{"paper_id":"61bddc6a-ce63-4cb2-97cf-5064823df02a","body_text":"Hybrid short- and long-read assembly of an experimentally evolved Sodalis 1 \nglossinidius strain SgGmmC1* 2 \n 3 \nPoppy Pes c o d 1, 2* , L ee R . H aines 2,4* , Al istair C D arby 3 , Ian Goodhead 1# 4 \n 5 \n1 School of Sc ience, En gineering and E nvironmen t, Univ er s it y  of  S alfo rd, Sal ford, UK 6 \n2 Depar t ment  o f Vector  Bio logy, Liverpool School of Tr opic al Medic ine, Live rpool, UK 7 \n3  Institu te o f Integr ative Biol ogy, U niv ersity of Liv er pool, Li v er pool, U K 8 \n4  Cur rent  a d dr ess : Universi t y of N ot re D a me, N ot re Dame, IN, U SA .  9 \n 10 \n*  These author s  contr ibuted equal ly to t his  work.  11 \n 12 \n# Addr es s corresponden ce to Ian Goo dhead, i.b.goodh e ad @sal f ord.ac.u k  13 \n 14 \n 15 \nABSTRA CT 16 \nBacterial symbionts of inse c t s  under g o dramatic genome r educ t ion dur ing t heir 17 \nevoluti onar y  t ransition fr o m fre e- l i vin g to  h os t -dependent  l if e styles, but th e d y nami c s of 18 \nthis proces s  r emain poorly under s to od due to th e  d i f f iculty of observing these proce sses in 19 \nreal-time. So dali s gloss inidius, a  f ac ul t a t iv e endos ymbion t of tsetse flies , pr ovi d es  an 20 \nexc ep tional opport un ity to study t his  tr ans it i o n ex perimental ly: U n l ike high ly s pe cialis ed 21 \nobligate sy mbionts, S. glo s si nidius  ca n be cult ured in vi tro  and r e tains a large genome  ( 4 22 \nMbp ) wi t h extensi ve pseudogene content  (49%), s u gge sting a recent evolution a r y 23 \ntr ans it i o n. He re, we pr es ent  a compar ative g en omic  analysi s  of S. gloss inidius  s tr ains  24 \nis o l at ed from l abo rato r y colony-d erived G los s ina mor s it ans  mor s it ans , co mparing one aft e r 25 \nten years of serial passa ging i n  labor ator y  cultur e (SgG mmC 1*) to a counter part isolated at 26 \nthe same time (SgGmmB4).  H ybrid genome ass embly  u sin g O xfo rd N ano pore and Illumina 27 \ntechnologies produced a high-quality 4.29 Mbp genome comprisin g one c ir c u l ar  28 \nchromo s ome and f our plasmids  similar. Comp a r ative analy sis r evea led a si gnific ant  deletion 29 \n(17,209 bp)  c ont aining 31 g en es, in cluding thiM  (inv olved in thiam ine bios ynthesi s) and 30 \ngenes en coding sulfur t ransport ers. Additional ly, we identified multiple small-sca le 31 \n.CC-BY 4.0 International licenseavailable under a \n(which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made \nThe copyright holder for this preprintthis version posted June 27, 2025. ; https://doi.org/10.1101/2025.06.26.659281doi: bioRxiv preprint \n\nmut a t ions ( 8 delet ions, 39 i n s er tions,  10 S NP s )  resulting in frameshifts in g enes  including a 32 \nhemolysin precur sor ( shlA ).  Our  f indi ng s  demo ns t rate th a t , under stable la borat ory 33 \nconditions without  t he s elective pressures of the host environment, S. gl os sinidius  cont i n ues  34 \nto und ergo genome degradat ion. The  los s of thiM  s upp orts previous hypot hese s  of  35 \ncomplement a r y metabol ic pathway s  bet w een  S. gloss inidius  and the pr i ma ry symbion t 36 \nW ig g le s w o r t h i a  g lo s s i n i d ia  f or thiami n e b i o s ynth es i s. Th i s stud y provides ins ight s into t he 37 \nevoluti onar y  t rajectory of  f ac ult ative symbiont s  and has implic at ions  fo r parat ransg en i c 38 \nappro a ches u s ing S. glo ssinidius  f or trypano s o me c on trol. 39 \n 40 \nKEY WO RD S: S od al is glossinidius , t s et se s y mbiont, s y mbios is , ex p erimental evoluti on , who l e-41 \ngenome sequencing  42 \n 43 \nDA TA SU M MAR Y 44 \nRaw N ano pore and MiSeq r ea d s  are available i n  t he European Nucleotide A r chiv e und e r  45 \nPro je ct  P RJEB323 21 (R e ad  ac ce ssions : ER X332 1202 and ERX332 1201 r espe ctively). A full 46 \nGBK forma t ted annot a tion o f SgGmmC1* has b een depos it ed in F ig share at 47 \nhtt ps : / /doi.org/1 0.178 66/ r d. sa lf ord.8052437.v 1 . 48 \n 49 \nIMP ACT S TATEME NT  50 \nIn this s t ud y we p rovide new i n s ights into t he ongoing ge n ome degradatio n  o f S odali s  51 \ngloss inidius, a  f acult a t iv e end os ymbiont  of tsetse flies  impl icated in thei r  ab ili t y t o t r a n s mit  52 \ntr ypanos omia si s, d uring prolon g ed  la bor ator y  cultur e. Through high- quali ty c omparative 53 \ngenomics we reveal si gn i f ic ant  gene los s and  mutational changes, including disrupt i o n of 54 \npathway s i n v olved in nut r ient b iosynt hesis. The se findings en han c e our  understanding of  55 \ns y mbiont genome evolu tion  and  inf or m t he dev el o pmen t of S. gloss inidius  as  a poten tial 56 \ntool for parat ransge n i c strat egi es in c ontro l l ing t rypanosome transmis s ion.  57 \n  58 \n.CC-BY 4.0 International licenseavailable under a \n(which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made \nThe copyright holder for this preprintthis version posted June 27, 2025. ; https://doi.org/10.1101/2025.06.26.659281doi: bioRxiv preprint \n\nIN TRO D U C TI O N  59 \nTset s e f l ies (Genu s : Glossina) are unu s ual insect s  that , li ke keds and bat flies , r eprodu c e b y 60 \ngivi n g b i r th to  l ive young (adenot rop hic viv ipar ity). As obligate haemat ophag es , tset se 61 \nharbo urs a limit ed bac t er ial microbi o me due t o their r es t ricted diet, f eed i ng solely on blood 62 \nin the adul t  li f e stage and on pr oteina ceous mil k fr om the femal e ’ s  spec ialised a c ce ssor y  63 \nglands d uring the larval s t age (Minchi n, 1905). At  l east fo ur spec ies of ba cteria play 64 \nimpor tan t r oles  in the  nu tri t i o n, fecu ndity and vec t orial ca p a city of  t s et s e flies: 65 \nW ig g le s w o r t h i a  g lo s s i n i d ia , an ob l igat e p rima ry s ymbiont is ubiquitous;  Sodalis  gloss inidius , 66 \na facultative commensal symbiont , and  Wolbachia  and S pi r op lasma, paras i t ic  r eproduct i ve 67 \nmanipulato rs, all have va ried in fectio n r a t es  acro ss t he wide dis tr ibution o f  ts et s e acro s s 68 \nsub-Saharan Af rica  ( Doudou mis  et al , 2017) . S.  glos s inidius  is  a G r am-negative sec on dar y 69 \nbacterial endosymbiont t hat has been l inked t o mod ifying tsetse fly su s c e pt ibil i t y to 70 \ntr ypanos ome i n fec t ion by parasi t es  b elonging to  t he genus  Tr ypanos om a  ( Trappeniers et al , 71 \n2019; Makhulu e t  a l,  2021 ; K al lu et  al, 2023) . Trypano somes  cau se a fat al d i sease in hu man s  72 \nand animals called tr y p a n os omi a sis  ( Maudlin and Ellis , 1985).  73 \n 74 \nOblig at e i n s e ct endos ymbiont  geno mes  demo ns t rate spe c ialisation to t he ir  host 75 \nenvironme nt by being non culture- v ia ble and having s ma ll, compact genomes relative to 76 \nfr ee- liv ing r el a t iv es (Moran e t al., 2008). S. gloss inidius  i s notable fo r being the fir s t 77 \nendosymbiont dis covered to be bo t h amenable to in vitro culture (Dale & Maudlin, 1999)  78 \nand for  hav ing a rela t i vely large geno me: The S. gloss inidius geno me is  4 Mbp , similar in siz e 79 \nto fr ee -li ving bac t eria, but large in comparison to  t he 0.7 Mbp genome of  W . glos s inidia ; 80 \nboth  o f th ese featur es  sug gest a r ec ent  s witch to ho s t a s s ociation and s ymbiosis  (Toh et  a l., 81 \n2006).  82 \n 83 \nThe tr ans ition  o f bac t erial  genomes from f ree -liv ing to endosymbiosis  i s  po orly understood . 84 \nEvidence su g ges t s  endo symbiont  ev olut ion i nvolv es inactivation of funct i onal genes via 85 \nsmall  insert ions  or  deleti ons to pro du ce “p seudogenes ” . The se p s eud ogen es  are 86 \nsub s equ ently l o s t  in lar g e delet ion e v e n ts f ac ilitat ed by  f requent  populat io n bot tlen eck s  87 \n(Goodhead and D a r by, 2 015). As mut ation s  ac c u mulate in non-esse n tial g en es , fr ames hif ts  88 \nand large number s  of  i n s er tions o r de letions (indels) result in los s of gene funct i o n i n  a 89 \n.CC-BY 4.0 International licenseavailable under a \n(which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made \nThe copyright holder for this preprintthis version posted June 27, 2025. ; https://doi.org/10.1101/2025.06.26.659281doi: bioRxiv preprint \n\npro c es s  known a s p s eudo g en i sation; pseudogene s may or may not r etain f unction 90 \ndepending on t he l e vel o f degr adatio n (Goodhead et al, 2020). 91 \n 92 \nS. glos sinidius  r epresents a  un i q ue mid-way point in the ev oluti on a r y transition t owa r ds 93 \nobligate endosymbiosi s , evidenced b y t he high propor tion o f pseudogenes  in its geno me: 94 \n49% compared to  t he usual  1% pr es ent  i n  f r ee-l iving bac t eria (Goodhead e t al., 2017; Toh et 95 \nal., 2006). Ho w ever , w h i lst th e o bli gat e en dos ymb i o nt W . glos si nidia  is pre sent in all 96 \nindividuals  of  t h e G los s ina genus , S. glossinidius  is found at r ates  r anging f rom 1% - 94% in 97 \nwild tsetse  po pulations ( Farikou et al 2011; Den nis  et al, 201 4; Mf opit et al ,  2023). Thi s lack 98 \nof consi s t e n cy implies S . glos s inidius retains a facultative re lationship with i ts host and 99 \nimpacts our ability to study ba cter i al genome degr adation in natu r al s ett in g s . Indeed,  100 \ngenomic informati on for  S . glossi nidi us  ha s  lar ge ly been collected fro m is olates  derived f r om 101 \nGl o s sina  u nder labo rat ory conditions (Toh et al., 2006; Belda et  al., 2010; G oodhead et al., 102 \n2017).  103 \n 104 \nS. glos sinidius  ha s s everal att ribut es  t h a t  make it  a p rom ising mic r oorganis m for  u s e in 105 \ntsetse and trypano some contr ol s t rategies – i t  is  amenable to cultur e in v itro , has a 106 \ndemonstr able a b i l it y t o inf ect the t a rget vector, and can be genetically  man i p ula t ed t o 107 \nexpress  contr ol factors (Bear d et al, 1 993; W elbur n et al, 1987). A r ec o mbinant strain of S. 108 \ngloss inidius ex pr essing fun ctional anti -t rypanosome Nanobodies ®  ha s  been used to deliver 109 \nthe Nanobodies to tsetse in an in  v i v o  lab-based proof  o f  c o nc ept  s t ud y ( De Voogh t et al  110 \n2012). Whil s t  promising, the effector  molecule w as only s t able wit hin the t setse ho s t  when 111 \nits native S.  glossi n i dius  population  w as s ignificantly suppre s s ed. Thi s s u g g ests fitnes s  112 \ndifferences  bet w een nat iv e and lab- reared strains of th e b a ct e r ium, mean i ng fur ther  stud i es 113 \ninto t he fitn ess  of  l ab- derived S. g los s inidi us  versu s n atural populat i ons  ar e war ranted. A  114 \nthor ough under s t anding of the pathw ay of genome d eg r adation in lab-ba sed s y stems wi ll 115 \nals o  b e  vital fo r  paratr ans genic appro a che s, to pr edic t  i mpac t s  on ba cter ia l  f i t nes s or t heir 116 \nability to succes s f ull y infect t he tsetse ho s t . 117 \n 118 \nHer e we p rese n t a high-qua lity whole  genome sequence a s se mbly and  an n otation o f  a 119 \nstrain of S. gloss inidius  i solated f rom a lab colony o f G lo s s ina mor s it ans  mo r s it ans  th a t  w as  120 \nserially  pas saged in vitr o f or t en  year s. We h av e compar ed i t s  genome t o th at  o f a 121 \n.CC-BY 4.0 International licenseavailable under a \n(which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made \nThe copyright holder for this preprintthis version posted June 27, 2025. ; https://doi.org/10.1101/2025.06.26.659281doi: bioRxiv preprint \n\ncounter part isolated at t he s ame time and  f rom t he s ame colony a s  t he original s t rain, with 122 \nthe aim o f  d i r ectly obs er vi n g  genome r eduction under  la b o rator y s et tings . We identify both  123 \nsmall  and  lar ge-s cale deletions as  well as  smaller scale mutat ions , pr oduce a b etter  124 \nunder s t a n ding of pathway s  und er s el ection in labor at ory settings , and hyp othesis  t he 125 \nimpact on natu ral S . g lossini d i us populat i o ns . 126 \n 127 \nMETH OD S  128 \nIn vitro cultur e of S . gl ossini d i us  129 \nS. glos sinidius  wa s i solated fr om two male G. m. mor sitans from t he  colony at  the Liv er pool 130 \nSchool of Tropical Medicine in 2007 following previously  des cribed met ho ds ( Matt hew et  131 \nal., 2005). The strains w er e named Sg G mmC1 and Sg GmmB4,  and  wer e i mmediately s t ored 132 \nat -80° C. The SgGmmB 4 i solate was s equenced following a s ingle pas s a ge fro m a glycerol 133 \nstoc k a s previously d es cribed using P ac ific Bios c ience s RSII sequen cing ( Goodhead et al, 134 \n2020). Beginning in 2009 , cultur es  of  Sg GmmC1 were incubated at  27°C in a microaeroph i l ic 135 \natmospher e a n d pas s aged ev er y 1 0-1 4 day s  for  ten years, al t ernating bet wee n  solid-phase 136 \ngrowt h on Columbia agar base supple ment ed with d efibrinated ho r s e bloo d (Oxoid/Thermo 137 \nFis her ) and liquid-phase growth in serum-fre e insect media (Sigma Aldr ich). We here ref er 138 \nto t he “ evo lved” s t rain of  Soda lis  gloss inidius  as  “ Sg G mm C 1*” .  139 \n 140 \nCompar ative genome an alys is  141 \nDNA wa s e xt racted fro m a cell pel let of SgGmmC 1* usin g the Zymo Q u ick DNA kit and 142 \nquant i f ie d us ing a Qubit™ 3.0. The S gGmmC1 * geno me w a s s equ enc ed u si ng both Oxford  143 \nNanopore MinION and Illumina MiSeq platforms. A library for  the MinI O N wa s generat ed 144 \nusing the SQ K- L S K108 1D  l igat i o n k it  a n d s equenced on a R9.4 flow cell, producing 14 9,84 7 145 \nreads of mean length 3,47 7.62bp and  length N50 of 14,4 68bp. A t otal of 52 1,111 ,234 ba se s  146 \nwere sequenced giv ing an e stimated 118x genome coverage. For Illumina MiSeq 147 \nsequenci n g , a QIA seq FX D NA library pr ep k it  a n d v 2 150b p s equencin g c artr idge wer e us ed 148 \npro duc ing a to tal of 755,405 paired- e nd reads. MiSeq reads were a ssessed and adaptor s  149 \ntr imm ed using F a stQC (v0.1 1.4) (A ndrews, 2010). MiSeq read s were tr imm ed for  qual ity 150 \nusing fa s t p w it h default settings (v0.12.5) (Chen  et  al . , 20 18).  151 \n 152 \n.CC-BY 4.0 International licenseavailable under a \n(which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made \nThe copyright holder for this preprintthis version posted June 27, 2025. ; https://doi.org/10.1101/2025.06.26.659281doi: bioRxiv preprint \n\nA hybr i d  geno me for SgGmmC1* wa s  built  u s ing Unicycler (v0.4.8-bet a) (Wi c k et al., 2017) 153 \nby a ssembling the Mi Seq r eads  de no vo and mapping th e lon g  MinIO N rea ds onto t his  154 \nassembly for  s tr uctur a l corr ection. Th e hybrid as s embly wa s ann otated usi ng PR OK KA 155 \n(v1.13.3) (Seeman n, 2014) u s ing t he referen c e strain SgGmmB4 genome (G enBank 156 \nAcce ssion LN 8 54557 .1) , whi ch wa s  de po s ited in 2016. The annot ated Sg G mmC1* assembly 157 \nwas  compar ed to Sg G mmB4 us ing t h e Artemis  Comparis o n Tool (ACT) (v13.0.0)  (Carver et  158 \nal., 2005) to ident ify s tr uctur a l chang es and  lar ge d el et ions , and  Snippy (v3 .1) ( S eemann, 159 \n2017) t o i d entify s ingle-n uc leot ide po lymorphisms (S NP s) and insertions/d el et ions. A n 160 \nassembly graph of  t he as sembly w as  pr oduced us ing Banda ge (v0.8.1) (Wi c k  et  a l., 2015)  to 161 \nasse s s  its quality (Dat a not shown).  162 \n 163 \nRESUL T S  164 \nCompar ative genomic s of SgGmmC1*  and SgG mm B4 165 \nThe assembled Sg G mmC1* genome was  4 ,28 8,97 5b p i n  size wit h 54. 43% GC and no 166 \nunident i fied bas e pairs (N s ) . Th e a s s e mbly  con sis t ed of s ix contig s: one c ir cular  c hr omosome 167 \n(4,149, 326b p), four circular p l asmid s (pSG 1 , 81, 522bp ; pSG 2, 26 ,425bp; p S G3 , 19,5 12bp; 168 \nand pSG4 / 3, 1 0,81 6bp)  and  o ne pla s mid fr ag ment (pSG2 pla s mid fr ag, 1,3 74bp) were 169 \npro duc ed . Sg GmmC1* w as iden tified with 99.99% si mil ar ity to the r efe rence strain 170 \nSg G mmB4 us in g BLAST N (v2.8.1) (Fig ure 1) (Zhang et  al., 2000) . One large deletion o f 17, 171 \n209bp wa s iden tified, containing 13 c oding g en es  and 1 8 non-coding gene s  (Table 1). This  172 \ndeleted regi o n i n c luded t he c o ding gene th i M , which enc od es  a prot ein inv olv ed in t hia mine  173 \nbiosynthe sis  and one of t wo c opies of  esar that  encode s a prot ei n  r e lated t o t he quorum-174 \nsen s ing pr otein Lux R . Fur ther  d e le t ed  genes  in cluded th ree s ulf ur and one  sulfite 175 \ntr ans membrane tr ans po rter  protei n s ; one copy of th e p e p tidas e yxeP  (the ot her copy of 176 \nwhich i s p seudogeni s ed elsewhere in the genome); and mul tipl e sugar pho sphotr ans f erase s  177 \n(Figure 1 and Table 1). 178 \n 179 \nSnippy als o ident ified ei ght  s ma ll del e t i o ns  (r ange of 1-27bp, mean 11.6b p, s tand a r d 180 \ndeviation 6.08), 39 insert i o ns  (1- 8bp, mean 3.1bp, std 4. 72) and 10 S N P s . I n to tal, 12/ 32 181 \n(37.5%) insert i o ns  in a codin g region resulted in a frames hift ; Putati vely pseudogenis ed 182 \ngenes in cluded t he putati ve tr ans por t  pr otein gene HsrA , one of fiv e c op i es of th e p r oline 183 \ntr ans po rter  gene pr op and a he molys in pr ec u rsor shl A  (Table 2).  184 \n.CC-BY 4.0 International licenseavailable under a \n(which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made \nThe copyright holder for this preprintthis version posted June 27, 2025. ; https://doi.org/10.1101/2025.06.26.659281doi: bioRxiv preprint \n\n 185 \nRaw N ano pore and MiSeq r eads  are available f rom the European Nucleotide Ar c hive under  186 \nPro je ct  P RJEB323 21 (R e ad  ac ce ssions : ER X332 1202 and ERX332 1201 r espe ctively). A full 187 \nGBK forma t ted annot a tion o f SgGmmC1* has been depos it ed in F ig share at 188 \nhtt ps : / /doi.org/1 0.178 66/ r d. sa lf ord.8052437.v 1 . 189 \n 190 \nDISCU SSI ON  191 \nThe pseudogeni s at ion and deletion of  genes de scribed  suppor t the hypo thesi s of s ymbiont 192 \ngenome siz e r educ t ion thr ough ps eu d ogenis ation and  sub sequent delet i on of genes  193 \nunneces s ar y to  sur v ival in a s table host enviro nment. In the c ase of Sg G mmC 1*, 10 year s  of 194 \npas s a ging in a minimal med ia w it h c ar efully cont rolled  tempe ratur e and a t mospher e  has 195 \nresulted in inactivati on or  de leti on of genes unne cessar y to th i s environment. W it hin the 17 196 \nkbp deletion most coding genes a n d ps eudo g en es were involved in transmembr ane 197 \ntr ans po rt of or ganic  and inor ganic  co mpou nds , mainly of carboh y d rate s, o r were 198 \nmembr ane-b ound  t ransferases .  In  t s et s e, a s i n  o ther or ganis ms, thiamine is a n  es sential 199 \ncofactor f or amino acid and c ar bohyd rate metab olis m that is not present i n bloo d meals . 200 \nThe tsetse fly l a ck s  t he c apa city for  t h iamine biosy n thesi s but  carr ies  gene s  for thia mine 201 \ntr ans po rter s  (Int ernation a l  Glossina G en ome Initiative, 2012). W. glossinidia and  S. 202 \ngloss inidius c ont ribut e metabol ical ly complement ary cofac t ors to pr oduce an  int ac t  203 \nthiamine b iosynthesi s  pat hwa y : W. glossinidia has  t he c apacity for  B v it ami n bios ynt he s is 204 \nand the synthe sis  of  t hiamine monop ho s phat e from which S. glos sinidius  a lso benefits 205 \n(Belda et a l ., 2010; Sn yder  et  al. , 201 2; Hall et a l ., 2019).  206 \n 207 \nSerially passa gi n g  S. glos s inidius i n  t hi amine-ri ch media in vi tr o, wit h the re peat ed 208 \npopulat i o n bo ttl eneck s cau sed b y pa ssa gi n g , ha s re sulted in the de let i on of  the th i M  gene, a 209 \nputat iv e hydoxyethylt hia zole kinas e t h a t  is  an integr al  par t o f t he thiamin s al vage II pathway 210 \nthat  a llows S. glossini d i us to s ynthesi se th i azole phosphate carboxylate (TH Z-P; Belda et al  211 \n2010). ThiM  h a d  b een retained by th e or iginal  S. g loss inidius genome desp ite the loss  of  212 \nother  c o fac t ors in the thiam ine- p rod u c t ion pathway. That thiM  is deleted in the 213 \nexperimen t all y evolved S. g l ossinidi u s  strain SgCmmC 1*, i n  t he abs ence of W. glossinidia but 214 \nin the pr es ence of th i amine- rich medi a, support s  t he h y po thesi s that  thiM  i s maintained in 215 \nS. glos sinidius  despite ongoing genome degr adation due to p os itive selec tiv e pr es sure, likely 216 \n.CC-BY 4.0 International licenseavailable under a \n(which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made \nThe copyright holder for this preprintthis version posted June 27, 2025. ; https://doi.org/10.1101/2025.06.26.659281doi: bioRxiv preprint \n\ndue to t hia mine b iosy nt hes i s being e ssential for  t s et s e survival a n d a chieved thr ough co-217 \noper a t ion wi th the wide r  mic r obio me  (H all , 2019) .  218 \n 219 \nS. glos sinidius  also maintains a  f unc t i onal a cylated homos er ine l acto ne ( A HL)- bas ed quo rum 220 \nsen s ing s ystem t hat puta t iv ely modu l at es  gene expres s ion  ac cording t o dens it y  t hat is  221 \nhypoth es i sed to aid c o ordination bet ween symbiot ic  bacter ia d uring host tis s ue invasion 222 \n(Pont es  et  al, 2008;  R enoz  et  al 2023) . Sg GmmB4 cont a ins two fun ct i o nal  copies of es a R, 223 \nwhich enc o de s  an ohlL-responsive t rans c r iptional regulat o r, with the de leted c o py in 224 \nSg G mmC2* being proximal to th iM . It  is t herefor e lik ely th a t  the e volved s train mainta ins 225 \nquor um s en s ing fun ctions due to t he presence of the function al  copy, how ever whether  226 \ndensity-dependent  gene ex pr es sion h as b een a f fected will  r equi re further  s tu dy.  227 \n 228 \nIn Sg G mmC1* multiple sulfur- a ssoci a t ed transmembr ane p rot ei n s  were de let ed; s ulfu r is  229 \nused in the biosy n thesis and modification of  s ulfur-containing amino acids , me t hion ine and 230 \nc ysteine (Mbaye et al , 2019). Met hio nine and cysteine are bot h pro d uc ed by W . glossinidia 231 \nfor  the tsetse hos t  (Bing, et al , 2017) and th e d e gr adation o f t he ir pro duc t i on pathway s in S. 232 \ngloss inidius hav e been pr edic t ed as  it mov es towar ds  a sy mbiotic l if es tyle (Belda , et  al,  233 \n2010). A ssuming the deleted sulfur  a nd s ulfit e transmembr ane p rot e ins  facilitated sulfur  234 \ncompound up take, their deleti o n a d d s  evidence to su ggest degr adation of met hion ine and 235 \nc ysteine biosynthesi s  is o ccur ring.  236 \n 237 \nCarb ohydrate t ransmembrane t ransporter s  wer e t he m ost common gr oup of genes 238 \ncontaining small  mutations between th e evolv ed strain and t he a n c estor  s t rain fro m w h i ch 239 \nit was d e r ived, i n c luding t hos e involv ed in tran s por t o f pr ol ine, mannose, a n d a  Glc N A c 240 \nrepr es sor . Fur ther  gene f unc t ions p otentially impacted by va r iation included prot ein 241 \nmodification, D N A  transcription  and  regulation, membr ane p rodu c t ion, two phage lytic 242 \nphase as soc iat ed ge n e s , energ y pr oduction and haemolysis  ( Table 2). One notewor thy 243 \npseudogeni s at ion identi fied i n SgG m mC1* oc c ur red in shlA , a haemolys is precursor ( Ta b l e 244 \n2). D uring axenic  cultur e it was  not ed that when grown on  C olumbia blo od agar plates, 245 \nSg G mmC1* did not exhibit th e same l evel of hae molys is a s  t he original S . gl os si nidius is olat e 246 \ncounter part ( data no t shown), su g ge s ting pheno typic  conf i r mation of  an effect of this 247 \npseudogeni s at ion ev en t. This  implies either t hat haemo lysis is an imp ortant  trai t in t he  248 \n.CC-BY 4.0 International licenseavailable under a \n(which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made \nThe copyright holder for this preprintthis version posted June 27, 2025. ; https://doi.org/10.1101/2025.06.26.659281doi: bioRxiv preprint \n\ntsetse ho s t , but not  one to tho s e grown ou ts ide of  i t s  nat iv e habi t at, or tha t  the conditions 249 \nfor  haemo lys i s gene ex pr ession are not  met in vi t ro , r e sult ing in a lack  of  pos it i ve selec t iv e 250 \npressure for  maintaining the activi ty of this gene enabling i t s  delet i on. 251 \n 252 \nThe serial passa g ing of  S . glos s inidius imitat es the r epeated p opulati on  b ot tlenecks in 253 \nnatur al hos t -s ymbiont sy stems, whi ch  ar e k n own to contribu te t o symbiont genome 254 \nredu c t i o n. The in  v i t r o  pas sage s  were r epeated ev ery 10-14 day s , wher eas  t he l ab- rear ed 255 \ntsetse life c y cle take s  mor e than four  weeks. The higher fr equency of pa ssa ging i n vitr o than 256 \ntr ans mis si o n of S. gl ossinidius  between t s et s e in viv o  may have increas ed  t he bott l e n ec king 257 \neffect on genome degradatio n. 258 \n 259 \nThe Black Queen H y p othesi s  de s c r ibes  t he c o- dependencies  bet w een  a sso c iated  o rganis ms 260 \nin s ymbiotic r elationships ;  t hes e co-depen denc ies are for med by reductive  genomic 261 \nevoluti on dr iv en b y  r epeated popula tion bottl en e cks in a  r e s t ricted enviro nmen t (Mo r ris et 262 \nal., 2012). The large deletion in SgGmmC1*, ac c ompanied by  ina ctivation a nd 263 \npseudogeni s at ion of genes  via indels , suppor ts the con cept the S. glos sinidus i s capable o f 264 \nongoing genome r educ t ion by s t eps  a s de s cribed pr evi o us ly ( Goodhead and Dar by, 201 5). 265 \nFutur e w o rk wil l addr es s the eff e ct  o f  additional s election  p r ess ur e s  mor e c losel y replica t ing 266 \nthose found  in experimenta l systems fro m is olates fro m tsetse h os t s  to examine such 267 \npro c es s e s in real-wor l d  sy s t ems and  t hereby examine the tr ajec t ory of S. gl os s inidius 268 \ntowar ds  obligate s ymbiosis.  269 \n 270 \nLong-ter m ex p er imen t a l evo lution  of S. gl ossinidius  isolates  un der varyin g selection 271 \npressure s, s u ch a s  increa sed oxidative s tr ess or nu trient  d e privat ion, may alter t h e 272 \nevoluti onar y  t rajectory of  S. gloss inidius and eluc idat e the roles of gene s y s tems and 273 \nsel ect i ve pr essures thereo n. S imi lar ly , long- term passaging of S. glos s inidiu s thro ugh tse t s e 274 \nexpose d  t o tr y pano s omes ma y result in genetic, epigenetic or tr ans cript ion c h a n g e s in the 275 \npro duc ed  S. gloss inidius s t rain, increasi n g  ou r understand i n g  of  the tr i p a r t i t e  int e raction 276 \nbetween symbiont, ho s t, and par a s ite. Similarly, studyin g the ability of the  ev olved lab strain 277 \nSg G mmC1* to infect t s etse flies wou l d  allow for  ex am inat i on of t he r ol e S . glossini d i us  278 \ngenes h a ve on t rypano some vec t or c ompetence. 279 \n 280 \n.CC-BY 4.0 International licenseavailable under a \n(which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made \nThe copyright holder for this preprintthis version posted June 27, 2025. ; https://doi.org/10.1101/2025.06.26.659281doi: bioRxiv preprint \n\nACK NO W LED G EME N TS  281 \nThe author s  ac knowledge s upp ort fr o m the University of Salfo rd f or genom ics and  282 \ncomput a t ional i nfrastru c t ure and the Liverpool  School of Tr opic al Med i cine for  s upp orting 283 \nthe t s et s e colony work.  284 \n 285 \nFUNDI N G I NF ORMATI O N  286 \nPP wa s  suppor ted by a U niversit y of Salfor d a n d U nive rsity Alliance Doctoral Tr aining 287 \nScholarship. This  work wa s  f unded by  a Wellcome Trust S EED award t o IG : 200690 /Z / 16/ Z.  288 \n 289 \nAUTHOR C O N TR IBUTI O N S  290 \nSodalis  gloss inidius is olat e s were coll ec t ed by ACD and  mai n tained by L RH .  G en ome 291 \nsequenci n g  and analy sis wa s  per formed by P P  and  I G. Furt her analy s is and manuscr i p t 292 \nprep a r ation and r ev iew was  per formed by P P, L RH , ACD  and IG.  293 \n 294 \nCO NFLIC TS O F IN TER ES T 295 \nThe author s  declare t hat ther e are no  conflicts o f interest.296 \n.CC-BY 4.0 International licenseavailable under a \n(which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made \nThe copyright holder for this preprintthis version posted June 27, 2025. ; https://doi.org/10.1101/2025.06.26.659281doi: bioRxiv preprint \n\nFI G URES  297 \nFigure 1: Large deletion in the SgGmmC1* genome. Top:  Full geno me al ig nmen t of Soda lis  298 \ngloss inidius ev olved strain Sg GmmC1 * agains t  a n cestor  s tr ain Sg GmmB4 s howing 99.99 % 299 \nsi mil ar ity, with the single lar ge d el et i on expanded to show details  of  t he g enes  delet ed. The 300 \ntop f i gu re s ho w s the alignment  of th e  ent i r e c h romosome o f each strain, s howing the high 301 \nlevel of s imilari t y between the seque ncies, with t he l ar ge del etion ci r cle d . T h e b ottom 302 \ndiagram show s  t he c od i n g genes (or ange)  an d ps eudogene s (yellow) flanking and inc lud e d  303 \nin the del et i o n. G enome po s itio n s  (bp)  a r e rela tive to t he S gGmmB4 r efer ence genome. 304 \nFigures were produ c ed u sing ea syfig.  305 \n 306 \n 307 \n  308 \n.CC-BY 4.0 International licenseavailable under a \n(which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made \nThe copyright holder for this preprintthis version posted June 27, 2025. ; https://doi.org/10.1101/2025.06.26.659281doi: bioRxiv preprint \n\nTable 1  – G ene s within the 17,209  b p  d el et ed region of Sg GmmC1* ac c o rd ing to th e SgGmmB4 r eference (Goodhead et al , 2020). Gene names 309 \nand pro ducts  wer e ann otated using P ROKKA; con served ort hol ogous gr oup s wer e as sig n ed us in g the STRI N G  dat abas e; gene onto l o g y a nd 310 \ncon s er ved regions w er e assigned u s in g the Int erpr o database. K e y: C D S – coding D N A  s equen ce; p seudo – ps eu d ogene; C O G – c lu ster ed 311 \nort hologous group; G O  – gene ont ology. Yellow – pseudogene s ; blue – CD S . 312 \n 313 \n 314 \nCDS/ \npseudo \nCOG GO terms Conserved regions \nCode Description Biological Molecular Cellular \nba eS_ 1  S i g n a l  tr a ns d u c t io n  hi s t id in e - pr o te i n k in a se  BaeS \nps e u d o  COG 0 6 42 S i gn al t ransducti o n histi d i ne k i n a s e - - - - \nba eS_ 2  S i g n a l  tr a ns d u c t io n  hi s t id in e - pr o te i n k in a se  BaeS \nps e u d o  COG 0 6 42 S i gn al t ransducti o n histi d i ne k i n a s e GO :000 7 1 65  s ig n a l \nt r a n s du c t i o n  \nGO: 000 0 1 55  p hosp ho r e lay \nse n s or k ina s e  acti vity  \nGO: 001 6 0 21  i n t e g r a l  \nc o m p on e nt  of  \nme m br an e \nIPR0 0 3 661  S ign a l t ra n sd uc t i on  h i s t i d ine  \nki na s e , d i m e r i s a t i on / ph o s ph oa c c ep t o r  \ndo m ai n  \nIPR0 0 3 660  H AM P  dom ain (s e n so r a nd  \nc hem o t ax i s) \nba eS_ 3  S i g n a l  tr a ns d u c t io n  hi s t id in e - pr o te i n k in a se  BaeS \nps e u d o  COG 0 6 42 S i gn al t ransducti o n histi d i ne k i n a s e - - - IPR0 0 5 467  H istid in e ki na se d o ma i n \nS y s t em ati c G en e ID : 0 3996 Hy po th eti ca l prot ei n \nps e u d o  COG 0 8 26 C ol l ag en a s e-li ke pr ot ea s e, Prt C f a mi ly  - - - - \ny hbU _2 P u tativ e pr ot e as e YhbU p r e cu rs o r  \nps e u d o  COG 0 8 26 C ol l ag en a s e-li ke pr ot ea s e, Prt C f a mi ly  - - - IPR0 0 1 539  Pe p t i d a se  U 32 \nS y s t em ati c G en e ID : 0 3998 Pe p t i da se  f a m il y  U 3 2 \nps e u d o  COG 0 8 26 C ol l ag en a s e-li ke pr ot ea s e, Prt C f a mi ly  - - - IPR0 0 1 539  Pe p t i d a se  U 32 \nthi M  Hyd rox y et hy l thi azol e ki n a s e  \nCDS  COG 2 1 45 Hyd roxy e t hylth iaz ol e k ina se, s u g ar k i n a s e \nfa mi ly  \nGO :000 9 2 28  th iam in e  \nbiosynth e t i c p roc e ss  \nGO: 000 4 4 17  \nhy dr ox y eth y l thia zol e \nk i n a se ac t iv it y \n- IPR0 0 0 417  H y dr ox y eth y l thi azol e k i n as e  \nes aR _2 T r an sc r ip t i on a l  a c t iva t o r p ro te i n  E sa R \nCDS  COG 2 1 97 \nDNA -bindi ng r esp o n s e  r eg ulato r, \nN a rL /F ix J  f am i ly , co n t a in s  REC  a nd  H T H  \ndo m ai ns   \nGO :000 6 3 55  r e g ul ati on of \ntrans cri p t i on , DN A-\ntem p l a t ed  \nGO: 000 3 6 77  DNA  b indi ng  - IPR0 0 0 792  T rans cri p ti on r e gul a t or L u x R, C-\nte r mi n al  \n.CC-BY 4.0 International licenseavailable under a \n(which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made \nThe copyright holder for this preprintthis version posted June 27, 2025. ; https://doi.org/10.1101/2025.06.26.659281doi: bioRxiv preprint \n\nCDS/ \npseudo \nCOG GO terms Conserved regions \nCode Description Biological Molecular Cellular \nS y s t em ati c G en e ID : 0 4001 P u tativ e a c etyl tra n sf era s e  \nCDS  COG 0 4 54  N - a ce ty l t r an s fe ra se , GN AT  s u pe rf a mil y  \n(in clu des histo ne ac etylt ran sfer as e H P A 2 )  - GO: 000 80 8 0  N -\na c et y l t r an s f e r a s e a c t i v i t y  - IPR0 0 0 182  G NA T d om a i n \nS y s t em ati c G en e ID : 0 4002 Hy po th eti ca l prot ei n \nC D S  C O G 04 2 5  Tu s A - r el at e d  s u l f u r t r a ns f er a s e  -  -  -  IPR0 0 1 455  T us A -li k e  dom ain  \nS y s t em ati c G en e ID : 0 4003 Hy po th eti ca l prot ei n \nCDS  COG 2 3 91 \nU n c h ar a c te rize d  mem b r a ne  p ro te i n  \nYed E / Y e eE, c o nta ins t wo  sulfu r tr a ns p o rt \ndo m ai ns  \n- - - IPR0 0 7 272  Su lphur t ran sp o rt d omai n \nS y s t em ati c G en e ID : 0 4004 P u tativ e i n n e r m e m b rane p r ote in  \nCDS  COG 2 3 91 \nU n c h ar a c te rize d  mem b r a ne  p ro te i n  \nYed E / Y e eE, c o nta ins t wo  sulfu r tr a ns p o rt \ndo m ai ns  \n- - - IPR0 0 7 272  Su lphur t ran sp o rt d omai n \nyeeE _1 L ys R  su b str at e bi nding d o ma i n  p r ot ein  \nps e u d o  COG 0 5 83 DNA -bindi ng tr ans c ripti on a l r e gulato r, \nLy s R fa mil y  - - - IPR0 0 5 119  Ly s R , substrat e-bi nd i ng  \nyeeE _2 L ys R  su b str at e bi nding d o ma i n  p r ot ein  \nps e u d o  COG 0 5 83 DNA -bindi ng tr ans c ripti on a l r e gulato r, \nLy s R fa mil y  - - - G3D SA: 3.40. 1 9 0.1 0  Per ipla sm ic b i ndin g \np rot ei n-li ke I I  \ndt ha dh  D-thr e o -3-hy d ro xy as par ta t e de h yd rata s e  \nps e u d o  COG 3 6 16 D - s er i ne de am i na s e , p y r i d ox a l  ph os p h a te -\nde p e n d e n t  - - - IPR0 4 2 208  D -s eri ne deh y drat a s e-l ik e  d o main  \nS y s t em ati c G en e ID : 0 4008 Hy po th eti ca l prot ei n \nps e u d o  COG 1 2 88 U n c h ar a c te rize d  mem b r a ne  p ro te i n  Y f cC , \nion  tr an s port er su p erf a mi ly  - - \nGO: 001 6 0 21  i n t e g r a l  \nc o m p on e nt  of  \nme m br an e \nIPR0 1 838 5  C 4 - d i carboxyl a t e anae ro b i c \nc a rri er- lik e  \nIPR0 1 838 7  Un ch a rac te r ise d  p ro tei n  \nYc g A / Y f c C  \nS y s t em ati c G en e ID : 0 4009 Hy po th eti ca l prot ei n \nCDS  COG 1 2 88 U n c h ar a c te rize d  mem b r a ne  p ro te i n  Y f cC , \nion  tr an s port er su p erf a mi ly  - - \nGO: 001 6 0 21  i n t e g r a l  \nc o m p on e nt  of  \nme m br an e \nIPR0 1 838 5  C 4 - d i carboxyl a t e anae ro b i c \nc a rri er- lik e  \nIPR0 1 838 7  Un ch a rac te r ise d  p ro tei n  \nYc g A / Y f c C  \nyx e P _2  P ut ativ e hydr ola s e  Yx e P  \nCDS  COG 1 4 73 Metal-d e pen den t  - GO: 001 6 7 87  h y d r o la s e  -  IPR0 0 2 933  Pe p t i d a se  M2 0 \n.CC-BY 4.0 International licenseavailable under a \n(which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made \nThe copyright holder for this preprintthis version posted June 27, 2025. ; https://doi.org/10.1101/2025.06.26.659281doi: bioRxiv preprint \n\nCDS/ \npseudo \nCOG GO terms Conserved regions \nCode Description Biological Molecular Cellular \na mi das e/ a mi noacyl as e/ carb oxy p e p ti das e  activity  IPR0 1 7 439  A min o hydr olas e  \nSys t e ma t ic  Ge ne  I D:  _ 04 01 1  H y po t h et i c al  p r ot ei n  \nCDS  -  - - - - - \nya b J_ 2  Enami n e /i min e  dea minas e  \nps e u d o  COG 0 2 51 \nE na mine  dea m i nas e Ri dA, hous e  cl e a ni n g \no f r eacti ve en am i ne i n t er m ed i at es, \nY j gF/Y E R057 c/U K 1 1 4 fam il y \n- - - IPR0 0 6 175  Yj gF/ YE R05 7c/U K11 4  famil y  \nS y s t em ati c G en e ID : 0 4013 Hy po th eti ca l prot ei n \nps e u d o  -  - - - - - \nS y s t em ati c G en e ID : 0 4014 S u lf i te  e xp o r te r  T au E /S a f E \np se ud o C O G 0 73 0  U n c h ar a c te rize d  mem b r a ne  p ro te i n  Y f c A -  -  \nGO: 001 6 0 21  i n t e g r a l  \nc o m p on e nt  of  \nme m br an e \nIPR0 0 2 781  T ransm e mbran e p r ot ein Ta u E-li k e \nS y s t em ati c G en e ID : 0 4015 S u lf i te  e xp o r te r  T au E /S a f E \nC D S C O G 0 73 0  U n c h ar a c te rize d  mem b r a ne  p ro te i n  Y f c A -  -  \nGO: 001 6 0 21  i n t e g r a l  \nc o m p on e nt  of  \nme m br an e \nIPR0 0 2 781  T ransm e mbran e p r ot ein Ta u E-li k e \nS y s t em ati c G en e ID : 0 4016 Hy po th eti ca l prot ei n \nCDS  -  - - - - - \nS y s t em ati c G en e ID : 0 4017 PT S  s y s tem , l ac to se /ce llo b i o s e s pec if ic  I IB s ub u ni t  \nCDS  COG 3 4 14 P h o sp ho t ran sfer a s e s sy st em, g ala ctito l-\ns p ec i f ic  I I B  c om p o ne n t \n \nGO :000 9 4 01  \np h os p hoe no lp yr u v a t e -\ndep end e nt s ugar \nph o sp h otransf era s e  \nsyst em \n \nGO: 000 89 8 2  p r ot ein-N ( PI )-\nph o sp h o hi st idi ne-su gar \np ho s ph ot r a n s f e r a s e \nactivity  \n- IPR0 0 3 501  Ph o sp h otr an sf er as e syst em, EII B \nc om p o ne n t , t yp e  2/ 3 \nga tC  Gal a ctitol per m e as e  I I C  co m pon en t  \nps e u d o  COG 3 7 75 P h o sp ho t ran sfer a s e s sy st em, g ala ctito l-\ns p ec i f ic  I I C  c o mp o ne n t - -  - \nIPR0 0 4 703 P h os ph o t ra n s fe ra se  s y s te m , \ns u g a r - sp ec i f ic  pe rme ase  c o m p o ne n t \nIPR0 1 3 853 G a l a cti t o l  p er me as e  II C  \nc o mpon e nt  \nS y s t em ati c G en e ID : 0 4019 Hy po th eti ca l prot ei n \n.CC-BY 4.0 International licenseavailable under a \n(which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made \nThe copyright holder for this preprintthis version posted June 27, 2025. ; https://doi.org/10.1101/2025.06.26.659281doi: bioRxiv preprint \n\nCDS/ \npseudo \nCOG GO terms Conserved regions \nCode Description Biological Molecular Cellular \nCDS  -  - - - - - \nS y s t em ati c G en e ID : 0 4020 P T S syst e m gal actit ol-sp e cifi c tra n s po r t er s u bunit  II A  \nCDS  COG 1 7 62 \nPh os p h ot r a n s f e r as e  s y s t e m  \nm anni tol/fr u cto s e-s p eci fic I IA  domai n \n(Nt r-t y p e)  \n- - - IPR0 0 2 178  PT S  EI IA  type-2 domai n \nS y s t em ati c G en e ID : 0 4021 Hy po th eti ca l prot ei n \nCDS  N OG 33 1 0 3  n on  su p e rvis e d  orth o l og ou s gr oup - - - - \nfa b G _4 3 -o xo a cyl-[ acyl- car ri er- p rot e in] r e ductas e  FabG  \nps e u d o  COG 1 0 28 NAD ( P )-dep end e nt d eh y drog e n as e , sh ort-\nch a in al c ohol de hy d ro gen a s e f a m i ly  - GO: 001 6 4 91  \nox i dor e d u ctas e  acti vity  - IPR0 0 2 347  Sh o rt- c hai n \nd e h yd rog e na s e /r edu cta s e SD R \nS y s t em ati c G en e ID : 0 4023 T ra n sposas e IS 116/IS 1 1 0/I S9 0 2 f a m i ly p ro t ei n \np s eu do  C O G 35 4 7  Tr a n s po s a s e  GO :000 6 3 13  t ransp o si ti on , \nDN A-m edi at e d  \nGO: 000 3 6 77  DNA  b indi ng  \nGO: 000 4 803  tr a n s po s ase  \nactivity  \n- IPR0 0 3 346  T ransp o sa s e, I S116/I S110 /IS 9 0 2  \nS y s t em ati c G en e ID : 0 4024 T ra n sposas e \np s eu do  C O G 35 4 7  Tr a n s po s a s e  GO :000 6 3 13  t ransp o si ti on , \nDN A-m edi at e d  \nGO: 000 3 6 77  DNA  b indi ng  \nGO: 000 4 803  tr a n s po s ase  \nactivity  \n- IPR0 0 3 346  T ransp o sa s e, I S116/I S110 /IS 9 0 2  \nS y s t em ati c G en e ID : 0 4025 H yp o t he t ic al  p r o te in  \nps e u d o  COG 3 5 47 T ra ns p os ase  - - - - \n  315 \n.CC-BY 4.0 International licenseavailable under a \n(which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made \nThe copyright holder for this preprintthis version posted June 27, 2025. ; https://doi.org/10.1101/2025.06.26.659281doi: bioRxiv preprint \n\nTable 2 - Snippy output  w it h al l insertions , delet ions  and single-nucleotide polymor phis ms in Sodalis glossinidius  s t rain SgG mmC1*, in 316 \ncomparison with compar a t or strain SgGmmB4. Gene names  and  p rodu cts were annotat ed us ing PRO KKA;  con se rved ort ho logous group s were 317 \nassigned u sing t he STRI NG databa se; g en e  on tology and c o n s er v ed r egions were as s igned u sing the Int erpro databas e. Ke y: *** - Tier 1 318 \npseudogeni s at ion (mor e  t han two-thi rds of the gene as a n notated in SgGmmB4 is r emoved); ** - Tier 2 pseudogenisation (disrupt ion o f a 319 \ncon s er ved region by tru nc at ion); * - T ier 3 pseudogenisa t ion (disrupt i o n of a c on served r egi o n by  a mis s en s e mu t a t ion). CD S – C od in g  DN A  320 \nsequence. P seudo – pseudogene. In s – insert ion. Del –  dele tion. Co m – c o mplex  polymor phism. SN P – si n gl e nu cleotide po l ymo rphism. A A – 321 \namino acid. B – biol ogic al pr oc es s. M – molec u l ar  process . C – cell u l ar  proces s . Yellow – ps eud o ge n es ; blue – CD S ;  o range – potent i al n ovel 322 \npseudogene s, shaded b y tier.  323 \n 324 \nB 4  p os i t ion Ty p e CD S/ \np s eu do GO  Te rm s COG s  SN P eff ec t  Disru pte d c on ser v e d re gi on s  \n ⎯ Chromosome ⎯  \n* ** S y s te mati c  G e n e I D: 0 0 037 h y po t h et i c al  p r ot ei n  \n26 ,5 7 7  ins  CDS  - -  - - Fra m e s hif t truncat e s at A A \n59/ 1 9 9 \n- \n* ** h srA _1 pu tati ve  transp o rt p rot e in H sr A \n35 1 ,610  ins  CDS  GO:00 5 5 085  B: tr an s m e mb r an e  \ntr a n sp o r t \nC O G 0477  MFS fam i ly p e rm eas e  Fra m e s hif t r em o v es hs rA_ 1 \non an t i s e n s e st rand, r e p l ac e s \nwi th t wo over l a p pin g  \nh y po t h et i c al s  o n s en s e  \nstran d  ( no  c o ns erv ed  r egi ons \non ne w gen es)  \nIP R 0 1170 1 ; P F 0 7690 (m ajor fa cil it ator \nsupe rfamil y )  \nIP R 0 2084 6 ; PS 5 0850  ( majo r fac il itato r \nsupe rfamil y  p r ofi l e)  \nIP R 0 3625 9 ; SS F 1 03 47 3  (M F S gen e ra l  \ns u b s tr a te  tr a ns p or te r) \nPT H R 23 50 1 : S F 180  (m u lt id r u g  r e sis t a nce  \npro t ei n B h omol og )  \nG 3 D S A : 1. 20 . 1 7 20 . 1 0  (m u l tid ru g  r esi stanc e \npro t ei n D )  \nGO:00 2 2 857  M: tr an s m em b r an e  \nt ra ns port er a cti vi ty  \nSys t e ma t ic  Ge ne  I D:  00 81 0  Tai l fib e r  p r ot ein g p37 C t e r mina l  \n58 8 ,917  ins  ps e u d o  - -  C O G 4675  Mi cr o c ysti n -d epen dent pro t e in \n(functi on  unkno wn)  \nFra m e s hif t truncat e s at A A \n17 1 /2 60 \nIP R 0 2224 6 ;  PF12 6 0 4  (b act eri o pha g e tail  \nf ib re  pr o te i n  g p 37  C  te rm i na l f a m ily )  \nnn r_ 3  NA D( P )H- h ydr a t e r epai r en zyme  N n r  \n61 9 ,593  ins  ps e u d o  GO:00 5 2 855  M: AD P -d epe nde n t  \nN AD ( P) H-hy dr at e \nde h yd ratas e a ctivi ty \nC O G 0062  Ca r bohydr a t e t r a ns port a nd  \nme ta bo l i sm ;  N A D H X  ep i me ra se  \nactivity  \nFra m e s hif t r em o v es fi rst \n53/ 1 1 9 AA s an d  a l t ers AA s \n54-73  \nIP R 0 0063 1 ;  PF01 2 5 6  ( ca rb o hy d rat e k inas e \nfamil y )  PS 5 13 83  ( Yj eF C-te rm ina l  domain )  \nIP R 0 2905 6  (r i bo k in a se - l ik e s u pe r fa m il y) \nC O G 0063  Ca r bohydr a t e t r a ns port a nd  \nme ta bo l i sm \n \n \n \npr o P _2/ proP _1 Pr oli ne /beta in e tr anspo rt e r  \n86 9 ,745  ins  CDS  GO:00 5 5 085  M: AD P -d epe nde n t  C O G 0477  Ca r bohydr a t e t r a ns port a nd  R e m ov es pr o p _2 st op Al l cons erv ed r e g i ons i n ta ct  \n.CC-BY 4.0 International licenseavailable under a \n(which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made \nThe copyright holder for this preprintthis version posted June 27, 2025. ; https://doi.org/10.1101/2025.06.26.659281doi: bioRxiv preprint \n\nB 4  p os i t ion Ty p e CD S/ \np s eu do GO  Te rm s COG s  SN P eff ec t  Disru pte d c on ser v e d re gi on s  \nN AD ( P) H-hy dr at e \nde h yd ratas e a ctivi ty \nm e t a b olis m, m a j o r  fa cil i tator \nsupe rfamil y  \nc od o n , merg i ng  p r op_ 2  a n d  \npro p_1 \nGO:00 2 2 857  M: tr an s m em b r an e  \nt ra ns port er a cti vi ty  \nGO:00 1 6 021  C: int eg ral comp o n en t o f \nme m b ra ne  \n**  p g l_2  Po l ygala ctu ronas e \n1 ,04 8 , 5 09 i n s C D S G O : 0 00 59 7 5 B :  c a r b oh y dr a te  me t a b o l i c  \npro c ess  \nC O G 2706  Ca r bohydr a t e t r a ns port a nd  \nm e t a b olis m, 6-\nph o sp h ogl uco n ol act on a s e \nFra m e s hif t truncat e s at A A \n17 1 /1 82 \nIP R 0 0074 3 ;  PF00 2 9 5  ( gly c osi de h yd rol a s e \nf a m i ly  2 8 ) \nIP R 0 1105 0 ;  S S F 51 1 2 6  (pe c t in  ly a se - like  \nsupe rfamil y )  \nGO:00 0 4 650  M: poly gal act u r onas e \nacti v i ty  \n* S y st em ati c Gen e ID : 0 160 3  B a c te r io ph age  l ys i s  p ro te in  \n1 ,21 5 , 3 24 c o m C D S -  - N O G 2 69 7 2 D NA - p ac k ag ing  p r o te i n gp 3  M is se nse  a t A A 23 ,  R  t o  K IP R 0 3206 6 ;  PF16 6 7 7  (D NA-pa ck a g in g \npro t ei n gp 3 )  \n1,2 15 , 3 8 7  SN P  CDS  - -  NOG 2 6 972  DN A-pa c ka gi n g  prot ein  g p3 Synony m ous - \n* n ag C _ 2 N -a cetyl gl uc osami ne r ep r e s so r  \n1 ,42 4 , 4 93 S NP C D S G O : 0 00 63 5 5 B :  re g u l a ti o n  o f  \nt ra ns c ripti on, D NA-\nte mp l a ted  \nC O G 1940  Ca r bohydr a t e t r a ns port a nd  \nme ta bo l i sm , R O K  f a m ily \nMi ss en s e  a t  AA  2 6 3/ 419, C  \nto F  \nIP R 0 0060 0 ;  PF00 4 80  (ROK  f a m ily ) \nIP R 0 4312 9 ;  S S F 53 0 6 7  (ac t i n - l ike  ATP a se \ndo m ain )  \nGO:00 0 3 700  M: D N A-bin d i ng \nt ra ns c ripti on fa ctor a c t ivity  \n* pt sG _ 1  PTS syst e m glu cos e-sp eci fi c EII CBA c o m po n e n t  \n1,4 27 , 6 7 6  SN P  CDS  GO:00 0 9 401  B: phosph oenolpy ruv at e-\nde pen den t  s ugar \nphosphotra n sf e ra s e \nsyst em  \nC O G 1263  \n \nc ar b oh y dr a te  t ra n sp o r t a n d  \nm e t a b olis m, PTS s yst em  \nM i s s e n s e at  A A  2 8/ 6 7 8, M  t o  \nI \nIP R 0 1097 4 ;  TIG R 0 1 998  ( N-\nac e tyl g l ucos am i ne-sp eci fi c PTS tr anspo rt er \ns u bu ni t  I I B C )  \nIP R 0 1301 3 ;  P S 5 1103  (phos p ho enol py r uvat e -\ndep end e nt s ugar phosphotr an sf e ras e  \nsyst em  ( PTS) )  \nIP R 0 0335 2 ;  PF02 3 7 8  ( ph os p h o t r an s fe ra se  \nsyst em , EI I C)  \nPT H R 30 00 9  ( cytoch rom e c-t y p e sy n t hesi s \npro t ei n a n d  P T S tr ans m e m br ane \nco m p onent )  \nGO:00 0 8 982  M: pr o t ei n-N ( PI) -\nphosphohi stidi n e-suga r \nphosphotra n sf e ra s e \nacti v i ty  \nC O G 1264  ca rb o h yd rat e  tr an s port  and \nm e t a b olis m, PTS s yst em  \nGO:00 1 5 572  M: N-ac etyl glu c o sam in e  \nt r an s m em b r a n e  \nt ra ns port er a cti vi ty  \nC O G 2190  ca rb o h yd rat e  tr an s port  and \nm e t a b olis m, PTS s yst em  \nGO:00 1 9 866  C: org anel l e in ner \nme m b ra ne  \nGO:00 1 6 021  C: int eg ral comp o n en t o f \nme m b ra ne  \nGO:00 1 6 020  C: m emb rane  \n \n* s uc B  Co m p one nt of 2-ox ogluta rat e  dehy d ro ge na s e  \n1,4 52 , 0 5 3  SN P  CDS  GO:00 0 6 099  B: tr ic ar bo xy li c aci d  cycl e C O G 0508  Ene r gy pr o d u ct i o n and  c o nve rsi o n, \ndehydro g enas e \nMi ss en s e  a t  AA  2 7 4/ 396, P  \nto S  \nIP R 0 0625 5 ;  TIG R 0 1 347  ( di hy d r ol i p oy l l y s i n e-\nr esi due s u cc inylt ransf eras e )  G O : 0 01 67 4 6 M :  ac y ltr a n s f er ase  a c t iv i t y \n.CC-BY 4.0 International licenseavailable under a \n(which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made \nThe copyright holder for this preprintthis version posted June 27, 2025. ; https://doi.org/10.1101/2025.06.26.659281doi: bioRxiv preprint \n\nB 4  p os i t ion Ty p e CD S/ \np s eu do GO  Te rm s COG s  SN P eff ec t  Disru pte d c on ser v e d re gi on s  \nGO:00 0 4 149  M: d i h yd rol ipoyl lys in e -\nre s i d ue  su c ci n y l tr a ns f er a se \nacti v i ty  \nIP R 0 0107 8;  PF00 1 9 8  ( 2-ox o ac id \ndehydro g enas es acyl t ra n sfe ras e ( ca t a l y t ic \ndo m ain )  \nGO:00 4 5 252  C: o x o g l utarat e \nde h yd rog e n as e c ompl ex  \n**  yd c V  I nne r m e mbr a n e t ransp ort er  pr o t ein Yd c V  \n1 ,53 5 , 7 73 i n s C D S G O : 0 05 50 8 5 B :  t ra n smem b ra ne  \ntr a n sp o r t \nC O G 1177  Ino r gani c i o n tr ans p o rt an d  \nm e t a b olis m; b i n di ng-p ro t e in-\ndep end e nt t r a ns port sy st e ms i n n e r \nm e mb ra ne com p o nent  \nM is se nse  a t A A 26 4  (V  t o  G)  \nand  tr u n cat ed at AA  266/ 2 8 1  \nIP R 0 3590 6  ( M e tI -li ke sup e r fami ly )  \nPT H R 43 84 8  ( Pu t r es cin e  tra n sp ort s yst em  \np e r mea se  p ro te i n)   \nGO:00 1 6 020  C: m emb rane  \nS y s t em ati c G en e ID : 0 2100 hy p otheti cal p r ot ein  \n1,5 66 , 2 6 0  ins  CDS  - -  C O G 5464  Trans po s as e, fun cti on  u nkno wn Tw o hypothet i cal  g e n e s \nme rge d \nAl l cons erv ed r e g i ons i n ta ct  \n* w c aJ U D P- g l uc os e:und eca p r enyl-phosp h at e glu c o s e-1- p ho sp hat e tra n sf era s e  \n1,6 38 , 8 6 4  SN P  CDS  - -  C O G 2148  Cel l wal l / m emb ra ne / e nv el op e \nbiog en esi s , su gar tr an sf e ras e \nMi ss en s e  a t  AA  3 7 4/ 464, G \nto A  \nIP R 0 1747 3 ;  TIG R 0 3 023  ( u nde ca p r e ny l -\nph o sp hat e  g lu cos e phos p hot ransfe ra s e )  \nIP R 0 1747 5 ;  TIG R 0 3 025  ( exopoly sa ccha rid e \nb i o s y n t he s is  p o ly p ren y l \ngly cosyl phosph o tra nsf e r a s e )  \nIP R 0 0336 2 ;  PF02 3 9 7  (b act eri a l  s ugar \ntransf eras e )  \nPT H R 30 57 6  ( c o lo n i c  b i o s y n t h e s i s  U D P -\ng l uco se  l i p id  ca rr ie r  tra ns fe r a se ) \nza p C_ 2  Ce ll  di visi o n p ro t e i n  Z ap C  \n1,7 01 , 4 6 2  ins  ps e u d o  - -  NOG 0 1 298  Cel l divi si on Fra m e s hif t r em o v ed  ge n e IP R 0 0980 9 ;  PF07 1 2 6 (Ce ll - d iv is io n  p r o tei n  \nZapC )  \n* ** m a n C 1 M an n o s e-1-p h o sp h at e gu a ny l ylt ransf eras e  1  \n1,8 67 , 8 9 8  ins  CDS  GO:00 0 9 058  B: bi o s yntheti c p ro c e ss C O G 0662  Ca r bohydr a t e t r a ns port a nd  \nme ta bo l i sm \nFra m e s hif t truncat e s at A A \n1 2 9/ 4 7 1 an d  m ov e s  A A s  \n13 9 -47 1  onto n ext fram e \nw it h  c lo se s t  p r o mo te r  > 5 0b p  \nupstr eam f r o m s e cond h a l f \nIP R 0 0637 5 ;  TIG R 0 1 479  ( m a nn ose - 1 -\nph o sp hat e  g u an y l yltr an sf eras e /mann o s e-6-\nph o sp hat e  is om eras e )  \nIP R 0 0583 5 ;  PF00 4 8 3 (n ucl e oti dyl \ntransf eras e )  \nIP R 0 2904 4  ( n u cl eoti d e - d i phosph o -su gar \ntransf eras e s)  \nGO:00 0 0 271  B: poly sa ccha rid e \nbiosy n th eti c pro c ess  \nC O G 0836  Cel l wal l / m emb ra ne / e nv el op e \nbiog en esi s  \nGO:00 0 5 976  B: poly sa ccha rid e \nm etaboli c p roc es s  \nGO:00 1 6 779  M: nuc l eoti dy l tr an sf er as e \nacti v i ty  \nS y s t em ati c G en e ID : 0 2481 hy p otheti cal p r ot ein  \n1,8 96 , 4 2 1  ins  CDS  - -  - - Fra m e s hif t at A A 2 20 /246, \na l te r s  re ma i ni ng  AA s  a nd  \ne x ten d s  to  25 0  \n- \nS y s t em ati c G en e ID : 0 2937 hy p otheti cal p r ot ein  \n2,2 05 , 2 0 6  ins  ps e u d o  - -  - - Fra m e s hif t alt er s f rom AA \n4 2 / 46 , e x t e n d s  t o 49  \nAl l cons erv ed r e g i ons i n ta ct  \nsh l A  H e m ol ys i n pr e c u r s or  \n.CC-BY 4.0 International licenseavailable under a \n(which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made \nThe copyright holder for this preprintthis version posted June 27, 2025. ; https://doi.org/10.1101/2025.06.26.659281doi: bioRxiv preprint \n\nB 4  p os i t ion Ty p e CD S/ \np s eu do GO  Te rm s COG s  SN P eff ec t  Disru pte d c on ser v e d re gi on s  \nS y s t em ati c G en e ID : 0 3153 hy p otheti cal p r ot ein  \n2,3 60 , 3 2 6  ins  CDS  - -  C O G 3210  Intra c e l lula r tr affi ckin g ,  s e cr e ti o n \nan d v e si cula r tr ans p o rt \nFra m e s hif t at A A 2 33 /248 o f  \nshlA  alt e rs l ast 1 5  AA s and \nadd s PR O K K A _ 0 315 3 \nunchang e d  onto end  \nAl l cons erv ed r e g i ons i n ta ct  \nCDS  - -  - - Al l cons erv ed r e g i ons i n ta ct  \nS y s t em ati c G en e ID : 0 3155 hy p otheti cal p r ot ein  \nS y s t em ati c G en e ID : 0 3156 hy p otheti cal p r ot ein  \n2,3 61 , 5 1 5  ins  CDS  - -  - - Fra m e s hif t m er ge s \nP RO KKA _0 3 1 55  and \nP RO KKA _0 3 1 56  with 17  \next ra A As b et w e en th em \nAl l cons erv ed r e g i ons i n ta ct  \nCDS  - -  - - Al l cons erv ed r e g i ons i n ta ct  \n* p h e S P heny l alani ne--tR NA  li gas e al pha  subu ni t  \n2,3 67 , 3 0 9  SN P  CDS  GO:00 4 3 039  B: t RN A am ino a c ylati o n  C O G 0016  Transl ati on, ri bo s om a l  st ru ctur e a n d  \nbiog en esi s  \nM i s s e n s e at  A A  5 9/ 3 2 7, V  t o  \nG \nIP R 0 2291 1 ;  MF_ 00281  ( P hen y l a l anin e —\ntR NA li g as e  alp h a s u b u nit ph eS)  \nIP R 0 0452 9 ; x  ( Phe n yl a l an in e— tR NA  l iga se  \nalph a  su bu nit )  \nIP R 0 04188 ;  PF02 9 1 2  ( Am in oa cy l tR N A \nsyn t heta s e  cla ss II , N -t er minal  d o ma in)  \nIP R 0 1097 8;  S S F 46 5 8 9  ( tRN A -b in d i ng  a rm ) \nG 3 D S A : 3. 30 . 9 3 0. 1 0  (Bi r a b if u nc t io n a l  \npro t ei n; doma in 2 )  \nGO:00 0 6 432  B: ph en y lala nyl-t RNA  \namin oacylat i o n  \nGO:00 0 5 524  M: AT P bin d i ng  \nGO:00 0 0 049  M: t RN A b i ndin g  \nG O : 0 00 48 1 2 M :  am in o ac y l- tR N A  lig a se  \nacti v i ty  \nG O : 0 00 01 6 6 M :  n u cle o t ide  b i n d in g \nGO:00 0 4 826  M: ph en yl al anin e-t R N A \nli ga se  a c t ivi t y \nGO:00 0 5 737  C: c y topl as m  \nsl yA _2 Tr an s c ri p ti o na l r e g u l ator Sl yA \n2 ,40 7 , 9 04 i n s C D S G O : 0 00 63 5 5 B :  re g u l a ti o n  o f  \nt ra ns c ripti on, D NA-\nte mp l a ted  \nC O G 1846  Trans cr ipti on r e gul a t or  I n s e rtion 50bp i n fr ont of \nge n e, u na l te red  A A  \nse que n c e  but move s n ear est \nstart codo n f r o m 50 b p a w ay \nt o  9 3b p  aw ay \nAl l cons erv ed r e g i ons i n ta ct  \nGO:00 0 3 700  M: D N A-bin d i ng \nt ra ns c ripti on fa ctor a c t ivity  \n* f nr Fu m a r a te  a n d  n it ra t e  re d uc ti on  re g ul a t o ry  p r o te in  \n2 ,46 7 , 5 83 S NP C D S G O : 0 00 63 5 5 B :  re g u l a ti o n  o f  \nt ra ns c ripti on, D NA-\nte mp l a ted  \nC O G 0664  Si gn al  t ransdu cti on  me chanis ms, \nt r a n scr ip t i o n al  re g ul a t o r, c rp  f n r  \nfamil y \nMi ss en s e  a t  AA  5 2 /2 51 , P  to \nT \nIP R 0 0059 5 ;  cd00 0 3 8  ( effe ctor  domai n of th e \nCAP fa mil y o f  tran scri p t ion  facto rs ), P F 000 27  \n(cy cl i c  n u cl e otid e - b i ndi ng d o mai n), P S 50 04 2  \n( c A M P /c GMP  b i n d ing  mo t i f  p ro f ile ) \nIP R 0 1471 0  (R m l C - li ke  je ll y  r o ll f o l d) \nIP R 0 1849 0  (c yc lic  n uc le o t i d e - b i nd in g - li ke) \nPT H R 24 56 7  ( C R P  f a m i ly  t r a n s c r ip t i o n a l  \nreg u la to r y  p ro t e in ) \nGO:00 0 3 700  M: D N A-bin d i ng \nt ra ns c ripti on fa ctor a c t ivity  \nGO:00 0 3 677  M: D N A bi ndin g \nqu iA_ 1  Qui n at e/ shi kim at e  deh y dr o genas e ( q ui- )  \n2,5 62 , 2 8 2  ins  ps e u d o  - -  C O G 4993  Ca r bohydr a t e t r a ns port a nd  \nm e t a b olis m, de hydro g enase  \nFra m e s hif t ad d s 2 2 AA s to \nstart o f g e n e  \nAl l cons erv ed r e g i ons i n ta ct  \nS y s t em ati c G en e ID : 0 3881 hy p otheti cal p r ot ein  \n2,8 73 , 8 8 5  ins  ps e u d o  - -  NOG 2 9 7256  Non sup ervi s ed  o rth ol o g ous gr o ups  Fra m e s hif t at 23 2 / 844 - \n.CC-BY 4.0 International licenseavailable under a \n(which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made \nThe copyright holder for this preprintthis version posted June 27, 2025. ; https://doi.org/10.1101/2025.06.26.659281doi: bioRxiv preprint \n\nB 4  p os i t ion Ty p e CD S/ \np s eu do GO  Te rm s COG s  SN P eff ec t  Disru pte d c on ser v e d re gi on s  \nbr ea k i ng i n t o t w o  prot ei ns  \nch a A _2 S od i u m/ pr o ton an t iport e r Cha A  \nch a A _3  \n3,2 07 , 2 6 2  ins  ps e u d o  - -  C O G 0387  Ino r gani c i o n tr ans p o rt an d  \nm e t a b olis m, ca l ci um p ro t on  \nFra m e s hif t m er ge s chaA _ 2  \nand  ch aA _ 3 wi th  an ext ra 7 \nAA s  b e twee n \nAl l cons erv ed r e g i ons i n ta ct  \nps e u d o  - -  Al l cons erv ed r e g i ons i n ta ct  \nS y s t em ati c G en e ID : 0 4953 N M T1 / THI 5  lik e pr ot e i n  \n3,6 32 , 8 6 3  ins  CDS  - -  - - Fra m e s hif t truncat e s at A A \n26 9 /2 91 \nAl l cons erv ed r e g i ons i n ta ct  \n* S y st em ati c Gen e ID : 0 520 4 h y po t h e t ic a l p r o te i n  \n3 ,83 0 , 3 47 S NP C D S G O : 0 00 59 7 5 B :  c a r b oh y dr a te  me t a b o l i c  \npro c ess  \nC O G 0726  Ca r bohydr a t e t r a ns port a nd  \nm e t a b olis m, 4-ami no-4-deo xy -\nalph a- L-ara b i nopyran o syl  \nun d eca pr enyl pho s pha t e \nbiosynth e ti c p roc e ss  \nMi ss en s e  a t  AA  2 5 2/ 324, A \nto E  \nG 3 D S A : 3. 20 . 2 0 . 37 0  ( gl y c osi d e  \nhy dr ola s e /d e a c et ylas e )  \ncd 1 0 935  (puta t i v e  ca tal y t i c domain of \nli p o po l ysa c ch a ri de bio syn th e sis  pr ot e i n  \nWa l W  a n d  it s  ba c te r ia l ho m ol og s) \nG O : 0 00 38 2 4 M :  c a ta ly ti c  a c ti v it y  \nyji R _ 3  putativ e H TH- ty pe t ran s cri ptional r e gulat or Yji R  \n3,9 62 , 5 8 4  ins  ps e u d o  GO:00 0 3 824  M: ca t a l yt ic act ivi ty C O G 1167  Trans cr ipti ona l  r e gu l a to r, gn t R \nfamil y \nFra m e s hif t r em o v es fi rst \n46/ 1 0 3 AA s  \nIP R 0 1542 2 ;  G 3D SA: 3 . 9 0. 1150. 10  (as p a rtat e \namin o tra nsf er a s e, d o main 1)  \nIP R 0 1542 4 ;  S S F 53 3 83  ( P L P -d epe nde n t  \ntransf eras e s)  \nPT H R 42 79 0 : S F 7  (t r a ns c ripti onal  r e gulato r-\nre l a te d ) \n* *  S yst e m a t i c G en e  ID:  05385  h y po t he t ic al  p r o te i n  \n3,9 67 , 5 4 7  ins  CDS  GO:00 3 2 506  B: cy to kin eti c p ro ce ss  C O G 3266  Fu n ctio n  unkn o w n, D a mX-r el at ed \npro t ei n  \nFra m e s hif t r em o v es fi rst \n38/ 2 8 8 AA s  \nIP R 0 3289 9 ;  MF_ 0202 1  ( C e l l div i si o n prot ei n \nDam X )  GO:00 4 2 834  M: pe ptid ogl ycan bindin g  \nGO:00 3 0 428  C: c el l s e ptum  \n ⎯ pSG4/3 ⎯ \nS y s t em ati c G en e ID : 0 5911 hy p otheti cal p r ot ein  \n636  ins  CDS  - -  - - Fra m e s hif t at A A 5 8/ 64 , \ne x ten d s  to  17 3  \n- \n* *  S yst e m a t i c G en e  ID:  0591 2  h y po t he t ic al  p r o te i n \n3 ,68 0 i n s C D S G O : 0 00 82 3 7 M :  me ta llo p e p t id a se \nacti v i ty  \nC O G 3291  P K D  r epe at  Fra m e s hif t truncat e s at A A \n56 0 /5 73 \nIP R 0 2407 9 ; G3D SA : 3 . 4 0.3 90.10  \n(c oll ag e n as e , c atalyti c dom a i n)  \nS SF55 4 86  (Me t a llo p ro te a se s  (\"z i nc i n s\") , \ncatal ytic do ma i n )  \nPF13 6 88  ( M etal lo-p eptida s e  fami ly M1 2)  \nS y s t em ati c G en e ID : 0 5921 hy p otheti cal p r ot ein  \n10 ,1 4 0  del  CDS  - -  - - Fra m e s hif t truncat e s at A A \n92/ 1 0 6 \n- \n1 0 ,2 69 S NP Mi ss en s e  a t  AA  5 0 , T  to P  \n.CC-BY 4.0 International licenseavailable under a \n(which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. 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