Serological investigation and genotyping of Mycobacterium avium subsp. paratuberculosis in sheep and goats in Inner Mongolia, China.

Paratuberculosis a contagious and chronic disease in domestic and wild ruminants, is caused by Mycobacterium avium subspecies paratuberculosis (MAP). Typical clinical signs include intractable diarrhea, progressive emaciation, proliferative enteropathy, and mesenteric lymphadenitis. Paratuberculosis is endemic to many parts of the world and responsible for considerable economic losses. In this study, different types of paratuberculosis and MAP in sheep and goats were investigated in Inner Mongolia, a northern province in China contiguous with two countries and eight other provinces. A total of 4434 serum samples were collected from six cities in the western, central, and eastern regions of Inner Mongolia and analyzed using the ELISA test. In addition, tissue samples were collected from seven animals that were suspected to be infected with MAP. Finally, these tissues samples were analyzed by histopathological examination followed by polymerase chain reaction (PCR), IS1311 PCR-restriction enzyme analysis (PCR-REA), and a sequence analysis of five genes. Among all 4434 ruminant serum samples collected from the six cities in the western, central, and eastern regions of Inner Mongolia, 7.60% (337/4434) measured positive for the MAP antibody. The proportions of positive MAP antibody results for serum samples collected in the western, central, and eastern regions were 5.10% (105/2058), 6.63% (85/1282), and 13.44% (147/1094), respectively. For the seven suspected infected animals selected from the herd with the highest rate of positivity, the gross pathology and histopathology of the necropsied animals were found to be consistent with the pathological features of paratuberculosis. The PCR analysis further confirmed the diagnosis of paratuberculosis. The rest of the results demonstrated that herds of sheep and goats in Inner Mongolia were infected with both MAP type II and type III. To the best of our knowledge, this is the first study of the two subtypes of MAP strains in sheep and goats in Inner Mongolia.

Evaluation of IS1245 LAMP in Mycobacterium avium and the influence of host-related genetic diversity on its application.

Early detection and treatment are paramount for the timely control of Mycobacterium avium infections. Herein, we designed a LAMP assay targeting a widely used species-specific marker IS1245 for the rapid detection of M. avium and evaluated its applicability using human (n = 137) and pig (n = 91) M. avium isolates from Japan. The developed assay could detect as low as 1 genome copy of M. avium DNA within 30 minutes. All 91 (100%) M. avium isolates from pigs were detected positive while all other tested bacterial species were negative. Interestingly, among the 137 clinical M. avium isolates, 41 (30%) were undetectable with this LAMP assay as they lacked IS1245, the absence of which was revealed by PCR and whole-genome sequencing. These findings highlighted genotypic differences in M. avium strains from humans and pigs in Japan and how this diversity can influence the applicability of a detection tool across different geographic areas and hosts.

Difference in drug susceptibility distribution and clinical characteristics between Mycobacterium avium and Mycobacterium intracellulare lung diseases in Shanghai, China.

Introduction. Mycobacterium avium complex (MAC) has been reported as the most common aetiology of lung disease involving nontuberculous mycobacteria.Hypothesis. Antimicrobial susceptibility and clinical characteristics may differ between Mycobacterium avium and Mycobacterium intracellulare.Aim. We aimed to evaluate the differences in antimicrobial susceptibility profiles between two major MAC species (Mycobacterium avium and Mycobacterium intracellulare) from patients with pulmonary infections and to provide epidemiologic data with minimum inhibitory concentration (MIC) distributions.Methodology. Between January 2019 and May 2020, 45 M. avium and 242 M. intracellulare isolates were obtained from Shanghai Pulmonary Hospital. The demographic and clinical characteristics of patients were obtained from their medical records. The MICs of 13 antimicrobials were determined for the MAC isolates using commercial Sensititre SLOWMYCO MIC plates and the broth microdilution method, as recommended by the Clinical and Laboratory Standards Institute (CLSI; Standards M24-A2). MIC50 and MIC90 values were derived from the MIC distributions.Results. M. intracellulare had higher resistance rates than M. avium for most tested antimicrobials except clarithromycin, ethambutol, and ciprofloxacin. Clarithromycin was the most effective antimicrobial against both the M. avium (88.89 %) and M. intracellulare (91.32 %) isolates, with no significant difference between the species (P=0.601). The MIC90 of clarithromycin was higher for M. avium (32 µg ml-1) than M. intracellulare (8 µg ml-1). The MIC50 of rifabutin was more than four times higher for M. intracellulare (1 µg ml-1) than M. avium (≤0.25 µg ml-1). The percentages of patients aged >60 years and patients with sputum, cough, and cavitary lesions were significantly higher than among patients with M. intracellulare infection than M. avium infections.Conclusions. The pulmonary disease caused by distinct MAC species had different antimicrobial susceptibility, symptoms, and radiographic findings.

Chest wall pseudotumor: a case of non-tuberculous mycobacterial infection.

BACKGROUND: Non-tuberculous mycobacterial (NTM) infections are increasing worldwide, making them an international public health problem. Surgical management is often indicated for localized infectious disease; however, most surgeons are unaware of the potential risks of transmission during surgery. CASE PRESENTATION: An 88-year-old Asian female was referred to our hospital for a tumor in the right lateral thoracic region. One month prior, she had a feeling of fullness and complained of localized pain and warmth in the right lateral thoracic wall. Pain and warmth gradually resolved without intervention; however, the fullness was getting worse. Computed tomography (CT) scan showed a mass of approximately 65 × 30 mm with an osteolytic change, involving the right 8th rib. Based on the rapid growth rate and CT findings, we strongly suspected a malignant chest wall tumor, and en bloc tumor resection with the 8th rib was performed. When the specimen was cut, a large amount of viscous pus was drained and its culture showed growth of Mycobacterium avium. Microscopically, the non-caseating epithelioid cell granuloma extended into the rib, infiltrating the bone cortex. On follow-up 1 month after discharge, there were no signs of infection or other adverse events associated with the surgery. CONCLUSIONS: Herein, we report about a patient with a mass diagnosed as an NTM abscess involving the rib cage, which was confused with a malignant tumor and eventually diagnosed following surgical excision. This report emphasizes the need to be aware of the possibility of NTM infection and take appropriate precautions if the patient has a rapidly growing mass in the chest wall.

Different clinical features of patients with pulmonary disease caused by various Mycobacterium avium-intracellulare complex subspecies and antimicrobial susceptibility.

OBJECTIVES: Characteristics of the Mycobacterium avium-intracellulare complex pulmonary disease (MAC-PD) caused by distinct subspecies remain uncertain. METHODS: This study was conducted from 2013-2015 in three hospitals in Taiwan. RESULTS: Among the 144 patients with MAC-PD, 57 (39.6%), 37 (25.7%), 37 (25.7%), and 13 (9.0%) were infected with Mycobacterium intracellulare subspecies intracellulare (MIsI), Mycobacterium avium subspecies hominissuis (MAsH), Mycobacterium intracellulare subspecies chimaera (MIsC), and others, respectively. Patients with MAsH-PD were younger (p = 0.010) with higher human immunodeficiency virus infection rates (27.0%, 0.0%, 0.0%, and 7.7% for MAsH-PD, MIsC-PD, MIsI-PD, and others, respectively; p < 0.001). Twenty-two (15.3%) patients reported spontaneous culture-negative conversion, but 15 (10.4%) and 33 (22.9%) patients developed radiographic progression and unfavorable outcomes, especially MAsH-PD. The susceptibility rates to clarithromycin and inhaled amikacin were both 98.6%. MAsH demonstrated the lowest rate of resistance to moxifloxacin (66.7%, 97.3%, 89.1%, and 92.3% for MAsH-PD, MIsC-PD, MIsI-PD, and others, respectively; p = 0.001) and MIsI isolates had the highest rate of resistance to intravenous amikacin (25%, 13.5%, 38.2%, and 15.4% for MAsH-PD, MIsC-PD, MIsI-PD, and others, respectively; p = 0.024). CONCLUSIONS: Pulmonary disease caused by distinct MAC subspecies had different outcomes and drug susceptibility. The local prevalence of species needs to be monitored.

Nontuberculous mycobacteria in patients with suspected tuberculosis and the genetic diversity of Mycobacterium avium in the extreme south of Brazil.

OBJECTIVE: Nontuberculous mycobacteria (NTM) are a heterogeneous group of bacteria that are widely distributed in nature and associated with opportunistic infections in humans. The aims of this study were to identify NTM in patients with suspected tuberculosis who presented positive cultures and to evaluate the genetic diversity of strains identified as Mycobacterium avium. METHODS: We studied pulmonary and extrapulmonary samples obtained from 1,248 patients. The samples that tested positive on culture and negative for the M. tuberculosis complex by molecular identification techniques were evaluated by detection of the hsp65 and rpoB genes and sequencing of conserved fragments of these genes. All strains identified as M. avium were genotyped using the eight-locus mycobacterial interspersed repetitive unit-variable-number tandem-repeat method. RESULTS: We found that NTM accounted for 25 (7.5%) of the 332 mycobacteria isolated. Of those 25, 18 (72%) were M. avium, 5 (20%) were M. abscessus, 1 (4%) was M. gastri, and 1 (4%) was M. kansasii. The 18 M. avium strains showed high diversity, only two strains being genetically related. CONCLUSIONS: These results highlight the need to consider the investigation of NTM in patients with suspected active tuberculosis who present with positive cultures, as well as to evaluate the genetic diversity of M. avium strains.

Prevalence of Opportunistic Pathogens and Diversity of Microbial Communities in the Water System of a Pulmonary Hospital.

OBJECTIVE: Our objective was to investigate the occurrence of opportunistic pathogens and characterize the bacterial community structures in the water system of a pulmonary hospital. METHODS: The water samples were collected from automatic and manual faucets in the consulting room, treatment room, dressing room, respiratory ward, and other non-medical rooms in three buildings of the hospital. Quantitative polymerase chain reaction was used to quantify the load of several waterborne opportunistic pathogens and related microorganisms, including Legionella spp., Mycobacterium spp., and M. avium. Illumina sequencing targeting 16S rRNA genes was performed to profile bacterial communities. RESULTS: The occurrence rates of Legionella spp., Mycobacterium spp., and M. avium were 100%, 100%, and 76%, respectively in all samples. Higher occurrence rates of M. avium were observed in the outpatient service building (building 1, 91.7%) and respiration department and wards (building 2, 80%) than in the office building (building 3), where no M. avium was found. M. avium were more abundant in automatic faucets (average 2.21 × 10 4 gene copies/L) than in manual faucets (average 1.03 × 10 4 gene copies/mL) ( P < 0.01). Proteobacteria, Actinobacteria, Bacteroidetes, Cyanobacteria, Firmicutes, and Acidobacteria were the dominant bacterial phyla. Disinfectant residuals, nitrate, and temperature were found to be the key environmental factors driving microbial community structure shifts in water systems. CONCLUSION: This study revealed a high level of colonization of water faucets by opportunistic pathogens and provided insight into the characteristics of microbial communities in a hospital water system and approaches to reduce risks of microbial contamination.

Outbreak of avian mycobacteriosis in a commercial turkey breeder flock.

Avian mycobacteriosis (AM) is a chronic and contagious disease of pet birds, captive exotic, wild and domestic fowl, and mammals. Mycobacterium avium subsp. avium is the most common cause of AM in poultry. For the first time, we report a chronic outbreak of AM in an Iranian breeder flock of 250 45-week-old turkeys (Meleagris gallopavo) with a morbidity and mortality rate of 91.6% and 80%, respectively. A well-defined clinical feature of the outbreak included a progressive weight loss, decreased egg production, listlessness, and lameness. Tuberculous nodules were seen on liver, spleen, ovary, and ribs. Granulomatous inflammation and acid-fast bacilli were confirmed by using Ziehl-Neelsen method on hepatic lesions. M. avium subsp. avium was identified by polymerase chain reaction techniques based on the presence of 16S ribosomal RNA gene and insertion elements IS1245 and IS901. In this report, we not only describe the epidemiological, pathological, and molecular characteristics of the outbreak in detail, but we also discuss multiple factors influencing the introduction and development of AM critically. In this case, wild feral pigeons might have been the source of infection, but further molecular-epidemiology studies are needed to understand the role of wild birds in the persistence and transmission of Mycobacterium.RESEARCH HIGHLIGHTS First report of avian mycobacteriosis in an Iranian commercial turkey flock is described in detail.Risk factors intrinsic to the bird and mycobacteria, as well as extrinsic factors influencing the introduction and development of avian mycobacteriosis in birds, are critically discussed.

The impact of the coexistence of mycobacterium avium with mycobacterium tuberculosis on the result of GeneXpert and MGIT susceptibility test.

Rapid tests to diagnose tuberculosis relies on molecular detection of the M. tuberculosis. GeneXpert MTB/RIF test identifies M. tuberculosis and rifampicin resistance. We present a case of simultaneous coinfection with M. tuberculosis and M. avium. M. tuberculosis was detected in the sputum by PCR GeneXpert method. Unrecognized coexistence of M. tuberculosis and M. avium modified the results of drug susceptibility tests making the primary identification of M. tuberculosis as multi-drug resistant strain. We performed in vitro experiments to investigate the effect of the coexistence of M. avium with M. tuberculosis on the results of GeneXpert method, and drug susceptibility test.

Extrapulmonary Nontuberculous Mycobacteriosis in a Lung Transplant Recipient / Micobacteriosis no tuberculosa extrapulmonar en un receptor de transplante de pulmón

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Successful treatment of an AIDS patient with prolonged Mycobacterium avium bacteremia, high HIV RNA, HBV infection, Kaposi's sarcoma and cytomegalovirus retinitis.

We report an AIDS patient with a high HIV RNA copy number in the plasma who was successfully treated for prolonged Mycobacterium avium bacteremia and other complications. An HIV-infected patient with high fever, anemia, high alkaline phosphatase, cystic lung lesions, hepatitis B virus infection and Kaposi's sarcoma was referred to our hospital. PCR of the blood revealed Mycobacterium avium bacteremia and the time to blood culture positivity was 8 days. The HIV-1 RNA copy number in the plasma was more than ten million copies/ml and the CD4-positive T cell count was 21 cells/µL. Although the high fever resolved five days after therapy for Mycobacterium avium was started, the fever recurred just before starting anti-retroviral therapy (ART) including dolutegravir. The patient experienced repeated but self-limiting bouts of severe inflammation. Mycobacteremia was intermittently detected up to 79 days, suggesting that the recurrent episodes of inflammation were due to the intermittent dissemination of mycobacteria, and that persistent treatment is needed. Five months after the beginning of ART, the HIV-1 RNA copy number in the plasma was still 28,000 copies/ml. An HIV drug-resistance test revealed sensitivity to all anti-retroviral drugs. Eleven months after the initiation of ART, the HIV RNA copy number in the plasma decreased to 45 copies/mL and the CD4-positive T cell count recovered to 205 cells/µL. Our case also suggests that dolutegravir can be effective in cases with prolonged high levels of HIV RNA. Our findings emphasize that prompt diagnosis and persistent therapy for mycobacterial infection are important for successful treatment.

Acute lameness in a cat with disseminated mycobacteriosis.

Mycobacterium avium infection was diagnosed in an adult cat showing acute lameness of the right hind limb, enlargement of the right popliteal lymph node and two cutaneous nodular lesions of the right chest wall. Conventional radiography of the proximal tibia showed a proliferative osteolytic lesion. Cytological examination of the right popliteal lymph node and the nodular skin lesions fine needle aspiration smears, demonstrated granulomatous inflammation with many negative staining bacilli within macrophages or in smears background. The diagnosis was confirmed by Ziehl­Neelsen staining of the smears and the identification of mycobacteria was performed by microbiological and molecular methods. Histopathology performed after the necropsy revealed disseminated mycobacteriosis with granulomatous mesenteric lymphadenitis, granulomatous pneumonia, hepatitis and tibial osteomyelitis. M. avium is a well­known agent of gastro­enteric, respiratory or disseminated disease in immunocompromised cats but there are few cases reported in literature of bone involvement in systemic mycobacteriosis.

Nontuberculous mycobacteria in patients with suspected tuberculosis and the genetic diversity of Mycobacterium avium in the extreme south of Brazil / Micobactérias não tuberculosas em pacientes com suspeita de tuberculose e a diversidade genética de Mycobacterium avium no extremo sul do Brasil

ABSTRACT Objective: Nontuberculous mycobacteria (NTM) are a heterogeneous group of bacteria that are widely distributed in nature and associated with opportunistic infections in humans. The aims of this study were to identify NTM in patients with suspected tuberculosis who presented positive cultures and to evaluate the genetic diversity of strains identified as Mycobacterium avium. Methods: We studied pulmonary and extrapulmonary samples obtained from 1,248 patients. The samples that tested positive on culture and negative for the M. tuberculosis complex by molecular identification techniques were evaluated by detection of the hsp65 and rpoB genes and sequencing of conserved fragments of these genes. All strains identified as M. avium were genotyped using the eight-locus mycobacterial interspersed repetitive unit-variable-number tandem-repeat method. Results: We found that NTM accounted for 25 (7.5%) of the 332 mycobacteria isolated. Of those 25, 18 (72%) were M. avium, 5 (20%) were M. abscessus, 1 (4%) was M. gastri, and 1 (4%) was M. kansasii. The 18 M. avium strains showed high diversity, only two strains being genetically related. Conclusions: These results highlight the need to consider the investigation of NTM in patients with suspected active tuberculosis who present with positive cultures, as well as to evaluate the genetic diversity of M. avium strains.

RESUMO Objetivo: As micobactérias não tuberculosas (MNT) são um grupo heterogêneo de bactérias amplamente distribuídas na natureza e relacionadas com infecções oportunistas em seres humanos. Os objetivos deste estudo foram identificar MNT em pacientes com suspeita de tuberculose e culturas positivas e avaliar a diversidade genética de cepas identificadas como Mycobacterium avium. Métodos: Foram estudadas amostras pulmonares e extrapulmonares provenientes de 1.248 pacientes. As amostras que apresentaram resultado positivo em cultura e negativo para o complexo M. tuberculosis na identificação molecular foram avaliadas por meio da detecção dos genes hsp65 e rpoB e de sequenciamento de fragmentos conservados desses genes. Todas as cepas identificadas como M. avium foram genotipadas pelo método mycobacterial interspersed repetitive unit-variable-number tandem-repeat com oito loci. Resultados: Das 332 micobactérias isoladas, 25 (7,5%) eram MNT. Dessas 25, 18 (72%) eram M. avium, 5 (20%) eram M. abscessus, 1 (4%) era M. gastri e 1 (4%) era M. kansasii. As 18 cepas de M. avium apresentaram alta diversidade, e apenas duas eram geneticamente relacionadas. Conclusões: Esses resultados mostram a necessidade de considerar a investigação de MNT em pacientes com suspeita de tuberculose ativa e culturas positivas e de avaliar a diversidade genética de cepas de M. avium.

Clinical findings and treatment of disseminated 'Mycobacterium avium subspecies hominissuis' infection in a domestic cat.

A cat was referred because of diffuse parenchymal lung disease. Close examinations revealed a swollen abdominal lymph node and multiple nodules of the liver. Mycobacterium avium subspecies hominissuis infection was confirmed by culture and single nucleotide polymorphism analysis of samples recovered from the liver and bronchoalveolar lavage. After administration of combination antibiotics for 6 months, culture results were negative. Though atonic seizures were observed during the treatment, it disappeared after isoniazid discontinuation and pyridoxal phosphate administration. On day 771 of illness, no clinical signs, lung diseases, or obvious swelling of lymph nodes was observed. This is the first report to confirm Mycobacterium avium subspecies hominissuis infection in cats through gene analysis and to completely cure it with combination antibiotics.

Moxifloxacin resistance and genotyping of Mycobacterium avium and Mycobacterium intracellulare isolates in Japan.

BACKGROUND: Although fluoroquinolones are considered as alternative therapies of pulmonary Mycobacterium avium complex (MAC) disease, the association between fluoroquinolone resistance and MAC genotypes in clinical isolates from individuals not previously treated for MAC infection is not fully clear. METHODS: Totals of 154 M. avium isolates and 35 Mycobacterium intracellulare isolates were obtained from treatment-naïve patients with pulmonary MAC disease at the diagnosis of MAC infection at 8 hospitals in Japan. Their susceptibilities of moxifloxacin were determined by broth microdilution methods. Moxifloxacin-resistant isolates were examined for mutations of gyrA and gyrB. Variable numbers of tandem repeats (VNTR) assay was performed using 15 M. avium VNTR loci and 16 M. intracellulare VNTR loci. RESULTS: Moxifloxacin susceptibility was categorized as resistant and intermediate for 6.5% and 16.9%, respectively, of M. avium isolates and 8.6% and 17.1% of M. intracellulare isolates. Although the isolates of both species had amino acid substitutions of Thr 96 and Thr 522 at the sites corresponding to Ser 95 in the M. tuberculosis GyrA and Gly 520 in the M. tuberculosis GyrB, respectively, these substitutions were observed irrespective of susceptibility and did not confer resistance. The VNTR assays showed revealed three clusters among M. avium isolates and two clusters among M. intracellulare isolates. No significant differences in moxifloxacin resistance were observed among these clusters. CONCLUSIONS: Although resistance or intermediate resistance to moxifloxacin was observed in approximately one-fourth of M. avium and M. intracellulare isolates, this resistance was not associated with mutations in gyrA and gyrB or with VNTR genotypes.

Unusual presentation of canine Mycobacterium avium infection.

This short communication describes the clinical and morphological findings, diagnosis and treatment of a case of Mycobacterium avium infection in a golden retriever that presented with a progressive nasal swelling and lymphadenopathy. Although well documented in cats, where cutaneous lesions are frequently recognised, canine M avium infection is less commonly reported, and cutaneous lesions are rare. To the authors' knowledge this is the first documented case of canine M avium infection that presented with a cutaneous lesion but no systemic clinical signs. It occurred in a dog with no previously reported breed predisposition and highlights that in cases of cutaneous histiocytic infiltrate in dogs mycobacterial infection should remain a differential diagnosis, even in the absence of suggestive organisms on histopathological examination.

Genotyping of swine Mycobacterium avium subsp. hominissuis isolates from Kyushu, Japan.

The incidence of diseases caused by nontuberculous mycobacteria (NTM) is increasing annually worldwide, including Japan. Mycobacterium avium subsp. hoiminissuis (MAH) is one of the most common NTM species responsible for chronic lung diseases in animals and humans. In the current study, mycobacterial interspersed repetitive unit-variable number tandem repeat (MIRU-VNTR) typing was employed to characterize the genetic diversity of swine MAH isolates from Kyushu, Japan. In total, 309 isolates were obtained from the lymph nodes of 107 pigs not displaying any clinical signs of disease, of which 307 were identified as MAH, comprising 173 strains. Based on eight established MIRU-VNTR loci, the MAH strains represented 50 genotypes constituting three lineages, and 29 had not been described in the Mac French National Institute for Agricultural Research Nouzilly MIRU-VNTR (Mac-INMV) database. MAH was the dominant M. avium complex (MAC) in pigs from Kyushu, and there was high genetic diversity among genotype profiles of MAH from Kyushu. We identified three predominant genotype profiles in the tested area sharing high relatedness with genotype profiles of strains isolated in European countries. MAH was the most common NTM in pigs from Kyushu and exhibited high diversity, with new strain-derived genotypes.