Mycobacterium Nonchromogenicum Research Articles

Bovine Tuberculosis in Wild Boar (Sus scrofa) in Slovenia.

Mycobacteria of the Mycobacterium tuberculosis complex (MTC) are capable of infecting a wide variety of animals. Wild boar (Sus scrofa) has been recognized as an important wildlife reservoir for bovine tuberculosis. We screened wild boar in Slovenia for the presence of (1) Mycobacterium bovis in tissues and (2) antibodies to M. bovis in blood samples. In 2016 and 2017, 1284 tissue samples from 676 wild boar were subjected to cultivation. In 2018 and 2019, blood samples from 132 wild boar were examined using an ELISA kit. None of the MTC species were isolated from the tissue samples, and no antibodies to M. bovis were detected in the blood samples. Several nontuberculous mycobacteria (NTM), identified by 16S rRNA sequencing and/or matrix-assisted laser desorption ionization time-of-flight mass spectrometry, were found in the tissues of 9.8% of the wild boar: Mycobacterium nonchromogenicum, Mycobacterium peregrinum/Mycobacterium septicum, Mycobacterium avium, Mycobacterium engbaekii, Mycobacterium arupense, Mycobacterium algericum, Mycobacterium bohemicum, Mycobacterium confluentis, Mycobacterium flavescens, Mycobacterium fortuitum, Mycobacterium thermoresistibile, and Mycobacterium vaccae. Species-level identification was not possible for 21.2% of the isolates. At the time of the study, wild boar in Slovenia were not at risk from bTB; the significance of the presence of NTM in wild boar remains to be clarified and evaluated from a One Health perspective.

Prevalence of nontuberculous mycobacteria and the emergence of rare species in Henan Province, China.

Nontuberculous mycobacteria (NTM) is a large group of mycobacteria other than the Mycobacterium tuberculosis complex and Mycobacterium leprae. Epidemiological investigations have found that the incidence of NTM infections is increasing in China, and it is naturally resistant to many antibiotics. Therefore, studies of NTM species in clinical isolates are useful for understanding the epidemiology of NTM infections. The present study aimed to investigate the incidence of NTM infections and types of NTM species. Of the 420 samples collected, 285 were positive for M. tuberculosis, 62 samples were negative, and the remaining 73 samples contained NTM, including 35 (8.3%) only NTM and 38 (9%) mixed (M. tuberculosis and NTM). The most prevalent NTM species were Mycobacterium intracellulare (30.1%), followed by Mycobacterium abscessus (15%) and M. triviale (12%). M. gordonae infection was detected in 9.5% of total NTM-positive cases. Moreover, this study reports the presence of Mycobacterium nonchromogenicum infection and a high prevalence of M. triviale for the first time in Henan. M. intracellulare is the most prevalent, accompanied by some emerging NTM species, including M. nonchromogenicum and a high prevalence of M. triviale in Henan Province. Monitoring NTM transmission and epidemiology could enhance mycobacteriosis management in future.

Atypical systemic mycobacteriosis in Hermann's tortoises (Testudo hermanni)

ABSTRACT Background: Hermann's tortoise (Testudo hermanni) is a near threatened species found throughout southern Europe. Captive breeding of endangered species is subject to health risks, such as the stress of captivity and ambient conditions that may favor the emergence of infectious diseases. An episode of systemic atypical mycobacteriosis was investigated in six Hermann's tortoises from a captive population. The symptoms appeared in the form of anorexia, weakness, and lethargy, with inflammatory and edematous lesions of the hind limbs and tail. Four of the affected tortoises were euthanized and necropsied. Methods: Blood samples and joint aspirates were obtained for assessment. Euthanasia of four affected animals was performed. Postmortem examination included necropsy, histopathology, and mycobacterium detection by culture and polymerase chain reaction (PCR). In the other two animals, only hematology and blood biochemistry was carried out. Results: Hematological and blood biochemistry analyses gave inconsistent results. In joint aspirates, mononuclear cells with phagocytozed bacillary structures were observed. Ziehl-Neelsen stain revealed the presence of acid-fast bacteria. Granulomatous lesions were observed in liver, lungs, spleen, heart, muscle, kidney, and ovaries. Culture and identification were positive for mycobacteria and molecular sequencing led to the identification of Mycobacterium nonchromogenicum. Conclusions and clinical relevance: Although mycobacteriosis is rare in reptile collections, strict hygienic and prophylactic measures in species of high ecological value must be performed, especially if the animals are to be used for reintroduction projects.

Mycobacterium nonchromogenicum en un paciente con neumoconiosis: reporte de un caso en atención primaria

La neumoconiosis es una patología pulmonar crónica producida por el daño tisular que originan algunos tóxicos al ser inhalados durante ciertos periodos. El caso presentado corresponde a un paciente inmunocompetente con silicoantracosis, un tipo de neumoconiosis, en quien se detecta infección pulmonar por Mycobacterium nonchromogenicum en un centro de atención primaria en Bogotá, Colombia, poco común y de gran interés médico.

Correction for Vasireddy et al., Mycobacterium arupense, Mycobacterium heraklionense, and a Newly Proposed Species, "Mycobacterium virginiense" sp. nov., but Not Mycobacterium nonchromogenicum, as Species of the Mycobacterium terrae Complex Causing Tenosynovitis and Osteomyelitis.

Volume 54, no. 5, p. [1340–1351][1], 2016, . Page 1349, column 1: The last three paragraphs preceding the Acknowledgments section should be replaced with the following. Mycobacterium virginiense (vir.gi.ni.en′se. N.L. neut. adj. virginiense, of or

Comparative Genomics and Proteomic Analysis of Four Non-tuberculous Mycobacterium Species and Mycobacterium tuberculosis Complex: Occurrence of Shared Immunogenic Proteins.

The Esx and PE/PPE families of proteins are among the most immunodominant mycobacterial antigens and have thus been the focus of research to develop vaccines and immunological tests for diagnosis of bovine and human tuberculosis, mainly caused by Mycobacterium bovis and Mycobacterium tuberculosis, respectively. In non-tuberculous mycobacteria (NTM), multiple copies of genes encoding homologous proteins have mainly been identified in pathogenic Mycobacterium species phylogenically related to Mycobacterium tuberculosis and Mycobacterium bovis. Only ancestral copies of these genes have been identified in nonpathogenic NTM species like Mycobacterium smegmatis, Mycobacterium sp. KMS, Mycobacterium sp. MCS, and Mycobacterium sp. JLS. In this study we elucidated the genomes of four nonpathogenic NTM species, viz Mycobacterium komanii sp. nov., Mycobacterium malmesburii sp. nov., Mycobacterium nonchromogenicum, and Mycobacterium fortuitum ATCC 6841. These genomes were investigated for genes encoding for the Esx and PE/PPE (situated in the esx cluster) family of proteins as well as adjacent genes situated in the ESX-1 to ESX-5 regions. To identify proteins actually expressed, comparative proteomic analyses of purified protein derivatives from three of the NTM as well as Mycobacterium kansasii ATCC 12478 and the commercially available purified protein derivatives from Mycobacterium bovis and Mycobacterium avium was performed. The genomic analysis revealed the occurrence in each of the four NTM, orthologs of the genes encoding for the Esx family, the PE and PPE family proteins in M. bovis and M. tuberculosis. The identification of genes of the ESX-1, ESX-3, and ESX-4 region including esxA, esxB, ppe68, pe5, and pe35 adds to earlier reports of these genes in nonpathogenic NTM like M. smegmatis, Mycobacterium sp. JLS and Mycobacterium KMS. This report is also the first to identify esxN gene situated within the ESX-5 locus in M. nonchromogenicum. Our proteomics analysis identified a total of 609 proteins in the six PPDs and 22 of these were identified as shared between PPD of M.bovis and one or more of the NTM PPDs. Previously characterized M tuberculosis/M. bovis homologous immunogenic proteins detected in one or more of the nonpathogenic NTM in this study included CFP-10 (detected in M. malmesburii sp. nov. PPD), GroES (detected in all NTM PPDs but M. malmesburii sp. nov.), DnaK (detected in all NTM PPDs), and GroEL (detected in all NTM PPDs). This study confirms reports that the ESX-1, ESX-3, and ESX-4 regions are ancestral regions and thus found in the genomes of most mycobacteria. Identification of NTM homologs of immunogenic proteins warrants further investigation of their ability to cause cross-reactive immune responses with MTBC antigens.

Detection of wide genetic diversity and several novel strains among non-avium nontuberculous mycobacteria isolated from farmed and wild animals in Hungary.

Besides Mycobacterium avium numerous nontuberculous Mycobacterium (NTM) species exist, which pose constant health risk to both humans and animals. The aim of our study was to identify non-avium NTM isolates from veterinary origin in Hungary, and to detect the occurrence of rifampicin resistance among them. Two hundred and twenty-five strains isolated between 2006 and 2013 from domestic and wild animals and veterinary important samples were identified on the basis of partial DNA sequences of different structural or coding genes, besides commercial kits and multiplex PCR. From 14 different sources, 28 NTM strains and 8 hitherto unidentified strain types were detected. Mycobacterium nonchromogenicum was the most frequently occurring strain (25·78%). Besides, new hosts and mycobacteria-related pathological symptoms were detected. Noticeable rifampicin resistance (42·76%) was found among 159 strains from six different host species. Furthermore, we described the problematics of strain-misidentifications using commercial kits. Our study identified the most common non-avium NTM strains in Hungary, and provided account of their occurrence, host range, and pathogenicity. The detected high rifampicin resistance among the strains isolated mainly from fallow and red deer clearly shows that more attention should be paid to the examination of wild animals especially to those ones which may have contact or shared territory with farmed animals. In domestic animal husbandry the maintenance of tuberculosis free status is of primary importance. As immunological cross-reactions due to NTM hamper the diagnosis of bovine tuberculosis, the precise identification of NTM strains would be essential in the veterinary diagnostics, especially for potentially zoonotic strains. This is the first study investigating the strain diversity of non-avium NTM in Hungary.

Distribution of non-tuberculosis mycobacteria strains from suspected tuberculosis patients by heat shock protein 65 PCR-RFLP.

The genus Mycobacterium contains more than 150 species. Non-tuberculosis mycobacteria (NTM) often cause extrapulmonary and pulmonary disease. Mycobacteria detection at species level is necessary and provides useful information on epidemiology and facilitates successful treatment of patients. This retrospective study aimed to determine the incidence of the NTM isolates and Mycobacterium tuberculosis (Mtb) in clinical specimens collected from Iranian patients during February 2011–December 2013, by PCR–restriction fragment length polymorphism analysis (PRA) of the hsp65 gene. We applied conventional biochemical test and hsp65–PRA identification assay to identify species of mycobacteria in specimens from patients suspected of having mycobacterial isolates. This method was a sensitive, specific and effective assay for detecting mycobacterial species and had a 100% sensitivity and specificity for Mtb and Mycobacterium avium complex (MAC) species. Using PRA for 380 mycobacterial selected isolates, including 317 Mtb, four Mycobacterium bovis and of the 59 clinical isolates, the most commonly identified organism was Mycobacterium kansasii (35.6%), followed by Mycobacterium simiae (16.9%), Mycobacterium gordonae (16.9%), Mycobacterium fortuitum (5.1%), Mycobacterium intracellulare (5.1%), Mycobacterium avium (5.1%), Mycobacterium scrofulaceum (3.4%), Mycobacterium gastri (3.4%), Mycobacterium flavescens (3.4%), Mycobacterium chelonae (3.4%) and Mycobacterium nonchromogenicum (1.7%). PRA method, in comparison with classical methods, is rapid, useful and sensitive for the phylogenetic analysis and species detection of mycobacterial strains. Mycobacterium kansasii is the most common cause of infection by NTM in patients with non-HIV and HIV which demonstrated a high outbreak and diversity of NTM strains in our laboratory.

Presence of Mycobacterium avium Subspecies and Hepatitis E Virus in Raw Meat Products

Meat and meat products may be the source of various pathogenic and potentially pathogenic agents for humans. We ascertained the occurrence of Mycobacterium avium subsp. paratuberculosis, subsp. avium, and subsp. hominissuis, and hepatitis E virus in retail raw meat products. The DNA of at least one of the target M. avium subspecies was detected in 26 (29.2%) of 89 analyzed samples of meat products. Fourteen (15.7%), 1 (1.1%), and 17 (19.1%) samples contained the DNA of Mycobacterium avium subsp. paratuberculosis, subsp. avium, and subsp. hominissuis, respectively. The number of mycobacterial cells per gram of meat products determined by real-time quantitative PCR ranged from 1.15 × 10(2) to 6.97 × 10(3). Mycobacterium chitae and Mycobacterium nonchromogenicum were isolated from three (3.4%) samples. Culture examination was not positive for any M. avium subspecies. Hepatitis E virus RNA was not detected in any of the samples.

Prevalence and Distribution of Non-Tuberculous Mycobacteria (NTM) in Cattle, African Buffaloes (Syncerus caffer) and their Environments in South Africa

It has been hypothesized that a variety of non-tuberculous mycobacteria (NTM) species to which livestock and wildlife species are naturally exposed induce broadly cross-reactive anti-mycobacterial immune responses which interfere with current standard diagnostic assays. Non-tuberculous mycobacteria have also been implicated in Mycobacterium bovis-specific immune responsiveness, hence potentially the development of tuberculosis. Cattle and African buffaloes are both maintenance hosts of bovine tuberculosis (BTB) in South Africa, yet the effective diagnosis and control in these species may be hampered by adverse effects of NTM. As part of an investigation of the role of NTM in the immune responsiveness of cattle and African buffaloes to NTM, we conducted a countrywide survey to establish the prevalent NTM species and their distribution in the natural environments of these animals. A total of 1123 samples (water, soil, nasal and pharyngeal swabs) were collected for mycobacterium isolation. In addition, NTM isolated from tissue samples between 1991 and 2011 were included in the analysis. Mycobacteria were isolated from 56% of the samples from the countrywide survey. A total of 420 NTM isolates from soil, water, animal tissues and animal-derived swab samples were genotyped with the following results: 302 belonged to 40 known NTM species, 79 were found to be closely related to 23 known NTM species, and 38 isolates were found to be potentially novel species that are not currently listed in the RIDOM and NCBI BLAST databases. The four NTM species or closely related groups most frequently isolated in this survey included Mycobacterium terrae (11.2% of isolates), a group of mycobacteria closely related to Mycobacterium moriokaense (referred to as M. moriokaense-like) (8.1% of isolates), Mycobacterium nonchromogenicum (7.4% of isolates) and Mycobacterium vaccae/M. vanbaalenii (5.2% of isolates). The phylogenetic analysis of the M. moriokaense-like isolates, based on the 16S rRNA sequences, revealed at least eight clusters, possibly associated with eight different NTM species. Our findings provide account of NTM species diversity and associated prevalences in cattle and African buffaloes and their environments in South Africa. The identification of the most prevalent NTM species in this study will allow for a targeted investigation of their effects on host immune responses.

Identification and pathogenicity analysis of a novel non-tuberculous mycobacterium clinical isolate with nine-antibiotic resistance

With mycobacteriosis increasing, the study of non-tuberculous mycobacteria is imperative for clinical therapy and management. Nontuberculous mycobacteria are naturally resistant to most anti-tuberculosis drugs. Accordingly, it is important to decipher the biology of the novel non-tuberculous mycobacteria through complete genomic analysis of novel pathogenic mycobacteria. We describe Mycobacterium sinense JDM601, a novel, slow-growing mycobacterium of the Mycobacterium terrae complex resistant to nine antibiotics, by clinical presentation, cultural and biochemical characteristics, minimal inhibitory concentrations, and genome-sequencing analysis. JDM601 is closest to Mycobacterium nonchromogenicum according to mycolic acid composition, but closest to Mycobacterium algericum sp. nov according to 16S rDNA. JDM601 is resistant to isoniazid, streptomycin, rifampin, euteropas, protionamide, capromycin, ciprofloxacin, amikacin and levofloxacin but not ethambutol. The clinical information, mycolic acid composition, and virulence genes indicate that JDM601 is an opportunistic pathogen.

The evaluation of different molecular methods for rapid identification of non-tuberculous Mycobacterium

Objective To evaluate three molecular methods for rapid identification of nontuberculous Mycobacterium(NTM).Methods Forty-one clinical NTM isolates were collected and 16S rRNA gene sequencing was used as the standard method for NTM identification.Meanwhile,the restriction fragment length polymorphism of hsp65 PCR-RFLP and hsp65 gene sequencing were used to identify NTM strains and compared with 16S rRNA gene sequencing.Results The results of 16S rRNA sequencing showed that there were nine Mycobacterium chelonae complex strains,seven Mycobacteriumfortuitum strains,seven Mycobacterium intracellulare strains,three Mycobacterium avium strains,three Mycobacterium kansasii complex strains, three Mycobacterium smegmatis strains, three Mycobacterium terrae strains, two Mycobacterium phlei strains,two Mycobacterium nonchromogenicum strains,one Mycobacterium scrofulaceum strain and one Mycobacterium arupense strain.Compared with 16S rRNA gene sequencing,hsp65 PCR-RFLP could identify nine Mycobacterium chelonae complexes and three Mycobacterium kansasii complexes to subspecies Mycobacterium abscessus and Mycobacterium kansasii,respectively; One Mycobacterium fortuitum strain and one Mycobacterium nonchromogenicum strain were different from 16S rRNA gene sequencing results ,but other isolates were the same.The coincidence was 95.1%.By hsp65 gene sequencing,only one identification of Mycobacterium hiberniae strain was different from 16S rRNA gene sequencing and the coincidence was 97.6%.And hsp65 gene sequencing could further identify nine Mycobacterium chelonae complexes and three Mycobacterium kansasii complexes to subspecies Mycobacterium abscessus and Mycobacterium kansasii,respectively.Conclusions All three molecular methods can identify NTM strains rapidly.Compared with 16S rRNA gene sequencing,hsp65 gene sequencing and hsp65 PCR-RFLP are easier to identify clinical common NTM strains(such as Mycobacterium kansasii and Mycobacterium abscessus),and can be widely used in clinical practice. Key words: Mycobacterium; RNA,ribosomal,16S; Bacterial proteins; Heat-shock proteins; Polymerase chain reaction; Polymorphism,restriction fragment length

Nitrite-Ogawa egg medium for differentiating strains of Mycobacterium nonchromogenicum complex (M. nonchromogenicum, M. terrae and M. triviale) from other slowly growing mycobacteria (author's transl)

Nitrite-Ogawa egg medium for differentiating strains of Mycobacterium nonchromogenicum complex (M. nonchromogenicum, M. terrae and M. triviale) from other slowly growing mycobacteria (author's transl)

Occurrence of Mycobacterium spp. and other pathogens in lymph nodes of slaughtered swine and wild boars ( Sus scrofa)

Mycobacterium spp. and other pathogens were investigated in 258 swine lymph nodes (129 with and 129 without apparent lesions), and 120 lymph nodes (60 with and 60 without lesions) from wild boars ( Sus scrofa). A total of lymph nodes from swine and wild boars were collected of different animals. Submaxillar and mesenteric lymph nodes were submitted to microbiological examination and colonies suggestive of Mycobacterium spp. (alcohol–acid bacilli) were submitted to PCR Restriction Assay (PRA). In swine with lymphadenitis, Mycobacterium spp. (24.1%) and Rhodococcus equi (13.2%) were the most prevalent microorganisms, while in lymph nodes without lesions were identified a complex of microorganisms, including of environmental mycobacteria. In wild boars with lymphadenitis, ß-haemolytic Streptococcus (10.0%), Mycobacterium spp (8.4%) and R. equi (6.6%) were the most frequent. Among mycobacterias were identified predominantly Mycobacterium avium subspecies type 1 (48.3%) and M. avium subspecies type 2 (16.1%), followed by Mycobacterium intracellulare, Mycobacterium szulgai, Mycobacterium fortuitum, Mycobacterium gordonae, Mycobacterium simiae, Mycobacterium nonchromogenicum and Mycobacterium intracellulare type 2.

Mycobacterium senuense sp. nov., a slowly growing, non-chromogenic species closely related to the Mycobacterium terrae complex

A previously undescribed, slowly growing, non-chromogenic mycobacterium, isolated from a Korean patient with a symptomatic pulmonary infection, is described as representing a novel species. Its 16S rRNA gene sequence was unique and phylogenetic analysis based on 16S rRNA gene sequences showed that this organism belonged to the Mycobacterium terrae subclade. Phenotypically, the strain was generally similar to M. terrae and Mycobacterium nonchromogenicum, but its growth rate was slower than those of other M. terrae complex strains. A unique mycolic acid profile and phylogenetic analysis based on two different alternative chronometer molecules, hsp65 and rpoB, confirm the taxonomic status of this strain as a representative of a novel species. The name Mycobacterium senuense sp. nov. is proposed, with the type strain 05-832(T) (=DSM 44999(T) =KCTC 19147(T)).

Infectious tenosynovitis and osteomyelitis caused by Mycobacterium nonchromogenicum

Infections due to Mycobacterium terrae complex are rare. We report a severe case of chronic tenosynovitis and osteomyelitis of the hand caused by Mycobacterium nonchromogenicum requiring second ray finger amputation.

Mycobacterial Synovitis Caused by Slow-Growing Nonchromogenic Species: Eighteen Cases and a Review of the Literature

Slow-growing nonchromogenic mycobacterial species are an infrequent cause of soft tissue infection. Because these organisms are rare, they are not often initially considered in the differential diagnosis of synovitis. To evaluate the clinical and pathologic characteristics of patients with synovitis resulting from slow-growing nonchromogenic mycobacterial species. A 20-year retrospective review of records from The Methodist Hospital Microbiology Laboratory identified 18 culture-positive cases of synovitis that resulted from slow-growing nonchromogenic mycobacteria, including 14 caused by Mycobacterium avium complex, 1 caused by Mycobacterium malmoense, 1 caused by Mycobacterium haemophilum, and 2 caused by Mycobacterium nonchromogenicum isolates. In addition, a comprehensive literature search revealed an additional 48 cases of synovitis caused by slow-growing nonchromogenic mycobacteria. The historic literature described the majority of the 48 patients as previously healthy, elderly individuals with a several-month history of monoarticular pain and swelling in the small joints of the upper extremity. In contrast, the current series demonstrated the probable role of multiple chronic coexisting medical conditions in promoting disease susceptibility. These patients were also unique in their significantly younger age distribution and diversity of infection sites. Histologic examination and direct acid-fast bacteria stains generally did not aid the diagnosis. Amputation was performed in 2 patients because of delayed identification of disease. The current series demonstrates that difficult identification and infrequent occurrence cause these organisms to be overlooked by physicians and laboratory personnel. A heightened clinical suspicion for slow-growing nonchromogenic mycobacterial species is necessary when routine culture and histopathologic findings do not readily isolate an organism, or when the patient does not respond to antibiotic and anti-inflammatory treatment.

Mycobacterial synovitis caused by slow-growing nonchromogenic species: eighteen cases and a review of the literature.

Abstract Context.—Slow-growing nonchromogenic mycobacterial species are an infrequent cause of soft tissue infection. Because these organisms are rare, they are not often initially considered in the differential diagnosis of synovitis. Objective.—To evaluate the clinical and pathologic characteristics of patients with synovitis resulting from slow-growing nonchromogenic mycobacterial species. Design.—A 20-year retrospective review of records from The Methodist Hospital Microbiology Laboratory identified 18 culture-positive cases of synovitis that resulted from slow-growing nonchromogenic mycobacteria, including 14 caused by Mycobacterium avium complex, 1 caused by Mycobacterium malmoense, 1 caused by Mycobacterium haemophilum, and 2 caused by Mycobacterium nonchromogenicum isolates. In addition, a comprehensive literature search revealed an additional 48 cases of synovitis caused by slow-growing nonchromogenic mycobacteria. Results.—The historic literature described the majority of the 48 patients as prev...

A case of Mycobacterium nonchromogenicum pulmonary infection showing multiple nodular shadows in an immunocompetent patient

Mycobacterium nonchromogenicum is generally considered nonpathogenic. However, M. nonchromogenicum rarely causes human disease; particularly, pulmonary disease is extremely rare. The common finding of M. nonchromogenicum pulmonary infection on chest X-ray is a solitary cavity. The present report describes an unusual case of M. nonchromogenicum primary pulmonary infection showing multiple nodular shadows.

Molecular analyses of mycobacteria other than the M. tuberculosis complex isolated from Northern Ireland cattle

Mycobacteria other than the Mycobacterium tuberculosis complex (MOTT), isolated from Northern Ireland cattle, were identified by PCR amplification of the 16S rRNA gene, and subsequent reverse cross blot hybridisation and sequence analyses. Elucidation of the MOTT species was to facilitate specificity testing of new and existing diagnostic test reagents for bovine tuberculosis. The presence of the genes for potential diagnostic antigens: MPB70, MPB64, ESAT-6 and CFP-10 in the isolated MOTT species was investigated. Molecular analyses of cultured isolates from bovine lymph node specimens of 48 cattle identified a wide variety of mycobacterial species including Mycobacterium nonchromogenicum, Mycobacterium malmoense, Mycobacterium bohemicum, Mycobacterium paratuberculosis, Mycobacterium avium, Mycobacterium kansasii, Mycobacterium holsaticum, Mycobacterium palustre, Mycobacterium sp. IWGMT 90210, Mycobacterium sp. LIV-2129, a potentially novel mycobacterial species (EMBL/GenBank/DDBJ Accession Number AJ617495) and Rhodococcus equi. Apart from M. kansasii, the results of traditional (standard phenotypic and biochemical) and molecular identification methods did not correlate well, with traditional methods identifying fewer species. Most of the species identified were either recognised pathogenic or potential pathogenic species. The genes for ESAT-6, CFP-10 and, unusually, MPB64 were detected in M. kansasii only. The MPB70 gene was not detected in any of the species. This study supported restricted species distribution of these genes as well as identifying a different range of MOTT species that could be included in specificity testing of new diagnostic reagents for bovine tuberculosis.