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The genus Mycobacterium comprises more than 150 species, including important pathogens for humans which cause major public health problems. The vast majority of efforts to understand the genus have been addressed in studies with Mycobacterium tuberculosis. The biological differentiation between M. tuberculosis and nontuberculous mycobacteria (NTM) is important because there are distinctions in the sources of infection, treatments, and the course of disease. Likewise, the importance of studying NTM is not only due to its clinical significance but also due to the mechanisms by which some species are pathogenic while others are not. Mycobacterium avium complex (MAC) is the most important group of NTM opportunistic pathogens, since it is the second largest medical complex in the genus after the M. tuberculosis complex. Here, we evaluated the virulence and immune response of M. avium subsp. avium and Mycobacterium colombiense, using experimental models of progressive pulmonary tuberculosis and subcutaneous infection in BALB/c mice. Mice infected intratracheally with a high dose of MAC strains showed high expression of tumor necrosis factor alpha (TNF-α) and inducible nitric oxide synthase with rapid bacillus elimination and numerous granulomas, but without lung consolidation during late infection in coexistence with high expression of anti-inflammatory cytokines. In contrast, subcutaneous infection showed high production of the proinflammatory cytokines TNF-α and gamma interferon with relatively low production of anti-inflammatory cytokines such as interleukin-10 (IL-10) or IL-4, which efficiently eliminate the bacilli but maintain extensive inflammation and fibrosis. Thus, MAC infection evokes different immune and inflammatory responses depending on the MAC species and affected tissue.

Mycobacterium avium complex (MAC) infection causes disseminated disease in immunocompromised hosts, such as human immunodeficiency virus (HIV)-positive patients, and pulmonary disease in persons without systemic immunosuppression, which has been increasing in many countries. In Japan, the incidence of pulmonary MAC disease caused by M. avium is about 7 times higher than that caused by M. intracellulare. To explore the bacterial factors that affect the pathological state of MAC disease caused by M. avium, we determined the complete genome sequence of the previously unreported M. avium subsp. hominissuis strain TH135 isolated from a HIV-negative patient with pulmonary MAC disease and compared it with the known genomic sequence of M. avium strain 104 derived from an acquired immunodeficiency syndrome patient with MAC disease. The genome of strain TH135 consists of a 4,951,217-bp circular chromosome with 4,636 coding sequences. Comparative analysis revealed that 4,012 genes are shared between the two strains, and strains TH135 and 104 have 624 and 1,108 unique genes, respectively. Many strain-specific regions including virulence-associated genes were found in genomes of both strains, and except for some regions, the G+C content in the specific regions was low compared with the mean G+C content of the corresponding chromosome. Screening of clinical isolates for genes located in the strain-specific regions revealed that the detection rates of strain TH135-specific genes were relatively high in specimens isolated from pulmonary MAC disease patients, while, those of strain 104-specific genes were relatively high in those from HIV-positive patients. Collectively, M. avium strains that cause pulmonary and disseminated disease possess genetically distinct features, and it suggests that the acquisition of specific genes during strain evolution has played an important role in the pathological manifestations of MAC disease.

Neutrophils are the primary cells contributing to initial defense against mycobacteria. Yet, little is known about the potential of various mycobacterial strains to stimulate neutrophils. This study was focused to compare the differential capacity of vaccine strains, Mycobacterium bovis bacillus Calmette-Guerin (BCG) and Mycobacterium indicus pranii (Mw), and laboratory strain H37Rv to activate and enhance neutrophil functions. The expression of phenotypic markers like Fcγ receptor, toll-like receptor (TLR), and chemokine receptor; secretion of pro-inflammatory cytokines; and the rate of apoptosis were studied in infected neutrophils. Increased expression of CD32, CD64, TLR4, and CXCR3; increased TNF-α secretion; and downregulation of early apoptosis were observed in H37Rv-infected neutrophils. Among the vaccine strains, BCG increased the expression of only CD32 on neutrophils, while Mw was comparatively ineffective. To understand the paracrine role of neutrophils, the supernatants from infected neutrophils were used to stimulate monocytes and T helper cells. The secretory molecules from all infected neutrophils increased the expression of CCR5 on monocytes, whereas only H37Rv-infected supernatant increased the expression of CCR7 on monocytes and CD69 on T cells. Thus, H37Rv was more effective in activating neutrophils and in turn stimulating monocytes and T cells. By comparison, vaccine strains were less effective in modulating neutrophil functions., (© 2012 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.)

We report herein the draft genome sequence of Mycobacterium intracellulare clinical strain M.i.198, which consistently exhibits hypervirulence in human patients, human macrophages in vitro, and immunocompetent mice.

Introduction: Patients with lung cancer are sometimes found to have respiratory cultures growing Mycobacterium avium complex (MAC). This study describes the clinical, pathologic, and radiographic -characteristics of individuals who harbor concomitant lung cancer and MAC., Methods: Retrospective analysis of patients with positive respiratory cultures for MAC (370 men, 475 women) and with newly diagnosed lung cancer (792 men, 840 women) from 1995 to 2010., Results: Of the patients with respiratory cultures growing MAC, 8.6% of men and 6.3% of women had lung cancer. Twenty-five percent of patients with lung cancer and 3% with nonbronchiectatic benign lung disease grew MAC from their respiratory cultures. Significantly fewer women with both MAC and lung cancer were smokers than the control group of women with lung cancer and negative MAC cultures (68% versus 89%, p < 0.01). Squamous cell carcinoma occurred in 40% of women in the MAC/lung cancer group versus 28% of women in the lung cancer control group. Peripherally located squamous cell carcinomas were found in 71% of the MAC/lung cancer group versus 40% of the lung cancer control group (p = 0.01), Conclusions: The percentage of smokers among women with both MAC and lung cancer was lower than among the lung cancer control group who did not grow MAC. The presence of MAC in respiratory cultures of lung cancer patients was particularly associated with squamous cell carcinomas located in the periphery of the lung. Because MAC typically affects distal airways, this possible association between MAC infection and lung cancer warrants further study.

STAT1 is a key component of Interferon (IFN)-γ and IFN-α signaling and mediates protection against mycobacteria, fungal, viral infections, and cancer. Dominant negative inhibitory as well as gain of function heterozygous STAT1 mutations demonstrate that IFN-γ driven cellular responses need to be tightly regulated to control infections. We describe an autosomal dominant mutation in the SH2 domain of STAT1 that disrupts protein phosphorylation, c.1961T>A (M654K). The mutant allele does not permit STAT1 phosphorylation, and impairs STAT1 phosphorylation of the wild type allele. Protein dimerization is preserved but DNA binding activity, IFN-γ driven GAS-luciferase activity, and expression of IFN-γ target genes are reduced. IFN-α driven ISRE response, but not IFN-α driven GAS response, are preserved when cells are co-transfected with wild type and the mutant STAT1 constructs. M654K exerts a dominant negative effect on IFN-γ related immunity and is recessive for IFN-α induced immune function.

Pathogenesis by mycobacteria requires the exploitation of host-cell signaling pathways to enhance intracellular survival and persistence of the pathogen. Among patients with end-stage acquired immune-deficiency syndrome, disseminated infection with Mycobacterium avium, a member of the M. avium complex (MAC), is the most common bacterial infection. The virulence and intrinsic multidrug resistance of this pathogen has been attributed in part to its unique cell wall, which is a complex array of hydrocarbon chains containing the arabinogalactan-peptidoglycan mycolic acid core found in all mycobacteria, surrounded by a second electron-dense layer made up, in part, of serovar-specific glycopeptidolipids (GPLs) found only in MAC. Via cell-surface receptors, M. avium, an intra-macrophage (mφ) pathogen, can modulate various host signaling pathways such as the mitogen-activated protein kinase and nuclear factor κB pathways. The modulation of specific mφ signaling cascades can result in the regulation of pro- and anti-inflammatory cytokine production, and the process of phagolysosome fusion. The outcome of this M. avium-host mφ interaction could result in host disease or death of the invading pathogen. This review will focus on the immunomodulation aspects of M. avium pathogenesis as well as the role of GPLs as virulence factors.

Mycobacterium avium (Mav) lung infections, called nodular-bronchiectasis (NB)-type M. avium complex (MAC) disease, are globally increasing. To elucidate whether there are unusual populations of Mav, causing NB-type disease rather than cavitary (CA)-type disease, we compared the virulence of Mav isolates from patients with NB-type (NB-Mav) and those from CA-type (CA-Mav) diseases, based on intracellular growth in various types of human cells. Five strains each of NB-Mav and CA-Mav were compared with each other for their invasiveness and ability to intracellularly replicate in various types of cultured cells of human origin. The two types of Mav isolates showed a similar ability, on average, to replicate in macrophages and lung epithelial cells. Moreover, they showed a similar ability to induce the production of reactive nitrogen intermediates and reactive oxygen intermediates by macrophages and susceptibility to antimicrobial molecules. Therefore, it appears that there is no essential difference in virulence in terms of infectivity to human macrophages and lung cells between Mav strains isolated from NB-MAC disease and those from CA-MAC disease. These findings indicate the importance of further studies to elucidate the mechanism for the establishment of NB-type MAC diseases based on host immunological conditions rather than the pathogenic nature of MAC organisms themselves.

Purpose of Review: Although the incidence of tuberculosis has reduced in developed countries, there is a growing interest in nontuberculous mycobacteria (NTM) as a cause of lung disease. However, NTM are a heterogeneous group and most of the data come from only three species: Mycobacterium avium complex, Mycobacterium kansasii and Mycobacterium abscessus. Still, information about these three species is confusing because it is based mainly on retrospective studies and series of clinical cases performed in developed countries. In recent years, new information has appeared about other species and the pathogenesis of NTM., Recent Findings: Epidemiological studies show that NTM infection is a worldwide phenomenon with an increasing presence in developing countries perhaps because of the implementation of tap water. Women with characteristic phenotype are at higher risk of acquiring NTM infection along with patients with defects on cystic fibrosis transmembrane conductance regulators. New studies on Mycobacterium fortuitum, Mycobacterium xenopi, Mycobacterium szulgai and Mycobacterium simiae indicate that the American Thoracic Society criteria for diagnosing NTM disease may not be useful for all species of NTM., Summary: New multicentric and prospective studies are needed to clarify the pathogenesis and treatment of NTM. These organisms form a numerous and heterogeneous group and each species should be studied separately.

A 73-year-old woman was admitted to our hospital because of persistent fever, headache and fatigue for several weeks. On admission, she was diagnosed as having meningitis due to Mycobacterium intracellulare (M. intracellulare) detected in her cerebrospinal fluid (CSF) by polymerase chain reaction. Even though anti-tuberculous therapy improved her CSF findings, her condition was not restored. Brain MRI showed multifocal and asymmetrical increases in T2 signals involving white matter and cortical gray-white junction of cerebral hemispheres, cerebellum and brainstem. Based on the progression of clinical symptoms and radiological features, we diagnosed her illness as acute disseminated encephalomyelitis (ADEM) associated with meningitis due to M. intracellulare. Steroid therapy dramatically improved her condition. This is the first report of ADEM following meningitis due to M. intracellulare in a non-immunocompromized host.

Macrophages (M(phi)s) play a central role as anti-microbial effector cells in the expression of host resistance to mycobacterial infections. With respect to antimicrobial effector molecules of host M(phi) against mycobacterial pathogens, recent studies suggest the possibility that the reactive nitrogen intermediates (RNI)--and reactive oxygen intermediates-independent antimycobacterial mechanism(s) may be crucial for the antimycobacterial function of host M(phi). In this context, we previously found that free fatty acids (FFAs) such as arachidonic acid (AA) and linolenic acid exhibited potent antimicrobial activity against mycobacterial organisms, including Mycobacterium tuberculosis (MTB) and Mycobacterium avium complex (MAC). In addition, FFAs in combination with RNI played critical roles in manifestation of the activity of M(phi) against mycobacterial organisms. Moreover, our recent studies have shown the following findings. First, anti-MTB activity of IFN-gamma-activated M(phi)s was specifically blocked by arachidonyl trifluoromethylketone (aTFMK), an inhibitor of cytosolic phospholipase A2 (cPLA2). Second, ATP potentiated the anti-MAC bactericidal activity of M(phi)s cultivated in the presence of clarithromycin and rifamycin. This effect of ATP was closely related to intracellular Ca2+ mobilization and was specifically blocked by aTFMK. Third, intramacrophage translocation of membranous AA molecules to MAC-containing phagosomes was also specifically blocked by aTFMK. In the confocal microscopic observation of MAC-infected M(phi)s, ATP enhanced the intracellular translocation of cPLA2 into MAC-containing phagosomes. These findings suggest that FFAs (especially AA) produced by the enzymatic action of cPLA2 play important roles as antimycobacterial effectors in the expression of M(phi) antimicrobial activity against mycobacterial pathogens.

Little is known about the clinical relevance of nontuberculous mycobacteria (NTM) in the Arabian Peninsula. We assessed the prevalence and studied a random sample of isolates at a reference laboratory in Muscat, Oman. NTM cause disease in this region, and their prevalence has increased.

The Mycobacterium avium complex (MAC) is the most frequently isolated species among non-tuberculous mycobacteria (NTM) clinical isolates. Physicians pay attention to the differential diagnosis of the disease caused by MAC from tuberculosis because of their similar clinical presentations. Expression of the macrophage-induced gene (mig) is one of the virulence phenotypes in MAC, but it has not been determined whether the presence of the mig gene itself has any relationship with clinical disease or whether it is merely a marker for MAC. To uncover the significance of the mig gene among MAC clinical isolates, positive cultures from respiratory specimens from patients in a tertiary referral centre were identified by sequencing the 16S rRNA gene. The mig gene was also evaluated using PCR and sequence analysis. The medical records from the patients were reviewed retrospectively. The diagnostic criteria from the American Thoracic Association were adopted for the diagnosis of NTM lung disease. A total of 45 MAC clinical isolates were identified over a period of 1 year. Following 16S rRNA sequencing, all of the 23 M. avium isolates were categorized as sequevar I. Among the 22 Mycobacterium intracellulare isolates, 18 strains were identified as M. intracellulare sequevar I and the remaining four consisted of one each of sequevars II, III, IV and V. The proportion of cases that fitted the diagnostic criteria of NTM lung disease was 26.7 % (12/45). The mig PCR results were 100 % positive for the MAC isolates studied, irrespective of their species, sequevar or disease-causing properties. However, following bootstrap analysis of the mig sequences, we observed definite grouping between M. avium and M. intracellulare. Thus the mig gene is a species-specific marker with distinct sequence diversity between the two species M. avium and M. intracellulare, but there is poor correlation between disease-causing properties and specific mig sequences.

Mycobacterium avium complex (MAC) disease has been increasing worldwide not only in immunocompromised but also in immunocompetent humans. However, the relationship between mycobacterial strain virulence and disease progression in immunocompetent humans is unclear. In this study, we isolated 6 strains from patients with pulmonary MAC disease. To explore the virulence, we examined the growth in human THP-1 macrophages and pathogenicity in C57BL/6 mice. We found that one strain, designated 198, which was isolated from a patient showing the most progressive disease, persisted in THP-1 cells. In addition, strain 198 grew to a high bacterial load with strong inflammation in mouse lungs and spleens 16 weeks after infection. To our knowledge, strain 198 is the first isolated MAC strain that exhibits hypervirulence consistently for the human patient, human macrophages in vitro, and even for immunocompetent mice. Other strains showed limited survival and weak virulence both in macrophages and in mice, uncorrelated to disease progression in human patients. We demonstrated that there is a hypervirulent clinical MAC strain whose experimental virulence corresponds to the serious disease progression in the patients. The existence of such strain suggests the involvement of bacterial virulence in the pathogenesis of pulmonary MAC disease in immunocompetent status.

Objective: We present a case series and review of the literature of the management options in non-HIV-infected patients with Mycobacterium avium complex pulmonary disease (MAC-PD) with a focus on treatment failure and drug resistant disease., Case Series: Five case histories are presented, depicting various clinical scenarios necessitating different approaches to therapy and highlighting the limitations and complications of these options., Discussion: Mycobacterium avium complex (MAC) is well recognized as a significant cause of pulmonary disease in non-HIV infected patients and in those with intact immunity. Isolation of non-tuberculous mycobacteria (NTM) in culture is essential for the diagnosis of NTM lung disease. The typical presentation of MAC lung disease is apical fibrocavitary lung disease in men in their late 40s and early 50s who have a history of cigarette smoking and, frequently, excessive alcohol use. Other presentations of NTM lung disease include nodular bronchiectasis, solitary or multiple pulmonary nodules, and hypersensitivity pneumonitis. When indicated, the standard recommended treatment for most patients is a three-times-weekly regimen of clarithromycin or azithromycin, rifampin, and ethambutol with or without amikacin. Daily therapy is recommended for fibrocavitary disease. Based on published studies, macrolides are the only agents used for treatment of MAC disease for which there is a correlation between in vitro susceptibility and in vivo (clinical) response. Data regarding treatment of macrolide-resistant MAC (MRMAC) and multi-drug resistant MAC (MDRMAC) is sparse. Several drugs have been evaluated in drug-resistant MAC and have potential as effective therapy. Use of multiple drugs to which the isolate is susceptible is preferred to avoid development of future resistance. Surgery in mycobacterial disease is technically difficult, but selected patients with focal disease do benefit from resection of the involved lung., Conclusions: MAC has protean pulmonary manifestations, especially in those with no recognizable impairments in their immune system. Drug treatment, however, remains difficult with high failure rates and poor long-term sputum conversion. This case series is based on our clinical experience highlighting treatment options and the often unrecognized morbidity and mortality of severe, progressive MAC-PD. It underscores the need for increased awareness of MAC-PD and MDRMAC and the difficulties encountered in their management.

Organisms within the Mycobacterium avium complex (MAC) may have differential virulence. We compared 33 subjects with MAC pulmonary disease to 75 subjects with a single positive culture without disease. M. avium isolates were significantly more likely to be associated with MAC pulmonary disease (odds ratio = 5.14, 95% confidence interval = 1.25 to 22.73) than M. intracellulare.

We present the case of a 55-year-old-man with AIDS who had disseminated Mycobacterium avium-intracellulare (MAI) infection who was nonadherent to antiretroviral treatment and prophylaxis for opportunistic infections. The patient acquired HIV via intravenous drug sue; he had a CD4+ cell count of 4/microL (1%) and an HIV RNA level of 114,000 copies/mL at the time of his HIV diagnosis. He presented with MAI bacteremia and had rapidly evolving and resolving cutaneous lesions caused by MAI. Concomitant nonspecific brain lesions presented a diagnostic challenge, but they improved with MAI treatment. In persons with HIV/AIDS who have cutaneous lesions, biopsy of superficial lesions has proved helpful. MAI infection should be considered in the differential diagnosis of cutaneous and brain lesions in persons with HIV/AIDS.

We previously demonstrated that mice of the I/St strain are extremely susceptible to Mycobacterium tuberculosis, as well as to the taxonomically distant intracellular bacteria Chlamydia pneumoniae and Salmonella enterica. To broaden our knowledge about the control of susceptibility to intracellular pathogens, we studied the infection caused by Mycobacterium avium virulent strain 724 in a panel of inbred mouse strains and found that I/St mice are resistant to M. avium. By comparing I/St mice with B6 mice, we demonstrated that (i) B6 mice are much more susceptible to infection caused by M. avium in terms of bacterial multiplication in the lung tissue and severity of lung pathology; (ii) in B6 mice but not in I/St mice infection leads to prolonged leukocyte infiltration of the lung tissue, development of necrotic lung granulomata, and lethality; and (iii) the unfavorable infectious course in B6 mice is accompanied by elevated production of gamma interferon, tumor necrosis factor alpha, and especially interleukin-12 in the lungs. Importantly, M. avium-resistant I/St mice carry a functional r allele of the Slc11a1 (formerly Nramp1) gene, while B6 mice have the Slc11a1(s) genotype. Segregation genetic analysis of (I/St x B6) F2 hybrids demonstrated that susceptibility or resistance to infection caused by M. avium largely depended upon the Slc11a1 genotype and that other genetic traits had a relatively weak influence. This close-to-monogenic pattern differs sharply from the host control of many other intracellular bacterial infections, for which the involvement of numerous quantitative trait loci has been ubiquitously observed.

We report adjunct treatment of (interferon) IFN-alpha2b (Intron-A) in a patient with complete interferon-gamma receptor R1 (IFNGR1) deficiency suffering from disseminated infection with Mycobacterium avium complex (MAC) resistant to multiple anti-mycobacterial agents. A low dose of IFN-alpha2b (3 x 10(6) units/m(2) three times weekly subcutaneously) successfully attenuated progressive hepatosplenomegaly and abdominal/retroperitoneal/pelvic lymphadenopathy, although the patient continued to be mycobacteremic. This is the first report of a complete IFNGR1 deficiency treated with adjuvant IFN-alpha2b for disseminated MAC infection.

Background: Knowledge of the relationship between respiratory disorders and gastroesophageal reflux disease (GERD) is increasing. However, the association between GERD and pulmonary disease caused by nontuberculous mycobacteria (NTM) has not been studied in detail. We investigated the prevalence of GERD in patients with the nodular bronchiectatic form of NTM lung disease., Methods: Fifty-eight patients with the nodular bronchiectatic form of NTM lung disease underwent ambulatory 24-h esophageal pH monitoring. Of the 58 patients, 27 patients were identified as having Mycobacterium avium complex infection (15 with Mycobacterium intracellulare and 12 with M avium), and 31 patients had Mycobacterium abscessus pulmonary infection., Results: The prevalence of GERD in patients with the nodular bronchiectatic form of NTM lung disease was 26% (15 of 58 patients). Only 27% (4 of 15 patients) had typical GERD symptoms. No statistically significant differences were found between patients with GERD and those without GERD with regard to age, sex, body mass index, or pulmonary function test results. However, patients with GERD were more likely to have a sputum smear that was positive for acid-fast bacilli (12 of 15 patients, 80%), compared with patients without GERD (19 of 43 patients, 44%) [p = 0.033]. In addition, bronchiectasis and bronchiolitis were observed in more lobes in patients with GERD than in patients without GERD (p = 0.008 and p = 0.005, respectively)., Conclusions: Patients with the nodular bronchiectatic form of NTM lung disease have a high prevalence of increased esophageal acid exposure, usually without typical GERD symptoms.

A rapidly progressive, fatal recrudescence of pulmonary Kaposi sarcoma developed in an HIV-infected man who was receiving corticosteroids for treatment of an immune reconstitution syndrome secondary to Mycobacterium avium complex pulmonary infection. We discuss the implications for current diagnosis and management of HIV-associated pulmonary diseases.

The incidence of atypical mycobacterial infection among solid organ transplant recipients is increasing. While lung transplant recipients in particular are at greater risk of atypical mycobacterial infection than other solid organ transplant recipients, it is typically confined to the lung and disseminated infection remains quite rare. We describe a case of disseminated Mycobacterium avium-complex (MAC) in a lung transplant recipient presenting as granulomatous liver disease with signs of portal hypertension. After identification of the infection and institution of proper therapy, the patient had significant improvement in both clinical signs of portal hypertension and liver function tests. Current literature suggests a favorable prognosis in most cases of MAC infection in lung transplant recipients with appropriate treatment. This case highlights the need to maintain an elevated index of suspicion for atypical pathogens with unusual clinical presentations among the lung transplant population.

Purpose of Review: Nontuberculous mycobacterial disease, especially pulmonary disease, is increasingly encountered by clinicians. Therapy of the most common nontuberculous mycobacterial pathogen, Mycobacterium avium complex, improved with the introduction of macrolide-containing regimens, but treatment for this and most other nontuberculous mycobacterial pathogens remains difficult., Recent Findings: Treatment trials with macrolide-containing regimens for Mycobacterium avium complex lung disease have yielded generally favorable outcomes. Studies consistently show that in-vitro susceptibility to macrolides remains the only in-vitro susceptibility for Mycobacterium avium complex that correlates with in-vivo response. Patients who have macrolide-resistant Mycobacterium avium complex isolates are much harder to treat and have higher mortality than patients with macrolide-susceptible isolates. Studies also consistently show that patients who fail therapy, even those who remain macrolide susceptible in vitro, are more difficult to treat than patients without previous therapy., Summary: There have been no significant treatment advances for Mycobacterium avium complex lung disease, and nontuberculous mycobacterial disease in general, since the advent of the newer macrolides. It has become clear that the best opportunity for treatment success is the first treatment effort. It is also clear that protection against the emergence of macrolide-resistant Mycobacterium avium complex isolates is critically important. For further progress in the treatment of these pathogens, new and more active drugs must be developed.

Picolinic acid (PA) potentiates macrophage (MPhi) antimicrobial activity against intracellular Mycobacterium avium complex (MAC). Here, we studied the mechanisms of this phenomenon using human THP-1 MPhis. First, when PA-treated MAC-infected MPhis were cultured in the presence or absence of reactive oxygen intermediate (ROI) scavengers, nitric oxide synthase (NOS) inhibitors or phospholipase A(2) (PLA(2)) inhibitors, none of these agents blocked the activity of PA in potentiating MPhi anti-MAC activity. Second, when PA was added to the in vitro anti-MAC bactericidal system consisting of either ROIs, reactive nitrogen intermediates (RNIs) or free fatty acid (FFA) molecules, which are the major MPhi antimicrobial effectors, PA inhibited the activity of ROIs and conversely potentiated the activity of RNIs; PA did not affect the activity of FFAs. Third, PA reduced mRNA expression of NADPH oxidase and beta-defensin-1 by MAC-infected MPhis, whilst neither cytosolic PLA(2) nor CAP37 mRNA expression was affected. Notably, inducible NOS and secretory PLA(2) mRNA expression was not detected for MAC-infected MPhis even when given PA treatment. These findings suggest that ROIs, RNIs, FFAs and beta-defensin-1 do not play important roles in the PA-induced potentiation of MPhi anti-MAC activity.

The diagnosis and treatment of mycobacterial diseases are very important clinical issues. Among mycobacterial diseases, pulmonary tuberculosis remains an important cause of morbidity and mortality throughout the world. Pulmonary tuberculosis demonstrates a variety of clinical and radiological features. In addition, the prevalence of Mycobacterium avium complex (MAC) has increased, especially in elderly women without underlying diseases. Clinically, there is a significant difference between tuberculosis and atypical mycobacterium infection in terms of the infection control measures adopted and the choice of treatment. Therefore, it is very important to know the characteristic radiological findings of mycobacterial diseases. In the present review, key radiological points for diagnosing mycobacterial diseases are discussed.

There is evidence that drinking water may be a source of infections with pathogenic nontuberculous mycobacteria (NTM) in humans. One method by which NTM are believed to enter drinking water distribution systems is by their intracellular colonization of protozoa. Our goal was to determine whether we could detect a reduction in the prevalence of NTM recovered from an unfiltered surface drinking water system after the addition of ozonation and filtration treatment and to characterize NTM isolates by using molecular methods. We sampled water from two initially unfiltered surface drinking water treatment plants over a 29-month period. One plant received the addition of filtration and ozonation after 6 months of sampling. Sample sites included those at treatment plant effluents, distributed water, and cold water taps (point-of-use [POU] sites) in public or commercial buildings located within each distribution system. NTM were recovered from 27% of the sites. POU sites yielded the majority of NTM, with >50% recovery despite the addition of ozonation and filtration. Closely related electrophoretic groups of Mycobacterium avium were found to persist at POU sites for up to 26 months. Water collected from POU cold water outlets was persistently colonized with NTM despite the addition of ozonation and filtration to a drinking water system. This suggests that cold water POU outlets need to be considered as a potential source of chronic human exposure to NTM.

We report a 65-year-old man with tenosynovitis of the wrist and finger caused by Mycobacterium intracellulare. The diagnosis was made after subsequent recurrence, when a specimen of synovial fluid was subjected to specific polymerase chain reaction and mycobacterial culture. Synovectomy and susceptibility-guided antituberculous therapy permitted complete healing of tenosynovitis.

We previously reported that glycopeptidolipid (GPL) isolated from Mycobacterium avium serovar 4 inhibited phagosome-lysosome (P-L) fusion when macrophages phagocytosed heat-killed Staphylococcus aureus (SA). In the present study we analyzed the underlying inhibitory mechanism of GPL coated on SA. Elimination of oligosaccharide from GPL abrogated its inhibitory activity. GPL did not inhibit P-L fusion of opsonized SA phagocytosed via complement receptors. The inhibitory activity of GPL was competitively reduced by the presence of alpha-methyl-D-mannoside and anti-mannose receptor antibody, suggesting that inhibition of P-L fusion by GPL is mediated through mannose receptor. Recruitment of early endosome antigen 1 and Ca2+/calmodulin kinase II in human macrophage-like THP-1 cells were significantly suppressed by GPL, indicating that GPL inhibits steps for leading to the P-L fusion.

The clinical significance and prevalence of Mycobacterium avium and Mycobacterium intracellulare were analyzed in a cohort of 7,472 patients who, from 1999 to 2003, sought care at the University of Texas M.D. Anderson Cancer Center, Houston, and had cultures performed for mycobacteria. Patients were stratified for age, sex, and underlying diseases, and bacteria were identified by 16S rRNA gene sequencing. M. avium was isolated in 62 (0.83%) of 7,472 patients and M. intracellulare in 65 (0.87%). Clinically, only 10 of the 62 (16.2%) patients with M. avium had probable to definite evidence of infection, whereas the majority (83.8%) had weak evidence of infection. Sex and age did not affect the isolation or infection of M. avium. Hematological tumors predisposed to M. avium colonization but not infection. In contrast, 41 of the 65 (63.1%) patients with M. intracellulare had probable to definite infection, a level much higher than those with M. avium (P < 0.001). M. intracellulare was more prevalent in women (1.33% of 3,311) than in men (0.50% of 4,161) (P < 0.001), and underlying diseases had no effect in women. Men with lung cancer had a higher prevalence (1.37%) than men without (0.34%) (4.0-fold; P < 0.001), but it was similar to that in women. A marked age trend for the isolation of M. intracellulare among women was noted: 0.27% (1-fold) for ages of <50 years, 0.85% (3.1-fold) for ages 50 to 59 years, 1.50% (5.6-fold) for ages 60 to 69 years, and 3.74% (13.9-fold) for ages >/=70 years (trend, P < 0.001). The combined rate for women >/=50 was 1.86% (95% confidence interval [1.30 to 2.42%]) (6.9-fold). Together, these results suggest that, among non-AIDS patients, M. intracellulare is more pathogenic and tends to infect women increasingly beyond menopause (age >/=50 years) regardless of underlying disease. The prevalence rate of 1.86% in postmenopausal women suggests the need to further investigate the public health significance of M. intracellulare.

Mycobacterium avium is a major opportunistic pathogen of AIDS patients in the United States. The understanding of M. avium pathogenesis has been hampered by the inability to create gene knockouts by homologous recombination, an important mechanism for defining and characterizing virulence factors. In this study a functional methyltransferase D (mtfD) gene was deleted by allelic replacement in the M. avium strain 104. Methyltransferase D is involved in the methylation of glycopeptidolipids (GPLs); highly antigenic glycolipids found in copious amounts on the M. avium cell surface. Interestingly, the loss of mtfD resulted in M. avium 104 containing only the non-serotype specific GPL. Results also suggest that the mtfD encodes for a 3-O-methyltransferase. The absence of significant amounts of any serotype-specific GPLs as a consequence of mtfD deletion indicates that the synthesis of the core 3,4-di-O-methyl rhamnose is a prerequisite for synthesis of the serotype-specific GPLs. Macrophages infected with the mtfD mutant show elevated production of tumour necrosis factor-alpha (TNF-alpha) and RANTES compared to control infections. In addition, the M. avium 104 mtfD mutant exhibits decreased ability to survive/proliferate in mouse liver and lung compared to wild-type 104, as assessed by bacterial counts. Importantly, the mtfD mutant complemented with a wild-type mtfD gene maintained an infection level similar to wild-type. These experiments demonstrate that the loss of mtfD results in a M. avium 104 strain, which preferentially activates macrophages in vitro and shows attenuated virulence in mice. Together our data support a role for GPLs in M. avium pathogenesis.

Pathogens that are transmitted between the environment, wildlife, livestock and humans represent major challenges for the protection of human and domestic animal health, the economic sustainability of agriculture, and the conservation of wildlife. Among such pathogens, the genus Mycobacterium is well represented by M. bovis, the etiological agent of bovine tuberculosis, M. avium ssp. paratuberculosis (Map) the etiological agent of Johne disease, M. avium ssp. avium (Maa) and in a few common cases by other emergent environmental mycobacteria. Epidemiologic surveys performed in Europe, North America and New Zealand have demonstrated the existence and importance of environmental and wildlife reservoirs of mycobacterial infections that limit the attempts of disease control programmes. The aim of this review is to examine the zoonotic aspects of mycobacteria transmitted from the environment and wildlife. This work is focused on the species of two main groups of mycobacteria classified as important pathogens for humans and animals: first, M. bovis, the causative agent of bovine tuberculosis, which belongs to the M. tuberculosis complex and has a broad host range including wildlife, captive wildlife, domestic livestock, non-human primates and humans; the second group examined, is the M. avium-intracellulare complex (MAC) which includes M. avium ssp. avium causing major health problems in AIDS patients and M. avium ssp. paratuberculosis the etiological agent of Johne disease in cattle and identified in patients with Crohn disease. MAC agents, in addition to a broad host range, are environmental mycobacteria found in numerous biotopes including the soil, water, aerosols, protozoa, deep litter and fresh tropical vegetation. This review examines the possible reservoirs of these pathogens in the environment and in wildlife, their role as sources of infection in humans and animals and their health impact on humans. The possibilities of control and management programmes for these mycobacterial infections are examined with regards to the importance of their natural reservoirs.

Disseminated Mycobacterium avium infection is common in AIDS patients that do not receive anti-AIDS therapy and in patients for whom therapy fails. M. avium is commonly acquired by ingestion, and a large number of AIDS patients have M. avium in their intestinal tracts. To better understand the dynamics of the infection in patients with AIDS, we studied orally infected mice. To determine if immunocompetent mice challenged orally with M. avium can develop protection against the infection, and if so, which cell population(s) is responsible for the protection, we exposed wild-type as well as CD4(-/-), CD8(-/-), and gammadelta(-/-) knockout mice to low concentrations of M. avium strain 101 given orally, followed by treatment with azithromycin. After 1 month, the mice were challenged with kanamycin-resistant M. avium 104. Only CD4(+) T cells appeared to be required for protection against the second challenge. Both CD4(+) and CD8(+) T cells produced comparable amounts of gamma interferon after the first exposure to the bacterium. Tumor necrosis factor alpha was elevated in CD4(+) T cells but not in CD8(+) T cells. Following exposure to a small inoculum of mycobacteria orally, wild-type mice did not develop disseminated infection for approximately 4 months, although viable bacteria could be observed in the mesenteric lymph nodes. The ingestion of small numbers of M. avium cells induces a protective immune response in the intestines against subsequent infection. However, the bacteria remain viable in intestinal lymph nodes and might disseminate later.

PPE and PE gene families, which encode numerous proteins of unknown function, account for 10% of Mycobacterium tuberculosis genome. Mycobacterium avium genome has similar PPE and PE gene families. Using a temperature-sensitive phage phAE94 transposon mutagenesis system, a M. avium transposon library was created in the strain MAC109. Screening of individual mutants in human U937 macrophages for the ability to replicate intracellularly, we identified several attenuated clones. One of them, the 2D6 mutant, has a transposon interrupting a PPE gene (52% homologous to Rv 1787 in M. tuberculosis) was identified. The mutant and the wild-type strain had comparable ability to enter macrophages. Challenge of mice with the 2D6 mutant resulted in approximately 1 log and 2 log fewer bacteria in the spleen, at 1 and 3 weeks after infection, compared with the wild-type bacterium. The 2D6 mutant grows like the wild-type bacterium in vitro. Vacuoles containing the 2D6 mutant acidified to pH 4.8; whereas, vacuoles containing wild-type bacterium were only slightly acidic. It was also observed that, in contrast to the wild-type bacterium, the 2D6 mutant did not prevent phagosome-lysosome fusion, and it is only expressed within macrophage but not in 7H9 broth. These results revealed a role for this PPE gene in the growth of M. avium in macrophages and in virulence in mice.

The role that colonization with Mycobacterium avium plays in the development of disseminated disease is unclear. In this study, we determined whether all M. avium strains isolated from the portals of M. avium infection are capable of crossing the mucosal border and causing infection. The patients in this study were enrolled in AIDS Clinical Trial Group protocol 341. The patients were divided into 3 groups; 2 groups differed in their immunological and clinical risk for M. avium disease. A third group (n=22 patients) had culture-documented disseminated M. avium complex disease at the time of entry in the study. Eight of 22 patients had M. avium isolated from both a colonized site and blood or bone marrow specimens. All 8 patients had distinct M. avium strains; 2 patients had a polyclonal infection. The virulence properties of 13 strains were determined, including invasion of gastrointestinal cells and replication in macrophages. There were significant differences in the virulence properties, and these differences may provide insight into the interplay between microbial pathogenesis and host defense.

Mycobacterium avium complex (MAC) infection is the most common disseminated bacterial infection in untreated patients with acquired immunodeficiency syndrome (AIDS). We investigated the potential role of monocyte chemotactic protein-1 (MCP-1) in the pathogenesis of disseminated MAC, using the simian immunodeficiency virus (SIV)/macaque model of AIDS. Macaques were inoculated with SIV, followed by challenge with a pathogenic AIDS isolate of M. avium 14 days later. After challenge with M. avium, marked increases in serum MCP-1 levels were detected in SIV-infected macaques, a finding that was duplicated in coinoculated bronchoalveolar macrophages. MCP-1 levels were significantly higher in SIV-infected macaques than in non-SIV-infected controls (327.1 vs. 151.5 pg/mL, respectively; P=.04), suggesting that up-regulation of MCP-1 contributes to the development of progressive mycobacterial disease. Similarly, morphometric analysis revealed increased expression of MCP-1 in hepatic microgranulomas from SIV-infected macaques. We conclude that the pronounced increases in MCP-1 levels demonstrated in tissue and serum samples after M. avium inoculation may play a role in the development of disseminated mycobacterial disease.

Hemolysin was quantified in 58 isolates of Mycobacterium avium from human, animal, and environmental sources. Human Mav-A and Mav-B isolates were the strongest producers; in contrast, animal and environmental Mav-A isolates and human, animal, and environmental Mav-C organisms were low-level producers. Hemolysin production was not restricted to isolates causing invasive infections.

Whether infection with Mycobacterium avium complex (MAC) among patients with acquired immune deficiency syndrome results from recent exposure to virulent strains or reactivation of latent infection acquired years earlier is unknown. To address this question, tissue samples from 47 simian immunodeficiency virus (SIV)-infected and 63 SIV-uninfected rhesus macaques were cultured. MAC was cultured from 14 SIV-uninfected macaques (22.2%) and 32 SIV-infected macaques (68.1%); median bacterial burdens were 33.3 and 998.7 cfu/g, respectively. Genetically distinct strains of MAC were identified for 13 SIV-uninfected macaques (20.6%) and 15 SIV-infected macaques (31.9%). A genetically identical MAC strain (K128A) was identified for 25 SIV-infected macaques (53.2%) and 1 SIV-uninfected macaque (1.6%). Multivariate analysis identified infection with SIV/Delta(B670), diagnosis of an SIV-related tumor or opportunistic infection, and birth on site as risks for MAC infection. SIV-uninfected and SIV-infected macaques yielding unique strains of MAC were considered to have latent and reactivation infection, respectively, whereas animals infected with strain K128A were considered to have recent infection, demonstrating that both mechanisms occur among rhesus macaques.

A female patient with multiple osteomyelitis and pulmonary Mycobacterium avium disease visited an orthopaedic clinic with back pain. Systemic bone scan showed multiple sites of increased radioactivity in the vertebral bodies, right scapula, femurs and ribs. M. avium was isolated from sputum and a sample aspirated from the right scapula. The route of infection was unknown as there was no history of trauma or surgery. HIV testing was negative. As there was no underlying immunological disease she was diagnosed as disseminated M. avium complex (DMAC) disease in an immunocompetent adult. Cytokine production on several stimuli from peripheral blood mononuclear cells was similar to that in pulmonary M. avium patients. Sequence analysis of IFN-gamma receptor revealed no nucleotide substitution. We detected serotypes 1, 2 and 4 from mycobacteria cultured from the right scapula, and conclude that this case could be the result of undetected immune deficiency and/or unrecognised virulence of the infecting isolate.

Background: The pathogenesis of Mycobacterium avium complex and Mycobacterium tuberculosis in the respiratory tract is poorly understood, as are the reasons for their differing virulence. We have previously shown that their initial adherence to the mucosa is identical., Methods: The interaction of M avium complex, M tuberculosis, and M smegmatis with human respiratory tissue was investigated in an organ culture model with an air interface. Tissue was infected for intervals up to 14 days and assessed by scanning electron microscopy for adherent bacteria or cultured for recoverable bacteria., Results: The mean number of adherent bacteria/mm(2) (and the viable count of macerated tissue, cfu/ml) at 15 minutes, 3 and 24 hours, 7 and 14 days were: M avium complex 168 (153), 209 (136), 289 (344), 193 (313), 14140 (16544); M tuberculosis 30 (37), 39 (23), 48 (53), 1 (760), 76 (2186); M smegmatis 108 (176), 49 (133), 97 (81), 114 (427), 34 (58), (n=6). There was no significant change in morphology between infected and uninfected tissue or tissue infected with the different species over 14 days. The number of M avium complex on the mucosa and recovered from tissue increased over time (p=0.03). M tuberculosis decreased on the surface, but recoverable bacteria increased (p=0.01). M smegmatis numbers on the mucosa and recovered from tissue decreased. Sectioned tissue showed M avium complex and M tuberculosis in submucosal mucus glands and M tuberculosis penetrating epithelial cells in one experiment., Conclusions: The initial adherence to the mucosa of the three species was similar, but after 14 days they varied in their interaction with the tissue in a manner compatible with their pathogenicity.

Increasing numbers of patients have presented with a hypersensitivity pneumonitis-type course in association with hot tub exposure. Mycobacterium avium complex (MAC) organisms have been isolated from both patient specimens and hot tub water with matching fingerprints by restricted fragment length polymorphism and electrophoresis when performed. Review of the clinical, microbiologic, and radiographic presentations of 9 patients to the Mayo Clinic with this diagnosis are compared with 32 patients in the published literature. The diagnosis, treatment, and prognosis of MAC hot tub lung are reviewed., (Copyright 2003 Elsevier, Inc. All rights reserved.)

Three children, a girl aged 2.5 years and two boys aged 2 and 3 years respectively, presented with unilateral cervical lymphadenitis. The first patient had acute bacterial lymphadenitis due to group A Streptococcus, characterised by a painful cervical swelling of acute onset. The second patient had painless cervical lymphadenitis caused by Mycobacterium avium-intracellulare, which drained spontaneously. The third patient developed a non-tender, cervical swelling within a day. He too was systemically ill with fever and a headache. The lymphadenitis was caused by Bartonella henselae. After drainage, dissection and/or antibiotic therapy, all three recovered. A cervical mass in a young child is most frequently caused by an infectious lymphadenopathy. It rarely represents a malignant or other systemic disease. In many cases the diagnosis of infectious lymphadenitis can be made on the basis of the case history and clinical characteristics. However, when malignancy cannot be excluded tissue examination is always indicated.

The Mycobacterium avium complex (MAC) encompasses important pathogens in both animals and humans, yet little information is available on the factors required for MAC virulence. An animal isolate, M. avium strain 724 was found to be considerably more virulent in Balb/c mice than a human isolate, M. avium strain A5. To identify the genetic basis of this difference subtractive hybridization was applied, which resulted in the isolation of six DNA fragments unique to strain 724. BLAST searches showed that three sequences belonged to a large gene cluster responsible for biosynthesis of M. avium glycopeptidolipids (GPLs). To reveal the nature of variation between strains in the GPL cluster 27.5kb of a clone containing the A5 serotype-specific GPL (ssGPL) cluster was isolated, sequenced and compared to the corresponding region in other M. avium strains. The ssGPL cluster was highly conserved in the 5' region between all strains and serotypes tested; the 3' region reflects extensive divergence among serotypes including whole gene deletions and insertions of sequences containing open reading frames but lacking identity to any known genes.

There has not yet been systematic studies which attempt to elucidate detailed profiles of the interaction between antimicrobial drugs and macrophage microbicidal mechanisms. We examined the effects of antisense oligo DNAs (AsDNAs) against oxyR and ahpC on the susceptibility of Mycobacterium avium complex (MAC) to the H2O2-halogenation system and combined antimycobacterial drugs [clarithromycin (CAM) + rifampicin (RFP)], both separately and in combination. It was found that AsDNA treatment of MAC did not affect the susceptibility of the organisms to any of the antimicrobial systems tested. Since the present AsDNAs did not efficiently reduce the expression of AhpC mRNA, attempts to increase bacterial uptake of AsDNAs are necessary to achieve significant increase in the drug susceptibility of MAC organisms due to AsDNA treatment.