Variable-Number Tandem-Repeat Analysis of Respiratory and Household Water Biofilm Isolates of “Mycobacterium avium subsp. hominissuis” with Establishment of a PCR Database

Abstract

d"Mycobacterium avium subsp. hominissuis" is an important cause of pulmonary disease. It is acquired from environmental sources, but there is no methodology for large population studies. We evaluated the potential of variable-number tandemrepeat (VNTR) analysis. Clinical and household biofilm M. avium isolates underwent molecular identification. Testing for IS901 was done to separate M. avium subsp. avium from M. avium subsp. hominissuis. VNTR types were defined using VNTR loci, and subtyping was performed using 3= hsp65 and internal transcribed spacer (ITS) sequencing. Forty-nine VNTR types and eight subtypes of M. avium subsp. hominissuis (IS901 negative) were identified among 416 isolates of M. avium from 121 patients and 80 biofilm sites. Of those types, 67% were found only among patient isolates, 11% only among household water isolates, and 23% among both. Of 13 VNTR types that included >4 patients, the majority (61.5%) represented geographic clustering (same city). Most VNTR types with multiple patients belonged to the same 3= hsp65 sequence code (sequevar). A total of 44 isolates belonging to four M. avium subsp. hominissuis VNTR types (8%), including three with the rare Mav-F ITS sequence and 0/8 subspecies, produced amplicons with IS901 PCR primers. By sequencing, all 44 amplicons were not IS901 but ISMav6, which was recently observed in Japan but had not been previously described among U.S. isolates. VNTR analysis of M. avium subsp. hominissuis isolates is easier and faster than pulsed-field gel electrophoresis. Seven VNTR loci separated 417 isolates into 49 types. No isolates of M. avium subsp. avium were identified. The distributions of the VNTR copy numbers, the allelic diversity, and the low prevalence of ISMav6 differed from the findings for respiratory isolates reported from Japan. The species "Mycobacterium avium subsp. hominissuis" is an important cause of chronic lung disease in the setting of bronchiectasis and disseminated disease in patients with AIDS. The species is acquired from environmental sources, with recent studies focusing on household water. It is present worldwide, with all areas of the United States (especially the southern United States) being considered high-risk areas for the disease.Efforts to compare isolates of M. avium have been difficult. Three subspecies are currently recognized, namely, M. avium subsp. avium, M. avium subsp. paratuberculosis, and M. avium subsp. silvaticum. A fourth subspecies, M. avium subsp. hominissuis (including human isolates), has also been proposed (1, 2). IS901 is present in all isolates of M. avium subsp. avium but is not present in isolates of M. avium subsp. hominissuis (1); this difference has been used to separate the two subspecies (1, 2). Strain comparisons within M. avium (as well as with other members of the Mycobacterium avium complex [MAC], especially Mycobacterium intracellulare) have most frequently utilized pulsed-field gel electrophoresis (PFGE) (3, 4), which is expensive, technically difficult, time-consuming, and useful only fo...

Population Structure and Local Adaptation of MAC Lung Disease Agent Mycobacterium avium subsp. hominissuis

“…Epidemiological analyses of Mycobacteria routinely include a microsatellite copy number analysis (MLVA) to investigate the genetic relatedness of clinical and environmental isolates ( Supply etal. 2006 ; Iakhiaeva etal. 2016 ).…”

Randomized Trial of Liposomal Amikacin for Inhalation in Nontuberculous Mycobacterial Lung Disease

“…A microbiologic recurrence was defined as one or more positive cultures after culture conversion. Cultures positive for M. avium complex occurring after culture conversion were molecularly compared with pretreatment isolates using partial 16S ribosomal RNA (rRNA) gene sequencing and variable number tandem repeat (VNTR) to determine if the isolates belonged to the same M. avium complex species and genotype (relapse) (29)(30)(31) or if their sequencing or VNTR differed (new infection) (31).…”

Synergistic Interactions in Microbial Biofilms Facilitate the Establishment of Opportunistic Pathogenic Fungi in Household Dishwashers

“…Survival of microorganisms in extreme environments is often associated with formation of complex biofilms attached on a suitable surface ( Davey and O’toole, 2000 ). In domestic environments, biofilms were examined in tap water supply systems ( Mullis and Falkinham, 2013 ; Burkowska-But et al, 2015 ; Lührig et al, 2015 ; Babič et al, 2016 ; Iakhiaeva et al, 2016 ; Ling et al, 2016 ) and in wet niches such as shower heads ( Abe et al, 2016 ). In this study, we focussed on the isolation of microorganisms from biofilms formed on rubber seals of four dishwashers (DWs) and used these isolates to determine their biofilm forming abilities in vitro .…”