https://onlinelibrary.wiley.com/doi/pdf/10.1111/j.1469-0691.2008.02070.x

Then and now: use of 16S rDNA gene sequencing for bacterial identification and discovery of novel bacteria in clinical microbiology laboratories

A significant knowledge gap exists concerning the geographical distribution of nontuberculous mycobacteria (NTM) isolation worldwide. To provide a snapshot of NTM species distribution, global partners in the NTM-Network European Trials Group (NET) framework (www.ntm-net.org), a branch of the Tuberculosis Network European Trials Group (TB-NET), provided identification results of the total number of patients in 2008 in whom NTM were isolated from pulmonary samples. From these data, we visualised the relative distribution of the different NTM found per continent and per country. We received species identification data for 20 182 patients, from 62 laboratories in 30 countries across six continents. 91 different NTM species were isolated. Mycobacterium avium complex (MAC) bacteria predominated in most countries, followed by M. gordonae and M. xenopi. Important differences in geographical distribution of MAC species as well as M. xenopi, M. kansasii and rapid-growing mycobacteria were observed. This snapshot demonstrates that the species distribution among NTM isolates from pulmonary specimens in the year 2008 differed by continent and differed by country within these continents. These differences in species distribution may partly determine the frequency and manifestations of pulmonary NTM disease in each geographical location.

https://erj.ersjournals.com/content/erj/42/6/1604.full.pdf

The geographic diversity of nontuberculous mycobacteria isolated from pulmonary samples: an NTM-NET collaborative study

Section 4: What is the evidence for transmission of NTM between individuals? 

Recommendation 

Section 5: How should the lung disease attributable to NTM infection be defined? 

Recommendation 

Good practice point 

Section 6: What samples should be used to detect pulmonary non-tuberculous mycobacterial infection? 

Recommendations 

Good practice points 

Section …

/pdf/british-thoracic-society-guidelines-for-the-management-of-35wnkt4897.pdf

British Thoracic Society guidelines for the management of non-tuberculous mycobacterial pulmonary disease (NTM-PD)

Summary: Within the past 10 years, treatment and diagnostic guidelines for nontuberculous mycobacteria have been recommended by the American Thoracic Society (ATS) and the Infectious Diseases Society of America (IDSA). Moreover, the Clinical and Laboratory Standards Institute (CLSI) has published and recently (in 2011) updated recommendations including suggested antimicrobial and susceptibility breakpoints. The CLSI has also recommended the broth microdilution method as the gold standard for laboratories performing antimicrobial susceptibility testing of nontuberculous mycobacteria. This article reviews the laboratory, diagnostic, and treatment guidelines together with established and probable drug resistance mechanisms of the nontuberculous mycobacteria.

Antimicrobial susceptibility testing, drug resistance mechanisms, and therapy of infections with nontuberculous mycobacteria.

The application of phylogenetic taxonomic procedures led to improvements in the classification of bacteria assigned to the phylum Actinobacteria but even so there remains a need to further clarify relationships within a taxon that encompasses organisms of agricultural, biotechnological, clinical, and ecological importance. Classification of the morphologically diverse bacteria belonging to this large phylum based on a limited number of features has proved to be difficult, not least when taxonomic decisions rested heavily on interpretation of poorly resolved 16S rRNA gene trees. Here, draft genome sequences of a large collection of actinobacterial type strains were used to infer phylogenetic trees from genome-scale data using principles drawn from phylogenetic systematics. The majority of taxa were found to be monophyletic but several orders, families, and genera, as well as many species and a few subspecies were shown to be in need of revision leading to proposals for the recognition of 2 orders, 10 families, and 17 genera, as well as the transfer of over 100 species to other genera. In addition, emended descriptions are given for many species mainly involving the addition of data on genome size and DNA G+C content, the former can be considered to be a valuable taxonomic marker in actinobacterial systematics. Many of the incongruities detected when the results of the present study were compared with existing classifications had been recognized from 16S rRNA gene trees though whole-genome phylogenies proved to be much better resolved. The few significant incongruities found between 16S/23S rRNA and whole genome trees underline the pitfalls inherent in phylogenies based upon single gene sequences. Similarly good congruence was found between the discontinuous distribution of phenotypic properties and taxa delineated in the phylogenetic trees though diverse non-monophyletic taxa appeared to be based on the use of plesiomorphic character states as diagnostic features.

/pdf/genome-based-taxonomic-classification-of-the-phylum-4uc1zrzg88.pdf

Genome-Based Taxonomic Classification of the Phylum Actinobacteria

The possibility that the strains included within the Mycobacterium avium complex (MAC), but not belonging either to M. avium or to Mycobacterium intracellulare, may be members of undescribed taxa, has already been questioned by several taxonomists. A very homogeneous cluster of 12 strains characterized by identical nucleotide sequences both in the 16S rDNA and in the 16S–23S internal transcribed spacer was investigated. Similar strains, previously reported in the literature, had been assigned either to the species M. intracellulare on the basis of the 16S rDNA similarity or to the group of MAC intermediates. However, several phenotypical and epidemiological characteristics seem to distinguish these strains from all other MAC organisms. The unique mycolic acid pattern obtained by HPLC is striking as it is characterized by two clusters of peaks, instead of the three presented by all other MAC organisms. All of the strains have been isolated from humans and all but one came from the respiratory tract of elderly people. The clinical significance of these strains, ascertained for seven patients, seems to suggest an unusually high virulence. The characteristics of all the strains reported in the literature, genotypically identical to the ones described here, seem to confirm our data, without reports of isolations from animals or the environment or, among humans, from AIDS patients. Therefore, an elevation of the MAC variant was proposed and characterized here, with the name Mycobacterium chimaera sp. nov.; this increases the number of species included in the M. avium complex. The type strain is FI-01069T (=CIP 107892T=DSM 44623T).

Proposal to elevate the genetic variant MAC-A, included in the Mycobacterium avium complex, to species rank as Mycobacterium chimaera sp. nov.

INNO-LiPA MYCOBACTERIA (Innogenetics, Ghent, Belgium) is a reverse hybridization DNA probe assay that has been recently improved by increasing the number of identifiable mycobacterial species to 16. Our assessment, performed with 197 mycobacteria belonging to 81 taxa, revealed 100% specificity and sensitivity for 20 out of 23 probes. The probes specific for Mycobacterium fortuitum complex, for the Mycobacterium avium-intracellulare-scrofulaceum group, and for Mycobacterium intracellulare type 2 cross-reacted with several mycobacteria rarely isolated from clinical specimens. The overall sensitivity was 100%, and the overall specificity was 94.4%.

Evaluation of INNO-LiPA MYCOBACTERIA v2: improved reverse hybridization multiple DNA probe assay for mycobacterial identification.

We evaluated the BDProbeTec ET system (Becton Dickinson, Sparks, Md.), a strand displacement amplification-based technique, for direct detection of Mycobacterium tuberculosis in 867 clinical samples. Of 294 extrapulmonary specimens, 52 had positive results by both BDProbeTec ET and culture and 209 had negative results by both methods; sensitivity and specificity were 76.5 and 95.9%, respectively. After resolution of discrepancies, the sensitivity rose to 77.8%.

Evaluation of the BDProbeTec ET system for direct detection of Mycobacterium tuberculosis in pulmonary and extrapulmonary samples: a multicenter study.

Eight mycobacterial strains isolated during an 11 year period from the sputum of independent patients with various pulmonary disorders and, in one case, from a lymph node of a young girl, were found to present identical features. Phenotypic and genotypic characteristics revealed that the most closely related species to these test isolates were Mycobacterium triplex and Mycobacterium lentiflavum. However, the lipids of the cell wall of the test isolates differed from those of the latter species by TLC and presented unique profiles by both GC and HPLC. Genotypic analysis showed that they had unique 16S rRNA gene and internal transcribed spacer (ITS) sequences, and could be differentiated from all other mycobacterial strains by PCR restriction analysis of hsp65. The strains presented high resistance to antimycobacterial drugs. The name Mycobacterium florentinum sp. nov. is proposed for this taxon, with strain FI-93171(T) (=DSM 44852(T) = CIP 108409(T)) as the type strain.

Mycobacterium florentinum sp. nov., isolated from humans

We report on four cases of infection by the recently described species Mycobacterium parascrofulaceum. In two cases the mycobacterium was isolated from AIDS patients, while in the others it was responsible for pulmonary disease in elderly men. Our findings suggest that M. parascrofulaceum is an opportunistic pathogen, like many other nontuberculous mycobacterial species.

Gianna Mazzarelli

Papers

Proposal to elevate the genetic variant MAC-A, included in the Mycobacterium avium complex, to species rank as Mycobacterium chimaera sp. nov.

Evaluation of INNO-LiPA MYCOBACTERIA v2: improved reverse hybridization multiple DNA probe assay for mycobacterial identification.

Evaluation of the BDProbeTec ET system for direct detection of Mycobacterium tuberculosis in pulmonary and extrapulmonary samples: a multicenter study.

Mycobacterium florentinum sp. nov., isolated from humans

Infections Due to the Newly Described Species Mycobacterium parascrofulaceum