Mycobacterium intracellulare among TB suspected patients in Bulgaria - microbiological aspects.

INTRODUCTION: Nontuberculous mycobacteria (NTM) are representatives of the genus Mycobacterium with a worldwide distribution, associated mainly with water, soil, and biofilms. Some of NTMs, such as Mycobacterium avium complex (MAC), are etiological agents of human diseases - disseminated or with different localization, most often pulmonary. AIM: In the present study, we analyzed Mycobacterium intracellulare isolates recovered from clinical specimens of tuberculosis (TB) suspected patients in Bulgaria, 2018-2020. MATERIALS AND METHODS: The cultures were grown on solid and liquid media. For species identification, we used immune chromatographic (TB Ag MPT64) test and Line Probe Assay (LPA) from the positive cultures. RESULTS: M. intracellulare was identified in 32 patients from 82,780 specimens. It was predominantly isolated in females - 62.5% vs. 37.5% in males. The most affected age group was 65 years and over (38%). The distribution of the isolates in Bulgaria was uneven. Most of them (65.6%) were concentrated in two districts of the country: Plovdiv and Sofia-city. All strains were sensitive to macrolides and aminoglycosides except one with macrolide resistance. NTM pulmonary disease was confirmed in 16 patients with M. intracellulare isolate. CONCLUSIONS: Analysing the 32 M. intracellulare isolates identified among TB suspected patients in Bulgaria between 2018 and 2020, we found that only half of them met the American Thoracic Society (ATS) diagnostic criteria for NTM pulmonary disease. For the remaining patients with M. intracellulare isolates we did not have sufficient data to support this diagnosis. Efforts by Bulgarian respiratory and microbiological societies are needed for adherence to the international guidelines.

Whole genome sequencing of Bulgarian rifampicin resistant Mycobacterium tuberculosis strains.

INTRODUCTION: The transmission of drug-resistant tuberculosis is one of the greatest challenges facing the global tuberculosis control. AIM: The aim of the study was to investigate the resent transmission of rifampicin resistant tuberculosis in Bulgaria and to describe the mutations related to the antimicrobials' resistance using whole genome sequencing. MATERIALS AND METHODS: As part of an ECDC funded pilot study for evaluation of the systematic use of whole genome sequencing (WGS) of Mycobacterium tuberculosis (MTB) surveillance (EUSeqMyTB), Bulgaria provided 65 rifampicin resistant isolates over a three years' timeframe (2017-2019) representing 87.5% of the notified rifampicin resistant cases. Drug resistance prediction and relatedness analysis of the resistant isolates was performed in collaboration with San Raffaele Scientific Institute, Milan, Italy. RESULTS: Almost all of the isolates were identified as Euro-American lineage (96.9%); 18.5% of the isolates were found to be resistant to fluoroquinolones, but no mutations conferring resistance to bedaquiline or linezolid could be identified. Less than half (43.3%) of the isolates were clustered (<5 SNPs distance) into a total of seven national SNP-based clusters, while a total of six isolates were found to be part of different cross-border clusters. All clustered cases originated from Bulgaria. CONCLUSIONS: WGS has proven to be a reliable tool for surveillance and tracing of recent transmission of tuberculosis and has the potential for resistance prediction for most of the antituberculosis drugs.

First Etiologically Confirmed Cases of Mycobacterium Marinum Infection in Bulgaria.

This study aimed to describe the first two microbiologically confirmed cases of cutaneous and soft tissue Mycobacterium marinum infection in Bulgaria. The isolation of the Nontuberculous Mycobacteria (NTM) strains and their species identification was performed at NRL TB, NCIPD using specific media and cultivation conditions, and PCR based Line Probe Assay (LPA) from the positive cultures. The two patients had closely related jobs to fishes and water reservoirs and both of them had a similar clinical manifestation of M. mari-num infection known as "swimming pool" or "fish tank" granuloma. The prolonged specific treatment with at least two-drug combina-tion of rifampicin plus ethambutol and some complications were a big challenge for clinicians as well as the patients.