،l, l B  s *޽h ? f333f33fMMMy___PPT10Y+D=' = @B +J? a>Y>mm0=(  08F 4 0 ,T 0 c $X99?g ( 0 6ܜH   Anabaena variabilis DdlA>  f3 f3Ff3@       HB 0B C 1# ~ ( 0 6ȣL Anabaena variabilis CphA>  f3 f3Ff3@       HB 0B C 1M 8 0 6M] Gloeobacter violaceus CphA>  f3 f3Ff3N        HB  0B C 1M sHB  0B C 1  M  0 6p` Acinetobacter sp. MurC>  f3 f3Ff34     HB  0B C 1{ HB  0B C 1  5 0 6l" Ralstonia eutropha MurC>  f3 f3Ff3N       HB 0B C 1G; 0 6.~ Bordetella parapertussis MurC>  f3 f3Ff3N        HB 0 C 1N]HB 0 C 1 HB 0B C 1 j  0 6+` Mannheimia succiniciproducens.  fFf4     HB 0B C 1m p  0 6|\ Pasteurella multocida.  fFf4      HB 0 C 1 2 mHB 0 C 1 m r 0 6   Haemophilus somnus.  fFf4     HB 0 C 1  rHB 0 C 1 r  0 60J  Actinobacillus pleuropneumonia.  fFf4    HB 0 C 1  HB 0 C 1 Z E 0 6@<  Enterococcus faecium.  fFf4     HB  0 C 1Y  \  !0 6PL  Enterococcus faecalis.  fFf4     HB "0 C 1  \HB #0 C 1 \Z EHB $0 C 1Z E u %0 6a! N Streptococcus agalactiae.  fFf&    HB &0 C 1 p G '0 6Q i  Streptococcus mutans.  fFf&   HB (0 C 1 r  )0 6   Streptococcus suis.  fFf&   HB *0 C 1  HB +0 C 1  GHB ,0 C 1 G uHB -0 C 1 u  .0 6@ lx  Leptospira interrogans.  fFfB       HB /0 C 1 c  00 6QC  Clostridium perfringens.  fFf&     HB 10 C 1O 'M 20 6`P  Desulfotalea psychrophila.  fFfB       HB 30 C 1O  HB 40 C 1 O  50 6 s" } Listeria innocua.  fFf  HB 60 C 1 70 6,g  Listeria monocytogenes.  fFf  HB 80 C 1HB 90 C 1.  :0 6  Clostridium acetobutylicum.  fFf&    HB ;0 C 1. HB <0 C 1 . HB =0 C 1  HB >0 C 1  HB ?0 C 1j  HB @0 C 1j  #HB A0 C 1 # 9 B0 6}    Bordetella pertussis.  FB       HB C0 C 1   D0 6#wQ    Neisseria meningitidis.  FB       HB E0 C 1 sn HB F0 C 1   G0 6) a   Nitrosomonas europaea.  FB      HB H0 C 1  HB I0 C 1\  J0 6`/T 8   Acinetobacter sp..  F(    HB K0 C 1} ! ]  L0 64\ 5   Pseudomonas syringae.  F&    HB M0 C 1   N0 69- "  o Escherichia coli.  F  HB O0 C 1 ,  P0 6=$  cv  Vibrio cholerae.  F4    HB Q0B C 1= , HB R0 C 1 , HB S0 C 1} ! HB T0 C 1o } !  U0 6C   Buchnera aphidicola.  F4     HB V0B C 1* o HB W0 C 1\ o HB X0 C 1 9\ HB Y0 C 1 9 F  Z0 6@J s  Escherichia coli DdlA>  f3 f3Ff3&   HB [0B C 1    \0 6PE   Bacillus subtilis DdlA>  f3 f3Ff3&   HB ]0B C 1( L  ^0 6W   Listeria monocytogenes DdlA>  f3 f3Ff34    HB _0B C 1( HB `0B C 1  ( HB a0B C 1 F T dW b0# 0 4BB c0 3 1tVuBB d0 3 1dBB e0 3 1VdW f0 6_| e0.12 g B  R g0 s *f3  +R h0 s *f 1R i0 s *.1 j0 0eV eGshBs"( 2F    k0 0tiV eGshFs"( 2F f   l0 0m { V [ ATP-grasp" ( 2 F f3  n m0 0qw  Distribution of fused GshFs based of the C-terminal domain sequences and mapped onto a tree of bacterial GSHB and ATP-Grasp proteins (ME analysis)2( 2DffF ff&  x H 0 0޽h ? 33___PPT10i.OfV+D=' = @B +1  0 HA(  H^ H S <   l H c $8l< 0  l 7In the absence of the full genome of the two serotypes a genomic comparison of the two is not possible. Therefore, subtractive hybridization of each against K12 led to the discovery that there are some 150 distinct sequences specific for each serotype. Only some three are the same illustrating that both serotypes use different subsets of virulence factors to cause very similar disease profiles. What leads to pathological presentation, and host specificity -- O78 is being sequenced. H H 0޽h ? 3380___PPT10.=PiT 0  @T(  Td T c $<   l  T s *l< 0  l RUTI  one of the most common infections. * ** H T 0޽h ? 3380___PPT10.'T), 0 D<Ph(  hd h c $<   l0 h s *I< 0  l n2002  full genome of CFT073 2001  full genome of O1578 87 H h 0޽h ? 3380___PPT10.sP 0 `t8(  td t c $<   l t s * l< @  l " H t 0޽h ? ̙33 0 p|8(  |d | c $<   l | s *8l< @  l " H | 0޽h ? ̙33r\lprt6t)?*wXyeN6g O&0<583;D OYRV6ޞ3Vn)\(   N:kashi@techunix.technion.ac.il>*racheli@ibexperts.com0ugophna@dal.ca0ugophna@dal.ca/ 00DArialNew Rot$    %On-screen ShowTAU" 'ArialComic Sans MSMS Sans SerifTimes New RomanDefault DesignAPathogenomics in Israel Eliora Z. Ron $Facilities for pathogenomic studiesMetagenomics Metagenomics 9LGT lateral gene transfer in relation to pathogenesis 9LGT lateral gene transfer in relation to pathogenesis PowerPoint Presentation2 Typing of bacterial strains and drug resistance .Identification of bacterial virulence factors+Identification of fungal virulence factorsWhole genome analysisVirulent E. coli strainsPowerPoint PresentationSepticemia (colibacillosis)Goals:PowerPoint PresentationDIdentification of virulence related sequences in septicemic strainsSubtractive hybridizationSubtractive hybridization)Search for unique septicemic sequences-Comparison of unique sequences of O2 and O78PowerPoint PresentationFComparison of unique sequences of two septicemic strains - O2 and O78Current-future research Thank you!!PowerPoint PresentationMetagenomics MLST of O78 strainsPowerPoint PresentationPowerPoint PresentationPowerPoint PresentationPowerPoint PresentationPowerPoint PresentationPowerPoint Presentation  Fonts UsedDesign Template Slide Titles" 8@ _PID_HLINKSA\<%mailto:kashi@techunix.technion.ac.ilmailto:racheli@ibexperts.commailto:ugophna@dal.ca"_҇ Eliora RonEliora Ron2 mailto:ugophna@dal.camailto:yaira@tauex.tau.ac.ilmailto:simay@tx.technion.ac.ilmailto:eliora@post.tau:kashi@techunix.technion.ac.il>*racheli@ibexperts.com0ugophna@dal.ca0ugophna@dal.ca/ 00DArialNew Rot$t\ )0tY 0"DComic Sans MS$t\ )0tY 0B DMS Sans Serif$t\ )0tY 00DTimes New Romant\ )0tY 0c .  @n?" dd@  @@`` 5( ~ n ($%&'  #! " _b$ ov飼kf#A2$%Gg_gDj#A2$nI8i v o&A2$0h,m5A @,7Ab$L-s$֠n:A 0AA ff33@@K2ʚ;n8ʚ;g4BdBd )0hppp@ <42d2d 0$t<4BdBd 0$t( <4!d!dy 0$0___PPT10 ppR___PPT94,&@ &@L@@4@@?  %RPathogenomics in Israel$ff"    Metagenomics LGT = lateral gene transfer Typing of bacterial strains and drug resistance Identification of virulence factors Whole genome analysis 33"   )%#Facilities for pathogenomic studies"$ $G$ff.    Good facilities for genomics, transcriptomics and proteomics in several universities and research institutes High level bioinformatics All facilities are also available as service E33.    Metagenomics ( $ff("   zMetagenomics of culturable and non-culturable microorganism populations present in biofilms of Acute Otitis Media (AOM, Middle Ear Infection) This infection involves a variety of bacterial species, found in the form of biofilms, which are inherently resistant to antibiotic treatment Detection of microbial biodiversity of AOM biofilm is limited due to current cultivation methods The group of Fauzi Silbaq (ArabQual) and Racheli Kreisberg-Zakarin (IBEX) racheli@ibexperts.com in involved in a study the biodiversity of AOM using a metagenomics approach, including microbiology, functional genomics and bioinformatics methodologies v P P33l33 33v3333         &     q  e  <               8   X  0Metagenomics $ff MMetagenomics of intestinal microflora in health and disease and the effect of the TLR mutations Toll-Like Receptors (TLRs) recognize pathogen specific patterns of microorganisms. Mutations in TLRs are associated with inflammatory bowel diseases (Crohn's disease and ulcerative colitis). Patients with those diseases have altered intestinal microflora Uri Gophna, currently with Ford Doolitle in Halifax ugophna@dal.ca performs culture-independent profiling of intestinal microflora of TLR knockout mice under normal conditions and after a challenge which models inflammatory bowel diseaseza Z Z ZaG33G33 G33G33     Q  G  2  W          6   l  0pLGT  lateral gene transfer in relation to pathogenesis (97$ff$8 Uri Gophna ugophna@dal.ca studies the role of lateral gene transfer (LGT) in the evolution of pathogens Uses bioinformatics for the identification of laterally acquired genes and pathways in pathogens \ DE33E33~E33E33N      0  pLGT  lateral gene transfer in relation to pathogenesis (97G$ff(8 Yair Aharonowitz, Ilya Borovok and Gerald Cohen from Tel Aviv University study gluthathione synthesis <yaira@tauex.tau.ac.il> Identified GshF orthologs, consisting of a -glutamylcysteine ligase (GshA) domain fused to an ATP-grasp domain, in 20 gram-positive and gram-negative bacteria. Remarkably, 95% of these bacteria are mammalian pathogens. Presumably, this fusion gene, once formed, spread between mammalian hosts most likely by horizontal gene transfer R~ ZP8 Z2 ZG33C        1                          q    1 Typing of bacterial strains and drug resistance 62(/$ff(1 -The group of Chezi Kashi kashi@techunix.technion.ac.il uses novel methods for molecular typing ofVibrio cholera in order to study emerging new pathogenic strains The group of Sima Yaron (Technion) is involved in typing the serovars of Salmonella enterica in respect to drug resistance and virulence l. aG33G33~G33G33.G33       )  J          -  06  #Identification of virulence factors$$G$ff# The group of Sima Yaron has developed a rapid, simple screen for real-time quantification of promoter - activity in S. enterica using a library of plasmids with GFP as a reporter The group of Nir   !"#$%&'()*+,-./0123456789:;<=>?@ABCDEFGHIJKLMNOPQRSTUVWXYZ[\]^_`abcdefghijklmnopqrstuvwxyz{|}~      !"#$%&'()*+,-./0123456789:;<=>?@ABCDEFGHIJKLMNOPQRSTUVWXYZ[\]^_`abcdefghijklmnopqrstuvwxyz{|}~      !"#$%&'()*+,-./0123456789:;<=>?@ABCDEFGHIJKLMNOPQRSTUVWXYZ[\]^_`abcdefghijklmnopqrstuvwxyz{|}~Root EntrydO)p2UoPicturesCurrent User>SummaryInformation(PowerPoint Document(DocumentSummaryInformation87p t\ )0tY 0"DComic Sans MS$t\ )0tY 0B DMS Sans Serif$t\ )0tY 00DTimes New Romant\ )0tY 0c .  @n?" dd@  @@`` 5( ~ n ($%&'  #! " _b$ ov飼kf#A2$%Gg_gDj#A2$nI8i v o&A2$0h,m5A @,7Ab$L-s$֠n:A 0AA ff33@@K2ʚ;n8ʚ;g4BdBd )0hppp@ <42d2d 0$t<4BdBd 0$t( <4!d!dy 0$0___PPT10 ppR___PPT94,&@ &@L@@4@@?  %.RPathogenomics in Israel$ff"    Metagenomics LGT = lateral gene transfer Typing of bacterial strains and drug resistance Identification of virulence factors Whole genome analysis 33"   )%#Facilities for pathogenomic studies"$ $G$ff.    Good facilities for genomics, transcriptomics and proteomics in several universities and research institutes High level bioinformatics All facilities are also available as service E33.    Metagenomics ( $ff("   zMetagenomics of culturable and non-culturable microorganism populations present in biofilms of Acute Otitis Media (AOM, Middle Ear Infection) This infection involves a variety of bacterial species, found in the form of biofilms, which are inherently resistant to antibiotic treatment Detection of microbial biodiversity of AOM biofilm is limited due to current cultivation methods The group of Fauzi Silbaq (ArabQual) and Racheli Kreisberg-Zakarin (IBEX) racheli@ibexperts.com in involved in a study the biodiversity of AOM using a metagenomics approach, including microbiology, functional genomics and bioinformatics methodologies v P P33l33 33v3333         &     q  e  <               8   X  0Metagenomics $ff MMetagenomics of intestinal microflora in health and disease and the effect of the TLR mutations Toll-Like Receptors (TLRs) recognize pathogen specific patterns of microorganisms. Mutations in TLRs are associated with inflammatory bowel diseases (Crohn's disease and ulcerative colitis). Patients with those diseases have altered intestinal microflora Uri Gophna, currently with Ford Doolitle in Halifax ugophna@dal.ca performs culture-independent profiling of intestinal microflora of TLR knockout mice under normal conditions and after a challenge which models inflammatory bowel diseaseza Z Z ZaG33G33 G33G33     Q  G  2  W          6   l  0pLGT  lateral gene transfer in relation to pathogenesis (97$ff$8 Uri Gophna ugophna@dal.ca studies the role of lateral gene transfer (LGT) in the evolution of pathogens Uses bioinformatics for the identification of laterally acquired genes and pathways in pathogens \ DE33E33~E33E33N      0  pLGT  lateral gene transfer in relation to pathogenesis (97G$ff(8 lYair Aharonowitz, Ilya Borovok and Gerald Cohen from Tel Aviv University study gluthathione synthesis Identified GshF orthologs, consisting of a -glutamylcysteine ligase (GshA) domain fused to an ATP-grasp domain, in 20 gram-positive and gram-negative bacteria. Remarkably, 95% of these bacteria are mammalian pathogens. Presumably, this fusion gene, once formed, spread between mammalian hosts most likely by horizontal gene transfer Rf ZP8 Z2 ZG33C(        1                  q    1 Typing of bacterial strains and drug resistance 62(/$ff(1 -The group of Chezi Kashi kashi@techunix.technion.ac.il uses novel methods for molecular typing ofVibrio cholera in order to study emerging new pathogenic strains The group of Sima Yaron (Technion) is involved in typing the serovars of Salmonella enterica in respect to drug resistance and virulence l. aG33G33~G33G33.G33       )  J          -  06  #Identification of virulence factors$$G$ff# The group of Sima Yaron has developed a rapid, simple screen for real-time quantification of promoter - activity in S. enterica using a library of plasmids with GFP as a reporter The group of Nir Asherov (Tel Aviv U.) studies Aspergillus fumigatus, which causes serious disease (around 60% mortality) in immunocompromised people. They concentrate on studying novel cell wall proteins and identified 68 such cwp s. They are studying these CWPs in vitro and in infected animals by the use of deletion mutants. They are currently in the process of preparing a Cwp-specific microarrays to study Cwp gene expression patternsl ZtG33 G33G33G33RG33G33uG33z     `   0          9  V    s           Whole genome analysis$ff Two genomes of bacterial pathogens are being sequenced and analyzed in Israel Bacillus anthracis (Shaefferman et al, Biological Institute, Nes Ziona) Septicemic Escherichia coli  strain of serotype O78 (Ron, Tel Aviv U.) N  NE33G 33AG 33G 33G 33G 33 G  33G 33G 33W             *  Virulent E. coli strainsF G$ffG$ff G$ff Most of the E. coli strains are commensal, but a small number are pathogenic Pathogenic E. coli strains are divided into two groups: Intestinal strains. These produce enterotoxins and constitute a major problem, especially in young children and travellers (Montesumu s revenge) Extraintestinal strains  ExPEC (Extraintestinal Pathogenic E. coli)  Z Z Z G33G33FG33G33&G33E33E33E33G33G33       B               Septicemia (colibacillosis)G$ff.    Colisepticemia is the major causes of mortality from community and hospital-acquired infections (more than 80%) Main cause of mortality in immuno-supressed patients (HIV, chemotherapy, old age) Colisepticemia is an emerging disease  83% increase 1980  1992, over 40% of the bacteremia cases in community acquired infectionsG E33e33e33e33e33e33 ~     '  D   ' Goals:6G$ffg ff Define virulence-essential ExPEC-specific genes Profile strains involved in UTI, NBM and sepsis using these ExPEC-specific genes Use the data to define potential targets for development of vaccines and/or antibacterial drugs.| /E33E33QE33_e33 e 33 E 33v    <  o  CIdentification of virulence related sequences in septicemic strainsDDG$ff.1    3Whole genome sequencing Subtractive hybridization >3  3E33E33 4 Subtractive hybridization  G$ff  Obtain pathogen specific sequences, absent from non-pathogenic K12 strain Excellent chance of  hitting pathogenicity islands which are pathogen specific and very large Faster (and much cheaper) than whole genome sequencing E33  Subtractive hybridization2 G$f3G$f3  QA way to study comparative genomics with organisms which have not been sequenced .R PE33E33 R PSearch for unique  septicemic sequences))G f32      PUsing suppression subtractive hybridization (SSH) we identified sequences unique to strain O78-9 and absent from the non-pathogenic strain K-12 Oover 80 O78-specific open reading frames were found (91 to 1473 bp in length) The same experiment was repeated with another septicemic strain O2-1772 117 unique O2 sequences were identified Q g6GgIg'ggZ  <  ;   9 ,Comparison of unique sequences of O2 and O78--G$f3,  ^Although the two strains cause the same disease, there is a high diversity between the SSH libraries of O2 and O78 strains, with only a few shared genes coding for virulence factors. Is this diversity serotype specific? To determine this we profiled additional septicemic strains of the same serotypes the presence of each of the unique sequences l_ Z|e9eeEe:   L   EComparison of unique sequences of two septicemic strains - O2 and O78FFG f32&     high level of genome plasticity there is a high diversity between the septicemic strainsunexpected for strains causing the same disease Septicemic strains of serogroups O2 and O78 contain a large pool of virulence genes which are used in a  mix and match fashionD  Ye0e33e##tG     &       ` #Current-future researchG$ff $Molecular-physiological studies of the newly-identified virulence genes Bioinformatic studies to determine the evolution of these virulence genes (many in PAIs with evidence of LGT) Whole genome sequencing  joint project with Prof. Joerg Hacker and Prof. Gerhard Gottschalk: E3333E33bH   F  J  $ ($ Thank you!!"  G$ff  TAU group Uri Gophna Diana Ideses Daphna Mokady Dr. Dvora Biran Collaborations Wuerzburg University (Prof. Joerg Hacker) Greifswald University (Prof. Michael Hecker) Goetingen University (Prof. Gerhard Gottschalk)   Z7 Z Z Z G337G33G33sG3333*                          ( "Metagenomics ( Uri Gophna, currently with Ford Doolitle in Halifax studies the effect of TLR mutations on micrflora in health and disease by culture-independent profiling of intestinal microflora of TLR knockout mice under normal conditions and after a challenge which models inflammatory bowel diseaseH  0@|    3   F   k MLST of O78 strains" G$f3  Multi Locus Sequence Typing 450  500 bp of 7  housekeeping genes Criteria for chosing genes: 97-98% homology to E. coli K-12 (from blast data) appear in pathogenic and non pathogenic strains map at considerable distance from each other several allels in the populationf_ Z Z_EEEEZ&  (       ! [The MLST results are compatible with the results from subtractive hybridization and sequencing The profile of virulence factors in ExPEC strains is independent of the host and independent of the serotype There is a high diversity of virulence genes in the various E. coli septicemic strains and each strain has its own profile of virulence genesP\ AE##=AAKA@     @ !"#  Both libraries contain many sequences associated with genomic plasticity - evolution by horizontal gene transfer Many sequences of O2 and O78 are homologous to virulence related sequences of human ExPEC strains ns  c  tecEA&      /$%&'0   0(   x  c $` `   x  c $i  H  0޽h ? 3380___PPT10.\ۇ0  0(  x  c $:'g  : x  c $:  : H  0޽h ? 3380___PPT10.\Yr^':_)!](   N89:;<=>?@ABCDEFGHIJKLMNOPQRSTUVWXZ[\]^_`abcde      !"#$%&'()*+,-./0123456789:;<=>?@ABCDEFGHIJKLMNOPQRSTUVWXYZ[\]^_`abcdefghijklmnopqrstuvwxyz{|}~"_ Eliora RonEliora Ronh  Pathogenomics in Israel eliora ronc Eliora Ronc67oMicrosoft PowerPointel@/@\@U2UoGhg  '?& &&#TNPP\2OMi & TNPP &&TNPP    --- !---&&j}w@ }ww0- &Gy& --0-- @BComic Sans MS }ww0- ff.2 j Pathogenomics% . ff.2 j- in Israel .--0-- ff@"Arialw@ }ww0- 33. 2 :.@BComic Sans MS }ww0- 33.2 ^ Metagenomics&! .@"Arialw@ }ww0- 33. 2 :.@BComic Sans MS }ww0- 33.02 ^LGT = lateral gene transfer    .@"Arialw@ }ww0- 33. 2 P:.@BComic Sans MS }ww0- 33.?2 P^%Typing of bacterial strains and drug          . 33.2 ^ resistance .@"Arialw@ }ww0- 33. 2 :.@BComic Sans MS }ww0- 33.=2 ^$Identification of virulence factors          .@"Arialw@ }ww0- 33. 2 :.@BComic Sans MS }ww0- 33.'2 ^Whole genome analysis-  !   .--"System 0-&TNPP &\ ՜.+,D՜.+,Asherov (Tel Aviv U.) studies Aspergillus fumigatus, which causes serious disease (around 60% mortality) in immunocompromised people. They concentrate on studying novel cell wall proteins and identified 68 such cwp s. They are studying these CWPs in vitro and in infected animals by the use of deletion mutants. They are currently in the process of preparing a Cwp-specific microarrays to study Cwp gene expression patternsl ZtG33 G33G33G33RG33G33uG33z     `   0          9  V    s           Whole genome analysis$ff Two genomes of bacterial pathogens are being sequenced and analyzed in Israel Bacillus anthracis (Shaefferman et al, Biological Institute, Nes Ziona) Septicemic Escherichia coli  strain of serotype O78 (Ron, Tel Aviv U.) N  NE33G 33AG 33G 33G 33G 33 G  33G 33G 33W             *  Virulent E. coli strainsF G$ffG$ff G$ff Most of the E. coli strains are commensal, but a small number are pathogenic Pathogenic E. coli strains are divided into two groups: Intestinal strains. These produce enterotoxins and constitute a major problem, especially in young children and travellers (Montesumu s revenge) Extraintestinal strains  ExPEC (Extraintestinal Pathogenic E. coli)  Z Z Z G33G33FG33G33&G33E33E33E33G33G33       B               Septicemia (colibacillosis)G$ff.    Colisepticemia is the major causes of mortality from community and hospital-acquired infections (more than 80%) Main cause of mortality in immuno-supressed patients (HIV, chemotherapy, old age) Colisepticemia is an emerging disease  83% increase 1980  1992, over 40% of the bacteremia cases in community acquired infectionsG E33e33e33e33e33e33 ~     '  D   ' Goals:6G$ffg ff Define virulence-essential ExPEC-specific genes Profile strains involved in UTI, NBM and sepsis using these ExPEC-specific genes Use the data to define potential targets for development of vaccines and/or antibacterial drugs.| /E33E33QE33_e33 e 33 E 33v    <  o  CIdentification of virulence related sequences in septicemic strainsDDG$ff.1    3Whole genome sequencing Subtractive hybridization >3  3E33E33 4 Subtractive hybridization  G$ff  Obtain pathogen specific sequences, absent from non-pathogenic K12 strain Excellent chance of  hitting pathogenicity islands which are pathogen specific and very large Faster (and much cheaper) than whole genome sequencing E33  Subtractive hybridization2 G$f3G$f3  QA way to study comparative genomics with organisms which have not been sequenced .R PE33E33 R PSearch for unique  septicemic sequences))G f32      PUsing suppression subtractive hybridization (SSH) we identified sequences unique to strain O78-9 and absent from the non-pathogenic strain K-12 Oover 80 O78-specific open reading frames were found (91 to 1473 bp in length) The same experiment was repeated with another septicemic strain O2-1772 117 unique O2 sequences were identified Q g6GgIg'ggZ  <  ;   9 ,Comparison of unique sequences of O2 and O78--G$f3,  ^Although the two strains cause the same disease, there is a high diversity between the SSH libraries of O2 and O78 strains, with only a few shared genes coding for virulence factors. Is this diversity serotype specific? To determine this we profiled additional septicemic strains of the same serotypes the presence of each of the unique sequences l_ Z|e9eeEe:   L   EComparison of unique sequences of two septicemic strains - O2 and O78FFG f32&     high level of genome plasticity there is a high diversity between the septicemic strainsunexpected for strains causing the same disease Septicemic strains of serogroups O2 and O78 contain a large pool of virulence genes which are used in a  mix and match fashionD  Ye0e33e##tG     &       ` #Current-future researchG$ff $Molecular-physiological studies of the newly-identified virulence genes Bioinformatic studies to determine the evolution of these virulence genes (many in PAIs with evidence of LGT) Whole genome sequencing  joint project with Prof. Joerg Hacker and Prof. Gerhard Gottschalk: E3333E33bH   F  J  $ ($ Thank you!!"  G$ff  TAU group Uri Gophna Diana Ideses Daphna Mokady Dr. Dvora Biran Collaborations Wuerzburg University (Prof. Joerg Hacker) Greifswald University (Prof. Michael Hecker) Goetingen University (Prof. Gerhard Gottschalk)   Z7 Z Z Z G337G33G33sG3333*                          ( "Metagenomics ( Uri Gophna, currently with Ford Doolitle in Halifax studies the effect of TLR mutations on micrflora in health and disease by culture-independent profiling of intestinal microflora of TLR knockout mice under normal conditions and after a challenge which models inflammatory bowel diseaseH  0@|    3   F   k MLST of O78 strains" G$f3  Multi Locus Sequence Typing 450  500 bp of 7  housekeeping genes Criteria for chosing genes: 97-98% homology to E. coli K-12 (from blast data) appear in pathogenic and non pathogenic strains map at considerable distance from each other several allels in the populationf_ Z Z_EEEEZ&  (       ! [The MLST results are compatible with the results from subtractive hybridization and sequencing The profile of virulence factors in ExPEC strains is independent of the host and independent of the serotype There is a high diversity of virulence genes in the various E. coli septicemic strains and each strain has its own profile of virulence genesP\ AE##=AAKA@     @ !"#  Both libraries contain many sequences associated with genomic plasticity - evolution by horizontal gene transfer Many sequences of O2 and O78 are homologous to virulence related sequences of human ExPEC strains ns  c  tecEA&      /$%&'   ((  (r ( S T :q< :  ( s :0e0e #" 0e `}  : H ( 0޽h ? 3380___PPT10.\`WQr dxg)C](  ^ N:kashi@techunix.technion.ac.il>*racheli@ibexperts.com0ugophna@dal.ca0ugophna@dal.ca>*yaira@tauex.tau.ac.il/ 00DArialNew Rot$t\ )0tY 0"DComic Sans MS$t\ )0tY 0B DMS Sans Serif$t\ )0tY 00DTimes New Romant\ )0tY 0c .  @n?" dd@  @@`` 5( ~ n ($%&'  #! " _b$ ov飼kf#A2$%Gg_gDj#A2$nI8i v o&A2$0h,m5A @,7Ab$L-s$֠n:A 0AA ff33@@K2ʚ;n8ʚ;g4BdBd )0hppp@ <42d2d 0$t<4BdBd 0$t( <4!d!dy 0$0___PPT10 ppR___PPT94,&@ &@L@@4@@?  %RPathogenomics in Israel$ff"    Metagenomics LGT = lateral gene transfer Typing of bacterial strains and drug resistance Identification of virulence factors Whole genome analysis 33"   )%#Facilities for pathogenomic studies"$ $G$ff.    Good facilities for genomics, transcriptomics and proteomics in several universities and research institutes High level bioinformatics All facilities are also available as service E33.    Metagenomics ( $ff("   zMetagenomics of culturable and non-culturable microorganism populations present in biofilms of Acute Otitis Media (AOM, Middle Ear Infection) This infection involves a variety of bacterial species, found in the form of biofilms, which are inherently resistant to antibiotic treatment Detection of microbial biodiversity of AOM biofilm is limited due to current cultivation methods The group of Fauzi Silbaq (ArabQual) and Racheli Kreisberg-Zakarin (IBEX) racheli@ibexperts.com in involved in a study the biodiversity of AOM using a metagenomics approach, including microbiology, functional genomics and bioinformatics methodologies v P P33l33 33v3333         &     q  e  <               8   X  0Metagenomics $ff MMetagenomics of intestinal microflora in health and disease and the effect of the TLR mutations Toll-Like Receptors (TLRs) recognize pathogen specific patterns of microorganisms. Mutations in TLRs are associated with inflammatory bowel diseases (Crohn's disease and ulcerative colitis). Patients with those diseases have altered intestinal microflora Uri Gophna, currently with Ford Doolitle in Halifax ugophna@dal.ca performs culture-independent profiling of intestinal microflora of TLR knockout mice under normal conditions and after a challenge which models inflammatory bowel diseaseza Z Z ZaG33G33 G33G33     Q  G  2  W          6   l  0pLGT  lateral gene transfer in relation to pathogenesis (97$ff$8 Uri Gophna ugophna@dal.ca studies the role of lateral gene transfer (LGT) in the evolution of pathogens Uses bioinformatics for the identification of laterally acquired genes and pathways in pathogens \ DE33E33~E33E33N      0  pLGT  lateral gene transfer in relation to pathogenesis (97G$ff(8 Yair Aharonowitz, Ilya Borovok and Gerald Cohen from Tel Aviv University study gluthathione synthesis yaira@tauex.tau.ac.il Identified GshF orthologs, consisting of a -glutamylcysteine ligase (GshA) domain fused to an ATP-grasp domain, in 20 gram-positive and gram-negative bacteria. Remarkably, 95% of these bacteria are mammalian pathogens. Presumably, this fusion gene, once formed, spread between mammalian hosts most likely by horizontal gene transfer R Z8 Z2 ZG33C@        1                    q   0f{  1 Typing of bacterial strains and drug resistance 62(/$ff(1 -The group of Chezi Kashi kashi@techunix.technion.ac.il uses novel methods for molecular typing ofVibrio cholera in order to study emerging new pathogenic strains The group of Sima Yaron (Technion) is involved in typing the serovars of Salmonella enterica in respect to drug resistance and virulence l. aG33G33~G33G33.G33       )  J          -  06  #Identification of virulence factors$$G$ff# The group of Sima Yaron has developed a rapid, simple screen for real-time quantification of promoter - activity in S. enterica using a library of plasmids with GFP as a reporter The group of Nir Asherov (Tel Aviv U.) studies Aspergillus fumigatus, which causes serious disease (around 60% mortality) in immunocompromised people. They concentrate on studying novel cell wall proteins and identified 68 such cwp s. They are studying these CWPs in vitro and in infected animals by the use of deletion mutants. They are currently in the process of preparing a Cwp-specific microarrays to study Cwp gene expression patternsl ZtG33 G33G33G33RG33G33uG33z     `   0          9  V    s           Whole genome analysis$ff Two genomes of bacterial pathogens are being sequenced and analyzed in Israel Bacillus anthracis (Shaefferman et al, Biological Institute, Nes Ziona) Septicemic Escherichia coli  strain of serotype O78 (Ron, Tel Aviv U.) N  NE33G 33AG 33G 33G 33G 33 G  33G 33G 33W             *  Virulent E. coli strainsF G$ffG$ff G$ff Most of the E. coli strains are commensal, but a small number are pathogenic Pathogenic E. coli strains are divided into two groups: Intestinal strains. These produce enterotoxins and constitute a major problem, especially in young children and travellers (Montesumu s revenge) Extraintestinal strains  ExPEC (Extraintestinal Pathogenic E. coli)  Z Z Z G33G33FG33G33&G33E33E33E33G33G33       B               Septicemia (colibacillosis)G$ff.    Colisepticemia is the major causes of mortality from community and hospital-acquired infections (more than 80%) Main cause of mortality in immuno-supressed patients (HIV, chemotherapy, old age) Colisepticemia is an emerging disease  83% increase 1980  1992, over 40% of the bacteremia cases in community acquired infectionsG E33e33e33e33e33e33 ~     '  D   ' Goals:6G$ffg ff Define virulence-essential ExPEC-specific genes Profile strains involved in UTI, NBM and sepsis using these ExPEC-specific genes Use the data to define potential targets for development of vaccines and/or antibacterial drugs.| /E33E33QE33_e33 e 33 E 33v    <  o  CIdentification of virulence related sequences in septicemic strainsDDG$ff.1    3Whole genome sequencing Subtractive hybridization >3  3E33E33 4 Subtractive hybridization  G$ff  Obtain pathogen specific sequences, absent from non-pathogenic K12 strain Excellent chance of  hitting pathogenicity islands which are pathogen specific and very large Faster (and much cheaper) than whole genome sequencing E33  Subtractive hybridization2 G$f3G$f3  QA way to study comparative genomics with organisms which have not been sequenced .R PE33E33 R PSearch for unique  septicemic sequences))G f32      PUsing suppression subtractive hybridization (SSH) we identified sequences unique to strain O78-9 and absent from the non-pathogenic strain K-12 Oover 80 O78-specific open reading frames were found (91 to 1473 bp in length) The same experiment was repeated with another septicemic strain O2-1772 117 unique O2 sequences were identified Q g6GgIg'ggZ  <  ;   9 ,Comparison of unique sequences of O2 and O78--G$f3,  ^Although the two strains cause the same disease, there is a high diversity between the SSH libraries of O2 and O78 strains, with only a few shared genes coding for virulence factors. Is this diversity serotype specific? To determine this we profiled additional septicemic strains of the same serotypes the presence of each of the unique sequences l_ Z|e9eeEe:   L   EComparison of unique sequences of two septicemic strains - O2 and O78FFG f32&     high level of genome plasticity there is a high diversity between the septicemic strainsunexpected for strains causing the same disease Septicemic strains of serogroups O2 and O78 contain a large pool of virulence genes which are used in a  mix and match fashionD  Ye0e33e##tG     &       ` #Current-future researchG$ff $Molecular-physiological studies of the newly-identified virulence genes Bioinformatic studies to determine the evolution of these virulence genes (many in PAIs with evidence of LGT) Whole genome sequencing  joint project with Prof. Joerg Hacker and Prof. Gerhard Gottschalk: E3333E33bH   F  J  $ ($ Thank you!!"  G$ff  TAU group Uri Gophna Diana Ideses Daphna Mokady Dr. Dvora Biran Collaborations Wuerzburg University (Prof. Joerg Hacker) Greifswald University (Prof. Michael Hecker) Goetingen University (Prof. Gerhard Gottschalk)   Z7 Z Z Z G337G33G33sG3333*                          ( "Metagenomics ( Uri Gophna, currently with Ford Doolitle in Halifax studies the effect of TLR mutations on micrflora in health and disease by culture-independent profiling of intestinal microflora of TLR knockout mice under normal conditions and after a challenge which models inflammatory bowel diseaseH  0@|    3   F   k MLST of O78 strains" G$f3  Multi Locus Sequence Typing 450  500 bp of 7  housekeeping genes Criteria for chosing genes: 97-98% homology to E. coli K-12 (from blast data) appear in pathogenic and non pathogenic strains map at considerable distance from each other several allels in the populationf_ Z Z_EEEEZ&  (       ! [The MLST results are compatible with the results from subtractive hybridization and sequencing The profile of virulence factors in ExPEC strains is independent of the host and independent of the serotype There is a high diversity of virulence genes in the various E. coli septicemic strains and each strain has its own profile of virulence genesP\ AE##=AAKA@     @ !"#  Both libraries contain many sequences associated with genomic plasticity - evolution by horizontal gene transfer Many sequences of O2 and O78 are homologous to virulence related sequences of human ExPEC strains ns  c  tecEA&      /$%&'   ((  (r ( S T :q< :  ( s :0e0e #" 0e `}  : H ( 0޽h ? 3380___PPT10.\`WQrg g)](   N:kashi@techunix.technion.ac.il>*racheli@ibexperts.com0ugophna@dal.ca0ugophna@dal.ca>*yaira@tauex.tau.ac.ilB.simay@tx.technion.ac.il/ 00DArialNew Rot$t\ )0tY 0"DComic Sans MS$t\ )0tY 0B DMS Sans Serif$t\ )0tY 00DTimes New Romant\ )0tY 0c .  @n?" dd@  @@`` 5( ~ n ($%&'  #! " _b$ ov飼kf#A2$%Gg_gDj#A2$nI8i v o&A2$0h,m5A @,7Ab$L-s$֠n:A 0AA ff33@@K2ʚ;n8ʚ;g4BdBd )0hppp@ <42d2d 0$t<4BdBd 0$t( <4!d!dy 0$0___PPT10 ppR___PPT94,&@ &@L@@4@@?  %RPathogenomics in Israel$ff"    Metagenomics LGT = lateral gene transfer Typing of bacterial strains and drug resistance Identification of virulence factors Whole genome analysis 33"   )%#Facilities for pathogenomic studies"$ $G$ff.    Good facilities for genomics, transcriptomics and proteomics in several universities and research institutes High level bioinformatics All facilities are also available as service E33.    Metagenomics ( $ff("   zMetagenomics of culturable and non-culturable microorganism populations present in biofilms of Acute Otitis Media (AOM, Middle Ear Infection) This infection involves a variety of bacterial species, found in the form of biofilms, which are inherently resistant to antibiotic treatment Detection of microbial biodiversity of AOM biofilm is limited due to current cultivation methods The group of Fauzi Silbaq (ArabQual) and Racheli Kreisberg-Zakarin (IBEX) racheli@ibexperts.com in involved in a study the biodiversity of AOM using a metagenomics approach, including microbiology, functional genomics and bioinformatics methodologies v P P33l33 33v3333         &     q  e  <               8   X  0Metagenomics $ff MMetagenomics of intestinal microflora in health and disease and the effect of the TLR mutations Toll-Like Receptors (TLRs) recognize pathogen specific patterns of microorganisms. Mutations in TLRs are associated with inflammatory bowel diseases (Crohn's disease and ulcerative colitis). Patients with those diseases have altered intestinal microflora Uri Gophna, currently with Ford Doolitle in Halifax ugophna@dal.ca performs culture-independent profiling of intestinal microflora of TLR knockout mice under normal conditions and after a challenge which models inflammatory bowel diseaseza Z Z ZaG33G33 G33G33     Q  G  2  W          6   l  0pLGT  lateral gene transfer in relation to pathogenesis (97$ff$8 Uri Gophna ugophna@dal.ca studies the role of lateral gene transfer (LGT) in the evolution of pathogens Uses bioinformatics for the identification of laterally acquired genes and pathways in pathogens \ DE33E33~E33E33N      0  pLGT  lateral gene transfer in relation to pathogenesis (97G$ff(8 Yair Aharonowitz, Ilya Borovok and Gerald Cohen from Tel Aviv University study gluthathione synthesis yaira@tauex.tau.ac.il Identified GshF orthologs, consisting of a -glutamylcysteine ligase (GshA) domain fused to an ATP-grasp domain, in 20 gram-positive and gram-negative bacteria. Remarkably, 95% of these bacteria are mammalian pathogens. Presumably, this fusion gene, once formed, spread between mammalian hosts most likely by horizontal gene transfer R Z8 Z2 ZG33C@        1                    q   0f{  1 Typing of bacterial strains and drug resistance 62(/$ff(1 9The group of Chezi Kashi kashi@techunix.technion.ac.il uses novel methods for molecular typing ofVibrio cholera in order to study emerging new pathogenic strains The group of Sima Yaron is involved in typing the serovars of Salmonella enterica in respect to drug resistance and virulence simay@tx.technion.ac.ill: aG33G33sG33G33EG33        )  J          -     06 0"9  #Identification of virulence factors$$G$ff# The group of Sima Yaron has developed a rapid, simple screen for real-time quantification of promoter - activity in S. enterica using a library of plasmids with GFP as a reporter The group of Nir Asherov (Tel Aviv U.) studies Aspergillus fumigatus, which causes serious disease (around 60% mortality) in immunocompromised people. They concentrate on studying novel cell wall proteins and identified 68 such cwp s. They are studying these CWPs in vitro and in infected animals by the use of deletion mutants. They are currently in the process of preparing a Cwp-specific microarrays to study Cwp gene expression patternsl ZtG33 G33G33G33RG33G33uG33z     `   0          9  V    s           Whole genome analysis$ff Two genomes of bacterial pathogens are being sequenced and analyzed in Israel Bacillus anthracis (Shaefferman et al, Biological Institute, Nes Ziona) Septicemic Escherichia coli  strain of serotype O78 (Ron, Tel Aviv U.) N  NE33G 33AG 33G 33G 33G 33 G  33G 33G 33W             *  Virulent E. coli strainsF G$ffG$ff G$ff Most of the E. coli strains are commensal, but a small number are pathogenic Pathogenic E. coli strains are divided into two groups: Intestinal strains. These produce enterotoxins and constitute a major problem, especially in young children and travellers (Montesumu s revenge) Extraintestinal strains  ExPEC (Extraintestinal Pathogenic E. coli)  Z Z Z G33G33FG33G33&G33E33E33E33G33G33       B               Septicemia (colibacillosis)G$ff.    Colisepticemia is the major causes of mortality from community and hospital-acquired infections (more than 80%) Main cause of mortality in immuno-supressed patients (HIV, chemotherapy, old age) Colisepticemia is an emerging disease  83% increase 1980  1992, over 40% of the bacteremia cases in community acquired infectionsG E33e33e33e33e33e33 ~     '  D   ' Goals:6G$ffg ff Define virulence-essential ExPEC-specific genes Profile strains involved in UTI, NBM and sepsis using these ExPEC-specific genes Use the data to define potential targets for development of vaccines and/or antibacterial drugs.| /E33E33QE33_e33 e 33 E 33v    <  o  CIdentification of virulence related sequences in septicemic strainsDDG$ff.1    3Whole genome sequencing Subtractive hybridization >3  3E33E33 4 Subtractive hybridization  G$ff  Obtain pathogen specific sequences, absent from non-pathogenic K12 strain Excellent chance of  hitting pathogenicity islands which are pathogen specific and very large Faster (and much cheaper) than whole genome sequencing E33  Subtractive hybridization2 G$f3G$f3  QA way to study comparative genomics with organisms which have not been sequenced .R PE33E33 R PSearch for unique  septicemic sequences))G f32      PUsing suppression subtractive hybridization (SSH) we identified sequences unique to strain O78-9 and absent from the non-pathogenic strain K-12 Oover 80 O78-specific open reading frames were found (91 to 1473 bp in length) The same experiment was repeated with another septicemic strain O2-1772 117 unique O2 sequences were identified Q g6GgIg'ggZ  <  ;   9 ,Comparison of unique sequences of O2 and O78--G$f3,  ^Although the two strains cause the same disease, there is a high diversity between the SSH libraries of O2 and O78 strains, with only a few shared genes coding for virulence factors. Is this diversity serotype specific? To determine this we profiled additional septicemic strains of the same serotypes the presence of each of the unique sequences l_ Z|e9eeEe:   L   EComparison of unique sequences of two septicemic strains - O2 and O78FFG f32&     high level of genome plasticity there is a high diversity between the septicemic strainsunexpected for strains causing the same disease Septicemic strains of serogroups O2 and O78 contain a large pool of virulence genes which are used in a  mix and match fashionD  Ye0e33e##tG     &       ` #Current-future researchG$ff $Molecular-physiological studies of the newly-identified virulence genes Bioinformatic studies to determine the evolution of these virulence genes (many in PAIs with evidence of LGT) Whole genome sequencing  joint project with Prof. Joerg Hacker and Prof. Gerhard Gottschalk: E3333E33bH   F  J  $ ($ Thank you!!"  G$ff  TAU group Uri Gophna Diana Ideses Daphna Mokady Dr. Dvora Biran Collaborations Wuerzburg University (Prof. Joerg Hacker) Greifswald University (Prof. Michael Hecker) Goetingen University (Prof. Gerhard Gottschalk)   Z7 Z Z Z G337G33G33sG3333*                          ( "Metagenomics ( Uri Gophna, currently with Ford Doolitle in Halifax studies the effect of TLR mutations on micrflora in health and disease by culture-independent profiling of intestinal microflora of TLR knockout mice under normal conditions and after a challenge which models inflammatory bowel diseaseH  0@|    3   F   k MLST of O78 strains" G$f3  Multi Locus Sequence Typing 450  500 bp of 7  housekeeping genes Criteria for chosing genes: 97-98% homology to E. coli K-12 (from blast data) appear in pathogenic and non pathogenic strains map at considerable distance from each other several allels in the populationf_ Z Z_EEEEZ&  (       ! [The MLST results are compatible with the results from subtractive hybridization and sequencing The profile of virulence factors in ExPEC strains is independent of the host and independent of the serotype There is a high diversity of virulence genes in the various E. coli septicemic strains and each strain has its own profile of virulence genesP\ AE##=AAKA@     @ !"#  Both libraries contain many sequences associated with genomic plasticity - evolution by horizontal gene transfer Many sequences of O2 and O78 are homologous to virulence related sequences of human ExPEC strains ns  c  tecEA&      /$%&'   0(  x  c $m: Y  : x  c $dn:` : H  0޽h ? 33___PPT10i.\ +D=' = @B +r%W()^(   N:kashi@techunix.technion.ac.il>*racheli@ibexperts.com0ugophna@dal.ca0ugophna@dal.ca>*yaira@tauex.tau.ac.ilB.simay@tx.technion.ac.ilB.nosherov@post.tau.ac.il/ 00DArialNew Rot$t\ )0tY 0"DComic Sans MS$t\ )0tY 0B DMS Sans Serif$t\ )0tY 00DTimes New Romant\ )0tY 0c .  @n?" dd@  @@`` 5( ~ n ($%&'  #! " _b$ ov飼kf#A2$%Gg_gDj#A2$nI8i v o&A2$0h,m5A @,7Ab$L-s$֠n:A 0AA ff33@@K2ʚ;n8ʚ;g4BdBd )0hppp@ <42d2d 0$t<4BdBd 0$t( <4!d!dy 0$0___PPT10 ppR___PPT94,&@ &@L@@4@@?  %SPathogenomics in Israel$ff"    Metagenomics LGT = lateral gene transfer Typing of bacterial strains and drug resistance Identification of virulence factors Whole genome analysis 33"   )%#Facilities for pathogenomic studies"$ $G$ff.    Good facilities for genomics, transcriptomics and proteomics in several universities and research institutes High level bioinformatics All facilities are also available as service E33.    Metagenomics ( $ff("   zMetagenomics of culturable and non-culturable microorganism populations present in biofilms of Acute Otitis Media (AOM, Middle Ear Infection) This infection involves a variety of bacterial species, found in the form of biofilms, which are inherently resistant to antibiotic treatment Detection of microbial biodiversity of AOM biofilm is limited due to current cultivation methods The group of Fauzi Silbaq (ArabQual) and Racheli Kreisberg-Zakarin (IBEX) racheli@ibexperts.com in involved in a study the biodiversity of AOM using a metagenomics approach, including microbiology, functional genomics and bioinformatics methodologies v P P33l33 33v3333         &     q  e  <               8   X  0Metagenomics $ff MMetagenomics of intestinal microflora in health and disease and the effect of the TLR mutations Toll-Like Receptors (TLRs) recognize pathogen specific patterns of microorganisms. Mutations in TLRs are associated with inflammatory bowel diseases (Crohn's disease and ulcerative colitis). Patients with those diseases have altered intestinal microflora Uri Gophna, currently with Ford Doolitle in Halifax ugophna@dal.ca performs culture-independent profiling of intestinal microflora of TLR knockout mice under normal conditions and after a challenge which models inflammatory bowel diseaseza Z Z ZaG33G33 G33G33     Q  G  2  W          6   l  0pLGT  lateral gene transfer in relation to pathogenesis (97$ff$8 Uri Gophna ugophna@dal.ca studies the role of lateral gene transfer (LGT) in the evolution of pathogens Uses bioinformatics for the identification of laterally acquired genes and pathways in pathogens \ DE33E33~E33E33N      0  pLGT  lateral gene transfer in relation to pathogenesis (97G$ff(8 Yair Aharonowitz, Ilya Borovok and Gerald Cohen from Tel Aviv University study gluthathione synthesis yaira@tauex.tau.ac.il Identified GshF orthologs, consisting of a -glutamylcysteine ligase (GshA) domain fused to an ATP-grasp domain, in 20 gram-positive and gram-negative bacteria. Remarkably, 95% of these bacteria are mammalian pathogens. Presumably, this fusion gene, once formed, spread between mammalian hosts most likely by horizontal gene transfer R Z8 Z2 ZG33C@        1                    q   0f{  1 Typing of bacterial strains and drug resistance 62(/$ff(1 TThe group of Chezi Kashi in the Technion kashi@techunix.technion.ac.il uses novel methods for molecular typing ofVibrio cholera in order to study emerging new pathogenic strains The group of Sima Yaron (Technion) is involved in typing the serovars of Salmonella enterica in respect to drug resistance and virulence simay@tx.technion.ac.illU qG33G33~G33G33EG33@          )  J            -     0)F 0=T  #Identification of virulence factors$$G$ff# The group of Sima Yaron has developed a rapid, simple screen for real-time quantification of promoter - activity in S. enterica using a library of plasmids with GFP as a reporter The group of Nir Asherov (Tel Aviv U.) nosherov@post.tau.ac.il studies Aspergillus fumigatus, which causes serious disease (around 60% mortality) in immunocompromised people. They concentrate on studying novel cell wall proteins and identified 68 such cwp s. They are studying these CWPs in vitro and in infected animals by the use of deletion mutants. They are currently in the process of preparing a Cwp-specific microarrays to study Cwp gene expression patterns ZtG33 G33G33G33jG33G33uG33     `   0             9  V    s           0 Whole genome analysis$ff Two genomes of bacterial pathogens are being sequenced and analyzed in Israel Bacillus anthracis (Shaefferman et al, Biological Institute, Nes Ziona) Septicemic Escherichia coli  strain of serotype O78 (Ron, Tel Aviv U.) N  NE33G 33AG 33G 33G 33G 33 G  33G 33G 33W             *  Virulent E. coli strainsF G$ffG$ff G$ff Most of the E. coli strains are commensal, but a small number are pathogenic Pathogenic E. coli strains are divided into two groups: Intestinal strains. These produce enterotoxins and constitute a major problem, especially in young children and travellers (Montesumu s revenge) Extraintestinal strains  ExPEC (Extraintestinal Pathogenic E. coli)  Z Z Z G33G33FG33G33&G33E33E33E33G33G33       B               Septicemia (colibacillosis)G$ff.    Colisepticemia is the major causes of mortality from community and hospital-acquired infections (more than 80%) Main cause of mortality in immuno-supressed patients (HIV, chemotherapy, old age) Colisepticemia is an emerging disease  83% increase 1980  1992, over 40% of the bacteremia cases in community acquired infectionsG E33e33e33e33e33e33 ~     '  D   ' Goals:6G$ffg ff Define virulence-essential ExPEC-specific genes Profile strains involved in UTI, NBM and sepsis using these ExPEC-specific genes Use the data to define potential targets for development of vaccines and/or antibacterial drugs.| /E33E33QE33_e33 e 33 E 33v    <  o  CIdentification of virulence related sequences in septicemic strainsDDG$ff.1    3Whole genome sequencing Subtractive hybridization >3  3E33E33 4 Subtractive hybridization  G$ff  Obtain pathogen specific sequences, absent from non-pathogenic K12 strain Excellent chance of  hitting pathogenicity islands which are pathogen specific and very large Faster (and much cheaper) than whole genome sequencing E33  Subtractive hybridization2 G$f3G$f3  QA way to study comparative genomics with organisms which have not been sequenced .R PE33E33 R PSearch for unique  septicemic sequences))G f32      PUsing suppression subtractive hybridization (SSH) we identified sequences unique to strain O78-9 and absent from the non-pathogenic strain K-12 Oover 80 O78-specific open reading frames were found (91 to 1473 bp in length) The same experiment was repeated with another septicemic strain O2-1772 117 unique O2 sequences were identified Q g6GgIg'ggZ  <  ;   9 ,Comparison of unique sequences of O2 and O78--G$f3,  ^Although the two strains cause the same disease, there is a high diversity between the SSH libraries of O2 and O78 strains, with only a few shared genes coding for virulence factors. Is this diversity serotype specific? To determine this we profiled additional septicemic strains of the same serotypes the presence of each of the unique sequences l_ Z|e9eeEe:   L   EComparison of unique sequences of two septicemic strains - O2 and O78FFG f32&     high level of genome plasticity there is a high diversity between the septicemic strainsunexpected for strains causing the same disease Septicemic strains of serogroups O2 and O78 contain a large pool of virulence genes which are used in a  mix and match fashionD  Ye0e33e##tG     &       ` #Current-future researchG$ff $Molecular-physiological studies of the newly-identified virulence genes Bioinformatic studies to determine the evolution of these virulence genes (many in PAIs with evidence of LGT) Whole genome sequencing  joint project with Prof. Joerg Hacker and Prof. Gerhard Gottschalk: E3333E33bH   F  J  $ ($ Thank you!!"  G$ff  TAU group Uri Gophna Diana Ideses Daphna Mokady Dr. Dvora Biran Collaborations Wuerzburg University (Prof. Joerg Hacker) Greifswald University (Prof. Michael Hecker) Goetingen University (Prof. Gerhard Gottschalk)   Z7 Z Z Z G337G33G33sG3333*                          ( "Metagenomics ( Uri Gophna, currently with Ford Doolitle in Halifax studies the effect of TLR mutations on micrflora in health and disease by culture-independent profiling of intestinal microflora of TLR knockout mice under normal conditions and after a challenge which models inflammatory bowel diseaseH  0@|    3   F   k MLST of O78 strains" G$f3  Multi Locus Sequence Typing 450  500 bp of 7  housekeeping genes Criteria for chosing genes: 97-98% homology to E. coli K-12 (from blast data) appear in pathogenic and non pathogenic strains map at considerable distance from each other several allels in the populationf_ Z Z_EEEEZ&  (       ! [The MLST results are compatible with the results from subtractive hybridization and sequencing The profile of virulence factors in ExPEC strains is independent of the host and independent of the serotype There is a high diversity of virulence genes in the various E. coli septicemic strains and each strain has its own profile of virulence genesP\ AE##=AAKA@     @ !"#  Both libraries contain many sequences associated with genomic plasticity - evolution by horizontal gene transfer Many sequences of O2 and O78 are homologous to virulence related sequences of human ExPEC strains ns  c  tecEA&      /$%&'   0(  x  c $m: Y  : x  c $dn:` : H  0޽h ? 33___PPT10i.\ +D=' = @B +}  04$(  4r 4 S $H: `  : r 4 S H:D : H 4 0޽h ? 33___PPT10i.\@+D=' = @B +r(  o()_(  8  N:kashi@techunix.technion.ac.il>*racheli@ibexperts.com0ugophna@dal.ca0ugophna@dal.ca>*yaira@tauex.tau.ac.ilB.simay@tx.technion.ac.ilB.nosherov@post.tau.ac.il> *eliora@post.tau.ac.il/ 00DArialNew Rot$t\ )0tY 0"DComic Sans MS$t\ )0tY 0B DMS Sans Serif$t\ )0tY 00DTimes New Romant\ )0tY 0c .  @n?" dd@  @@`` 5( ~ n ($%&'  #! " _b$ ov飼kf#A2$%Gg_gDj#A2$nI8i v o&A2$0h,m5A @,7Ab$L-s$֠n:A 0AA ff33@@K2ʚ;n8ʚ;g4BdBd )0hppp@ <42d2d 0$t<4BdBd 0$t( <4!d!dy 0$0___PPT10 ppR___PPT94,&@ &@L@@4@@?  %GTPathogenomics in Israel$ff"    Metagenomics LGT = lateral gene transfer Typing of bacterial strains and drug resistance Identification of virulence factors Whole genome analysis 33"   )%#Facilities for pathogenomic studies"$ $G$ff.    Good facilities for genomics, transcriptomics and proteomics in several universities and research institutes High level bioinformatics All facilities are also available as service E33.    Metagenomics ( $ff("   zMetagenomics of culturable and non-culturable microorganism populations present in biofilms of Acute Otitis Media (AOM, Middle Ear Infection) This infection involves a variety of bacterial species, found in the form of biofilms, which are inherently resistant to antibiotic treatment Detection of microbial biodiversity of AOM biofilm is limited due to current cultivation methods The group of Fauzi Silbaq (ArabQual) and Racheli Kreisberg-Zakarin (IBEX) racheli@ibexperts.com in involved in a study the biodiversity of AOM using a metagenomics approach, including microbiology, functional genomics and bioinformatics methodologies v P P33l33 33v3333         &     q  e  <               8   X  0Metagenomics $ff MMetagenomics of intestinal microflora in health and disease and the effect of the TLR mutations Toll-Like Receptors (TLRs) recognize pathogen specific patterns of microorganisms. Mutations in TLRs are associated with inflammatory bowel diseases (Crohn's disease and ulcerative colitis). Patients with those diseases have altered intestinal microflora Uri Gophna, currently with Ford Doolitle in Halifax ugophna@dal.ca performs culture-independent profiling of intestinal microflora of TLR knockout mice under normal conditions and after a challenge which models inflammatory bowel diseaseza Z Z ZaG33G33 G33G33     Q  G  2  W          6   l  0pLGT  lateral gene transfer in relation to pathogenesis (97$ff$8 Uri Gophna ugophna@dal.ca studies the role of lateral gene transfer (LGT) in the evolution of pathogens Uses bioinformatics for the identification of laterally acquired genes and pathways in pathogens \ DE33E33~E33E33N      0  pLGT  lateral gene transfer in relation to pathogenesis (97G$ff(8 Yair Aharonowitz, Ilya Borovok and Gerald Cohen from Tel Aviv University study gluthathione synthesis yaira@tauex.tau.ac.il Identified GshF orthologs, consisting of a -glutamylcysteine ligase (GshA) domain fused to an ATP-grasp domain, in 20 gram-positive and gram-negative bacteria. Remarkably, 95% of these bacteria are mammalian pathogens. Presumably, this fusion gene, once formed, spread between mammalian hosts most likely by horizontal gene transfer R Z8 Z2 ZG33C@        1                    q   0f{  1 Typing of bacterial strains and drug resistance 62(/$ff(1 TThe group of Chezi Kashi in the Technion kashi@techunix.technion.ac.il uses novel methods for molecular typing ofVibrio cholera in order to study emerging new pathogenic strains The group of Sima Yaron (Technion) is involved in typing the serovars of Salmonella enterica in respect to drug resistance and virulence simay@tx.technion.ac.illU qG33G33~G33G33EG33@          )  J            -     0)F 0=T  #Identification of virulence factors$$G$ff# The group of Sima Yaron has developed a rapid, simple screen for real-time quantification of promoter - activity in S. enterica using a library of plasmids with GFP as a reporter The group of Nir Asherov (Tel Aviv U.) nosherov@post.tau.ac.il studies Aspergillus fumigatus, which causes serious disease (around 60% mortality) in immunocompromised people. They concentrate on studying novel cell wall proteins and identified 68 such cwp s. They are studying these CWPs in vitro and in infected animals by the use of deletion mutants. They are currently in the process of preparing a Cwp-specific microarrays to study Cwp gene expression patterns ZtG33 G33G33G33jG33G33uG33     `   0             9  V    s           0 Whole genome analysis$ff Two genomes of bacterial pathogens are being sequenced and analyzed in Israel Bacillus anthracis (Shaefferman et al, Biological Institute, Nes Ziona) Septicemic Escherichia coli  strain of serotype O78 (Ron, Tel Aviv U.) eliora@post.tau.ac.il N   NE33G 33AG 33G 33G 33G 33 G  33G 33G 33W             *       0Virulent E. coli strainsF G$ffG$ff G$ff Most of the E. coli strains are commensal, but a small number are pathogenic Pathogenic E. coli strains are divided into two groups: Intestinal strains. These produce enterotoxins and constitute a major problem, especially in young children and travellers (Montesumu s revenge) Extraintestinal strains  ExPEC (Extraintestinal Pathogenic E. coli)  Z Z Z G33G33FG33G33&G33E33E33E33G33G33       B               Septicemia (colibacillosis)G$ff.    Colisepticemia is the major causes of mortality from community and hospital-acquired infections (more than 80%) Main cause of mortality in immuno-supressed patients (HIV, chemotherapy, old age) Colisepticemia is an emerging disease  83% increase 1980  1992, over 40% of the bacteremia cases in community acquired infectionsG E33e33e33e33e33e33 ~     '  D   ' Goals:6G$ffg ff Define virulence-essential ExPEC-specific genes Profile strains involved in UTI, NBM and sepsis using these ExPEC-specific genes Use the data to define potential targets for development of vaccines and/or antibacterial drugs.| /E33E33QE33_e33 e 33 E 33v    <  o  CIdentification of virulence related sequences in septicemic strainsDDG$ff.1    3Whole genome sequencing Subtractive hybridization >3  3E33E33 4 Subtractive hybridization  G$ff  Obtain pathogen specific sequences, absent from non-pathogenic K12 strain Excellent chance of  hitting pathogenicity islands which are pathogen specific and very large Faster (and much cheaper) than whole genome sequencing E33  Subtractive hybridization2 G$f3G$f3  QA way to study comparative genomics with organisms which have not been sequenced .R PE33E33 R PSearch for unique  septicemic sequences))G f32      PUsing suppression subtractive hybridization (SSH) we identified sequences unique to strain O78-9 and absent from the non-pathogenic strain K-12 Oover 80 O78-specific open reading frames were found (91 to 1473 bp in length) The same experiment was repeated with another septicemic strain O2-1772 117 unique O2 sequences were identified Q g6GgIg'ggZ  <  ;   9 ,Comparison of unique sequences of O2 and O78--G$f3,  ^Although the two strains cause the same disease, there is a high diversity between the SSH libraries of O2 and O78 strains, with only a few shared genes coding for virulence factors. Is this diversity serotype specific? To determine this we profiled additional septicemic strains of the same serotypes the presence of each of the unique sequences l_ Z|e9eeEe:   L   EComparison of unique sequences of two septicemic strains - O2 and O78FFG f32&     high level of genome plasticity there is a high diversity between the septicemic strainsunexpected for strains causing the same disease Septicemic strains of serogroups O2 and O78 contain a large pool of virulence genes which are used in a  mix and match fashionD  Ye0e33e##tG     &       ` #Current-future researchG$ff $Molecular-physiological studies of the newly-identified virulence genes Bioinformatic studies to determine the evolution of these virulence genes (many in PAIs with evidence of LGT) Whole genome sequencing  joint project with Prof. Joerg Hacker and Prof. Gerhard Gottschalk: E3333E33bH   F  J  $ ($ Thank you!!"  G$ff  TAU group Uri Gophna Diana Ideses Daphna Mokady Dr. Dvora Biran Collaborations Wuerzburg University (Prof. Joerg Hacker) Greifswald University (Prof. Michael Hecker) Goetingen University (Prof. Gerhard Gottschalk)   Z7 Z Z Z G337G33G33sG3333*                          ( "Metagenomics ( Uri Gophna, currently with Ford Doolitle in Halifax studies the effect of TLR mutations on micrflora in health and disease by culture-independent profiling of intestinal microflora of TLR knockout mice under normal conditions and after a challenge which models inflammatory bowel diseaseH  0@|    3   F   k MLST of O78 strains" G$f3  Multi Locus Sequence Typing 450  500 bp of 7  housekeeping genes Criteria for chosing genes: 97-98% homology to E. coli K-12 (from blast data) appear in pathogenic and non pathogenic strains map at considerable distance from each other several allels in the populationf_ Z Z_EEEEZ&  (       ! [The MLST results are compatible with the results from subtractive hybridization and sequencing The profile of virulence factors in ExPEC strains is independent of the host and independent of the serotype There is a high diversity of virulence genes in the various E. coli septicemic strains and each strain has its own profile of virulence genesP\ AE##=AAKA@     @ !"#  Both libraries contain many sequences associated with genomic plasticity - evolution by horizontal gene transfer Many sequences of O2 and O78 are homologous to virulence related sequences of human ExPEC strains ns  c  tecEA&      /$%&'  P0(  x  c $d6: `}  : x  c $: ` : H  0޽h ? 33___PPT10i.\iZ+D=' = @B +rߌ :)b`(  8  N:kashi@techunix.te.ac.il mailto:ashaefferman@iibr.gov.ilmailto:nosherov@post.tau.ac.ilmailto:GilS@tauex.tau.ac.ilchnion.ac.il>*racheli@ibexperts.com0ugophna@dal.ca0ugophna@dal.ca>*yaira@tauex.tau.ac.ilB.simay@tx.technion.ac.ilB.nosherov@post.tau.ac.il> *eliora@post.tau.ac.il/ 00DArialNew Rot$t\ )0tY 0"DComic Sans MS$t\ )0tY 0B DMS Sans Serif$t\ )0tY 00DTimes New Romant\ )0tY 0c .  @n?" dd@  @@`` 5( ~ n ($%&'  #! " _b$ ov飼kf#A2$%Gg_gDj#A2$nI8i v o&A2$0h,m5A @,7Ab$L-s$֠n:A 0AA ff33@@K2ʚ;n8ʚ;g4BdBd )0hppp@ <42d2d 0$t<4BdBd 0$t( <4!d!dy 0$0___PPT10 ppR___PPT94,&@ &@L@@4@@?  %TPathogenomics in Israel$ff"    Metagenomics LGT = lateral gene transfer Typing of bacterial strains and drug resistance Identification of virulence factors Whole genome analysis 33"   )%#Facilities for pathogenomic studies"$ $G$ff.    Good facilities for genomics, transcriptomics and proteomics in several universities and research institutes High level bioinformatics All facilities are also available as service E33.    Metagenomics ( $ff("   zMetagenomics of culturable and non-culturable microorganism populations present in biofilms of Acute Otitis Media (AOM, Middle Ear Infection) This infection involves a variety of bacterial species, found in the form of biofilms, which are inherently resistant to antibiotic treatment Detection of microbial biodiversity of AOM biofilm is limited due to current cultivation methods The group of Fauzi Silbaq (ArabQual) and Racheli Kreisberg-Zakarin (IBEX) racheli@ibexperts.com in involved in a study the biodiversity of AOM using a metagenomics approach, including microbiology, functional genomics and bioinformatics methodologies v P P33l33 33v3333         &     q  e  <               8   X  0Metagenomics $ff MMetagenomics of intestinal microflora in health and disease and the effect of the TLR mutations Toll-Like Receptors (TLRs) recognize pathogen specific patterns of microorganisms. Mutations in TLRs are associated with inflammatory bowel diseases (Crohn's disease and ulcerative colitis). Patients with those diseases have altered intestinal microflora Uri Gophna, currently with Ford Doolitle in Halifax ugophna@dal.ca performs culture-independent profiling of intestinal microflora of TLR knockout mice under normal conditions and after a challenge which models inflammatory bowel diseaseza Z Z ZaG33G33 G33G33     Q  G  2  W          6   l  0pLGT  lateral gene transfer in relation to pathogenesis (97$ff$8 Uri Gophna ugophna@dal.ca studies the role of lateral gene transfer (LGT) in the evolution of pathogens Uses bioinformatics for the identification of laterally acquired genes and pathways in pathogens \ DE33E33~E33E33N      0  pLGT  lateral gene transfer in relation to pathogenesis (97G$ff(8 Yair Aharonowitz, Ilya Borovok and Gerald Cohen from Tel Aviv University study gluthathione synthesis yaira@tauex.tau.ac.il Identified GshF orthologs, consisting of a -glutamylcysteine ligase (GshA) domain fused to an ATP-grasp domain, in 20 gram-positive and gram-negative bacteria. Remarkably, 95% of these bacteria are mammalian pathogens. Presumably, this fusion gene, once formed, spread between mammalian hosts most likely by horizontal gene transfer R Z8 Z2 ZG33C@        1                    q   0f{  1 Typing of bacterial strains and drug resistance 62(/$ff(1 TThe group of Chezi Kashi in the Technion kashi@techunix.technion.ac.il uses novel methods for molecular typing ofVibrio cholera in order to study emerging new pathogenic strains The group of Sima Yaron (Technion) is involved in typing the serovars of Salmonella enterica in respect to drug resistance and virulence simay@tx.technion.ac.illU qG33G33~G33G33EG33@          )  J            -     0)F 0=T  #Identification of virulence factors$$G$ff# The group of Sima Yaron has developed a rapid, simple screen for real-time quantification of promoter - activity in S. enterica using a library of plasmids with GFP as a reporter The group of Nir Asherov (Tel Aviv U.) nosherov@post.tau.ac.il studies Aspergillus fumigatus, which causes serious disease (around 60% mortality) in immunocompromised people. They concentrate on studying novel cell wall proteins and identified 68 such cwp s. They are studying these CWPs in vitro and in infected animals by the use of deletion mutants. They are currently in the process of preparing a Cwp-specific microarrays to study Cwp gene expression patterns ZtG33 G33G33G33jG33G33uG33     `   0             9  V    s           0 Whole genome analysis$ff "Two genomes of bacterial pathogens are being sequenced and analyzed in Israel Bacillus anthracis (Shaefferman et al, Biological Institute, Nes Ziona) <ashaefferman@iibr.gov.il> Septicemic Escherichia coli  strain of serotype O78 (Ron, Tel Aviv U.) eliora@post.tau      !"#$%&'()*+,-./0123456789:;<=>?@ABCDEFGHIJKLMNOPQRSTUVWXYZ[\]^_`abcdefghijklmnopqrstuvwxyz{|}~.ac.il N   NE33G 33]G 33G 33G 33G 33 G  33G 33G 33>W                      *       0Virulent E. coli strainsF G$ffG$ff G$ff Most of the E. coli strains are commensal, but a small number are pathogenic Pathogenic E. coli strains are divided into two groups: Intestinal strains. These produce enterotoxins and constitute a major problem, especially in young children and travellers (Montesumu s revenge) Extraintestinal strains  ExPEC (Extraintestinal Pathogenic E. coli)  Z Z Z G33G33FG33G33&G33E33E33E33G33G33       B               Septicemia (colibacillosis)G$ff.    Colisepticemia is the major causes of mortality from community and hospital-acquired infections (more than 80%) Main cause of mortality in immuno-supressed patients (HIV, chemotherapy, old age) Colisepticemia is an emerging disease  83% increase 1980  1992, over 40% of the bacteremia cases in community acquired infectionsG E33e33e33e33e33e33 ~     '  D   ' Goals:6G$ffg ff Define virulence-essential ExPEC-specific genes Profile strains involved in UTI, NBM and sepsis using these ExPEC-specific genes Use the data to define potential targets for development of vaccines and/or antibacterial drugs.| /E33E33QE33_e33 e 33 E 33v    <  o  CIdentification of virulence related sequences in septicemic strainsDDG$ff.1    3Whole genome sequencing Subtractive hybridization >3  3E33E33 4 Subtractive hybridization  G$ff  Obtain pathogen specific sequences, absent from non-pathogenic K12 strain Excellent chance of  hitting pathogenicity islands which are pathogen specific and very large Faster (and much cheaper) than whole genome sequencing E33  Subtractive hybridization2 G$f3G$f3  QA way to study comparative genomics with organisms which have not been sequenced .R PE33E33 R PSearch for unique  septicemic sequences))G f32      PUsing suppression subtractive hybridization (SSH) we identified sequences unique to strain O78-9 and absent from the non-pathogenic strain K-12 Oover 80 O78-specific open reading frames were found (91 to 1473 bp in length) The same experiment was repeated with another septicemic strain O2-1772 117 unique O2 sequences were identified Q g6GgIg'ggZ  <  ;   9 ,Comparison of unique sequences of O2 and O78--G$f3,  ^Although the two strains cause the same disease, there is a high diversity between the SSH libraries of O2 and O78 strains, with only a few shared genes coding for virulence factors. Is this diversity serotype specific? To determine this we profiled additional septicemic strains of the same serotypes the presence of each of the unique sequences l_ Z|e9eeEe:   L   EComparison of unique sequences of two septicemic strains - O2 and O78FFG f32&     high level of genome plasticity there is a high diversity between the septicemic strainsunexpected for strains causing the same disease Septicemic strains of serogroups O2 and O78 contain a large pool of virulence genes which are used in a  mix and match fashionD  Ye0e33e##tG     &       ` #Current-future researchG$ff $Molecular-physiological studies of the newly-identified virulence genes Bioinformatic studies to determine the evolution of these virulence genes (many in PAIs with evidence of LGT) Whole genome sequencing  joint project with Prof. Joerg Hacker and Prof. Gerhard Gottschalk: E3333E33bH   F  J  $ ($ Thank you!!"  G$ff  TAU group Uri Gophna Diana Ideses Daphna Mokady Dr. Dvora Biran Collaborations Wuerzburg University (Prof. Joerg Hacker) Greifswald University (Prof. Michael Hecker) Goetingen University (Prof. Gerhard Gottschalk)   Z7 Z Z Z G337G33G33sG3333*                          ( "Metagenomics ( Uri Gophna, currently with Ford Doolitle in Halifax studies the effect of TLR mutations on micrflora in health and disease by culture-independent profiling of intestinal microflora of TLR knockout mice under normal conditions and after a challenge which models inflammatory bowel diseaseH  0@|    3   F   k MLST of O78 strains" G$f3  Multi Locus Sequence Typing 450  500 bp of 7  housekeeping genes Criteria for chosing genes: 97-98% homology to E. coli K-12 (from blast data) appear in pathogenic and non pathogenic strains map at considerable distance from each other several allels in the populationf_ Z Z_EEEEZ&  (       ! [The MLST results are compatible with the results from subtractive hybridization and sequencing The profile of virulence factors in ExPEC strains is independent of the host and independent of the serotype There is a high diversity of virulence genes in the various E. coli septicemic strains and each strain has its own profile of virulence genesP\ AE##=AAKA@     @ !"#  Both libraries contain many sequences associated with genomic plasticity - evolution by horizontal gene transfer Many sequences of O2 and O78 are homologous to virulence related sequences of human ExPEC strains ns  c  tecEA&      /$%&'  P0(  x  c $d6: `}  : x  c $: ` : H  0޽h ? 33___PPT10i.\iZ+D=' = @B +rv ORqR)Ib(    N:kashi@techunix.technion.ac.il>*racheli@ibexperts.com0ugophna@dal.ca0ugophna@dal.ca>*yaira@tauex.tau.ac.ilB.simay@tx.technion.ac.il> *eliora@post.tau.ac.ilD 0ashaefferman@iibr.gov.ilB .nosherov@post.tau.ac.il/ 00DArialNew Rot$t\ )0tY 0h"DComic Sans MS$t\ )0tY 0hB DMS Sans Serif$t\ )0tY 00DTimes New Romant\ )0tY 0hc .  @n?" dd@  @@``  6) ~ n ($%&'  #! " )_b$ ov飼kf#A2$%Gg_gDj#A2$nI8i v o&A2$0h,m5A @,7Ab$L-s$֠n:A 0AA ff33@@K2ʚ;n8ʚ;g4BdBd )0hppp@ <42d2d 0$t<4BdBd 0$t( <4!d!dy 0$0___PPT10 ppR___PPT94,&@ &@L@@4@@?  %zVPathogenomics in Israel$ff"    Metagenomics LGT = lateral gene transfer Typing of bacterial strains and drug resistance Identification of virulence factors Whole genome analysis 33"   )%#Facilities for pathogenomic studies"$ $G$ff.    Good facilities for genomics, transcriptomics and proteomics in several universities and research institutes High level bioinformatics All facilities are also available as service E33.    Metagenomics ( $ff("   zMetagenomics of culturable and non-culturable microorganism populations present in biofilms of Acute Otitis Media (AOM, Middle Ear Infection) This infection involves a variety of bacterial species, found in the form of biofilms, which are inherently resistant to antibiotic treatment Detection of microbial biodiversity of AOM biofilm is limited due to current cultivation methods The group of Fauzi Silbaq (ArabQual) and Racheli Kreisberg-Zakarin (IBEX) racheli@ibexperts.com in involved in a study the biodiversity of AOM using a metagenomics approach, including microbiology, functional genomics and bioinformatics methodologies v P P33l33 33v3333         &     q  e  <               8   X  0Metagenomics $ff MMetagenomics of intestinal microflora in health and disease and the effect of the TLR mutations Toll-Like Receptors (TLRs) recognize pathogen specific patterns of microorganisms. Mutations in TLRs are associated with inflammatory bowel diseases (Crohn's disease and ulcerative colitis). Patients with those diseases have altered intestinal microflora Uri Gophna, currently with Ford Doolitle in Halifax ugophna@dal.ca performs culture-independent profiling of intestinal microflora of TLR knockout mice under normal conditions and after a challenge which models inflammatory bowel diseaseza Z Z ZaG33G33 G33G33     Q  G  2  W          6   l  0pLGT  lateral gene transfer in relation to pathogenesis (97$ff$8 Uri Gophna ugophna@dal.ca studies the role of lateral gene transfer (LGT) in the evolution of pathogens Uses bioinformatics for the identification of laterally acquired genes and pathways in pathogens \ DE33E33~E33E33N      0  pLGT  lateral gene transfer in relation to pathogenesis (97G$ff(8 Yair Aharonowitz, Ilya Borovok and Gerald Cohen from Tel Aviv University study gluthathione synthesis yaira@tauex.tau.ac.il Identified GshF orthologs, consisting of a -glutamylcysteine ligase (GshA) domain fused to an ATP-grasp domain, in 20 gram-positive and gram-negative bacteria. Remarkably, 95% of these bacteria are mammalian pathogens. Presumably, this fusion gene, once formed, spread between mammalian hosts most likely by horizontal gene transfer R Z8 Z2 ZG33C@        1                    q   0f{  1 Typing of bacterial strains and drug resistance 62(/$ff(1 TThe group of Chezi Kashi in the Technion kashi@techunix.technion.ac.il uses novel methods for molecular typing ofVibrio cholera in order to study emerging new pathogenic strains The group of Sima Yaron (Technion) is involved in typing the serovars of Salmonella enterica in respect to drug resistance and virulence simay@tx.technion.ac.illU qG33G33~G33G33EG33@          )  J            -     0)F 0=T  -Identification of bacterial virulence factors..G$ff- The group of Sima Yaron has developed a rapid, simple screen for real-time quantification of promoter - activity in S. enterica using a library of plasmids with GFP as a reporter The group of Gil Segal (Tel Aviv U.) is involved in the study of hyper variable genes are found in the Legionella icm/dot pathogenesis region. They have demonstrated the role of these genes in virulence   tG33 G33G33G33dG33%ggQgCG33     `     W *&*Identification of fungal virulence factors++G$ff* The group of Nir Asherov (Tel Aviv U.) nosherov@post.tau.ac.il studies Aspergillus fumigatus, which causes serious disease (around 60% mortality) in immunocompromised people. They concentrate on studying novel cell wall proteins and identified 68 such cwp s. They are studying these CWPs in vitro and in infected animals by the use of deletion mutants. They are currently in the process of preparing a Cwp-specific microarrays to study Cwp gene expression patternsH ZGG33G33uG332            8  U    r            0'> Whole genome analysis$ff  Two genomes of bacterial pathogens are being sequenced and analyzed in Israel Bacillus anthracis (Shaefferman et al, Biological Institute, Nes Ziona) ashaefferman@iibr.gov.il Septicemic Escherichia coli  strain of serotype O78 (Ron, Tel Aviv U.) eliora@post.tau.ac.il N   NE33G 33\G 33G 33G 33G 33 G  33G 33G 33W                *       0  0Virulent E. coli strainsF G$ffG$ff G$ff Most of the E. coli strains are commensal, but a small number are pathogenic Pathogenic E. coli strains are divided into two groups: Intestinal strains. These produce enterotoxins and constitute a major problem, especially in young children and travellers (Montesumu s revenge) Extraintestinal strains  ExPEC (Extraintestinal Pathogenic E. coli)  Z Z Z G33G33FG33G33&G33E33E33E33G33G33       B               Septicemia (colibacillosis)G$ff.    Colisepticemia is the major causes of mortality from community and hospital-acquired infections (more than 80%) Main cause of mortality in immuno-supressed patients (HIV, chemotherapy, old age) Colisepticemia is an emerging disease  83% increase 1980  1992, over 40% of the bacteremia cases in community acquired infectionsG E33e33e33e33e33e33 ~     '  D   ' Goals:6G$ffg ff Define virulence-essential ExPEC-specific genes Profile strains involved in UTI, NBM and sepsis using these ExPEC-specific genes Use the data to define potential targets for development of vaccines and/or antibacterial drugs.| /E33E33QE33_e33 e 33 E 33v    <  o  CIdentification of virulence related sequences in septicemic strainsDDG$ff.1    3Whole genome sequencing Subtractive hybridization >3  3E33E33 4 Subtractive hybridization  G$ff  Obtain pathogen specific sequences, absent from non-pathogenic K12 strain Excellent chance of  hitting pathogenicity islands which are pathogen specific and very large Faster (and much cheaper) than whole genome sequencing E33  Subtractive hybridization2 G$f3G$f3  QA way to study comparative genomics with organisms which have not been sequenced .R PE33E33 R PSearch for unique  septicemic sequences))G f32      PUsing suppression subtractive hybridization (SSH) we identified sequences unique to strain O78-9 and absent from the non-pathogenic strain K-12 Oover 80 O78-specific open reading frames were found (91 to 1473 bp in length) The same experiment was repeated with another septicemic strain O2-1772 117 unique O2 sequences were identified Q g6GgIg'ggZ  <  ;   9 ,Comparison of unique sequences of O2 and O78--G$f3,  ^Although the two strains cause the same disease, there is a high diversity between the SSH libraries of O2 and O78 strains, with only a few shared genes coding for virulence factors. Is this diversity serotype specific? To determine this we profiled additional septicemic strains of the same serotypes the presence of each of the unique sequences l_ Z|e9eeEe:   L   EComparison of unique sequences of two septicemic strains - O2 and O78FFG f32&     high level of genome plasticity there is a high diversity between the septicemic strainsunexpected for strains causing the same disease Septicemic strains of serogroups O2 and O78 contain a large pool of virulence genes which are used in a  mix and match fashionD  Ye0e33e##tG     &       ` #Current-future researchG$ff $Molecular-physiological studies of the newly-identified virulence genes Bioinformatic studies to determine the evolution of these virulence genes (many in PAIs with evidence of LGT) Whole genome sequencing  joint project with Prof. Joerg Hacker and Prof. Gerhard Gottschalk: E3333E33bH   F  J  $ ($ Thank you!!"  G$ff  TAU group Uri Gophna Diana Ideses Daphna Mokady Dr. Dvora Biran Collaborations Wuerzburg University (Prof. Joerg Hacker) Greifswald University (Prof. Michael Hecker) Goetingen University (Prof. Gerhard Gottschalk)   Z7 Z Z Z G337G33G33sG3333*                          ( "Metagenomics ( Uri Gophna, currently with Ford Doolitle in Halifax studies the effect of TLR mutations on micrflora in health and disease by culture-independent profiling of intestinal microflora of TLR knockout mice under normal conditions and after a challenge which models inflammatory bowel diseaseH  0@|    3   F   k MLST of O78 strains" G$f3  Multi Locus Sequence Typing 450  500 bp of 7  housekeeping genes Criteria for chosing genes: 97-98% homology to E. coli K-12 (from blast data) appear in pathogenic and non pathogenic strains map at considerable distance from each other several allels in the populationf_ Z Z_EEEEZ&  (       ! [The MLST results are compatible with the results from subtractive hybridization and sequencing The profile of virulence factors in ExPEC strains is independent of the host and independent of the serotype There is a high diversity of virulence genes in the various E. coli septicemic strains and each strain has its own profile of virulence genesP\ AE##=AAKA@     @ !"#  Both libraries contain many sequences associated with genomic plasticity - evolution by horizontal gene transfer Many sequences of O2 and O78 are homologous to virulence related sequences of human ExPEC strains ns  c  tecEA&      /$%&'}  04$(  4r 4 S $H: `  : r 4 S H:D : H 4 0޽h ? 33___PPT10i.\@+D=' = @B +J  (  r  S h~ `    c 0e0e  `}   H  0޽h ? 33  P0(  x  c $d6: `}  : x  c $: ` : H  0޽h ? 33___PPT10i.\iZ+D=' = @B +r R չ *&Rf*b(    N:kashi@techunix.technion.ac.il>*racheli@ibexperts.com0ugophna@dal.ca0ugophna@dal.ca>*yaira@tauex.tau.ac.ilB.simay@tx.technion.ac.il> *eliora@post.tau.ac.ilD 0ashaefferman@iibr.gov.ilB .nosherov@post.tau.ac.il< (GilS@tauex.tau.ac.il/ 00DArialNew Rot$t\ )0tY 0h"DComic Sans MS$t\ )0tY 0hB DMS Sans Serif$t\ )0tY 00DTimes New Romant\ )0tY 0hc .  @n?" dd@  @@``  6) ~ n ($%&'  #! " )_b$ ov飼kf#A2$%Gg_gDj#A2$nI8i v o&A2$0h,m5A @,7Ab$L-s$֠n:A 0AA ff33@@K2ʚ;n8ʚ;g4BdBd )0hppp@ <42d2d 0$t<4BdBd 0$t( <4!d!dy 0$0___PPT10 ppR___PPT94,&@ &@L@@4@@?  %VPathogenomics in Israel$ff"    Metagenomics LGT = lateral gene transfer Typing of bacterial strains and drug resistance Identification of virulence factors Whole genome analysis 33"   )%#Facilities for pathogenomic studies"$ $G$ff.    Good facilities for genomics, transcriptomics and proteomics in several universities and research institutes High level bioinformatics All facilities are also available as service E33.    Metagenomics ( $ff("   zMetagenomics of culturable and non-culturable microorganism populations present in biofilms of Acute Otitis Media (AOM, Middle Ear Infection) This infection involves a variety of bacterial species, found in the form of biofilms, which are inherently resistant to antibiotic treatment Detection of microbial biodiversity of AOM biofilm is limited due to current cultivation methods The group of Fauzi Silbaq (ArabQual) and Racheli Kreisberg-Zakarin (IBEX) racheli@ibexperts.com in involved in a study the biodiversity of AOM using a metagenomics approach, including microbiology, functional genomics and bioinformatics methodologies v P P33l33 33v3333         &     q  e  <               8   X  0Metagenomics $ff MMetagenomics of intestinal microflora in health and disease and the effect of the TLR mutations Toll-Like Receptors (TLRs) recognize pathogen specific patterns of microorganisms. Mutations in TLRs are associated with inflammatory bowel diseases (Crohn's disease and ulcerative colitis). Patients with those diseases have altered intestinal microflora Uri Gophna, currently with Ford Doolitle in Halifax ugophna@dal.ca performs culture-independent profiling of intestinal microflora of TLR knockout mice under normal conditions and after a challenge which models inflammatory bowel diseaseza Z Z ZaG33G33 G33G33     Q  G  2  W          6   l  0pLGT  lateral gene transfer in relation to pathogenesis (97$ff$8 Uri Gophna ugophna@dal.ca studies the role of lateral gene transfer (LGT) in the evolution of pathogens Uses bioinformatics for the identification of laterally acquired genes and pathways in pathogens \ DE33E33~E33E33N      0  pLGT  lateral gene transfer in relation to pathogenesis (97G$ff(8 Yair Aharonowitz, Ilya Borovok and Gerald Cohen from Tel Aviv University study gluthathione synthesis yaira@tauex.tau.ac.il Identified GshF orthologs, consisting of a -glutamylcysteine ligase (GshA) domain fused to an ATP-grasp domain, in 20 gram-positive and gram-negative bacteria. Remarkably, 95% of these bacteria are mammalian pathogens. Presumably, this fusion gene, once formed, spread between mammalian hosts most likely by horizontal gene transfer R Z8 Z2 ZG33C@        1                    q   0f{  1 Typing of bacterial strains and drug resistance 62(/$ff(1 TThe group of Chezi Kashi in the Technion kashi@techunix.technion.ac.il uses novel methods for molecular typing ofVibrio cholera in order to study emerging new pathogenic strains The group of Sima Yaron (Technion) is involved in typing the serovars of Salmonella enterica in respect to drug resistance and virulence simay@tx.technion.ac.illU qG33G33~G33G33EG33@          )  J            -     0)F 0=T  -Identification of bacterial virulence factors..G$ff- The group of Sima Yaron has developed a rapid, simple screen for real-time quantification of promoter - activity in S. enterica using a library of plasmids with GFP as a reporter The group of Gil Segal (Tel Aviv U.) GilS@tauex.tau.ac.il is involved in the study of hyper variable genes are found in the Legionella icm/dot pathogenesis region. They have demonstrated the role of these genes in virulence  tG33 G33G33G33yG33%ggQgCG33     `   H  B  V   0*&*Identification of fungal virulence factors++G$ff* The group of Nir Asherov (Tel Aviv U.) nosherov@post.tau.ac.il studies Aspergillus fumigatus, which causes serious disease (around 60% mortality) in immunocompromised people. They concentrate on studying novel cell wall proteins and identified 68 such cwp s. They are studying these CWPs in vitro and in infected animals by the use of deletion mutants. They are currently in the process of preparing a Cwp-specific microarrays to study Cwp gene expression patternsH ZGG33G33uG332            8  U    r            0'> Whole genome analysis$ff  Two genomes of bacterial pathogens are being sequenced and analyzed in Israel Bacillus anthracis (Shaefferman et al, Biological Institute, Nes Ziona) ashaefferman@iibr.gov.il Septicemic Escherichia coli  strain of serotype O78 (Ron, Tel Aviv U.) eliora@post.tau.ac.il N   NE33G 33\G 33G 33G 33G 33 G  33G 33G 33W                *       0  0Virulent E. coli strainsF G$ffG$ff G$ff Most of the E. coli strains are commensal, but a small number are pathogenic Pathogenic E. coli strains are divided into two groups: Intestinal strains. These produce enterotoxins and constitute a major problem, especially in young children and travellers (Montesumu s revenge) Extraintestinal strains  ExPEC (Extraintestinal Pathogenic E. coli)  Z Z Z G33G33FG33G33&G33E33E33E33G33G33       B               Septicemia (colibacillosis)G$ff.    Colisepticemia is the major causes of mortality from community and hospital-acquired infections (more than 80%) Main cause of mortality in immuno-supressed patients (HIV, chemotherapy, old age) Colisepticemia is an emerging disease  83% increase 1980  1992, over 40% of the bacteremia cases in community acquired infectionsG E33e33e33e33e33e33 ~     '  D   ' Goals:6G$ffg ff Define virulence-essential ExPEC-specific genes Profile strains involved in UTI, NBM and sepsis using these ExPEC-specific genes Use the data to define potential targets for development of vaccines and/or antibacterial drugs.| /E33E33QE33_e33 e 33 E 33v    <  o  CIdentification of virulence related sequences in septicemic strainsDDG$ff.1    3Whole genome sequencing Subtractive hybridization >3  3E33E33 4 Subtractive hybridization  G$ff  Obtain pathogen specific sequences, absent from non-pathogenic K12 strain Excellent chance of  hitting pathogenicity islands which are pathogen specific and very large Faster (and much cheaper) than whole genome sequencing E33  Subtractive hybridization2 G$f3G$f3  QA way to study comparative genomics with organisms which have not been sequenced .R PE33E33 R PSearch for unique  septicemic sequences))G f32      PUsing suppression subtractive hybridization (SSH) we identified sequences unique to strain O78-9 and absent from the non-pathogenic strain K-12 Oover 80 O78-specific open reading frames were found (91 to 1473 bp in length) The same experiment was repeated with another septicemic strain O2-1772 117 unique O2 sequences were identified Q g6GgIg'ggZ  <  ;   9 ,Comparison of unique sequences of O2 and O78--G$f3,  ^Although the two strains cause the same disease, there is a high diversity between the SSH libraries of O2 and O78 strains, with only a few shared genes coding for virulence factors. Is this diversity serotype specific? To determine this we profiled additional septicemic strains of the same serotypes the presence of each of the unique sequences l_ Z|e9eeEe:   L   EComparison of unique sequences of two septicemic strains - O2 and O78FFG f32&     high level of genome plasticity there is a high diversity between the septicemic strainsunexpected for strains causing the same disease Septicemic strains of serogroups O2 and O78 contain a large pool of virulence genes which are used in a  mix and match fashionD  Ye0e33e##tG     &       ` #Current-future researchG$ff $Molecular-physiological studies of the newly-identified virulence genes Bioinformatic studies to determine the evolution of these virulence genes (many in PAIs with evidence of LGT) Whole genome sequencing  joint project with Prof. Joerg Hacker and Prof. Gerhard Gottschalk: E3333E33bH   F  J  $ ($ Thank you!!"  G$ff  TAU group Uri Gophna Diana Ideses Daphna Mokady Dr. Dvora Biran Collaborations Wuerzburg University (Prof. Joerg Hacker) Greifswald University (Prof. Michael Hecker) Goetingen University (Prof. Gerhard Gottschalk)   Z7 Z Z Z G337G33G33sG3333*                          ( "Metagenomics ( Uri Gophna, currently with Ford Doolitle in Halifax studies the effect of TLR mutations on micrflora in health and disease by culture-independent profiling of intestinal microflora of TLR knockout mice under normal conditions and after a challenge which models inflammatory bowel diseaseH  0@|    3   F   k MLST of O78 strains" G$f3  Multi Locus Sequence Typing 450  500 bp of 7  housekeeping genes Criteria for chosing genes: 97-98% homology to E. coli K-12 (from blast data) appear in pathogenic and non pathogenic strains map at considerable distance from each other several allels in the populationf_ Z Z_EEEEZ&  (       ! [The MLST results are compatible with the results from subtractive hybridization and sequencing The profile of virulence factors in ExPEC strains is independent of the host and independent of the serotype There is a high diversity of virulence genes in the various E. coli septicemic strains and each strain has its own profile of virulence genesP\ AE##=AAKA@     @ !"#  Both libraries contain many sequences associated with genomic plasticity - evolution by horizontal gene transfer Many sequences of O2 and O78 are homologous to virulence related sequences of human ExPEC strains ns  c  tecEA&      /$%&'}  04$(  4r 4 S $H: `  : r 4 S H:D : H 4 0޽h ? 33___PPT10i.\@+D=' = @B +r  "*b(    N:kashi@techunix.technion.ac.il>*racheli@ibexperts.com0ugophna@dal.ca0ugophna@dal.ca>*yaira@tauex.tau.ac.ilB.simay@tx.technion.ac.il> *eliora@post.tau.ac.ilD 0ashaefferman@iibr.gov.ilB .nosherov@post.tau.ac.il< (GilS@tauex.tau.ac.il/ 00DArialNew Rot$t\ )0tY 0h"DComic Sans MS$t\ )0tY 0hB DMS Sans Serif$t\ )0tY 00DTimes New Romant\ )0tY 0hc .  @n?" dd@  @@``  6) ~ n ($%&'  #! " )_b$ ov飼kf#A2$%Gg_gDj#A2$nI8i v o&A2$0h,m5A @,7Ab$L-s$֠n:A 0AA ff33@@K2ʚ;n8ʚ;g4BdBd )0hppp@ <42d2d 0$t<4BdBd 0$t( <4!d!dy 0$0___PPT10 ppR___PPT94,&@ &@L@@4@@?  %VPathogenomics in Israel$ff"    Metagenomics LGT = lateral gene transfer Typing of bacterial strains and drug resistance Identification of virulence factors Whole genome analysis 33"   )%#Facilities for pathogenomic studies"$ $G$ff.    Good facilities for genomics, transcriptomics and proteomics in several universities and research institutes High level bioinformatics All facilities are also available as service E33.    Metagenomics ( $ff("   zMetagenomics of culturable and non-culturable microorganism populations present in biofilms of Acute Otitis Media (AOM, Middle Ear Infection) This infection involves a variety of bacterial species, found in the form of biofilms, which are inherently resistant to antibiotic treatment Detection of microbial biodiversity of AOM biofilm is limited due to current cultivation methods The group of Fauzi Silbaq (ArabQual) and Racheli Kreisberg-Zakarin (IBEX) racheli@ibexperts.com in involved in a study the biodiversity of AOM using a metagenomics approach, including microbiology, functional genomics and bioinformatics methodologies v P P33l33 33v3333         &     q  e  <               8   X  0Metagenomics $ff MMetagenomics of intestinal microflora in health and disease and the effect of the TLR mutations Toll-Like Receptors (TLRs) recognize pathogen specific patterns of microorganisms. Mutations in TLRs are associated with inflammatory bowel diseases (Crohn's disease and ulcerative colitis). Patients with those diseases have altered intestinal microflora Uri Gophna, currently with Ford Doolitle in Halifax ugophna@dal.ca performs culture-independent profiling of intestinal microflora of TLR knockout mice under normal conditions and after a challenge which models inflammatory bowel diseaseza Z Z ZaG33G33 G33G33     Q  G  2  W          6   l  0pLGT  lateral gene transfer in relation to pathogenesis (97$ff$8 Uri Gophna ugophna@dal.ca studies the role of lateral gene transfer (LGT) in the evolution of pathogens Uses bioinformatics for the identification of laterally acquired genes and pathways in pathogens \ DE33E33~E33E33N      0  pLGT  lateral gene transfer in relation to pathogenesis (97G$ff(8 Yair Aharonowitz, Ilya Borovok and Gerald Cohen from Tel Aviv University study gluthathione synthesis yaira@tauex.tau.ac.il Identified GshF orthologs, consisting of a -glutamylcysteine ligase (GshA) domain fused to an ATP-grasp domain, in 20 gram-positive and gram-negative bacteria. Remarkably, 95% of these bacteria are mammalian pathogens. Presumably, this fusion gene, once formed, spread between mammalian hosts most likely by horizontal gene transfer R Z8 Z2 ZG33C@        1                    q   0f{  1 Typing of bacterial strains and drug resistance 62(/$ff(1 TThe group of Chezi Kashi in the Technion kashi@techunix.technion.ac.il uses novel methods for molecular typing ofVibrio cholera in order to study emerging new pathogenic strains The group of Sima Yaron (Technion) is involved in typing the serovars of Salmonella enterica in respect to drug resistance and virulence simay@tx.technion.ac.illU qG33G33~G33G33EG33@          )  J            -     0)F 0=T  -Identification of bacterial virulence factors..G$ff- The group of Sima Yaron has developed a rapid, simple screen for real-time quantification of promoter - activity in S. enterica using a library of plasmids with GFP as a reporter The group of Gil Segal (Tel Aviv U.) GilS@tauex.tau.ac.il is involved in the study of hyper variable genes are found in the Legionella icm/dot pathogenesis region. They have demonstrated the role of these genes in virulence  tG33 G33G33G33yG33%ggQgCG33     `   H  B  V   0*&*Identification of fungal virulence factors++G$ff* The group of Nir Asherov (Tel Aviv U.) nosherov@post.tau.ac.il studies Aspergillus fumigatus, which causes serious disease (around 60% mortality) in immunocompromised people. They concentrate on studying novel cell wall proteins and identified 68 such cwp s. They are studying these CWPs in vitro and in infected animals by the use of deletion mutants. They are currently in the process of preparing a Cwp-specific microarrays to study Cwp gene expression patternsH ZGG33G33uG332            8  U    r            0'> Whole genome analysis$ff  Two genomes of bacterial pathogens are being sequenced and analyzed in Israel Bacillus anthracis (Shaefferman et al, Biological Institute, Nes Ziona) ashaefferman@iibr.gov.il Septicemic Escherichia coli  strain of serotype O78 (Ron, Tel Aviv U.) eliora@post.tau.ac.il N   NE33G 33\G 33G 33G 33G 33 G  33G 33G 33W                *       0  0Virulent E. coli strainsF G$ffG$ff G$ff Most of the E. coli strains are commensal, but a small number are pathogenic Pathogenic E. coli strains are divided into two groups: Intestinal strains. These produce enterotoxins and constitute a major problem, especially in young children and travellers (Montesumu s revenge) Extraintestinal strains  ExPEC (Extraintestinal Pathogenic E. coli)  Z Z Z G33G33FG33G33&G33E33E33E33G33G33       B               Septicemia (colibacillosis)G$ff.    Colisepticemia is the major causes of mortality from community and hospital-acquired infections (more than 80%) Main cause of mortality in immuno-supressed patients (HIV, chemotherapy, old age) Colisepticemia is an emerging disease  83% increase 1980  1992, over 40% of the bacteremia cases in community acquired infectionsG E33e33e33e33e33e33 ~     '  D   ' Goals:6G$ffg ff Define virulence-essential ExPEC-specific genes Profile strains involved in UTI, NBM and sepsis using these ExPEC-specific genes Use the data to define potential targets for development of vaccines and/or antibacterial drugs.| /E33E33QE33_e33 e 33 E 33v    <  o  CIdentification of virulence related sequences in septicemic strainsDDG$ff.1    3Whole genome sequencing Subtractive hybridization >3  3E33E33 4 Subtractive hybridization  G$ff  Obtain pathogen specific sequences, absent from non-pathogenic K12 strain Excellent chance of  hitting pathogenicity islands which are pathogen specific and very large Faster (and much cheaper) than whole genome sequencing E33  Subtractive hybridization2 G$f3G$f3  QA way to study comparative genomics with organisms which have not been sequenced .R PE33E33 R PSearch for unique  septicemic sequences))G f32      PUsing suppression subtractive hybridization (SSH) we identified sequences unique to strain O78-9 and absent from the non-pathogenic strain K-12 Oover 80 O78-specific open reading frames were found (91 to 1473 bp in length) The same experiment was repeated with another septicemic strain O2-1772 117 unique O2 sequences were identified Q g6GgIg'ggZ  <  ;   9 ,Comparison of unique sequences of O2 and O78--G$f3,  ^Although the two strains cause the same disease, there is a high diversity between the SSH libraries of O2 and O78 strains, with only a few shared genes coding for virulence factors. Is this diversity serotype specific? To determine this we profiled additional septicemic strains of the same serotypes the presence of each of the unique sequences l_ Z|e9eeEe:   L   EComparison of unique sequences of two septicemic strains - O2 and O78FFG f32&     high level of genome plasticity there is a high diversity between the septicemic strainsunexpected for strains causing the same disease Septicemic strains of serogroups O2 and O78 contain a large pool of virulence genes which are used in a  mix and match fashionD  Ye0e33e##tG     &       ` #Current-future researchG$ff $Molecular-physiological studies of the newly-identified virulence genes Bioinformatic studies to determine the evolution of these virulence genes (many in PAIs with evidence of LGT) Whole genome sequencing  joint project with Prof. Joerg Hacker and Prof. Gerhard Gottschalk: E3333E33bH   F  J  $ ($ Thank you!!"  G$ff  TAU group Uri Gophna Diana Ideses Daphna Mokady Dr. Dvora Biran Collaborations Wuerzburg University (Prof. Joerg Hacker) Greifswald University (Prof. Michael Hecker) Goetingen University (Prof. Gerhard Gottschalk)   Z7 Z Z Z G337G33G33sG3333*                          ( "Metagenomics ( Uri Gophna, currently with Ford Doolitle in Halifax studies the effect of TLR mutations on micrflora in health and disease by culture-independent profiling of intestinal microflora of TLR knockout mice under normal conditions and after a challenge which models inflammatory bowel diseaseH  0@|    3   F   k MLST of O78 strains" G$f3  Multi Locus Sequence Typing 450  500 bp of 7  housekeeping genes Criteria for chosing genes: 97-98% homology to E. coli K-12 (from blast data) appear in pathogenic and non pathogenic strains map at considerable distance from each other several allels in the populationf_ Z Z_EEEEZ&  (       ! [The MLST results are compatible with the results from subtractive hybridization and sequencing The profile of virulence factors in ExPEC strains is independent of the host and independent of the serotype There is a high diversity of virulence genes in the various E. coli septicemic strains and each strain has its own profile of virulence genesP\ AE##=AAKA@     @ !"#  Both libraries contain many sequences associated with genomic plasticity - evolution by horizontal gene transfer Many sequences of O2 and O78 are homologous to virulence related sequences of human ExPEC strains ns  c  tecEA&      /$%&'}  04$(  4r 4 S $H: `  : r 4 S H:0y : H 4 0޽h ? 33___PPT10i.\@+D=' = @B +rT" 5 0"*c(    N:kashi@techunix.technion.ac.il>*racheli@ibexperts.com0ugophna@dal.ca0ugophna@dal.ca>*yaira@tauex.tau.ac.ilB.simay@tx.technion.ac.il> *eliora@post.tau.ac.ilD 0ashaefferman@iibr.gov.ilB .nosherov@post.tau.ac.il< (GilS@tauex.tau.ac.il/ 00DArialNew Rot$t\ )0tY 0h"DComic Sans MS$t\ )0tY 0hB DMS Sans Serif$t\ )0tY 00DTimes New Romant\ )0tY 0hc .  @n?" dd@  @@``  6) ~ n ($%&'  #! " )_b$ ov飼kf#A2$%Gg_gDj#A2$nI8i v o&A2$0h,m5A @,7Ab$L-s$֠n:A 0AA ff33@@K2ʚ;n8ʚ;g4BdBd )0hppp@ <42d2d 0$t<4BdBd 0$t( <4!d!dy 0$0___PPT10 ppR___PPT94,&@ &@L@@4@@?  %{W@Pathogenomics in Israel Eliora Z. Ron @A$ff& ff$ff             Metagenomics LGT = lateral gene transfer Typing of bacterial strains and drug resistance Identification of virulence factors Whole genome analysis 33"   )%#Facilities for pathogenomic studies"$ $G$ff.    Good facilities for genomics, transcriptomics and proteomics in several universities and research institutes High level bioinformatics All facilities are also available as service E33.    Metagenomics ( $ff("   zMetagenomics of culturable and non-culturable microorganism populations present in biofilms of Acute Otitis Media (AOM, Middle Ear Infection) This infection involves a variety of bacterial species, found in the form of biofilms, which are inherently resistant to antibiotic treatment Detection of microbial biodiversity of AOM biofilm is limited due to current cultivation methods The group of Fauzi Silbaq (ArabQual) and Racheli Kreisberg-Zakarin (IBEX) racheli@ibexperts.com in involved in a study the biodiversity of AOM using a metagenomics approach, including microbiology, functional genomics and bioinformatics methodologies v P P33l33 33v3333         &     q  e  <               8   X  0Metagenomics $ff MMetagenomics of intestinal microflora in health and disease and the effect of the TLR mutations Toll-Like Receptors (TLRs) recognize pathogen specific patterns of microorganisms. Mutations in TLRs are associated with inflammatory bowel diseases (Crohn's disease and ulcerative colitis). Patients with those diseases have altered intestinal microflora Uri Gophna, currently with Ford Doolitle in Halifax ugophna@dal.ca performs culture-independent profiling of intestinal microflora of TLR knockout mice under normal conditions and after a challenge which models inflammatory bowel diseaseza Z Z ZaG33G33 G33G33     Q  G  2  W          6   l  0pLGT  lateral gene transfer in relation to pathogenesis (97$ff$8 Uri Gophna ugophna@dal.ca studies the role of lateral gene transfer (LGT) in the evolution of pathogens Uses bioinformatics for the identification of laterally acquired genes and pathways in pathogens \ DE33E33~E33E33N      0  pLGT  lateral gene transfer in relation to pathogenesis (97G$ff(8 Yair Aharonowitz, Ilya Borovok and Gerald Cohen from Tel Aviv University study gluthathione synthesis yaira@tauex.tau.ac.il Identified GshF orthologs, consisting of a -glutamylcysteine ligase (GshA) domain fused to an ATP-grasp domain, in 20 gram-positive and gram-negative bacteria. Remarkably, 95% of these bacteria are mammalian pathogens. Presumably, this fusion gene, once formed, spread between mammalian hosts most likely by horizontal gene transfer R Z8 Z2 ZG33C@        1                    q   0f{  1 Typing of bacterial strains and drug resistance 62(/$ff(1 TThe group of Chezi Kashi in the Technion kashi@techunix.technion.ac.il uses novel methods for molecular typing ofVibrio cholera in order to study emerging new pathogenic strains The group of Sima Yaron (Technion) is involved in typing the serovars of Salmonella enterica in respect to drug resistance and virulence simay@tx.technion.ac.illU qG33G33~G33G33EG33@          )  J            -     0)F 0=T  -Identification of bacterial virulence factors..G$ff- The group of Sima Yaron has developed a rapid, simple screen for real-time quantification of promoter - activity in S. enterica using a library of plasmids with GFP as a reporter The group of Gil Segal (Tel Aviv U.) GilS@tauex.tau.ac.il is involved in the study of hyper variable genes are found in the Legionella icm/dot pathogenesis region. They have demonstrated the role of these genes in virulence  tG33 G33G33G33yG33%ggQgCG33     `   H  B  V   0*&*Identification of fungal virulence factors++G$ff* The group of Nir Asherov (Tel Aviv U.) nosherov@post.tau.ac.il studies Aspergillus fumigatus, which causes serious disease (around 60% mortality) in immunocompromised people. They concentrate on studying novel cell wall proteins and identified 68 such cwp s. They are studying these CWPs in vitro and in infected animals by the use of deletion mutants. They are currently in the process of preparing a Cwp-specific microarrays to study Cwp gene expression patternsH ZGG33G33uG332            8  U    r            0'> Whole genome analysis$ff  Two genomes of bacterial pathogens are being sequenced and analyzed in Israel Bacillus anthracis (Shaefferman et al, Biological Institute, Nes Ziona) ashaefferman@iibr.gov.il Septicemic Escherichia coli  strain of serotype O78 (Ron, Tel Aviv U.) eliora@post.tau.ac.il N   NE33G 33\G 33G 33G 33G 33 G  33G 33G 33W                *       0  0Virulent E. coli strainsF G$ffG$ff G$ff Most of the E. coli strains are commensal, but a small number are pathogenic Pathogenic E. coli strains are divided into two groups: Intestinal strains. These produce enterotoxins and constitute a major problem, especially in young children and travellers (Montesumu s revenge) Extraintestinal strains  ExPEC (Extraintestinal Pathogenic E. coli)  Z Z Z G33G33FG33G33&G33E33E33E33G33G33       B               Septicemia (colibacillosis)G$ff.    Colisepticemia is the major causes of mortality from community and hospital-acquired infections (more than 80%) Main cause of mortality in immuno-supressed patients (HIV, chemotherapy, old age) Colisepticemia is an emerging disease  83% increase 1980  1992, over 40% of the bacteremia cases in community acquired infectionsG E33e33e33e33e33e33 ~     '  D   ' Goals:6G$ffg ff Define virulence-essential ExPEC-specific genes Profile strains involved in UTI, NBM and sepsis using these ExPEC-specific genes Use the data to define potential targets for development of vaccines and/or antibacterial drugs.| /E33E33QE33_e33 e 33 E 33v    <  o  CIdentification of virulence related sequences in septicemic strainsDDG$ff.1    3Whole genome sequencing Subtractive hybridization >3  3E33E33 4 Subtractive hybridization  G$ff  Obtain pathogen specific sequences, absent from non-pathogenic K12 strain Excellent chance of  hitting pathogenicity islands which are pathogen specific and very large Faster (and much cheaper) than whole genome sequencing E33  Subtractive hybridization2 G$f3G$f3  QA way to study comparative genomics with organisms which have not been sequenced .R PE33E33 R PSearch for unique  septicemic sequences))G f32      PUsing suppression subtractive hybridization (SSH) we identified sequences unique to strain O78-9 and absent from the non-pathogenic strain K-12 Oover 80 O78-specific open reading frames were found (91 to 1473 bp in length) The same experiment was repeated with another septicemic strain O2-1772 117 unique O2 sequences were identified Q g6GgIg'ggZ  <  ;   9 ,Comparison of unique sequences of O2 and O78--G$f3,  ^Although the two strains cause the same disease, there is a high diversity between the SSH libraries of O2 and O78 strains, with only a few shared genes coding for virulence factors. Is this diversity serotype specific? To determine this we profiled additional septicemic strains of the same serotypes the presence of each of the unique sequences l_ Z|e9eeEe:   L   EComparison of unique sequences of two septicemic strains - O2 and O78FFG f32&     high level of genome plasticity there is a high diversity between the septicemic strainsunexpected for strains causing the same disease Septicemic strains of serogroups O2 and O78 contain a large pool of virulence genes which are used in a  mix and match fashionD  Ye0e33e##tG     &       ` #Current-future researchG$ff $Molecular-physiological studies of the newly-identified virulence genes Bioinformatic studies to determine the evolution of these virulence genes (many in PAIs with evidence of LGT) Whole genome sequencing  joint project with Prof. Joerg Hacker and Prof. Gerhard Gottschalk: E3333E33bH   F  J  $ ($ Thank you!!"  G$ff  TAU group Uri Gophna Diana Ideses Daphna Mokady Dr. Dvora Biran Collaborations Wuerzburg University (Prof. Joerg Hacker) Greifswald University (Prof. Michael Hecker) Goetingen University (Prof. Gerhard Gottschalk)   Z7 Z Z Z G337G33G33sG3333*                          ( "Metagenomics ( Uri Gophna, currently with Ford Doolitle in Halifax studies the effect of TLR mutations on micrflora in health and disease by culture-independent profiling of intestinal microflora of TLR knockout mice under normal conditions and after a challenge which models inflammatory bowel diseaseH  0@|    3   F   k MLST of O78 strains" G$f3  Multi Locus Sequence Typing 450  500 bp of 7  housekeeping genes Criteria for chosing genes: 97-98% homology to E. coli K-12 (from blast data) appear in pathogenic and non pathogenic strains map at considerable distance from each other several allels in the populationf_ Z Z_EEEEZ&  (       ! [The MLST results are compatible with the results from subtractive hybridization and sequencing The profile of virulence factors in ExPEC strains is independent of the host and independent of the serotype There is a high diversity of virulence genes in the various E. coli septicemic strains and each strain has its own profile of virulence genesP\ AE##=AAKA@     @ !"#  Both libraries contain many sequences associated with genomic plasticity - evolution by horizontal gene transfer Many sequences of O2 and O78 are homologous to virulence related sequences of human ExPEC strains ns  c  tecEA&      /$%&'0  0 0(   x  c $@   x  c $`0s  H  0޽h ? 3380___PPT10.\@ r҇*Oh+'0x `h  Pathogenomics in Israel eliora ronc Eliora Ronc67oMicrosoft PowerPointel@/@\@U2UoGhg  ?& &&#TNPP\2OMi & TNPP &&TNPP    --- !---&&j}w@  }ww0- &Gy& --@-- @BComic Sans MS b}ww0- ff.2 B Pathogenomics% . ff.2 B= in Israel .@BComic Sans MS }ww0- ff.2 dEliora . ff.2 Z. Ron <. ff.2 eliora . ff.2 (@post.(. ff. 2 taub. ff. 2 .ac.. ff. 2 Nil . ff. 2 e>l.--(-- ff@"Arialw@ }ww0- 33. 2 =2l.@BComic Sans MS }ww0- 33.2 =V Metagenomics&! .@"Arialw@ }ww0- 33. 2 z2l.@BComic Sans MS }ww0- 33.02 zVLGT = lateral gene transferx    .@"Arialw@ }ww0- 33. 2 2l.@BComic Sans MS }ww0- 33.?2 V%Typing of bacterial strains and drug          . 33.2 V resistance .@"Arialw@ }ww0- 33. 2 (2l.@BComic Sans MS }ww0- 33.=2 (V$Identification of virulence factors          .@"Arialw@ }ww0- 33. 2 f2l.@BComic Sans MS }ww0- 33.'2 fVWhole genome analysis-  !   .--"System 0-&TNPP &Root EntrydO)x PicturesCurrent User/SummaryInformation(89:;<=>?@ABCDEFGHIJKLMNOPQRSTUVWXZ[\]^_`abcde_rothmrothmora Ronh  Pathogenomics in Israel eliora ronc Eliora Ronc67oMicrosoft PowerPointel@/@\@U2UoGhg  '?& &&#TNPP\2OMi & TNPP &&TNPP