RESUMEN
This report describes the first case in South Australia, Australia, of Mycobacterium pinnipedii tuberculosis in a free-ranging Australian fur seal (Arctocephalus pusillus doriferus). Severe pyogranulomatous pleuropneumonia with intrahistocytic acid-fast beaded filamentous bacilli was seen on histology. M. pinnipedii was confirmed by full 24-loci mycobacterial interspersed repetitive-unit-variable-number tandem-repeat (MIRU-VNTR) typing. Spillover concerns for public health and cattle are discussed.
Asunto(s)
Lobos Marinos , Mycobacterium/clasificación , Mycobacterium/aislamiento & purificación , Tuberculosis Pulmonar/veterinaria , Animales , Resultado Fatal , Masculino , Tuberculosis Pulmonar/microbiología , Tuberculosis Pulmonar/patologíaRESUMEN
A Mycobacterium tuberculosis complex organism was isolated from a zoo resident rock hyrax (Procavia capensis) imported into Canada from South Africa. The strain was identified biochemically as Mycobacterium microti. The spoligotype pattern obtained for this isolate was found to be rare. This represents the first report of isolation and spoligotyping of M. microti in North America.
Asunto(s)
Damanes/microbiología , Mycobacterium tuberculosis/aislamiento & purificación , Tuberculosis/veterinaria , Animales , Animales de Zoológico/microbiología , Técnicas de Tipificación Bacteriana/veterinaria , Canadá , Femenino , Mycobacterium tuberculosis/clasificación , Filogenia , Sudáfrica/etnología , Tuberculosis/diagnóstico , Tuberculosis/microbiologíaRESUMEN
In a previous report, we described a PCR protocol for the differentiation of the various species of the Mycobacterium tuberculosis complex (MTC) on the basis of genomic deletions (R. C. Huard, L. C. de Oliveira Lazzarini, W. R. Butler, D. van Soolingen, and J. L. Ho, J. Clin. Microbiol. 41:1637-1650, 2003). That report also provided a broad cross-comparison of several previously identified, phylogenetically relevant, long-sequence and single-nucleotide polymorphisms (LSPs and SNPs, respectively). In the present companion report, we expand upon the previous work (i) by continuing the evaluation of known MTC phylogenetic markers in a larger collection of tubercle bacilli (n = 125), (ii) by evaluating additional recently reported MTC species-specific and interspecific polymorphisms, and (iii) by describing the identification and distribution of a number of novel LSPs and SNPs. Notably, new genomic deletions were found in various Mycobacterium tuberculosis strains, new species-specific SNPs were identified for "Mycobacterium canettii," Mycobacterium microti, and Mycobacterium pinnipedii, and, for the first time, intraspecific single-nucleotide DNA differences were discovered for the dassie bacillus, the oryx bacillus, and the two Mycobacterium africanum subtype I variants. Surprisingly, coincident polymorphisms linked one M. africanum subtype I genotype with the dassie bacillus and M. microti with M. pinnipedii, thereby suggesting closer evolutionary ties within each pair of species than had been previously thought. Overall, the presented data add to the genetic definitions of several MTC organisms as well as fine-tune current models for the evolutionary history of the MTC.
Asunto(s)
Mycobacterium tuberculosis/clasificación , Polimorfismo de Nucleótido Simple , Animales , Marcadores Genéticos , Genoma Bacteriano , Humanos , Mycobacterium tuberculosis/genética , Filogenia , Reacción en Cadena de la Polimerasa , Especificidad de la Especie , Tuberculosis/microbiologíaRESUMEN
The Mycobacterium avium complex consists of epidemiologically distinct subsets. The classification of these subsets is complicated by a number of factors, including the ambiguous results obtained with phenotypic and genetic assays and the recent appreciation that human and avian strains appear to be distinct. In previous work, sequencing based on a 441-bp portion of the hsp65 gene has proven to efficiently classify isolates within the Mycobacterium genus but provides low resolution for distinguishing among members of the M. avium complex. Therefore, in this study, we have targeted the more variable 3' region of the hsp65 gene to determine whether it can effectively discriminate M. avium complex isolates at the levels of species and subspecies. Primers designed for this target consistently generated amplicons for all organisms classified as M. avium complex. Sequences obtained indicate that M. intracellulare is genetically divergent from M. avium organisms, and distinct sequevars were obtained for M. avium subsets, including M. avium subsp. avium (bird type), M. avium subsp. hominissuis, and M. avium subsp. paratuberculosis. In addition, sequence differences served to distinguish bovine from ovine strains of M. avium subsp. paratuberculosis. A unique profile for M. avium subsp. silvaticum was not obtained. These results indicate that sequencing the 3' region of the hsp65 gene can simply and unambiguously distinguish species and subspecies of the M. avium complex.
Asunto(s)
Proteínas Bacterianas/genética , Técnicas de Tipificación Bacteriana , Chaperoninas/genética , Complejo Mycobacterium avium/clasificación , Análisis de Secuencia de ADN , Animales , Proteínas Bacterianas/química , Bovinos , Chaperonina 60 , Chaperoninas/química , Cartilla de ADN , Humanos , Datos de Secuencia Molecular , Complejo Mycobacterium avium/genética , Infección por Mycobacterium avium-intracellulare/microbiología , Reacción en Cadena de la Polimerasa/métodos , Polimorfismo de Nucleótido Simple/genéticaRESUMEN
Though careful consideration has been placed towards genetic characterization of tubercle bacillus isolates causing disease in humans, those causing disease predominantly among wild and domesticated mammals have received less attention. In contrast to Mycobacterium tuberculosis, whose host range is largely specific to humans, M. bovis and "M bovis-like" organisms infect a broad range of animal species beyond their most prominent host in cattle. To determine whether strains of variable genomic content are associated with distinct distributions of disease, the DNA contents of M. bovis or M. bovis-like isolates from a variety of hosts were investigated via Affymetrix GeneChip. Consistent with previous genomic analysis of the M. tuberculosis complex (MTC), large sequence polymorphisms of putative diagnostic and biological consequence were able to unambiguously distinguish interrogated isolates. The distribution of deleted regions indicates organisms genomically removed from M. bovis and also points to structured genomic variability within M. bovis. Certain genomic profiles spanned a variety of hosts but were clustered by geography, while others associated primarily with host type. In contrast to the prevailing assumption that M. bovis has broad host capacity, genomic profiles suggest that distinct MTC lineages differentially infect a variety of mammals. From this, a phylogenetic stratification of genotypes offers a predictive framework upon which to base future genetic and phenotypic studies of the MTC.
Asunto(s)
Evolución Molecular , Genoma Bacteriano , Mycobacterium bovis/fisiología , Mycobacterium bovis/clasificación , Mycobacterium bovis/genética , Fenotipo , Reacción en Cadena de la Polimerasa/métodosRESUMEN
Mycobacterium avium subsp. paratuberculosis is an emerging pathogen of mammals and is being actively investigated as a possible zoonotic agent. The lack of reliable diagnostic assays has hampered rational assessment of the prevalence of this organism in humans and animals. We have used a comparative genomic approach to reveal genomic differences between M. avium subsp. paratuberculosis and its close relative M. avium subsp. avium, a highly prevalent environmental organism. From computational and DNA microarray-based study of two prototype strains, M. avium subsp. avium strain 104 and M. avium subsp. paratuberculosis strain K10, we have uncovered two types of large sequence polymorphisms (LSPs): those present in the former but missing in the latter (LSP(A)s) and those only present in the latter (LSP(P)s). We examined the distribution of 3 LSP(A)s and 17 LSP(P)s across a panel of 383 M. avium complex isolates in order to determine their potential utility for the development of accurate diagnostic tests. Our results show that the absence of LSP(A)8 is 100% specific for the identification of M. avium subsp. paratuberculosis. Of the 17 LSP(P)s, 10 regions were not specific for M. avium subsp. paratuberculosis while 7 were shown to be highly specific (>98%) and, in some cases, highly sensitive as well (up to 95%). These data highlight the need to evaluate these regions across a diverse panel of clinical and environmental isolates and indicate the LSPs best suited for M. avium subsp. paratuberculosis diagnostics.
Asunto(s)
Mycobacterium avium subsp. paratuberculosis/genética , Polimorfismo Genético , Genoma Bacteriano , Complejo Mycobacterium avium/genética , Mycobacterium avium subsp. paratuberculosis/aislamiento & purificación , Reacción en Cadena de la PolimerasaRESUMEN
We have initiated comparative genomic analysis of Mycobacterium avium subspecies by DNA microarray, uncovering 14 large sequence polymorphisms (LSPs) comprising over 700 kb that distinguish M. avium subsp. avium from M. avium subsp. paratuberculosis. Genes predicted to encode metabolic pathways were overrepresented in the LSPs, and analysis revealed a polymorphism within the mycobactin biosynthesis operon that potentially explains the in vitro mycobactin dependence of M. avium subsp. paratuberculosis.
Asunto(s)
Cromosomas Bacterianos/genética , ADN Bacteriano/análisis , Genoma Bacteriano , Mycobacterium avium subsp. paratuberculosis/genética , Mycobacterium avium/genética , Polimorfismo Genético , ADN Bacteriano/genética , Genes Bacterianos , Mycobacterium avium/metabolismo , Mycobacterium avium subsp. paratuberculosis/metabolismo , Análisis de Secuencia por Matrices de Oligonucleótidos , Oxazoles/metabolismoRESUMEN
AmpliBASE MT is an online databank of high-resolution DNA fingerprints representing fluorescent amplified fragment length polymorphism (FAFLP) profiles or amplitypes developed for the Mycobacterium tuberculosis complex strains from 48 different countries. AmpliBASE MT is based on a relational database management system that is hyperlinked to visualize genotyping results in the form of DNA fingerprint images for individual strains. A flexible search system based on systematic comparisons of fragment sizes in base pairs allows inter-laboratory comparison of FAFLP profiles. Besides this, the database also displays previously published data on IS6110 profiles, spoligotypes, MIRU-VNTRs and large sequence polymorphisms along with the FAFLP records that will give the overall comparisons. Being the first of its kind, AmpliBASE MT is expected to be a very helpful tool in strengthening the concept of 'geographic genomics' and will be very helpful to molecular epidemiologists and those interested in diagnostic development for tuberculosis.
Asunto(s)
Dermatoglifia del ADN/métodos , Sistemas de Administración de Bases de Datos , Bases de Datos de Ácidos Nucleicos , Perfilación de la Expresión Génica/métodos , Almacenamiento y Recuperación de la Información/métodos , Mycobacterium tuberculosis/genética , Interfaz Usuario-Computador , Variación Genética/genética , Internet , Fenotipo , Programas InformáticosRESUMEN
Despite their remarkable genetic homology, members of the Mycobacterium tuberculosis complex express very different phenotypes, most notably in their spectra of clinical presentation. For example, M. tuberculosis is regarded as pathogenic to humans, whereas members having deleted RD1, such as Mycobacterium microti and Mycobacterium bovis BCG, are not. The dassie bacillus, an infrequent variant of the M. tuberculosis complex characterized as being most similar to M. microti, is the causative agent of tuberculosis (TB) in the dassie (Procavia capensis). Intriguingly, the dassie bacillus is not pathogenic to rabbits or guinea pigs and has never been documented to infect humans. Although it was identified more than a half-century ago, the reasons behind its attenuation are unknown. Because large sequence polymorphisms have presented themselves as the most obvious genomic distinction among members of the M. tuberculosis complex, the DNA content of the dassie bacillus was interrogated by Affymetrix GeneChip to identify regions that are absent from it but present in M. tuberculosis H37Rv. Comparison has led to the identification of nine regions of difference (RD), five of which are shared with M. microti (RDs 3, 7, 8, 9, and 10). Although the dassie bacillus does not share the other documented deletions in M. microti (RD1(mic), RD5(mic), MID1, MID2, and MID3), it has endured unique deletions in the regions of RD1, RD5, N-RD25, and Rv3081-Rv3082c (virS). RD1(das), affecting only Rv3874-Rv3877, is the smallest natural deletion of the RD1 region uncovered and points to genes within this region that are likely implicated in virulence. Newfound deletions from the dassie bacillus are discussed in relation to their evolutionary and biological significance.
Asunto(s)
Eliminación de Gen , Genoma Bacteriano , Damanes/microbiología , Mycobacterium tuberculosis/clasificación , Mycobacterium/clasificación , Mycobacterium/genética , Tuberculosis Pulmonar/veterinaria , Animales , Técnicas de Tipificación Bacteriana , Masculino , Mycobacterium/aislamiento & purificación , Mycobacterium tuberculosis/genética , Análisis de Secuencia por Matrices de Oligonucleótidos , Reacción en Cadena de la Polimerasa , Análisis de Secuencia de ADN , Tuberculosis Pulmonar/microbiologíaRESUMEN
Mycobacterium tuberculosis complex isolates recovered from goats were originally classified as Mycobacterium tuberculosis subsp. caprae; however, this subspecies was recently reclassified as Mycobacterium bovis subsp. caprae. Besides biochemical (sensitivity to pyrazinamide) and epidemiological features, strains of this unusual member of the M. tuberculosis complex show a special combination of pncA, oxyR, katG and gyrA gene polymorphisms. Sequence analysis of the gyrB gene in these strains reveals special nucleotide substitutions not found in other members of the M. tuberculosis complex that can be used to differentiate caprine mycobacterial strains from M. bovis and other members of the M. tuberculosis complex. M. tuberculosis subsp. caprae now appears not to be restricted to Spanish goats, as strains of this organism have been isolated from cattle, wild boar and pigs. Its occurrence has also been reported in France, Austria and Germany. Two studies on the evolution of the M. tuberculosis complex based on the presence/absence of regions of difference have shown that the group of caprine isolates (or its ancestor) is older than M. bovis (or its ancestor). These findings reinforce the original suggestion that the caprine mycobacterial strains are a taxon of the M. tuberculosis complex, independent of M. bovis. Within the current context of the existing nomenclature of the M. tuberculosis complex, it is proposed that M. tuberculosis subsp. caprae be elevated to species status, as Mycobacterium caprae comb. nov., sp. nov.
Asunto(s)
Mycobacterium tuberculosis/clasificación , Mycobacterium/clasificación , Filogenia , Animales , Secuencia de Bases , Cabras/microbiología , Mycobacterium/aislamiento & purificación , Mycobacterium tuberculosis/genética , Mycobacterium tuberculosis/aislamiento & purificación , Polimorfismo GenéticoRESUMEN
A comparison of Mycobacterium tuberculosis complex isolates from seals (pinnipeds) in Australia, Argentina, Uruguay, Great Britain and New Zealand was undertaken to determine their relationships to each other and their taxonomic position within the complex. Isolates from 30 cases of tuberculosis in six species of pinniped and seven related isolates were compared to representative and standard strains of the M. tuberculosis complex. The seal isolates could be distinguished from other members of the M. tuberculosis complex, including the recently defined 'Mycobacterium canettii' and 'Mycobacterium caprae', on the basis of host preference and phenotypic and genetic tests. Pinnipeds appear to be the natural host for this 'seal bacillus', although the organism is also pathogenic in guinea pigs, rabbits, humans, Brazilian tapir (Tapirus terrestris) and, possibly, cattle. Infection caused by the seal bacillus is predominantly associated with granulomatous lesions in the peripheral lymph nodes, lungs, pleura, spleen and peritoneum. Cases of disseminated disease have been found. As with other members of the M. tuberculosis complex, aerosols are the most likely route of transmission. The name Mycobacterium pinnipedii sp. nov. is proposed for this novel member of the M. tuberculosis complex (the type strain is 6482(T)=ATCC BAA-688(T)=NCTC 13288(T)).
Asunto(s)
Mycobacterium tuberculosis/patogenicidad , Phocidae/microbiología , Tuberculosis/veterinaria , Animales , ADN Bacteriano/genética , ADN Ribosómico/genética , Humanos , Datos de Secuencia Molecular , Mycobacterium tuberculosis/clasificación , Mycobacterium tuberculosis/genética , Mycobacterium tuberculosis/aislamiento & purificación , Ácidos Micólicos/análisis , Fenotipo , ARN Bacteriano/genética , ARN Ribosómico 16S/genética , Especificidad de la Especie , Tuberculosis/microbiología , VirulenciaRESUMEN
Tuberculosis in seals is caused by a member of the Mycobacterium tuberculosis complex referred to as the 'seal bacillus'. Fluorescent amplified-fragment length polymorphism (FAFLP) analysis was applied to isolates from four Australian and six Argentinean seals and compared with FAFLP pattern for standard strains belonging to the M. tuberculosis complex. The FAFLP profiles derived from EcoRI/MseI restricted fragments of blind coded DNA samples differentiated the seal bacillus from other members of the M. tuberculosis complex. According to the phylogenetic analysis performed using FAFLP data, seal bacilli appear to have diverged significantly from other members of the M. tuberculosis complex. We describe the suitability of a panel of 19 highly polymorphic markers for rapid identification and comparative genomic analyses of the seal bacillus strains. It is likely that these bacilli got separated from the M. tuberculosis lineage as a result of different insertion deletion events occurring on a genome wide scale. Our analysis reveals that the seal bacillus and M. bovis are genetically related and therefore, might have originated from a common ancestor. Our data additionally support the hypothesis that seal bacillus occupies a unique taxonomic position within the M. tuberculosis complex.
Asunto(s)
Mycobacterium tuberculosis/clasificación , Mycobacterium tuberculosis/genética , Polimorfismo Genético , Phocidae/microbiología , Análisis de Secuencia de ADN/métodos , Tuberculosis/veterinaria , Animales , Evolución Molecular , Genoma Bacteriano , Genotipo , Filogenia , Tuberculosis/microbiologíaRESUMEN
The present update on the global distribution of Mycobacterium tuberculosis complex spoligotypes provides both the octal and binary descriptions of the spoligotypes for M. tuberculosis complex, including Mycobacterium bovis, from >90 countries (13,008 patterns grouped into 813 shared types containing 11,708 isolates and 1,300 orphan patterns). A number of potential indices were developed to summarize the information on the biogeographical specificity of a given shared type, as well as its geographical spreading (matching code and spreading index, respectively). To facilitate the analysis of hundreds of spoligotypes each made up of a binary succession of 43 bits of information, a number of major and minor visual rules were also defined. A total of six major rules (A to F) with the precise description of the extra missing spacers (minor rules) were used to define 36 major clades (or families) of M. tuberculosis. Some major clades identified were the East African-Indian (EAI) clade, the Beijing clade, the Haarlem clade, the Latin American and Mediterranean (LAM) clade, the Central Asian (CAS) clade, a European clade of IS6110 low banders (X; highly prevalent in the United States and United Kingdom), and a widespread yet poorly defined clade (T). When the visual rules defined above were used for an automated labeling of the 813 shared types to define nine superfamilies of strains (Mycobacterium africanum, Beijing, M. bovis, EAI, CAS, T, Haarlem, X, and LAM), 96.9% of the shared types received a label, showing the potential for automated labeling of M. tuberculosis families in well-defined phylogeographical families. Intercontinental matches of shared types among eight continents and subcontinents (Africa, North America, Central America, South America, Europe, the Middle East and Central Asia, and the Far East) are analyzed and discussed.
Asunto(s)
Mycobacterium tuberculosis/genética , Mycobacterium tuberculosis/aislamiento & purificación , Técnicas de Tipificación Bacteriana , ADN Bacteriano/genética , ADN Intergénico/genética , Bases de Datos de Ácidos Nucleicos , Humanos , Epidemiología Molecular , Mycobacterium bovis/clasificación , Mycobacterium bovis/genética , Mycobacterium bovis/aislamiento & purificación , Mycobacterium tuberculosis/clasificación , Tuberculosis/epidemiología , Tuberculosis/microbiologíaRESUMEN
We present a short summary of recent observations on the global distribution of the major clades of the Mycobacterium tuberculosis complex, the causative agent of tuberculosis. This global distribution was defined by data-mining of an international spoligotyping database, SpolDB3. This database contains 11708 patterns from as many clinical isolates originating from more than 90 countries. The 11708 spoligotypes were clustered into 813 shared types. A total of 1300 orphan patterns (clinical isolates showing a unique spoligotype) were also detected.
Asunto(s)
Mycobacterium tuberculosis/clasificación , Mycobacterium tuberculosis/aislamiento & purificación , Tuberculosis/microbiología , Bases de Datos Factuales , Filogenia , Tuberculosis/epidemiologíaRESUMEN
To better understand the evolution of the Mycobacterium tuberculosis complex, subspecies were tested for large sequence polymorphisms. Samples with greater numbers of deletions, without exception, were missing all the same regions that were deleted from samples with lesser numbers of deletions. Principal genetic groups based on single-nucleotide polymorphisms were restricted to one of the deletion-based groups, and isolates that shared genotypes based on molecular epidemiological markers were assigned almost exclusively to the same deletion type. The data provide compelling evidence that human tuberculosis did not originate from the present-day bovine form. Genomic deletions present themselves as an attractive modality to study the evolution of the M. tuberculosis complex.
Asunto(s)
Mycobacterium/clasificación , Animales , Evolución Biológica , Bovinos , Humanos , Mycobacterium/genética , Polimorfismo Genético , Eliminación de SecuenciaRESUMEN
Several human pathogens (e.g., Bacillus anthracis, Yersinia pestis, Bordetella pertussis, Plasmodium falciparum, and Mycobacterium tuberculosis) have very restricted unselected allelic variation in structural genes, which hinders study of the genetic relationships among strains and strain-trait correlations. To address this problem in a representative pathogen, 432 M. tuberculosis complex strains from global sources were genotyped on the basis of 230 synonymous (silent) single nucleotide polymorphisms (sSNPs) identified by comparison of four genome sequences. Eight major clusters of related genotypes were identified in M. tuberculosis sensu stricto, including a single cluster representing organisms responsible for several large outbreaks in the United States and Asia. All M. tuberculosis sensu stricto isolates of previously unknown phylogenetic position could be rapidly and unambiguously assigned to one of the eight major clusters, thus providing a facile strategy for identifying organisms that are clonally related by descent. Common clones of M. tuberculosis sensu stricto and M. bovis are distinct, deeply branching genotypic complexes whose extant members did not emerge directly from one another in the recent past. sSNP genotyping rapidly delineates relationships among closely related strains of pathogenic microbes and allows construction of genetic frameworks for examining the distribution of biomedically relevant traits such as virulence, transmissibility, and host range.
Asunto(s)
Mycobacterium tuberculosis/genética , Alelos , Animales , Variación Genética , Genoma Bacteriano , Genotipo , Humanos , Epidemiología Molecular , Mycobacterium bovis/genética , Mycobacterium bovis/aislamiento & purificación , Mycobacterium tuberculosis/clasificación , Mycobacterium tuberculosis/aislamiento & purificación , Mycobacterium tuberculosis/patogenicidad , Filogenia , Polimorfismo de Longitud del Fragmento de Restricción , Polimorfismo de Nucleótido Simple , Especificidad de la Especie , Virulencia/genéticaRESUMEN
As part of a larger study investigating diversity and distribution of Mycobacterium spp. in Australia, multilocus enzyme electrophoresis was used to assess genetic relationships at 17 enzyme loci amongst a collection of reference strains and isolates initially identified on biochemical and other grounds as M. intracellulare (70), "X' mycobacteria (10), M. scrofulaceum (7), M. avium (8) and M. avium subsp. paratuberculosis (2). Two of the isolates initially identified as M. intracellulare were shown to be quite distinct from the others. Both gave negative results in a species-specific DNA probe test, whilst one was positive by PCR. These results emphasize the uncertainties involved in identifying members of this group. The other M. intracellulare isolates formed a cohesive but diverse group, being divided into 48 electrophoretic types (ETs), with a mean genetic diversity of 0.38. Forty-three of these ETs contained only single isolates. There was no clear relationship between the serovar and ET designation. The index of association calculated for M. intracellulare was significantly different from zero, suggesting that it is a clonal species. PFGE was also applied to selected isolates from the ETs containing multiple isolates, and some of these could be differentiated further. The strains of M. scrofulaceum and "X' mycobacteria were distinct from M. intracellulare, but themselves were highly heterogeneous, with mean genetic diversities of 0.66 and 0.65, respectively. Each of these groups may represent more than one species. M. avium strains were distinct from the two M. avium subsp. paratuberculosis strains, as well as from the other mycobacteria studied.