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Usoro nke genome emela ka ngalaba nyocha ọrịa na-efe efe dịghachi ndụ, na-ekpughe ọrịa ọrịa na-efe efe, pathogenesis, mmekọrịta ndị ọbịa-pathogen, na usoro evolushọn nke etinyere na nje. Ngwakọta ụkwara nta nke Mycobacterium (MTBC) na-ele Mycobacterium bovis anya dị ka otu n'ime ụmụ anụmanụ na-emegharị ahụ nke na-ebute ụkwara nta (TB) na anụ mammals nke ụwa, ma bụrụ ihe atụ nke evolushọn nje. Dị ka ndị otu MTBC ndị ọzọ, Mycobacterium bovis na-eche na ọ bụ ihe na-emechi emechi, na-eji nwayọọ nwayọọ na-etolite pathogen, na o doro anya na ọ dịghị ihe ịrịba ama nke recombination ma ọ bụ kehoraizin nnyefe. Na ọrụ a, anyị na-etinye comparative genomics na a dum genome sequence (WGS) dataset mejupụtara 70 ehi M. si dị iche iche pedigrees (Europe na Africa) iji nweta nghọta na mkpụrụ ndụ ihe nketa di iche iche nke ehi M. Evolutionary ike. A na-eji ụzọ atọ dị iche iche tụọ ihe ịrịba ama nke nhazigharị. N'ụwa nile, a chọpụtala na ọnụ ọgụgụ dị nta nke ihe nchịkọtaghachi ma kwadoro site na ụzọ abụọ nwere onwe ha na nkwado siri ike. Ka o sina dị, ma e jiri ya tụnyere mmụgharị, nchikota nwere mmetụta na-esighị ike na ụdị dị iche iche nke M. bovis (n'ozuzu r / m = 0.037). Ọdịiche r/m dị iche nke enwetara na clonal complex nke Mycobacterium bovis na nhazi data anyị kwekọrọ n'echiche izugbe na ogo nchikota nwere ike ịdịgasị iche n'etiti ahịrị ndị e kenyere otu ụdị taxonomic. Dabere na ọrụ a, a pụghị ikpochapụ recombination na Mycobacterium bovis, n'ihi ya, ọ ga-abụ isiokwu nke mgbalị ndị ọzọ na nyocha ihe gbasara genomics n'ọdịnihu, nke WGS nke nnukwu data data sitere na ọnọdụ dị iche iche na-efe efe na gburugburu ụwa dị oke mkpa. A na-eme nyocha ọzọ na obere data Mycobacterium bovis data set (n = 42) site na mgbasa ozi TB nke ọtụtụ ndị ọbịa, na ihe karịrị 1,800 loci ka achọpụtara, nke ọ dịkarịa ala otu ụdị gosipụtara otu nucleotide polymorphism (SNP) . Ọtụtụ (87.1%) dị na mpaghara nzuzo, na oke zuru ụwa ọnụ nke mgbanwe ndị na-abụghị nke mgbanwe (dN/dS) nke mgbanwe mgbanwe karịrị 1.5, na-egosi na nhọrọ dị mma bụ ike evolushọn dị mkpa etinyere na M. bovis. Achọpụtara ọnụ ọgụgụ dị elu nke SNP na mkpụrụ ndụ ihe nketa bara ụba na ụdị ọrụ nke "metabolism lipid", "mgbidi cell na usoro cellular", na "n'etiti metabolism na iku ume", na-ekpughe ikike ha na usoro ndu na evolushọn nke Mycobacterium bovis dị mkpa. Ilebakwu anya na mkpụrụ ndụ ihe nketa na ndị nna nna MTBC nke na-adịkarị mfe ịnyefe mkpụrụ ndụ kehoraizin na gụnyere na usoro 3R (Ndozi DNA, replication, na recombination) na-ekpughe ọnụ ahịa ọjọọ zuru ụwa ọnụ nke ule na-anọpụ iche nke Taijima D, nke na-egosi nyocha nyocha gara aga.
Mycobacterium ụkwara nta complex (MTBC) bụ otu n'ime ihe kasị aga nke ọma taxa nke nje pathogens na-ahụkarị nke nje evolushọn. Ndị otu ya na-egosi njirimara nucleotide dị elu dị ịtụnanya na ọkwa genomic (> 99%)1,2. Ecotypes MTBC dị iche iche nwere ike ịkpata ụkwara nta (TB), nke bụ ọrịa granulomatous na-efe efe, n'ụdị dịgasị iche iche nke ụdị ndị ọbịa sitere na micro-mammals na ụmụ mmadụ3,4,5. Ugbu a, mgbagwoju ahụ gụnyere mmadụ [M. Ụkwara nta (Mtb), Mycobacterium africanum] na ụmụ anụmanụ na-emegharị ahụ (Mycobacterium bovis, Mycobacterium capitum, Mycobacterium pinnipedum, Mycobacterium microtobacter, Mycobacterium mongee, Mycobacterium miysani, Mycobacterium surika, "Bacillus chimpanzee" na "dassie"). M. canettii (nke a makwaara dị ka "Nodobacter glabrata") Nkezi njirimara nucleotide na mycobacteria ndị e kwuru n'elu bụ 98%, na ọrụ comparative genomics egosiwo na M. canettii na ndị ọzọ nke MTBC agbawala n'oge na-adịbeghị anya site na nna ochie.7 N'ịtụle echiche a, ụfọdụ ndị edemede na-akpọ M. Canettii Onye òtù 8.
A kọwapụtara MTBC n'usoro dị ka ihe mgbagwoju anya clonal siri ike, na a na-achịkwa usoro ndị bi na ya nke ọma site na mbelata dị iche iche, mbelata, nyocha nhọrọ, na mkpụrụ ndụ ihe nketa9,10. N'iche na evolushọn siri ike nke clonal siri ike, dị ka polymorphisms efu enweghị ike iweghachi ya site na nchikota. Dabere na ntọala a, ihe omume sochirinụ nke mkpochapụ genomic nke mpaghara dị iche iche (RD) na TbD1 (Mtb kpọmkwem nhichapụ 1 mpaghara) ka atụpụtara dị ka akara molecular nke MTBC evolution2,5,11. Arụ ọrụ comparative genomics na dum genome sequencing (WGS) na-akwado nkewa nke mmadụ na-eme mgbanwe n'ime usoro itoolu (Mycobacterium ụkwara nta L1 ruo L4, L7, na L8; na Mycobacterium africanum L5, L6, na L9), lineages L2 ka L4 na-ekekọrịta ihichapụ TbD1 mpaghara2,131.12. Na mgbakwunye, a na-atụ aro ka ndị otu emeghariri anụmanụ ka ha kekọrịta otu nna ochie, nke akọwapụtara site na nhichapụ clade akọwapụtara na RD7, RD8, RD9 na RD102, 5, na 14.
A na-ewere na mbufe mkpụrụ ndụ kwụ ọtọ (HGT) na mmemme nchikota dị ụkọ ma na-eme na ndị nna nna nke MTBC, karịa akụkọ ihe mere eme dị iche iche nke ndị otu MTBC dum15,16,17. Akụkọ abụọ mbụ nke Hughes and collaborators (2002) na Gutacker and collaborators (2006) na-atụ aro na ihe omume nchikota nwere ike inye aka n'ichepụta polymorphisms nke na-egosi akara a kapịrị ọnụ na M. tuberculosis strains18,19. Ihe kpatara enweghi nchikota doro anya na MTBC bu: (1) usoro nhazi na enweghi ike nke HGT; (2) ihe omume HGT dị ụkọ; (3) enweghi ohere nke nchikota ihe omume na MTBC niche14,17. N'oge na-adịbeghị anya, ụfọdụ ihe ọmụmụ dum-genome sequencing (WGS) na-etinye aka na MTBC strain 20 na Mycobacterium bovis 21 ewepụtala ihe akaebe nke recombination, nke mbụ gosipụtara na ụdị MTBC na-agbanwekarị obere iberibe DNA, ma n'ihi njedebe nucleotide dị iche iche, ihe omume ndị a ka ahụbeghị.
Mycobacterium bovis bụ onye otu MTBC na-enwetakarị site na anụ ụlọ (karịsịa ehi), n'agbanyeghị na ọ nwekwara ike kewapụ ya na anụ ọhịa nwere onwe na nke gbara ogige4,22,23,24. M. bovis malitere n'ime nnukwu ụlọ ọrụ clonal ise [European 1 (Eu1), European 2 (Eu2), European 3 (Eu3), African 1 (Af1) na Africa 2 (Af2)], dị ka spoligotyping profaịlụ, kpọmkwem nhichapụ na otu nucleotide polymorphisms (SNPs) 22,7,26 .8. Mgbagwoju anya clonal ndị a na-egosipụta usoro dị iche iche nke ọnụ ọgụgụ ndị Mycobacterium bovis yana njikọ ya na mpaghara mpaghara. Tụkwasị na nke ahụ, ọrụ WGS na-adịbeghị anya nke Zimpel na ndị na-arụkọ ọrụ (2020) mere na-emepụta phylogeny dabeere na SNP nke Mycobacterium bovis, nke nwere ihe karịrị 1,900 genomes, na-egosi na e nwere ma ọ dịkarịa ala anọ dị iche iche lineages (aha ya Lb1 ka Lb1 ka Lb4), ha na-adịghị kpamkpam kwekọrọ na mgbagwoju anya, ọ bụ ezie na a ga-akọwapụta ya na mbụ. Ndị ode akwụkwọ a mere nyocha dị iche iche nke phylogeny na mkpakọrịta nwoke na nwanyị molecular, mana ha agụghị akwụkwọ recombination30.
Ọrụ gara aga na-eji usoro mkpụrụ ndụ dị iche iche, dị ka spoligotyping, MIRU-VNTR (mycobacterial interspersed repeat unit-variable tandem repeat number), na nso nso a SNP dee gosiri a ụfọdụ larịị nke mkpụrụ ndụ ihe nketa di iche iche n'etiti M. bovis strains 31,32,33, 34,35. Ọdịiche dị iche iche nke mkpụrụ ndụ ihe nketa aghọwo ngwá ọrụ dị mkpa n'ịmụ banyere ọrịa ọrịa na-efe efe, nke na-enye aka maka nghọta miri emi nke pathogenesis, ọrịa nje na mgbasa ọrịa. Mwepụta nke usoro WGS na-enye ohere ikpughe ihe ndị na-anya ụgbọ ala evolushọn nke Mycobacterium bovis genome tinyere n'usoro mgbanwe na nnọgidesi ike na ndị ọbịa dị iche iche na ọnọdụ ọrịa na-efe efe.
Na ọrụ a, anyị na-eji comparative genomic analysis na dị iche iche Mycoplasma bovis datasets (n=70), gụnyere iche na dị iche iche clonal complexes, iji nweta nghọta na evolushọn usoro nke Mycoplasma bovis, karịsịa iji dozie phylogenetic mmekọrịta na recombination omume. Dị ka ihe mgbakwunye na nke a analysis, a sub-data set nke M. bovis ekewapụ (n = 42) nwetara site na nke ọma-jiri mara multi-host ụkwara nta mpaghara na Portugal 31,36 gara n'ihu inyocha na-infer na-abụghị njirimara The itule n'etiti ikwu ruru nke uche (dN) ka synonymous (dS) nucleotide nnọchi, nakwa dị ka evolushọn nke a kpọtụrụ aha na evolushọn. 37,38 nwetara site na MTBC nna nna site HGT, na encode 3R (DNA mmezi, replication, na recombination) usoro gene components 39. Họrọ mkpụrụ ndụ ihe nketa enwetara site HGT n'ihi na ha nwere ike na-anọchi anya oge ochie polymorphisms, n'ihi ya, a na-atụ anya na ha nwere ike inwe a elu nkezi nke synonymous mgbanwe. A na-ahọrọ mkpụrụ ndụ ihe nketa gụnyere na usoro 3R n'ihi na ọrụ gara aga na ụdị ụkwara nta nke M. na-egosi nhọrọ ndị na-adịghị mma / dị ọcha nke na-arụ ọrụ na mkpụrụ ndụ ihe nketa ndị a, ha nwere ike rụọ ọrụ dị mkpa na evolushọn 39. Ihe mgbaru ọsọ ọzọ nke ọrụ a bụ iji chọpụta ịdị adị nke nhazi nhazi. N'ihi nke a, na-atụle na anyị dataset si Portugal naanị nwere genomes nke European clone complex 2 na nsogbu ndị na-enyefeghị mmepụta mmepụta ihe, anyị kpebiri na-agụnye ọha dị genome data n'ikpeazụ nweta onye nnọchiteanya nke niile mmepụta oyiri ogige, na-emeziwanye ike na obosara nke nsonaazụ.
42 ọhụrụ Mycoplasma bovis genomes sitere na mpaghara Portuguese endemic multi-host ụkwara nta (nkọwa n'okpuru), nke e jibu ụzọ mara site na epidemiological echiche36, bụ etiti ọrụ a. N'iburu n'uche na nchịkọta data sitere na Portugal naanị nwere ndị nnọchiteanya nke ụlọ ọrụ mmepụta ihe nke Europe 2 na nsogbu na-enweghị ụlọ ndị a họpụtara ahọpụta, agbakwunyere n'ihu ọha dị dum-genome sequencing data iji gbasaa data set nke na-agụnye ndị nnọchiteanya niile nke M. bovis clone complexes. Ya mere, a na-eji isi mmalite data sequencing atọ zuru ezu na-arụ ọrụ a: mgbakọ genome zuru ezu / draft, ruo 10 scaffolds echekwara na NCBI (National Center for Biotechnology Information) (n = 15 iche); echekwara na SRA ( The Illumina fastq faịlụ nke usoro na-agụ Archive) na-anọchi anya mgbagwoju anya di iche iche nke M. bovis clones (n = 12 iche) 30; yana mkpụrụ ndụ ihe nketa iri anọ na abụọ sitere na Pọtugal. Mycobacterium bovis BCG (Bacille Calmette-Guerin) ewepụrụ na nchọ NCBI. A na-ejikarị M. bovis AF2122/97 mee ihe dị ka genome ntụaka ka etinyere na dataset. N'ihi enweghị ọhaneze nke usoro genome dum nke African 1 cloning complex nọchitere anya ya, yana ọnụ ọgụgụ dị nta nke genomes sitere na ụdị ndị nnọchite anya Af2 na Eu1, ejiri data usoro izizi nke SRA nyere mee ihe n'okwu ndị a. Ọrụ Zimpel na ndị ya na ha na-arụkọ ọrụ (2020) nyere aka chọpụta genome sitere na ogige cloning ahụ e kwuru n'elu ma nyere aka họrọ Mycobacterium bovis maka ntinye na nhazi data. Maka Eu3, ọ bụ naanị otu ụdị genome ka akọwara (Branger et al., 2020), yabụ genome anyị gụnyere bụ onye nnọchi anya dị iche nke ogige Eu3.
N'ụwa nile, nke a data set na-agụnye 70 ehi M. bovis dịpụrụ adịpụ site na ụdị 8 ndị ọbịa, kesara na mba 12 site na 1985 ruo 2016. A na-ahọpụta ụdị 36 dị ka Eu2, ụdị 7 bụ Eu1, ụdị 1 bụ Eu3, ụdị 3 bụ Af1, ụdị 4 bụ Af2 na 19 adịghị mgbagwoju anya n'okpuru ebe ọ bụla). Ozi zuru ezu (gụnyere nọmba ntinye) nke Mycobacterium bovis ejiri mee ihe n'ọmụmụ ihe a ka egosiri na Tebụl 1 na Tebụl mgbakwunye 1.
42 ọhụrụ sequenced dum genomes nke Mycobacterium bovis si Portugal si anụmanụ ụkwara nta hotspot na ekesa ihe karịrị 12 afọ bụ isi nke ọmụmụ a, dị ka nwere ike anụ ọhịa-anụ ụlọ ọrịa usoro na-mgbe niile nyochaa 31,36 (Mgbakwunye fig. 1). Dị ka usoro ndị sochirinụ si dị, ụdị anụ ahụ dịpụrụ adịpụ na anụ ụlọ (n = 14), mgbada uhie (n = 16) na anụ ọhịa (n = 12) site na 2003 ruo 2015: na-anakọta ma na-ejikwa anụmanụ dị ka ụkpụrụ ụkpụrụ nduzi akwadoro. Nlere anụ ahụ dị na OIE Terrestrial Animal Handbook na a na-etinyekwa ya na Stein-Jent-Jent na Stroculated. mmiri mmiri ọkara. A na-etinye omenala ndị a na 37 Celsius C ma na-enyocha uto otu ugboro n'izu ma ọ dịkarịa ala izu iri na abụọ. A na-echekwa ógbè ndị ahụ ozugbo na ngwọta glycerol na -80ºC. N'ime usoro nhọrọ Mycobacterium (Middlebrook 7H9, BD Diagnostics), a na-agafe ihe nlele mbụ echekwara site na otu ụzọ in vitro in vitro iji nweta DNA nke mmemme WGS. Maka nke a, ejiri 5% sodium pyruvate na 10% ADS (50 g albumin, 20 g glucose, 8.5 g sodium chloride na 1 L mmiri) mee ka ihe ngwọta omenala oyi kpọnwụrụ na Middlebrook 7H9 na 37°C Retrain. Mgbe izu 4 gachara, a na-emegharị usoro ahụ ma na-eleba anya na omenala ahụ mgbe niile ruo mgbe a hụrụ uto. A na-egbute mkpụrụ ndụ ndị ahụ site na centrifugation, a na-eweghachi pellet ahụ na 500 µL phosphate buffered saline (PBS), na-ekpo ọkụ na 99 Celsius C maka nkeji 30, centrifuged, na-echekwa elu na -20 Celsius C ruo WGS. A na-eme usoro niile na ụlọ ọrụ biosafety ọkwa 3.
A na-akwado ọba akwụkwọ genome nke WGS jikọtara ọnụ site na iji ndeksi pụrụ iche nke ihe nlele DNA ọ bụla, ma na-eji Illumina MiSeq (2 × 250 pb) (40 samples) na HiSeq (2 × 150 pb) (abụọ iche) teknụzụ (Eurofins Genomics, Germany) maka nhazi usoro. Dịka ntuziaka onye nrụpụta si dị, jiri Illumina Genome Analyzer nwere mgbakwunye modul nwere ngwụcha abụọ iji hazie DNA mkpụrụ ndụ ihe nketa, ma jiri Nextera XT DNA Prep Kit si Illumina wuo ọbaakwụkwọ ahụ.
N'iburu n'uche data enwetara site na SRA (n = 12), njirimara nke mgbagwoju anya nke clone nwere ike iji mee ihe dị ka metadata nke mbipụta kwekọrọ na 30, 41, 43. Mgbe ị na-atụle genome zuru ezu, ma e wezụga maka Mycobacterium bovis AF2122/97 na Mycobacterium bovis 3601, nke a ghọtara na 12 na ndị E5, nke na-aghọta 12 na ndị E5. n'otu n'otu, ọ bụ otu ihe ahụ dị ka genome zuru oke nke Mycobacterium ụkwara nta H37Rv (NCBI accession number NC_000962.3). A na-eme nhazi genome site na iji MAFFT (usoro nhazi ọtụtụ amino acid ma ọ bụ usoro nucleotide, ụdị 7.458) na parameter -addfragments48. Mgbe ahụ, chọọ maka enweghị ogige clonal dị iche iche na / ma ọ bụ ọnụnọ nke njirimara SNP.
Mycobacterium bovis (n = 42) ọhụrụ a na-agụ na mbụ na-agụ nke genome draft (n = 3) kwadoro mgbagwoju anya na genome Mycobacterium tuberculosis H37Rv site na vSNP pipeline na ọnụnọ nke nhichapụ na / ma ọ bụ SNP atụmatụ nke clones dị iche iche A na-eme ọchụchọ.
Na-anakọta ozi site na enweghị njirimara na/ma ọ bụ ọnụnọ / enweghị SNP na profaịlụ spoligotyping iji kenye data genomic na mgbagwoju clonal kwekọrọ. Maka mgbakọ akwụkwọ anọ ahụ, enweghị ike ịpụta profaịlụ spoligotyping, yabụ etinyere ha na otu "enweghị mgbagwoju anya".
Usoro ọrụ bioinformatics na-esochi ọrụ a na-amalite site na mgbakọ de novo na nkewa na atụmatụ ntụnye aka, na ebumnuche nke inyocha ihe omume nchikota yana kpọmkwem genome polymorphisms. Onyonyo 1 na-enye eserese ngosi nke usoro ndị a na-eso. Maka nyocha nchikota, a na-eji genome niile mee ka ike nke ntinye aka na ihe ndị metụtara ya pụta.
Iji belata njehie na ọgbọ nke genome consensus usoro, anyị na-ebu ụzọ nweta mgbakọ de novo, wee nweta isi ọtụtụ nhazi. Pipeline Unicycler dị ugbu a na https://github.com/rrwick/Unicycler49 ma ejiri ya rụọ mgbakọ de novo nke genomes usoro 54 (faịlụ 42 ọhụrụ na 12 fastq enwetara na SRA). Na nkenke, tupu ịgbakọta site na ọkọ, a na-eme nyocha ịdị mma na ụdị FastQC 0.11.7 (https://github.com/s-andrews/FastQC), na ụdị Trimmomatic 0.36 (nhọrọ "ịcha ihe nkwụnye ọkụ na usoro ọkụ ndị ọzọ si na-agụ" na "Bepụ ntọala site na njedebe nke 2) ma ọ bụrụ na a na-etinye ya na njedebe nke 0. (http://www.usadellab.org/cms/?page= trimmomatic) 50. Mgbe ahụ, a na-eji SPAdes optimiser49 mee ihe maka mgbakọ genome, na nsụgharị Pilon 1.1851 na-eji maka njikarịcha mgbakọ na mwepụ. Ahọpụtara ụkpụrụ nchikota nke nchekwa iji zere mgbakọ na-ezighi ezi, wee chọọ nha k-mer wee họrọ n'etiti 20% na 95% nke ogologo ọgụgụ. Soro ntuziaka SPAdes ma tụlee nha ọgụgụ, hichapụ contigs dị obere karịa 300 bp, ma guzobe 20 na-agụ ihe mkpuchi mkpuchi omimi nke 52. N'ime usoro mgbakọ de novo, mpaghara genomic dị ka proline-glutamate (PE) na proline-proline glutamate (PPE) paralogs adịghị ewepụ ya.
A na-enyocha ogo nke mgbakọ de novo site na pipeline QUAST (http://quast.sourceforge.net/quast.html), nke na-eme ka mmeghari ohuru nke contig na M. bovis AF2122/97 genome (NCBI accession number LT708304.1) Mapping (lee Tebụl mgbakwunye 1 maka ogo mma).
Site n'enyemaka nke vSNP pipeline (https://github.com/USDA-VS/vSNP), FASTQ faịlụ nke ọhụrụ usoro M. bovis si Illumina sequencing tụnyere M. bovis AF2122/97 genome (LT708304.1)) . Dabere na ndụmọdụ omume kacha mma nke Genome Analysis Toolkit (GATK) 53, 54, 55 na-etinye parampat nzacha ọkọlọtọ ma ọ bụ akara oke dị iche iche maka nhazigharị. A na-enyocha nsonaazụ ya site na iji akara uka SAMtools kacha ala nke 150 yana AC = 2. Jirikwa Kraken (http://ccb.jhu.edu/software/kraken/) iji lelee ọgụgụ ndị ahụ iji wepụ mmetọ. Pipeline vSNP eji akọwapụta usoro usoro n'ọrụ anyị na-enyocha usoro akọwapụtara SNP na ebumnuche, ma wepụkwa ọnọdụ ọrịa agwakọta. Ihe mkpuchi genome agụ dị mma karịa 99% (Ntinye mgbakwunye 1).
Iji zere mmejọ eserese na SNP na-ezighi ezi, kpochapụ ihe dị iche iche n'ọnọdụ ndị a: (1) na-akwado ya site na ihe na-erughị 20 agụ, (2) a na-ahụ ya na ugboro ole na-erughị 0.9, (3) ọ bụ na ọ dịkarịa ala otu Strains, ma ọ dịkarịa ala enwere oghere na ụdị ọzọ. The integrated genomics viewer (IGV) ụdị 2.4.19 (http://software.broadinstitute.org/software/igv/)56 e ji anya nyochaa SNPs na ọnọdụ nwere maapụ ma ọ bụ alignment nsogbu. Ebe ọ bụ na proline-glutamate (PE) na proline-proline glutamate (PPE) genes bụ nke ukwuu oyiri na bụ akụkụ nke multi-gene ezinụlọ, ha na-adị mfe nghọta site Illumina sequencing na mismapping, n'ihi ya, ha na-ahọrọ The mycobacterial bioinformatics workflow wepụrụ ndị òtù nke ụkwara nta na mgbagwoju mgbe eji atụmatụ nke nkewa ka usoro SNP. Ya mere, anyị wepụrụ mkpụrụ ndụ ihe nketa PE/PPE na indel na nyocha.
Dịka Bovilist si kwuo (http://genolist.pasteur.fr/BoviList/), SNP niile na-ekewa n'ime ụdị ọrụ. A na-eji ọkpọkọ SnpEff (https://pcingola.github.io/SnpEff/) na-enye nsonaazụ SNP (mgbanwe ndị na-enweghị aha ma ọ bụ enweghị aha). Mepụtara nchekwa data ọhụrụ nke Mycobacterium bovis AF2122/97 genome (LT708304.1).
Ejiri genome multiple alignment rụọ ọrụ site na iji Parsnp v1.2, dị ugbu a na https://github.com/marbl/parsnp57, na-eji 69 zuru ezu genomes / draft assemblies (na nhọrọ -c) na M. bovis AF2122/97 (LT708304.1) na-eji dị ka ntụaka. A na-eme nhazi anọ dị iche iche: naanị ndị òtù Eu2 cloning complex (n = 37), gụnyere ndị niile so na European cloning complex (n = 44), gụnyere njikọ njikọ nke European na African cloning complex (n = 51), ma tinye Mycobacterium bovis niile n'ime ọmụmụ ihe a (n = 70).
A na-eji isi nhazi nke Parsnp mepụtara iji mee ka osisi phylogenetic kachasị (ML) na-eji CIPRES Science Gateway v3.3 (http://www.phylo.org/)58 site na iji RaxML, ma mee 1000 eduzi emegharị.
A na-eji algọridim dị iche iche atọ dị iche iche na ngwaọrụ bioinformatics iji chọpụta ọnụnọ nke mmemme nchikota n'otu n'otu: SplitsTree4 software, Gubbins (ahịrị na-enweghị mmasị site na nchikota n'usoro nucleotide) pipeline, na RDP4 (mmemme nchọpụta nyocha, ụdị beta 4.101).
A na-eji usoro ire ere nkewa nke etinyere na SplitsTree4 v4.15.1 (http://www.splitstree.org/) 59 iji gbakọọ netwọk phylogenetic na-enweghị mgbọrọgwụ, na-eji ule Phi maka nkwenye ndekọ ọnụ ọgụgụ, yana ọnụ ụzọ dị mkpa bụ p = 0.05. A na-eji isi multi-alignment analysis of Parsnp mee ihe dị ka ntinye, na nbibi nkewa dị ka ọkọlọtọ netwọk na-enweta.
Gubbins pipeline v2.3.1 (https://github.com/sanger-pathogens/gubbins60 na-agba ọsọ na ndabara paramita dị ka ụzọ ọzọ iji nyochaa mmetụta nke recombination na Mycobacterium bovis. The algọridim emejuputa atumatu na pipeline reconstructs mkpa clone lineage The zuru ezu genome/draft mgbakọ nke data anyị set na genome genome/AF721) LT708304.1).
N'ikpeazụ, iji kwado mmemme nhazigharị nke ọkpọkọ Gubbins tụrụ aro, algọridim isii ahụ etinyere na RDP467 (RDP61, GENECONV62, Bootscan63, Maxchi64, Chimaera65 na SiScan66) na-etinye aka na nhazi ọtụtụ Parsnp n'okpuru ntọala ndabara. Anyị kpebisiri ike na opekata mpe atọ n'ime algọridim emejuputa atumatu na RDP4 ga-abụrịrị na-egosipụta akara ngosi dị mkpa mgbe niile iji nyochaa mmemme nchikota nke ọ bụla.
N'iburu n'uche na ma Gubbins na ngwanrọ RDP na-achọ akara ngosi nchikota site na ịlele isi ọtụtụ nhazi na windo ruru 500 bp, na-akwado na ntinye nke mkpụrụ ndụ ihe nketa PE/PPE n'oge mgbakọ de novo agaghị egbochi ya na mgbama nrụgharị ahụ achọtara, a na-eme nyocha ọzọ site na homolinearity Lelee nso nso nke mkpụrụ ndụ ihe nketa na-achọpụta recomination. Ejiri MAUVE-multi-genome alignment (http://darlinglab.org/mauve/mauve.html) rụọ maapụ synlinear site na iji genome zuru ezu iji wepụ ntụgharị genome mpaghara ma ọ bụ ntụgharị. Na mgbakwunye, ejiri genome dum mee nyocha homolinearity na usoro amino acid site na sava weebụ SyntTax (https://archaea.i2bc.paris-saclay.fr/SyntTax/).
Ntụle dị omimi karị nke data genome nke enwetara site na usoro ụkwara nta nke ọtụtụ ndị ọbịa na Portuguese bụ ịlele polymorphism nke mkpụrụ ndụ ihe nketa ndị a kpọtụrụ aha na akwụkwọ. Mkpụrụ ndụ ihe nketa ndị a bụ 37,38 na mkpụrụ ndụ ihe nketa 3R nke ndị nna nna MTBC nwetara site na HGT (Ndozi DNA, replication na recombination) usoro components 39. Jiri ClustalX v2.1 (http://www.clustal.org/clustal2/) wee jiri DnaSP v6.12.03 (http://wwwsp.ub.) (π) na ntinye paramita na-anọpụ iche nke Tajima D.
O yikarịrị ka osisi phylogenetic (ML) dabere na 69 Mycoplasma bovis kewapụrụ na mkpụrụ ndụ ihe nrụtụ aka enwetara (Njirimara 2A). E jiri ya tụnyere otu mkpụrụ ndụ ihe nketa ma ọ bụ osisi ndị nwere ọtụtụ mpaghara, atụmatụ a na-enye ohere maka ọgbọ nke osisi ndị dị ike karị nke na-adịghị ejide mgbanwe nke genome dum na ya mere na-egosipụta ikike dị ala nke ịkpa ókè n'etiti ụdị 68,69. Ọdịdị topological nke osisi ML na-adịkarị na nhazi mgbagwoju anya nke clones. A na-agbakọta genome nke Eu2 n'otu alaka, na genome nke Af1 na-agbakọtakwa ọnụ (Figure 2A). Ihe si na ya pụta na-adaba na mmekọrịta evolushọn a maara nke Mycobacterium bovis, ya bụ, enwere nnukwu ọdịiche dị n'etiti onye òtù Eu1 na otu nke mejupụtara ogige clonal ndị ọzọ na genome, mana akọwapụtaghị ihe mgbagwoju anya clonal 30. Enwere ike ịkọwa ntakịrị ihe na-ekwekọghị n'etiti ihe mgbagwoju anya nke clonal na mmekọrịta a na-ahụ na osisi phylogenetic site n'eziokwu ahụ bụ na a na-akọwa ihe mgbagwoju anya nke na-adabere na mpaghara genomic a kapịrị ọnụ, ebe osisi phylogenetic na-adabere n'ọtụtụ nhazi nke mkpụrụ ndụ ihe nketa na-anọchi anya genome dum.
A na-ewu osisi phylogenetic kacha (GTR) dabere na nhazi mkpụrụ ndụ ihe nketa nke Mycobacterium bovis genome tupu (A) na mgbe (B) mwepụ nke saịtị nchikota. Agba alaka ahụ na-anọchi anya ihe mgbagwoju anya Mycobacterium bovis: Europe 1 bụ odo odo, Europe 2 na-acha uhie uhie, Europe 3 bụ acha anụnụ anụnụ, Africa 1 bụ oroma, Africa 2 bụ akwụkwọ ndụ akwụkwọ ndụ. Osisi ahụ gbanyere mkpọrọgwụ ma dọta ya na ọnụ ọgụgụ, a na-atụkwa ogologo alaka ahụ ka ọ bụrụ nnọchi maka saịtị ọ bụla.
A na-akọwa mgbagwoju anya ụkwara nta nke Mycobacterium dị ka nke sitere na clonally, na ọtụtụ n'ime ihe akaebe a na-agbakọ kemgbe ọtụtụ afọ na-akwado echiche ahụ na HGT na-aga n'ihu na ihe nchịkọtaghachi agaghị eme na ọkwa nke MTBC15,17,18.
Ọrụ gara aga egosila na enwere ike ịnwe oke nchikota n'etiti MTBC strains20,21, ebe ndị ọzọ achọpụtaghị ihe omume nchikota nke enwere ike ịtụnye70,71. Tụgharịa okwu a na-elekwasị anya na Mycobacterium bovis, nke dị iche na ọrụ gara aga nke na-atụle naanị Mycobacterium ụkwara nta 70,71; ma ọ bụ tụlee MTBC n'ozuzu, na ọ fọrọ nke nta ka ọ dịghị M. bovis na-anọchi anya 20; ma ọ bụ tụlee naanị ụmụ irighiri ehi na-amachibidoro. Ihe ndekọ data nke mycobacteria, na ọrụ a, enwere ngụkọta nke ụdị 70, na-anọchite anya ogige clonal niile, nke a na-eji nyocha maka nhazigharị. A na-atụba ntọala data ahụ dịka ọkwa nchịkọta anọ si dị: (1) Ndị òtù Eu2, (2) ndị otu mgbagwoju anya nke Europe (ie Europe), (3) Europe na African clone complex (Eu + Af) na (4) mkpokọta data (gụnyere genomes ndị na-adịghị etinye na ogige cloning ọ bụla akọwara).
Iji mụọ echiche a n'ihu, a na-eme netwọk nkewa nkewa iji chọpụta enweghị ihe omume nchikota n'etiti genomes, n'ihi na usoro a nwere ike iji anya nke uche hụ mmekọrịta ndị nna ochie n'etiti ndị mmadụ n'otu n'otu ma gosipụta akara ngosi phylogenetic na-emegiderịta onwe ya. Ihe ndekọ data anọ niile na nyocha ahụ kwadoro ịdị adị nke loops na netwọk (ya bụ, mpaghara ndị na-adịghị abanye n'ime otu osisi), ma ule Phi enweghị nkwado ndekọ aha (Eu2, p = 0.0956; Europe, p = 0.1637; Eu + Af p = 0.2774; data dum set p = 0.245) na-enye ihe akaebe na-adịghị mma maka ihe omume reorganization. 3A-D).
Na Europe 2 genomes (n = 37) (A), European genomes (n = 44) (B), European na African genomes (n = 51) (C) na dum data set (n = 70) (D).
Mgbe nyocha a gasịrị, ma na-eburu n'uche ihe nlele cyclic na netwọk niile, a na-etinye usoro nrụgharị ahụ na-arụ ọrụ na pipeline Gubbins iji wughachi usoro nke clonal na iji gbakwunye atụmatụ nke mmetụta nke recombination na M. bovis genome. Wepụta ọnụ ọgụgụ mmụkọ nke mmemme nchikota, nke ọtụtụ n'ime ha mere na ngalaba njedebe (ya bụ, n'ime otu genome) (Table 2). Ihe ngosi ndị a na-egosi nkwekọ nke data niile ma na-egosi na ugboro ole ihe omume nchikota bụ ugboro 200 ruo 300 nke mgbanwe. Ozugbo paramita rho / theta na-anọchite anya ọnụ ọgụgụ ndị ikwu nke recombination na ngbanwe dị na alaka ụlọ ọrụ na-egosi na ọ dị n'etiti 0.0037 na 0.0056 (Table 3). N'oge na-adịbeghị anya, ọrụ e bipụtara nke 38 M. bovis strain gosipụtara ọnụ ahịa rho / theta dị elu (rho / theta = 0.1) karịa nke enwetara na dataset a, ma ọrụ nke Patané na ndị ọrụ ibe na-eji mgbakọ na-ezo aka na-eme ka e nwee mgbakọ na-eme ka ọ bụrụ ihe nchịkọta recombination .
Na-esote, paramita r / m na-anọchite anya ọnụọgụ dị iche iche nke nchịkọtaghachi na mmeghe mmụgharị, na nkezi uru ya dị n'etiti 0.025 na 0.037, na-egosi na e jiri ya tụnyere mgbanwe mgbanwe, nchikota nwere mmetụta dị ala n'ozuzu ya na mkpụrụ ndụ ihe nketa dị iche iche nke M. bovis (Table 3)). Maka ntụnyere sara mbara, a na-eji usoro yiri nke ahụ tụọ r / m parameter maka MTBC dataset nke mejupụtara 23 genomes, na-egosi nkezi uru nke 0.48620, ebe maka Patane na ndị ọrụ ibe 38 M. bovis dataset, ọ gosipụtara nkezi uru bụ 0.98. N'ime ọmụmụ ihe mbụ, ọ bụ naanị abụọ n'ime 23 genomes gụnyere na ọrụ nke M. bovis (M. bovis BCG na nrụtụ aka), n'ihi ya, uru a na-enweta nwere ike bụrụ nke na-adịghị mma n'ihi oke nke M. tuberculosis genome. N'ime akụkọ nke abụọ, ndị Mycobacterium bovis nyochara enwetara nke ọma na United States na ndị ọbịa anụ ụlọ. N'ụzọ dị iche, na nhazi data anyị, a na-anọchi anya ọnọdụ mpaghara na ụdị ndị ọbịa, a na-ejikwa genomes gbakọta n'ime ogige clonal dị iche iche nwere njirimara mkpụrụ ndụ dị iche iche nke ọnụ ọgụgụ mmadụ, si otú a na-enweta ihe ọmụma miri emi na nke sara mbara karị. Ihe dị iche r / m nkezi uru enwetara na data data anyị kwekọrọ na echiche na ogo nke recombination dịgasị iche iche dị iche iche n'etiti usoro ọmụmụ e kenyere otu ụdị taxonomic, ya mere, nsonaazụ ndị a na-egosi na M. bovis clone complex nwere ike igosi ọdịiche dị iche iche recombination Mmetụta bụkwa dị ka a tụrụ aro Didelot & Maiden72. Ka o sina dị, ịgbasawanye data a nke ukwuu site n'itinye ọnụ ọgụgụ ka ukwuu nke mkpụrụ ndụ ihe nketa M. bovis ga-enye ohere maka nkọwakwuo nke isi ihe a. Ma r / m na rho / theta parameters na-egosi mgbanwe dị n'etiti alaka, na nke a na-adabere na akụkọ banyere ụdị nje ndị ọzọ72,73.
N'ikpeazụ, iji kwado mmemme nhazigharị nke ọkpọkọ Gubbins mara, ejirila algọridim isii dị iche iche na ngwa RDP4 iji nwalee ihe atụ dị iche iche dị iche iche. N'ụwa niile, ihe na-erughị ọkara nke ihe omume Gubbins mara ọkwa bụ nke RDP4 kwadoro (Table 4 na 5). N'iburu n'uche ihe niile data setịpụrụ, a kwadoro ihe omume nchikota atọ, abụọ metụtara oghere ime na nke ọzọ metụtara otu genome n'ime alaka ụlọ ọrụ, nke enweghị ike ikenye ogige clonal (Table 4 na 5). Nchọpụta ihe omume na ngalaba ọnụ nwere ike igosi na nchikota a ka na-aga n'ihu na ụdị M. bovis nke oge a ma ọ bụ nsonaazụ na-ezighi ezi70. Na mpaghara nchikota echiche a, ihe dịka 20% nke ọnọdụ nwere nucleotides na-akọwapụtaghị (N), si otú a na-emetụta akara ngosi recombination (Mgbakwunye 2). Na mgbakwunye, mpaghara a na-emetụta mkpụrụ ndụ rrs, na-edobe 16S ribosomal RNA nke a na-atụ anya na a ga-echekwa ya nke ukwuu, yabụ mgbama nchikota putative a nwere ike ịbụ nsonaazụ nke njehie n'usoro ma ọ bụ enweghị nhazi. Mgbe ahụ dum genome alignment n'etiti Mb0003 na Mycobacterium bovis AF2122/97 e mere, na ịdị adị nke undefined nucleotides na SNPs kwadoro, n'ihi ya, nsogbu ndị nwere ike metụtara na-ezighị ezi alignment abụghị n'ihi na ndu ozi emejuputa atumatu na ọrụ a pụtara mgbe ịmụta usoro.
Enweghị oghere ma ọ bụ nucleotides na-akọwapụtaghị n'ime mpaghara nchịkọta nke ọnụ ọnụ dị n'ime (Figure 4 na 5). Banyere ihe omume ndị a, otu nwere naanị Eu2 genome ma na-emetụta mkpụrụ ndụ pks12, nke na-eme ka polyketide synthase nwere ike; ebe nke ọzọ edebanyere aha na Eu1 genome ma na-emetụta mkpụrụ ndụ narX nke na-eme ka ọ bụrụ nitrate reductase (Table 4) . N'ozuzu, nyocha nke nchikota na-egosi na e nwere ọnụ ọgụgụ dị nta nke nchịkọta nchịkọta na nkwado ndekọ ọnụ ọgụgụ, na ihe ndị na-egosi na-egosi na nchịkọtaghachi nwere mmetụta dị ala na usoro M. bovis. A na-atụ anya na mgbama nchikota ga-adị ala, mana ọ dị mkpa ịmata ọdịiche dị n'ezi evolushọn sitere na mkpọtụ azụ, nke bụ ọrụ siri ike. Iji belata ụda mkpọtụ ewepụtara site na mgbakọ na-adabere na nrụtụ aka na nsogbu 70, 71, ndị ọzọ niile ma e wezụga genome zuru ezu na-agbakọta site na ọkọ, na-enyocha àgwà mgbakọ ahụ ma hụ na nyocha QUAST pipeline (Nkwupụta 1) . Tụkwasị na nke ahụ, e mere usoro nyocha nke mgbakwunye iji nye ike na izi ezi nke nyocha n'ozuzu ya. Ya mere, a tụlere usoro nhazi nke mkpụrụ ndụ ihe nketa narX na pks12 site na eserese ọgụgụ megide Mycobacterium bovis AF2122/97. A kwadoro ọnọdụ SNP akwadoro na mpaghara recombination site n'itinye njirisi a kpọtụrụ aha na ngalaba usoro (ọ dịkarịa ala 20 na-agụ na 0.9 mgbanwe ugboro ugboro). A kwadoro polymorphism nke mkpụrụ ndụ narX n'ụzọ zuru ezu na genome abụọ (Mb1792361 na Mb7240415; 2.3%) na mkpụrụ ndụ ihe nketa nke pks12 genome: Mb0891, Mb1711, Mb1789, Mb1870, Mb17046, Mb1256, na Mb1256. Otú ọ dị, maka genome Mb2043, isii n'ime ọnọdụ asatọ ahụ adịghị ezute ihe omimi nke ịgụ ihe n'ihi na SNP na-akwado 17 na-agụ, nke dị n'okpuru ọnụ ahịa cutoff guzobere nke 20. Ya mere, enwere ike ịkwado recombination nke genomes isii (8.6%) na saịtị genome a (Njirimara 4 na 5).
Ọhụụ zuru ezu nke nhazi mpaghara nhazigharị nke Mycobacterium bovis dataset na-emetụta mkpụrụ ndụ narX nke na-etinye koodu nitrate reductase ga-ekwe omume. Enweghị oghere ma ọ bụ nucleotides na-akọwapụtaghị nke a chọtara na mpaghara nchikota nke ọnụ ime. Edere ihe omume a na genome nke Eu1. A tụlere ogo usoro nke mkpụrụ ndụ ihe nketa narX site n'ichepụta ọgụgụ nke Mycobacterium bovis AF2122/97. Kwado ebe SNP akwadoro na mpaghara nchikota site n'itinye njirisi a kpọtụrụ aha na ngalaba usoro (opekata mpe 20 na-agụ na 0.9 mgbanwe ugboro). A kwadoro polymorphism nke mkpụrụ ndụ narX n'ụzọ zuru ezu na genome nke Mb1792361 na Mb7240415 (2.3%).
Nleta zuru oke nke nhazi mpaghara nchikota nke Mycoplasma bovis dataset na-emetụta mkpụrụ ndụ pks12. Enweghị oghere ma ọ bụ nucleotides na-akọwapụtaghị nke a chọtara na mpaghara nchikota nke ọnụ ime. Banyere mmemme na-emetụta mkpụrụ ndụ pks12 na-etinye koodu polyketide synthase, ọ nwere naanị genome Eu2. A tụlere ogo usoro nke pks12 site na maapụ ọgụgụ Mycobacterium bovis AF2122/97. Kwado ebe SNP akwadoro na mpaghara nchikota site n'itinye njirisi a kpọtụrụ aha na ngalaba usoro (opekata mpe 20 na-agụ na 0.9 mgbanwe ugboro). A kwadoro polymorphisms nke genome Mb0891, Mb1711, Mb1789, Mb1870, Mb1758, Mb2043, na Mb1960.
Mkpụrụ ndụ PE na PPE nwere mpaghara ugboro ugboro nke na-adị mfe nghọta site na Illumina sequencing na mismapping, n'ihi ya, a na-ehichapụkarị ha na bioinformatics workflow nke M. ụkwara nta naanị mgbe ị na-eji atụmatụ nkewa-na-usoro. Ntinye nke ihe nchikota nke etinyere na oru a dabere na nzuko de novo na-eweputa PE/PPE. Anyị kwenyere na site n'itinye usoro nkwado atọ dị iche iche na algọridim site na SplitsTree, Gubbins pipeline na RDP4 software, atụmatụ ndị etinyere na-esi ike na nhazi na nzacha mpaghara ahazigharịrị site na mgbaama njehie kpatara. Otú ọ dị, iji wepụ nnyonye anya nke PE / PPE gene na Gubbins na RDP4 software iji chọpụta ụyọkọ SNP, ya mere, njirimara nke mpaghara recombination na-atụ aro ka ọ na-emetụta mkpụrụ ndụ narX na pks12, a na-enyocha agbata obi nke mkpụrụ ndụ ihe nketa ndị a (Mgbakwunye 3-5). Na M. bovis AF2122/97, a na-ekewa mkpụrụ ndụ narX site na narK2 na Mb1764c, ebe pks12 gbara ya gburugburu Mb2075c e Mb2073c (Ngbakwunye 3-5). Maapụ a na-emepụta site na iji maapụ synline MAUVE nke genome zuru ezu na-enye ozi gbasara nchekwa na nhazigharị usoro mkpụrụ ndụ ihe nketa, na-egosi blocks anọ, na enweghị akara nke ntụgharị genome ma ọ bụ ntụgharị. Na mgbakwunye, nyocha nke mmeju na usoro amino acid gosipụtara homology n'ime genomy niile zuru oke, ọ nweghịkwa PE/PPE achọtara na mpaghara dị nso nke narX ma ọ bụ pks12. Maka narX, otu genome (Mb0030) nwere akara ntukọ dị ala n'ihi na a chọpụtara na mkpụrụ ndụ narX dị ka iberibe abụọ (iberibe 1891 na 1890). Maka pks12, n'ihi myirịta, Mb0030 na Mb003 gosipụtara akara synlinearity dị ala, ebe a chọpụtara pks12 n'iberibe abụọ na atọ, n'otu n'otu, na-anọchite anya ngalaba dị iche iche nke protein (Nkọwapụta 3-5). N'iburu n'uche ozi a, na Gubbins na RDP4 software na-eme nyocha, lelee isi nhazi nke 500 bp kacha elu na windo, anyị kwadoro na PE/PPE gene agaghị egbochi mgbaama nchikota nke na-emetụta narX na pks12.
Ọ bụ ezie na a na-ewere akara ngosi recombination achọpụtara na data data a dị ka ihe fọdụrụnụ, ọ bụ ezie na a pụghị ịchịkwa recombination na M. bovis, n'ihi ya, ọ ga-anọgide na-abụ isiokwu nke nyocha ọzọ, bụ nke a na-edepụta genome niile sitere na ọnọdụ dị iche iche nke ọrịa epidemiological dị mkpa.
N'iji atụnyere osisi phylogenetic ML enwetara tupu na mgbe emezigharị mmezigharị (Njirimara 2A, B) edugaghị na mgbanwe dị ukwuu na mmekọrịta phylogenetic nke a na-eche, na ụdị M. bovis na-agbakọta n'otu otu.
Ka emechara eserese 42 ọhụrụ usoro M. bovis na-agụ ya na genome nke M. bovis AF2122/97, enwetara nhazi SNP nwere ọnọdụ 1816 polymorphic. Ọtụtụ SNPs (87.1%) dị na mpaghara nzuzo, na mkpụrụ ndụ ihe nketa emetụtara dị ka ụdị ọrụ egosipụtara na Bovilist (Nyocha 6A, B). N'iburu n'uche ngụkọta ọnụ ọgụgụ nke mkpụrụ ndụ ihe nketa na ụdị ọrụ ọ bụla, mkpụrụ ndụ ihe nketa dị na ụdị "metabolism lipid" gosipụtara SNP ndị ọzọ, na-esote "mgbidi cell na cell" na "n'etiti metabolism na respiration", na-ekpughe na ha nọ na M. bovis evolushọn.
Nyocha usoro nhazi nke dataset M. bovis sitere na Portugal (n = 42). Ngụkọta ọnụ ọgụgụ SNP edebanyere aha yana mkpụrụ ndụ ihe nketa metụtara maka ụdị ọrụ ọ bụla (A). Ngụkọta ọnụ ọgụgụ nke mgbanwe mgbanwe na enweghị aha debara aha site na ngalaba ọrụ (B).
N'ọ̀tụ̀tụ̀ zuru ụwa ọnụ, nkezi dN/dS dị mma karịa 1.5, nke na-egosi na nrụgide evolushọn zuru ụwa ọnụ bụ ikpochapụ ala nna ochie ma na-anọchi anya ọnọdụ dị mma (dị iche iche ma ọ bụ eduzi) na/ma ọ bụ ntụsara ahụ dị ọcha. N'ụdị nke "virulence, detoxification, adaptation", "usoro ntinye na phages", na "protein na-achịkwa", ihe karịrị ụzọ abụọ n'ụzọ atọ nke SNP bụ ndị na-enweghị aha (Nyocha 6B).
N'ime edemede niile, enwere mkpụrụ ndụ ihe nketa nwere ọtụtụ SNP, na-ebute ọnụọgụ ngbanwe nkezi (ya bụ, nkezi SNP kwa mkpụrụ ndụ ihe nketa) karịrị 1 (Figure 6A). Pks12 (Mb2074c) nwere 15 SNP na fas (Mb2553c) nwere 8 SNP nwere ụkpụrụ ngbanwe dị elu. Mkpụrụ ndụ ihe nketa abụọ a na-etinye aka na metabolism fatty acid. Mkpụrụ ndụ pks na-etinye koodu polyketide synthase (PKS), nke bụ enzyme multifunctional na-etinye aka na mycobacterial cell wall lipid biosynthesis74,75. Mkpụrụ ndụ ihe nketa a na-etinye polypeptide multifunctional nke na-etinye aka na njikọ nke mycoketides74,76. Mkpụrụ ndụ fas na-etinye aka na njikọ nke mycolic acid. Mkpụrụ ndụ ihe nketa abụọ a na-arụ ọrụ dị mkpa na biosynthesis nke mgbidi cell na kọntaktị na onye ọbịa.
Iji mụọ banyere mmalite nke Mycobacterium bovis, a nyochara ụdị mkpụrụ ndụ ihe nketa abụọ. Ọrụ ndị e bipụtara na mbụ na-eji usoro nhazi na usoro phylogenetic chọpụtara mkpụrụ ndụ ihe nketa nke ndị nna nna MTBC nwetara site na HGT tupu mgbanwe dị iche iche37,38. Edepụtara mkpụrụ ndụ ihe nketa ndị a na tebụl mgbakwunye 2. A na-enyocha nkesa SNP nke ngụkọta nke mkpụrụ ndụ ihe nketa 77 nwere ike jikọta na HGT, na ebe 26 polymorphic achọpụtara, nke n'ọtụtụ ọnọdụ (78%) mere ka mgbanwe ndị na-abụghị synonymous (NS) gbanwee (Nkwupụta 2). Ọrụ gara aga na genome MTBC gosipụtara na mpaghara HGT etinyere na-egosipụta oke NS SNP dị elu ma e jiri ya tụnyere genome ndị ọzọ. Ọ bụrụ na mmadụ na-eche na mpaghara recombination ndị a bụ ndị nna nna MTBC nwetara, ya mere, ha na-anọchi anya polymorphisms oge ochie, mgbe ahụ, a na-atụ anya na mgbanwe mgbanwe nke NS ga-adị elu, n'ihi na a na-atụ anya na a ga-ewepụ NS nnọchi site na nhọrọ na-adịghị mma n'ihi mgbanwe amino acid nwere ike ịgbanwe ọrụ nke protein. Ya mere, nsonaazụ anyị na-egosi na nsonaazụ arụ ọrụ nwere ike ịmalite site na nnọchi nke mkpụrụ ndụ ihe nketa HGT, nke gosipụtara mkpa ha dị maka ụdị mkpụrụ ndụ ihe nketa bara uru.
N'otu aka ahụ na nyocha a, a na-enyocha mkpụrụ ndụ ihe nketa nke na-etinye ihe mejupụtara nke sistemu 3R (Ndozi DNA, Nrụgharị, na Ntugharị) nke ọma dịka ndepụta nke dos Vultos and collaborators bipụtara na mbụ (2008)39. A pụghị ịhụ mgbanwe nke iberibe DNA ndị yiri ya ozugbo, ọ bụ ezie na ọ nwere ike bụrụ usoro a na-emekarị mgbe nje bacteria nwere njikọ chiri anya, dị ka n'ihe banyere nhazi data a; na mgbakwunye, usoro a nwere ike ịbụ isi ihe na usoro nrụzi DNA72, ya mere Kwuonụ na-ekere òkè n'ime nchikota homologous. Achọpụtara ngụkọta nke ọnọdụ 26 polymorphic nke mkpụrụ ndụ ihe nketa 54 kesara (Mgbakwunye 3). N'ime usoro mkpụrụ ndụ ihe nketa a, mgbanwe NS ruru ihe dị ka 65% nke nsonaazụ ya, nke dabara na akụkọ ndị gara aga gbasara ụdị ụkwara nta nke Mycobacterium.
Oge nzipu: Ọktoba 21-2021