Nontuberculous Mycobacterial Species Research Articles

Comparison of efficacies of different treatments for nontuberculous mycobacterial pulmonary disease in Anhui Province, China.

Although nontuberculous mycobacterial (NTM) infection is a common cause of pulmonary disease worldwide, few studies have focused on epidemiological and therapeutic factors related to NTM cases in Anhui Province, China. This retrospective study aimed to identify aetiological and clinical factors, and treatment outcomes of patients with NTM pulmonary disease (NTMPD) in Anhui. Retrospective clinical data obtained from medical records of NTMPD patients seeking care at Anhui Chest Hospital from July 2019 to June 2022 were analyzed. Treatment outcomes were compared between two patient groups: one receiving a standardised NTM treatment regimen and the other receiving precision treatment regimens. Genotypic analysis of 672 clinical NTMPD-associated isolates revealed that most were Mycobacterium intracellulare, while drug-susceptibility test results demonstrated diverse antibiotic resistance profiles for these isolates. Cough was the most common symptom for 101 NTMPD patients. After patients of both groups received treatment, symptoms improved, sputum culture conversion was observed for some patients, imaging findings stabilised; however, no statistically significant intergroup differences in treatment outcomes were found. In this study, M. intracellulare was the predominant NTM species identified in isolates obtained from NTMPD patients. Drug resistance profiles of our patient isolates were complex, highlighting the need for administration of timely, more effective, standardised treatments for patients with NTMPD in Anhui Province, China.

Nontuberculous mycobacterial disease in children: A systematic review and meta-analysis

BackgroundThe prevalence of nontuberculous mycobacterial (NTM) disease in children is increasing worldwide. The clinical manifestations of pediatric NTM patients are significantly different from those of adult patients, but the knowledge of the disease is generally poor. MethodsEnglish databases (PubMed, Web of Science, Embase, BIOSIS) and Chinese databases (CNKI, Wanfan, VIP) were searched on October 15th, 2022. All the articles of cross-sectional and cohort studies reporting the species composition and lesion site of the NTM disease in children using well-recognized NTM species identification methods were taken into account. Using a random effects model, we assessed the disease lesion sites and the prevalence of different NTM species in pediatric NTM disease. Sources of heterogeneity were analyzed using Cochran's Q and the I2 statistic. All analyses were performed using CMA V3.0. ResultsThe prevalence rates of NTM disease in children ranged between 0.6 and 5.36/100,000 in different countries, and Europe reported the highest prevalence rate. The most common clinical lesion site was lymph node, accounting for 71.1 % (55.0 %–83.2 %), followed by lung (19.3 %, 9.8%–34.4 %）and then skin and soft tissue (16.6 %,13.5%–20.3 %). Mycobacterium avium complex (MAC) was the most isolated NTM pathogen in children, accounting for 54.9 % (39.4%–69.6 %). Inconsistent with adult patients, Mycobacterium avium accounted for a dominant proportion in MAC than Mycobacterium intracellulare. ConclusionsThe lymph node was the most affected organ in pediatric NTM disease, while Mycobacterium avium was the most isolated pathogenic species in children.

Diagnostic Accuracy of the Abbott SD Bioline MPT64 antigen test for identification of MTB Complex in a U.S. Clinical Mycobacteriology Laboratory

Identification of the Mycobacterium tuberculosis complex (MTBC) from culture and differentiation from other non-tuberculous mycobacterial species is required for rapid diagnosis and accurate treatment. However, gaps exist in culture-based identification of acid-fast bacilli (AFB) positive cultures for rapid rule-out of MTBC in the United States. The SD Bioline™ MPT64 (Abbott Inc, South Korea) lateral flow assay (LFA) has high sensitivity and specificity for the detection of MTBC in liquid culture but has not been evaluated in a clinical mycobacteriology laboratory in the United States. We conducted a diagnostic accuracy study of this LFA for detection of MTBC versus NTMs on AFB positive cultures. A total of 362 tests were performed, with a sensitivity and specificity of 100 % (362/362) across all tests. The SD Bioline MPT64 assay provides accurate test results with AFB-positive liquid cultures and could fill the current gap for rapid rule-out of MTBC in U.S.-based clinical laboratories.

Nontuberculous Mycobacterial Infections in Pediatric Solid Organ Transplant and Hematopoietic Cell Transplant Recipients.

The diagnosis of nontuberculous mycobacterial infections is challenging in pediatric solid organ transplant and hematopoietic cell transplant recipients due to the absence of specific clinical manifestations, limitations of sampling, prolonged times for culture and identification, and difficulty discerning colonization from clinical disease. Treatment is dependent on the nontuberculous mycobacterial species, disease type, and pattern of drug resistance. Treatment of nontuberculous mycobacterial infections involves prolonged durations of therapy using multiple medications, which are limited by toxicities and drug-drug interactions.

Evaluation of the FluoroType Mycobacteria assay for identification of mycobacteria.

This study is the first clinical evaluation report of the novel FluoroType Mycobacteria assay. The assay has the potential to replace the established GenoType NTM product family in identification of culture-enriched mycobacteria. However, our research results suggest that the assay performs suboptimally and may not be feasible for use in all clinical settings.

Development of 2nd generation aminomethyl spectinomycins that overcome native efflux in Mycobacterium abscessus.

Mycobacterium abscessus (Mab), a nontuberculous mycobacterial (NTM) species, is an emerging pathogen with high intrinsic drug resistance. Current standard-of-care therapy results in poor outcomes, demonstrating the urgent need to develop effective antimycobacterial regimens. Through synthetic modification of spectinomycin (SPC), we have identified a distinct structural subclass of N-ethylene linked aminomethyl SPCs (eAmSPCs) that are up to 64-fold more potent against Mab over the parent SPC. Mechanism of action and crystallography studies demonstrate that the eAmSPCs display a mode of ribosomal inhibition consistent with SPC. However, they exert their increased antimicrobial activity through enhanced accumulation, largely by circumventing efflux mechanisms. The N-ethylene linkage within this series plays a critical role in avoiding TetV-mediated efflux, as lead eAmSPC 2593 displays a mere fourfold susceptibility improvement against Mab ΔtetV, in contrast to the 64-fold increase for SPC. Even a minor shortening of the linkage by a single carbon, akin to 1st generation AmSPC 1950, results in a substantial increase in MICs and a 16-fold rise in susceptibility against Mab ΔtetV. These shifts suggest that longer linkages might modify the kinetics of drug expulsion by TetV, ultimately shifting the equilibrium towards heightened intracellular concentrations and enhanced antimicrobial efficacy. Furthermore, lead eAmSPCs were also shown to synergize with various classes of anti-Mab antibiotics and retain activity against clinical isolates and other mycobacterial strains. Encouraging pharmacokinetic profiles coupled with robust efficacy in Mab murine infection models suggest that eAmSPCs hold the potential to be developed into treatments for Mab and other NTM infections.

Clinical Characteristics and Treatment Outcomes of Pulmonary Diseases Caused by Coinfections With Multiple Nontuberculous Mycobacterial Species.

Coinfections with multiple nontuberculous mycobacterial (NTM) species have not been widely studied. We aimed to evaluate the clinical characteristics and treatment outcomes in patients with NTM-pulmonary disease (PD) caused by coinfection with multiple NTM species. We retrospectively reviewed patients with NTM-PD at a tertiary referral hospital in Korea between March 2012 and December 2018. Coinfection was defined as two or more species of NTM pathogens isolated from the same respiratory specimen or different specimens within three months. Among 1,009 patients with NTM-PD, 147 (14.6%) NTM coinfections were observed (average age 64.7 years, 69.4% women). NTM species were identified more frequently (median 6 vs. 3 times, P < 0.001) in the coinfection group than in the single species group, and follow-up duration was also longer in the coinfection group (median 44.9 vs. 27.1 months, P < 0.001). Mycobacterium avium complex (MAC) and M. abscessus and M. massiliense (MAB) were the dominant combinations (n = 71, 48.3%). For patients treated for over six months in the MAC plus MAB group (n = 31), sputum culture conversion and microbiological cure were achieved in 67.7% and 41.9% of patients, respectively. We divided the MAC plus MAB coinfection group into three subgroups according to the target mycobacteria; however, no statistical differences were found in the treatment outcomes. In NTM-PD cases, a significant number of multiple NTM species coinfections occurred. Proper identification of all cultured NTM species through follow-up is necessary to detect multispecies coinfections. Further research is needed to understand the nature of NTM-PD in such cases.

Management Challenges of Extrapulmonary Nontuberculous Mycobacterial Infection: A Single-Center Case Series and Literature Review.

Extrapulmonary nontuberculous mycobacterial (NTM) disease remains largely enigmatic, yet these mycobacteria are increasingly acknowledged as important opportunistic pathogens in humans. Traditionally, NTM infections have been identified across various anatomical locations, with the respiratory system being the most affected and best understood. Historically, extrapulmonary NTM infection was predominantly associated with HIV/AIDS, with Mycobacterium avium lymphadenopathy being the most commonly reported. Today, however, because of the expanding utilization of immunosuppressive therapies and the demographic shift towards an aging population, an increasing number of NTM infections are expected and seen. Hence, a heightened index of suspicion is essential, necessitating a multifaceted approach to identification and drug sensitivity testing to improve treatment outcomes. In extrapulmonary NTM management, expert consultation is strongly recommended to determine the most efficacious treatment regimen, as individualized, patient-tailored therapies are often required. Furthermore, the economic burden of NTM disease is considerable, accompanied by high rates of hospitalization. To optimize the management of these intricate infections, there is an urgent need for comprehensive data on incidence, prevalence, and outcomes. This case-based series delves into the intricate nature of extrapulmonary NTM infections, focusing on both rapid and slow-growing NTM species, and explores therapeutic options, resistance mechanisms, and host-related immunological factors.

Trend of nontuberculous mycobacteria species and minimal inhibitory concentration in a referral laboratory in Korea from 2013 to 2019

BackgroundThis study investigated the trends of nontuberculous mycobacterial (NTM) isolates and the minimal inhibitory concentrations (MIC) of antimicrobial agents in Korea. MethodsData from 2013 to 2019 were collected from 69 medical institutions through 12 branches of the Korean Institute of Tuberculosis. NTM identification was conducted using the Advansure Mycobacteria Genoblot assay. The MIC of antibiotics against NTM species were measured using the broth microdilution method according to the Clinical and Laboratory Standards Institute guidelines. ResultsOver seven years, 86,194 NTM identifications were requested, with an annual increase from 8034 in 2013–17,229 in 2019. The most frequently identified NTM species were M. intracellulare (33,467; 47.3%) and M. avium (19,818; 27.2%), followed by M. abscessus (6858; 9.4%) and M. massiliense (3156; 4.3%). Regarding the antimicrobial agents, imipenem exhibited the greatest difference in MIC between M. intracellulare and M. avium, whereas clarithromycin showed the most significant difference between M. abscessus and M. massiliense. No notable changes were observed in the annual MIC distribution of most antibacterial agents, except for clarithromycin in M. abscessus. ConclusionsThe prevalence of NTM in Korea is gradually increasing, and follow-up studies on NTM isolates identified as the causative agents of infection are needed.

984. Detection of Mycobacterial Species from Specimens Submitted for Metagenomic Analysis

Abstract Background A national U.S. epidemic of non-tuberculous mycobacterial (NTM) infections is occurring at rates unseen over earlier decades, resulting in substantial numbers of pathogenic sequelae. Culture of mycobacteria is laborious, often requiring weeks for cultivation and speciation. Next generation sequencing (NGS) to detect NTM from clinical specimens has advantages over culture methods. From January 2018 to June 2022 we queried a large national NGS database to detect NTM rates and species. Methods DNA was isolated from 431,716 patient specimens collected from a variety of body sites: sputum, urine, sinus, wound or an unspecified swabs. Employing an Illumina’s 16S and ITS targeted-gene, deep sequencing approach, bacterial and/or fungal species resident in the specimen were determined. The identity and relative abundance of mycobacterial DNA recovered for each specimen were reported by species as the primary outcomes. Results Of the 431,716 specimens analyzed in this 4.5 year retrospective study, 0.001% (468) contained mycobacterial DNA. Forty different species of mycobacteria were detected with the most common frequency including: M abscessus, 40%; M mucogenicum, 13%; M. tuberculosis, 8.5% and M immunogenum 4.6%. M. avian and M. intracellulare were each detected in only 4% of the positive NTM specimens. 15% were other NTM species and 15% were not speciated NTM. Between 2018-2022 of the 468 NTM, the annual number of M. abscessus was 38, 53, 41 and 41, respectively. The concentration or relative abundance of mycobacterial DNA within each specimen varied from 4 to 100% of the total bacterial DNA recovered. Conclusion Over 4.5 years , NGS performed at a national molecular diagnostic laboratory detected 40 NTM species with a predominant fraction as M. abscessus (40%), supporting the clinical concern for an epidemic increase in M. abscessus infections during the last half decade. These data support the view that NGS metagenomics can uncover NTM epidemiological trends and implicate NTM infections. Disclosures Joseph F. John, Jr., MD, FACP, FIDSA, MicroGenDx: Advisor/Consultant Michael G. Schmidt, PhD, MicroGenDx: Advisor/Consultant Courtney L. Jarvis, PhD, MicroGenDX: Employee Niccole Diaz, BS, MicroGenDX: Advisor/Consultant Rick Martin, BS, MicroGenDX: Ownership Interest

Assessing residential activity in a home plumbing system simulator: monitoring the occurrence and relationship of major opportunistic pathogens and phagocytic amoebas.

Opportunistic premise plumbing pathogens (OPPPs) have been detected in buildings' plumbing systems causing waterborne disease outbreaks in the United States. In this study, we monitored the occurrence of OPPPs along with free-living amoeba (FLA) and investigated the effects of residential activities in a simulated home plumbing system (HPS). Water samples were collected from various locations in the HPS and analyzed for three major OPPPs: Legionella pneumophila, nontuberculous mycobacterial species (e.g., Mycobacterium avium, M. intracellulare, and M. abscessus), and Pseudomonas aeruginosa along with two groups of amoebas (Acanthamoeba and Vermamoeba vermiformis). A metagenomic approach was also used to further characterize the microbial communities. Results show that the microbial community is highly diverse with evidence of spatial and temporal structuring influenced by environmental conditions. L. pneumophila was the most prevalent pathogen (86% of samples), followed by M. intracellulare (66%) and P. aeruginosa (21%). Interestingly, M. avium and M. abscessus were not detected in any samples. The data revealed a relatively low prevalence of Acanthamoeba spp. (4%), while V. vermiformis was widely detected (81%) across all the sampling locations within the HPS. Locations with a high concentration of L. pneumophila and M. intracellulare coincided with the highest detection of V. vermiformis, suggesting the potential growth of both populations within FLA and additional protection in drinking water. After a period of stagnation lasting at least 2-weeks, the concentrations of OPPPs and amoeba immediately increased and then decreased gradually back to the baseline. Furthermore, monitoring the microbial population after drainage of the hot water tank and partial drainage of the entire HPS demonstrated no significant mitigation of the selected OPPPs. This study demonstrates that these organisms can adjust to their environment during such events and may survive in biofilms and/or grow within FLA, protecting them from stressors in the supplied water.

Silencing essential gene expression in Mycobacterium abscessus during infection.

Mycobacterium abscessus is a multi-drug-resistant non-tuberculous mycobacterial species causing tuberculosis-like lung infections. The functional characterization of the extensive repertoire of genes encoding virulence factors and drug targets has been largely restricted by the lack of powerful genetic tools. In this study, we evaluated the performances of a Tet-OFF system, previously optimized for M. tuberculosis and M. smegmatis. Fluorescence reporter M. abscessus strains were developed where mCherry was under the control of the Tet-OFF regulatory system on an integrative plasmid. The addition of anhydrotetracycline (ATc) correlated with a decrease in fluorescence intensity not only in solid and broth medium but also in colony biofilms, for both smooth and rough variants of M. abscessus. Next, unmarked mmpL3 conditional knockdown mutants were engineered in both smooth and rough M. abscessus. Biochemical studies indicated that the addition of ATc was associated with a dose-dependent decrease in (i) the production of the MmpL3 protein, (ii) the biosynthesis of trehalose dimycolate and mycolylated arabinogalactan, (iii) bacterial viability on agar and liquid medium as well as in biofilms. Importantly, intravenous injection of ATc in zebrafish embryos infected with the rough mmpL3 conditional mutant impeded bacterial growth and correlated with a significant gain in embryo survival. These results suggest that mmpL3 is essential for M. abscessus growth in vitro and in the infected host, thus validating MmpL3 as an attractive drug target. Together, this study demonstrates the potential of ATc-dependent repression to modulate the expression of M. abscessus genes in the infected host. IMPORTANCE Mycobacterium abscessus represents the most common rapidly growing mycobacterial pathogen in cystic fibrosis and is extremely difficult to eradicate. Essential genes are required for growth, often participate in pathogenesis, and encode valid drug targets for further chemotherapeutic developments. However, assessing the function of essential genes in M. abscessus remains challenging due to the limited spectrum of efficient genetic tools. Herein, we generated a Tet-OFF-based system allowing to knock down the expression of mmpL3, encoding the mycolic acid transporter in mycobacteria. Using this conditional mutant, we confirm the essentiality of mmpL3 in planktonic cultures, in biofilms, and during infection in zebrafish embryos. Thus, in this study, we developed a robust and reliable method to silence the expression of any M. abscessus gene during host infection.

Blocking ADP-ribosylation expands the anti-mycobacterial spectrum of rifamycins.

The clinical utility of rifamycins against non-tuberculous mycobacterial (NTM) disease is limited by intrinsic drug resistance achieved by ADP-ribosyltransferase Arr. By blocking the site of ribosylation, we recently optimized a series of analogs with substantially improved potency against Mycobacterium abscessus. Here, we show that a representative member of this series is significantly more potent than rifabutin against major NTM pathogens expressing Arr, providing a powerful medicinal chemistry approach to expand the antimycobacterial spectrum of rifamycins. IMPORTANCE Lung disease caused by a range of different species of non-tuberculous mycobacteria (NTM) is difficult to cure. The rifamycins are very active against Mycobacterium tuberculosis, which causes tuberculosis (TB), but inactive against many NTM species. Previously, we showed that the natural resistance of the NTM Mycobacterium abscessus to rifamycins is due to enzymatic inactivation of the drug by the bacterium. We generated chemically modified versions of rifamycins that prevent inactivation by the bacterium and thus become highly active against M. abscessus. Here, we show that such a chemically modified rifamycin is also highly active against several additional NTM species that harbor the rifamycin inactivating enzyme found in M. abscessus, including M. chelonae, M. fortuitum, and M. simiae. This finding expands the potential therapeutic utility of our novel rifamycins to include several currently difficult-to-cure NTM lung disease pathogens beyond M. abscessus.

What do the clinical features of positive nontuberculous mycobacteria isolates from patients with HIV/AIDS in China reveal? A systematic review and meta-analysis.

China has a high burden of nontuberculous mycobacterial (NTM) infections. Immunocompromised populations, such as those with human immunodeficiency virus/acquired immunodeficiency syndrome (HIV/AIDS), are at a higher risk of being infected with NTM than immunocompetent individuals. Yet, there is a paucity of information on the clinical features of positive NTM isolates from patients with HIV/AIDS in China. To address this gap, we conducted a systematic review and meta-analysis of existing studies, comparing them against current expert consensus to provide guidance for clinical practice. Two researchers independently searched eight databases (SinoMed, China National Knowledge Infrastructure, Wanfang, VIP, Cochrane Library, PubMed, Embase, and Web of Science) from inception to 26 December 2022 to retrieve published Chinese- and English-language studies reporting clinical features of NTM-positive isolates among patients with HIV/AIDS in China. We included 28 studies with 1861 patients. The rate of positive NTM isolates detected from men among all patients was 87.3%. NTM species distribution was mainly Mycobacterium avium complex (64.3%), which was predominant in different regions. The five most common clinical symptoms were fever (68.5%), cough or expectoration (67.0%), appetite loss (49.4%), weight loss (45.5%), and superficial lymphadenectasis (41.1%). The prevalence of laboratory tests were as follows: albumin <35 g/L (55.6%), erythrocyte sedimentation rate >20 mm/h (91.4%), anaemia (59.0%), predominantly mild, CD4+ T cell count ≤50 pieces/μL (70.3%), and CD4+ T cell count 51-200 pieces/μL (22.1%). Lesion manifestations in thoracic imaging mainly included bilateral lung involvement (83.8%), showed stripe shadows (60.3%), patchy shadows (42.9%), nodules (40.6%), and bronchiectasis (38.6%). Accompanied signs included thoracic lymph node enlargement (49.5%). Seventy per cent of symptoms improved after treatment. Focusing on clinical symptoms, laboratory tests, and thoracic imaging helps with initial screening for NTM infections. Physicians should raise awareness of the diagnosis and treatment of Mycobacterium avium complex, providing guidance for experimental treatment, screening of priority populations for NTM infections, and prophylactic treatment of NTM disease. PROSPERO CRD42023388185.

Metagenomic next-generation sequencing assistance in identifying non-tuberculous mycobacterial infections.

The advent of metagenomics next-generation sequencing (mNGS) has garnered attention as a novel method for detecting pathogenic infections, including Non-Tuberculous Mycobacterial (NTM) and tuberculosis (TB).However, the robustness and specificity of mNGS in NTM diagnostics have not been fully explored. In this retrospective study, we enrolled 27 patients with NTM genomic sequences via mNGS and conducted a comprehensive clinical evaluation. Pulmonary NTM disease was the most commonly observed presentation, with a subset of patients also presenting with extrapulmonary NTM infections.mNGS analysis identified six distinct NTM species, primarily Mycobacteriumavium complex (MAC), followed by Mycobacterium intracellulare andMycobacterium abscessus. Conventional routine culture methods encountered challenges, resulting in negative results for all available 22 samples. Among the 10 patients who underwent quantitative polymerase chain reaction (qPCR) testing, five tested positive for NTM. It is important to note that further species typing is necessary to determine the specific NTM type, as traditional pathogen detection methods serve as an initial step. In contrast, when supplemented with pathogen data, enables the identification of specific species, facilitating precise treatment decisions. In conclusion, mNGS demonstrates significant potential in aidingthe diagnosis of NTMdisease by rapidly detecting NTM pathogens and guiding treatment strategies. Its enhanced performance, faster turnaround time (TAT), and species identification capabilities make mNGS a promising tool for managing NTM infections.

Disseminated non-tuberculous mycobacterial infection caused by Mycobacterium obuense in an immunocompromised patient: a case report

BackgroundMycobacterium obuense (M. obuense) is a rapidly growing mycobacterium (RGM) which has been considered nonpathogenic. Here, we report a case of disseminated non-tuberculous mycobacterial (NTM) infection caused by M. obuense in an immunocompromised patient.Case presentationA 16-year-old boy was referred to our hospital due to acute myeloid leukemia. During the treatment of leukemia, the patient exhibited continuous fever, and diffuse miliary nodules with random distribution were found on chest computed tomography. Repeated examinations of bacterial culture tests revealed sputum and urine samples to be smear-positive for acid-fast bacillus, and blood culture from a peripherally inserted central catheter line showed the growth of NTM. The NTM species was identified as M. obuense by mass spectrometry and confirmed by genome sequencing. Combination therapy with amikacin, rifampicin, azithromycin, and moxifloxacin significantly improved the patient’s symptoms and radiological findings.ConclusionWe report a case of disseminated NTM infection caused by M. obuense for which combination anti-microbial therapy was effective. An immunocompromised host indwelling catheter is at risk of RGM bloodstream infections. Although relatively rare, M. obuense may be considered as a potential pathogen causing infectious diseases, especially in high-risk patients.

A Case of Mycobacterium avium Complex Vertebral Osteomyelitis in a Patient With Human Immunodeficiency Virus

Abstract Mycobacterium avium complex (MAC) infection is one of the most common bacterial opportunistic infections in persons living with human immunodeficiency virus (PLWH). Mycobacterium avium complex infections are caused by either M. avium or Mycobacterium intracellulare, which are nontuberculous mycobacterial species. Mycobacterium avium complex is prevalent in soil, water, and animals in the environment. Mycobacterium avium complex infections are commonly seen in persons with CD4 counts less than 50 cells/μL and are the most common cause of disseminated disease in immunocompromised patients. Vertebral osteomyelitis caused by MAC, however, remains uncommon in PLWH, with few cases reported in the literature (Lancet Infect Dis. 2004;4:557–565; Infect Dis Clin Pract. 2001;10:17–20). We present a case of vertebral osteomyelitis secondary to MAC in a person with human immunodeficiency virus and compare 10 other cases reported in the literature. Our case also highlights the importance of resuming secondary MAC prophylaxis when CD4 counts decline below 100 cells/μL in PLWH.

Molecular identification of non-tuberculous mycobacterial species isolated from extrapulmonary samples using real-time PCR and rpoB sequence analysis

Tuberculosis (TB) is one of the leading causes of mortality among infectious diseases and accounts for a serious health hazard wordwide. Apart from TB, the members of non-tuberculous mycobacteria (NTM), which includes around 170 species, may also cause different diseases in humans. Therefore this study aimed to investigate the distribution of NTM strains isolated from extrapulmonary (EP) samples by Real-Time PCR and PCR-sequencing methods in Southwest Iran. Three hundred and twenty-five suspected EP samples were collected from patients referred to the referral hospitals in Ahvaz, Iran. The isolates were initially screened by acid fast staining and identified by phenotypic culture and biochemical tests. The Real-Time PCR and rpoB- based PCR methods were performed followed by sequence analysis of rpoB gene. From 124 samples, 77 (62%) were positive for NTM by culture and rpoB sequence analysis. M. fortuitum was the most commonly isolated NTM in present study. In Real-Time PCR, only 69 (55.64%) isolates showed more homology with standard NTM isolates. In general, the growing trend of EPNTM infections in Iran needs specific programs and resources to get a better diagnosis. PCR sequencing is a reliable method, it can be used for definitive identification of positive cultures for identification of NTM species.

Analysis of time for diagnosis of nontuberculous mycobacterial lung disease and its associated factors in a tuberculosis-designated hospital in Shanghai

Objective: To investigate the pathogenic characteristics, bacteriological diagnosis time and its associated factors among patients with nontuberculous mycobacterial (NTM) lung disease in a large tuberculosis-designated hospital in Shanghai from 2020 to 2021, in order to improve diagnosis efficiency and formulate precision treatment. Methods: On the basis of the Tuberculosis Database in Shanghai Pulmonary Hospital, NTM patients diagnosed by the Department of Tuberculosis between January 2020 and December 2021 were screened. Demographic, clinical and bacterial information were retrospectively collected. Chi-square test, paired-sample nonparametric test and logistic regression model were used to analyze the factors associated with the diagnosis time of NTM lung disease. Results: A total of 294 patients with bacteriologically confirmed NTM lung disease were included in this study, 147 males and 147 females with a median age of 61(46, 69) years. Of them, 227 (77.2%) patients had comorbidity of bronchiectasis. Species identification results showed that Mycobacterium Avium-Intracellulare Complex was the main pathogen of NTM lung disease (56.1%), followed by Mycobacterium kansasii (19.0%) and Mycobacterium abscessus (15.3%). Species such as Mycobacterium xenopi and Mycobacterium malmoense were rarely identified, accounting for a total proportion of only 3.1%. Positive culture rates for sputum, bronchoalveolar lavage fluid and puncture fluid were 87.4%, 80.3% and 61.5%, respectively. Paired-sample analysis showed that the positive rate of sputum culture was significantly higher than that of smear microscopy (87.1% vs. 48.4%, P<0.01), while no statistical difference was observed between sputum and bronchoalveolar lavage fluid on positive culture rate (78.7% vs. 77.3%, P>0.05). Patients with cough or expectoration were observed with 4.04-fold (95%CI 1.80-9.05) or 2.95-fold (95%CI 1.34-6.52) higher probability of positive sputum culture, compared to those without. Regarding bronchoalveolar lavage fluid, female or patients with bronchiectasis had a 2.82-fold (95%CI 1.16-6.88) or 2.38-fold (95%CI 1.01-5.63) higher probability to achieve a positive culture. The median time to diagnosis of NTM lung disease was 32 (interquartile range: 26-42) days. The results of multivariable analysis showed that patients with symptom of expectoration (aOR=0.48, 95%CI 0.29-0.80) needed a shorter diagnosis time in comparison with patients without expectoration. With Mycobacterium Avium-Intracellulare Complex as a reference, lung disease caused by Mycobacterium abscessus needed shorter diagnosis time (aOR=0.43, 95%CI 0.21-0.88), whereas those caused by rare NTM species were observed to require a longer diagnosis time (aOR=8.31, 95%CI 1.01-68.6). Conclusion: The main pathogen causing NTM lung disease in Shanghai was Mycobacterium Avium-Intracellulare Complex. Sex, clinical symptoms and bronchiectasis had an impact on the positive rate of mycobacterial culture. The majority of patients in study hospital were timely diagnosed. Clinical symptoms and NTM species were associated with the bacteriological diagnosis time of NTM lung disease.

Antimicrobial Susceptibility Testing Using the MYCO Test System and MIC Distribution of 8 Drugs against Clinical Isolates of Nontuberculous Mycobacteria from Shanghai.

Given the increased incidence and prevalence of nontuberculous mycobacterial (NTM) diseases and the natural resistance of NTM to multiple antibiotics, in vitro susceptibility testing of different NTM species against drugs from the MYCO test system and new applied drugs is required. A total of 241 NTM clinical isolates were analyzed, including 181 slowly growing mycobacteria (SGM) and 60 rapidly growing mycobacteria (RGM). The Sensititre SLOMYCO and RAPMYCO panels were used for testing susceptibility to commonly used anti-NTM antibiotics. Furthermore, MIC distributions were determined against 8 potential anti-NTM drugs, including vancomycin (VAN), bedaquiline (BDQ), delamanid (DLM), faropenem (FAR), meropenem (MEM), clofazimine (CLO), cefoperazone-avibactam (CFP-AVI), and cefoxitin (FOX), and epidemiological cutoff values (ECOFFs) were analyzed using ECOFFinder. The results showed that most of the SGM strains were susceptible to amikacin (AMK), clarithromycin (CLA), and rifabutin (RFB) from the SLOMYCO panels and BDQ and CLO from the 8 applied drugs, while RGM strains were susceptible to tigecycline (TGC) from the RAPMYCO panels and also BDQ and CLO. The ECOFFs of CLO were 0.25, 0.25, 0.5, and 1 μg/mL for the mycobacteria M. kansasii, M. avium, M. intracellulare, and M. abscessus, respectively, and the ECOFF of BDQ was 0.5 μg/mL for the same four prevalent NTM species. Due to the weak activity of the other 6 drugs, no ECOFF was determined. This study on the susceptibility of NTM includes 8 potential anti-NTM drugs and a large sample size of Shanghai clinical isolates and demonstrates that BDQ and CLO had efficient activities against different NTM species in vitro, which can be applied to the treatment of NTM diseases. IMPORTANCE We designed customized panel that contains 8 repurposed drugs, including vancomycin (VAN), bedaquiline (BDQ), delamanid (DLM), faropenem (FAR), meropenem (MEM), clofazimine (CLO), cefoperazone-avibactam (CFP-AVI), and cefoxitin (FOX) from the MYCO test system. To better understand the efficacy of these 8 drugs against different NTM species, we determined the MICs of 241 NTM isolates collected in Shanghai, China. We attempted to define the tentative epidemiological cutoff values (ECOFFs) for the most prevalent NTM species, which is an important factor in setting up the breakpoint for a drug susceptibility testing. We used the MYCO test system as an automatic quantitative drug sensitivity test of NTM and extended the method to BDQ and CLO in this study. The MYCO test system complements commercial microdilution systems that currently lack BDQ and CLO detection.