Enhancement of mycobacterial pathogenesis by host interferon-γ.

The cytokine IFNγ is a principal effector of macrophage activation and immune resistance to mycobacterial infection; however, pathogenic mycobacteria are capable of surviving in IFNγ-activated macrophages by largely unknown mechanisms. In this study, we find that pathogenic mycobacteria, including M. bovis BCG and M. tuberculosis can sense IFNγ to promote their proliferative activity and virulence phenotype. Moreover, interaction with the host intracellular environment increases the susceptibility of mycobacteria to IFNγ through upregulating expression of mmpL10, a mycobacterial IFNγ receptor, thereby facilitating IFNγ-dependent survival and growth of mycobacteria in macrophages. Transmission electron microscopy analysis reveals that IFNγ triggers the secretion of extracellular vesicles, an essential virulence strategy of intracellular mycobacteria, while proteomics identifies numerous pivotal IFNγ-induced effectors required for mycobacterial infection in macrophages. Our study suggests that sensing host IFNγ is a crucial virulence mechanism used by pathogenic mycobacteria to survive and proliferate inside macrophages.

Improved detection of mycobacteria in CF and tissue samples grown in mycobacteria growth indicator tube incubated at 30°C compared to conventional growth conditions of liquid and solid media.

This study evaluates the growth of mycobacteria in samples from cystic fibrosis (CF) patients and tissue samples using the mycobacteria growth indicator tube (MGIT) incubated at 30°C in comparison to conventional MGIT cultures incubated at 37°C in a BACTEC MGIT 960 device and solid media incubated at 36°C and 30°C. A total of 1,549 samples were analyzed, of which 202 mycobacterial isolates were cultured from 197 positive specimens, including five mixed cultures. The highest detection rate was achieved from MGIT at 30°C, with 84.2% of mycobacterial isolates (170 of 202), which was significantly higher than any other culture condition (P < 0.0001 for any condition). MGIT at 37°C yielded 61.4% (124 of 202) of the recovered isolates, whereas Löwenstein Jensen (LJ) and Stonebrink at 36°C, and LJ and Stonebrink at 30°C retrieved 47.0% (95), 49.5% (100), 50.0% (101), and 53.0% (107) of the isolates, respectively. Of the 53 isolates that were grown exclusively under one culture condition, the highest number of isolates (36) was recovered from MGIT incubated at 30°C. MGIT at 37°C recovered eight of the 53 isolates, whereas LJ incubated at 30°C and Stonebrink incubated at 30°C and 36°C recovered five, three, and one isolate, respectively. No isolates were grown exclusively from LJ incubated at 36°C. In CF patients and tissue samples, MGIT cultivated at 30°C for 8 weeks increases the performance of mycobacterial culture. IMPORTANCE: Our study shows that the addition of mycobacteria growth indicator tube (MGIT) liquid culture incubated at 30°C improves the detection of mycobacteria from CF and tissue samples. MGIT incubated at 30°C recovered significantly more mycobacterial isolates than MGIT incubated at 37°C and significantly more isolates than either Lowenstein Jensen or Stonebrink solid media incubated at either 36°C or 30°C. Of 202 mycobacterial isolates recovered from 1,549 specimens, 170 were recovered from MGIT incubated at 30°C, followed by MGIT incubated at 37°C with 124 isolates and solid media culture conditions that recovered between 95 and 107 mycobacterial isolates. All conventional culture conditions combined without MGIT incubated at 30°C recovered 166 isolates. MGIT incubated at 30°C recovered the highest number of isolates detected exclusively by a single culture condition and recovered mycobacterial isolates of highly relevant mycobacterial species, including Mycobacterium abscessus and Mycobacterium tuberculosis.

Visualisation of in vivo protein synthesis during mycobacterial infection through [68Ga]Ga-DOTA-puromycin µPET/MRI.

Radiolabelled puromycin analogues will allow the quantification of protein synthesis through nuclear medicine-based imaging. A particularly useful application could be the non-invasive longitudinal visualisation of mycobacterial activity through direct quantification of puromycin binding. This study assesses the value of [68Ga]Ga-DOTA-puromycin in the visualisation of mycobacteria through positron emission tomography combined with magnetic resonance imaging (µPET/MRI). The radiopharmaceutical was produced by previously published and validated methods. [68Ga]Ga-DOTA-Puromycin imaging was performed on severe immunodeficient mice infected with Bacille Calmette-Guérin-derived M. Bovis (BCG). Acute and chronic infection stages were examined by µPET/MRI. A follow-up group of animals acted as controls (animals bearing S. aureus-derived infection and sterile inflammation) to assess tracer selectivity. [68Ga]Ga-DOTA-puromycin-µPET/MRI images revealed the acute, widespread infection within the right upper shoulder and armpit. Also, [68Ga]Ga-DOTA-puromycin signal sensitivity measured after a 12-week period was lower than that of [18F]FDG-PET in the same animals. A suitable correlation between normalised uptake values (NUV) and gold standard histopathological analysis confirms accurate tracer accumulation in viable bacteria. The radiopharmaceutical showed infection selectivity over inflammation but accumulated in both M. Bovis and S. Aureus, lacking pathogen specificity. Overall, [68Ga]Ga-DOTA-puromycin exhibits potential as a tool for non-invasive protein synthesis visualization, albeit without pathogen selectivity.

A Case of Disseminated Mycobacterium Haemophilum in a Kidney Transplant Recipient Presenting With Subcutaneous Nodules.

INTRODUCTION: Dermatologic manifestations of diseases in solid organ transplant recipients are common due to long-term immunosuppression. CASE PRESENTATION: We present the case of a 63-year-old man with a kidney transplant who exhibited subcutaneous nodules on lower extremities, cytopenia, and asymptomatic pulmonary infiltrate. Through a skin biopsy and 16S ribosomal RNA (rRNA) sequencing, Mycobacterium haemophilum was identified. His clinical course was complicated by empyema, septic arthritis, and recurrence of his skin manifestations, despite ongoing antimicrobial treatment. DISCUSSION: This case emphasizes the challenges and potential complications associated with M haemophilum infections in solid organ transplant recipients receiving long-term immunosuppressive therapy. It highlights the importance of employing advanced diagnostic techniques when evaluating dermatologic manifestations in these patients. The patient's complex clinical course also underscores the difficulties involved in effectively addressing and managing complications that may arise even after initiating therapy.

PDCD4 as a marker of mTOR pathway activation and therapeutic target in mycobacterial infections.

Programmed cell death protein 4 (PDCD4) is instrumental in regulating a range of cellular processes such as translation, apoptosis, signal transduction, and inflammatory responses. There is a notable inverse correlation between PDCD4 and the mammalian target of rapamycin (mTOR) pathway, which is integral to cellular growth control. Activation of mTOR is associated with the degradation of PDCD4. Although the role of PDCD4 is well established in oncogenesis and immune response regulation, its function in mycobacterial infections and its interplay with the mTOR pathway necessitate further elucidation. This study investigates the modulation of PDCD4 expression in the context of mycobacterial infections, revealing a consistent pattern of downregulation across diverse mycobacterial species. This observation underscores the potential utility of PDCD4 as a biomarker for assessing mTOR pathway activation during such infections. Building on this finding, we employed a novel approach using PDCD4-based mTOR (Tor)-signal-indicator (TOSI) reporter cells for the high-throughput screening of FDA-approved drugs, focusing on mTOR inhibitors. This methodology facilitated the identification of several agents, inclusive of known mTOR inhibitors, which upregulated PDCD4 expression and concurrently exhibited efficacy in impeding mycobacterial proliferation within macrophages. These results not only reinforce the significance of PDCD4 as a pivotal marker in the understanding of infectious diseases, particularly mycobacterial infections, but also illuminate its potential in the identification of mTOR inhibitors, thereby contributing to the advancement of therapeutic strategies. IMPORTANCE: This study emphasizes the critical role of the mammalian target of rapamycin (mTOR) pathway in macrophage responses to mycobacterial infections, elucidating how mycobacteria activate mTOR, resulting in PDCD4 degradation. The utilization of the (Tor)-signal-indicator (TOSI) vector for real-time monitoring of mTOR activity represents a significant advancement in understanding mTOR regulation during mycobacterial infection. These findings deepen our comprehension of mycobacteria's innate immune mechanisms and introduce PDCD4 as a novel marker for mTOR activity in infectious diseases. Importantly, this research laid the groundwork for high-throughput screening of mTOR inhibitors using FDA-approved drugs, offering the potential for repurposing treatments against mycobacterial infections. The identification of drugs that inhibit mTOR activation opens new avenues for host-directed therapies, marking a significant step forward in combating tuberculosis and other mycobacterial diseases.

Application of Genomic Epidemiology of Pathogens to Farmed Yellowtail Fish Mycobacteriosis in Kyushu, Japan.

To investigate mycobacterial cases of farmed yellowtail fish in coastal areas of western Japan (Kagoshima, Kyushu), where aquaculture fisheries are active, Mycobacterium pseudoshottsii, the causative agent, was isolated from six neighboring fishing ports in 2012 and 2013. A phylogenetic ana-lysis revealed that the strains isolated from one fishing port were closely related to those isolated from other regions of Japan, suggesting the nationwide spread of a single strain. However, strains from Japan were phylogenetically distinct from those from the Mediterranean and the United States; therefore, worldwide transmission was not observed based on the limited data obtained on the strains exami-ned in this study. The present results demonstrate that a bacterial genomic ana-lysis of infected cases, a mole-cular epidemiology strategy for public health, provides useful data for estimating the prevalence and transmission pathways of M. pseudoshottsii in farmed fish. A bacterial genome ana-lysis of strains, such as that performed herein, may play an important role in monitoring the prevalence of this pathogen in fish farms and possible epidemics in the future as a result of international traffic, logistics, and trade in fisheries.

Mycobacterium genavense granulomatous typhlocolitis in a horse.

A 23-y-old gelding was presented to a veterinary teaching hospital with a history of chronic, refractory diarrhea. Clinically, the horse was in poor body condition, with a thickened and corrugated large intestine identified by transcutaneous abdominal ultrasonography. At postmortem examination following euthanasia, the large colon and cecum had segmental thickening of the intestinal wall with innumerable mucosal ulcers and prominent polypoid mucosal masses. Many mesenteric and hepatic lymph nodes were enlarged. Histology revealed granulomatous and ulcerative typhlocolitis and granulomatous lymphadenitis with myriad acid-fast, variably gram-positive, intrahistiocytic bacilli that stained by immunohistochemistry for mycobacteria. Molecular testing by PCR and sequencing identified the causative agent as Mycobacterium genavense, which is an unusual presentation of infection in a horse.

[A case of HIV combined with Mycobacterium celatum infection].

Here, we reported the diagnosis and treatment of a case of HIV infected person complicated by an extremely rare infection with Mycobacterium celatum. Due to the similarity of homologous sequence regions between Mycobacterium celatum and Mycobacterium tuberculosis complex, the identification of conventional Mycobacterium species was incorrect, which was corrected after first-generation 16S rRNA sequencing. This report aimed to improve the clinical understanding of Mycobacterium celatum infection and the level of differential diagnosis between non-tuberculous mycobacterial disease and tuberculosis.

Mycobacterium haemophilum infection in immunocompetent adults: a literature review and case report.

BACKGROUND: Mycobacterium haemophilum has been increasingly found in severely immunocompromised patients but is scarcely reported in immunocompetent adults. METHODS: We systematically reviewed previous literature to identify studies on infection in immunocompetent adults. Articles reporting at least one case of M. haemophilum infection were included. We excluded articles involving patients who had immunosuppression-related diseases and routinely used glucocorticoids or immunosuppressants. We also reported a case of a young immunocompetent woman infected by M. haemophilum along the eyebrows, which was probably due to the use of an eyebrow pencil retrieved from a sink drain. RESULTS: Twelve qualifying articles reporting M. haemophilum infection in immunocompetent adults were identified. Among them, most cases report skin lesions along the eyebrows, and the remaining had cervicofacial lymphadenitis, lesions on the arm or fingers, inflammation in the eyeballs, or ulceration in the perineal region. Most cases were caused by tattoos, make-up, injury, or surgical operation. For diagnosis, specialized tissue culture sensitivity was roughly 75%, and polymerase chain reaction (PCR) test sensitivity was approximately 89%. Triple antibiotic therapy for 3 to 24 months, or surgical excision was effective in controlling infection. CONCLUSION: M. haemophilum infection should be considered if routine antibacterial and glucocorticoid treatments are ineffective against the disease, even in healthy adults. To definitively diagnose this infection, conditioned tissue culture or PCR testing is required. Treatment usually involves a combination of multiple antibiotics and, if necessary, surgical removal of infected tissue.

Activation of host nucleic acid sensors by Mycobacterium: good for us or good for them?

Although the importance of deoxyribonucleic acid (DNA) and ribonucleic acid (RNA) sensors in controlling viral infection is well established, their role in promoting an effective immune response to pathogens other than viruses is less clear. This is particularly true for infections with mycobacteria, as studies point to both protective and detrimental roles for activation of nucleic acid sensors in controlling a mycobacterial infection. Some of the contradiction likely stems from the use of different model systems and different mycobacterial species/strains as well as from which nucleic acid sensors were studied and what downstream effectors were evaluated. In this review, we will describe the different nucleic acid sensors that have been studied in the context of mycobacterial infections, and how the different studies compare. We conclude with a section on how nucleic acid sensor agonists have been used therapeutically and what further information is needed to enhance their potential as therapeutic agents.

Central nervous system mycobacteriosis caused by Mycobacterium genavense in degus (Octodon degus).

Degus (Octodon degus) that were kept at a breeding facility presented with neurological or respiratory symptoms and died. Necropsies were performed on 9 individuals, and no significant gross lesions were found. Histologically, spinal cord necrosis was observed in all 9 cases and granulomatous myelitis in 5 of the 9 cases. Locally extensive necrosis of the brain and encephalitis were observed in 7 of the 9 cases. Acid-fast bacteria were found in the spinal cords, brains, and lungs from all 9 cases. Immunohistochemically, Mycobacterium tuberculosis antigen was observed in the spinal cords, brains, and lungs from all 9 cases. Double-labeling immunofluorescence revealed M. tuberculosis antigen in IBA1- and myeloperoxidase-immunopositive cells. Extracted genomic DNA from 8 of the 9 cases was successfully amplified with the primers for Mycobacterium genavense ITS1 and hypothetical 21 kDa protein genes, and the polymerase chain reaction products were identified as M. genavense by DNA sequencing. This report highlights the susceptibility of degus to M. genavense infection in the central nervous system.

A case report of Mycobacterium fortuitum infection after muscle injection.

RATIONALE: Injection-related abscesses are a common complication in clinical practice, but the identification of infected bacteria might be difficult. PATIENT CONCERNS: A 51-year-old female patient was admitted to the hospital due to a lump on her right buttock that emerged after receiving intramuscular injections to treat left shoulder joint pain. The lump gradually enlarged into a 3.0 to 4.5 cm mass at the time of admission with symptoms such as skin redness, itching, and pain. DIAGNOSES: The patient received ultrasonic and other laboratory examinations. Laboratory results from the drainage indicated that the infection was caused by a rapidly growing mycobacteria and was confirmed as Mycobacterium fortuitum by matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry. INTERVENTIONS: The patient was treated with antibiotics for 12 days after incision and drainage of the abscess in the right buttock. Local dressings were changed regularly. A migration lesion that appeared 3 days after treatment was drained and cleaned when it matured. OUTCOMES: The lesion substantially decreased in size and the patient was discharged after 2 months of treatment. LESSONS: Rapidly growing mycobacteria are rare but important pathogens that should be considered in patients with injection-related abscesses. Early identification and appropriate treatment can result in a favorable prognosis.

Multicenter matched-pair study comparing BACT/ALERT® MP reagent systems for the recovery of mycobacteria from specimens other than blood.

The BACT/ALERT® MP Reagent System is a broth culture medium for optimal detection and recovery of mycobacteria from clinical samples. The MP formulation was recently modified to improve detection and recovery times. A multicenter prospective matched pair study design was conducted to validate the performance of improved MP (MP-I) versus current MP (MP-C) bottles utilizing nonsterile and normally sterile samples, except blood, from patients suspected of having mycobacterial infections. A total of 1488 clinical samples were collected to obtain 212 mycobacteria samples by either or both MP culture bottles. MP-I and MP-C sensitivities were 86.6% and 81.4%, respectively, but the difference was not significant (P = 0.163) while specificities were 96.8% and 93.8%, respectively, and that difference was significant (P = 0.002). Overall recovery was 94.34% for MP-I and 88.68% for MP-C (recovery was 100% for both bottles with 52 seeded samples). Overall performance of MP-I was better than MP-C for sensitivity, specificity, and recovery.

Healthcare-Associated Infections Caused by Mycolicibacterium neoaurum.

Mycolicibacterium neoaurum is a rapidly growing mycobacterium and an emerging cause of human infections. M. neoaurum infections are uncommon but likely underreported, and our understanding of the disease spectrum and optimum management is incomplete. We summarize demographic and clinical characteristics of a case of catheter-related M. neoaurum bacteremia in a child with leukemia and those of 36 previously reported episodes of M. neoaurum infection. Most infections occurred in young to middle-aged adults with serious underlying medical conditions and commonly involved medical devices. Overall, infections were not associated with severe illness or death. In contrast to other mycobacteria species, M. neoaurum was generally susceptible to multiple antimicrobial drugs and responded promptly to treatment, and infections were associated with good outcomes after relatively short therapy duration and device removal. Delays in identification and susceptibility testing were common. We recommend using combination antimicrobial drug therapy and removal of infected devices to eradicate infection.

DNA microarray chip assay in new use: early diagnostic value in cutaneous mycobacterial infection.

Introduction: The clinical practicability of DNA microarray chip in detecting the presence of mycobacterial species/isolates directly in the skin tissues has not been evaluated, nor the efficacy of DNA microarray chip as a novel diagnostic tool for the early diagnosis of cutaneous mycobacterial infections is known. Methods: The present study analyzed the incidence of cutaneous mycobacterial infections in Shanghai and explored the efficacy of a novel DNA microarray chip assay for the clinical diagnosis of the disease from skin tissue specimens compared to traditional detection methods. A total of 60 participants fulfilling the defined diagnostic criteria and confirmed positive for cutaneous mycobacterial infections from 2019 to 2021 were enrolled in the study. Subsequent to recording the participants' medical history and clinical characteristics, the skin tissue specimens were collected for analyses. The specimens underwent histopathological analyses, skin tissue culture, and DNA microarray chip assay. Results: Increased incidence of cutaneous mycobacterial infection was detected from 2019 to 2021. The most common infecting pathogen was M. marinum followed by M. abscessus. The sensitivity, specificity and accuracy of the skin tissue culture method were 70%, 100% and 76.62%, respectively, while that of the DNA microarray chip assay were 91.67%, 100% and 93.51%, respectively. The sensitivity and accuracy of the DNA microarray chip assay were significantly higher than those of the skin tissue culture method. The positive likelihood and diagnostic odds ratio were >10 and >1, respectively for both the methods. The negative likelihood ratio was significantly higher (30% vs 8.33%) and the Youden's index was significantly lower (70.00% vs 91.67%) in the skin culture method compared to that of the DNA microarray chip assay. There was a significant association of false negative results with a history of antibiotic use in the skin tissue culture method. Discussion: Given the increasing incidence of cutaneous mycobacterial infections, early diagnosis remains a prime clinical focus. The DNA microarray chip assay provides a simple, rapid, high-throughput, and reliable method for the diagnosis of cutaneous mycobacterial infections with potential for clinical application.

Immunological and metabolic characterization of environmental Mycobacterium chimaera infection in a murine model.

Mycobacterium chimaera causes pulmonary disease, but little is known of gradations in isolate virulence. Previously, 17 M. chimaera isolates were screened for survival in THP1 macrophages. "M. chimaera 1" was categorized as "more virulent" because it showed the greatest survival in macrophages, whereas "M. chimaera 2" was categorized as "less virulent" with reduced survival. Herein, we infected C3HeB/FeJ mice to compare the in vivo immune responses to M. chimaera 1 and 2. Unlike macrophages, significantly lower M. chimaera 1 counts were recovered from mouse lung tissue and BAL cells with less lung histopathologic changes compared to M. chimaera 2. Compared to M. chimaera 2, significantly more IL-1ß, IL-6, and TNFα was produced early after M. chimaera 1 infection. LC-MS metabolomics analyses of BAL fluid revealed divergence in sphingolipid, phospholipid metabolism between M. chimaera 1 versus M. chimaera 2 mice. From pan-GWAS analyses, virulence and organizing DNA/molecular structure genes were associated with more virulent M. chimaera isolates. Vigorous lung-specific immune responses to M. chimaera 1 may influence effective bacterial control, but for a different isolate M. chimaera 2, subvert immune control. Continued studies of the gradations in virulence among the same NTM species will advance our understanding of NTM pathogenesis.

Concurrent Mycobacterium genavense infection and intestinal B-cell lymphoma in a pet rabbit (Oryctolaguscuniculus).

A 6-year-old male intact pet rabbit was evaluated for chronic weight loss. A large mass was detected by palpation in the mid-abdomen and ultrasound examination suggested a jejunal location. Explorative laparotomy revealed a nodular mass within the jejunal wall. Histological examination of a biopsy revealed mycobacterial granulomatous enteritis with an atypical lymphoblastic proliferation suggestive of lymphoma. Neoplastic lymphocytes were immunopositive for Pax-5 but negative for CD3, which is diagnostic of a B-cell neoplasm. Numerous acid-fast bacteria were seen within histiocytes and identified by polymerase chain reaction as Mycobacterium genavense, which is a non-tuberculous and opportunistic mycobacterium with zoonotic potential. To the best of our knowledge, this is the first documented case of a concurrent B-cell lymphoma and M. genavense infection in a rabbit. Concomitant mycobacteriosis and lymphoma have been rarely described in animals and the coexistence of neoplasia and mycobacterial infection within the jejunum suggests a potential pathogenetic association. Interestingly, the rabbit owner worked in an anti-tuberculosis clinic, and an anthropic origin of the mycobacterial infection could not be excluded.

Disseminated Mycobacterium genavense infection in a patient with a history of sarcoidosis.

We present a case of Mycobacterium genavense infection in a man in his 60s with a history of sarcoidosis, treated for 24 years with systemic corticosteroids and later methotrexate as monotherapy. He presented with low grade fever, dyspnoea and right-sided thoracic pain and was admitted due to a treatment-refractory infection. After a prolonged period of symptoms and diagnostics, acid-fast bacilli were demonstrated in pleural fluid and PCR revealed M. genavense The patient was treated with intravenous amikacin, peroral azithromycin, rifampicin and ethambutol for a total of 18 months, with a good clinical and radiological treatment response. Infection with M. genavense is rare in HIV-negative immunocompromised hosts. Diagnosing and treating mycobacterial infections, especially for more rare species, remains a challenge as clinical evidence is sparse. Nonetheless, the disease-causing infection must be considered in symptomatic and immunocompromised patients.

Canine leproid granuloma caused by a member of the Mycobacterium tuberculosis complex.

Canine leproid granuloma (CLG) is a chronic form of dermatitis that has been associated with nontuberculous mycobacterial infections in Africa, Oceania, the Americas, and Europe. We report here a case of CLG associated with a member of the Mycobacterium tuberculosis complex (MTBC), which could be of public health concern. An 8-y-old pet dog developed 0.5-1-cm diameter, raised, firm, nonpruritic, alopecic, painless skin nodules on the external aspects of both pinnae. Histologic examination revealed severe pyogranulomatous dermatitis with intracellular Ziehl-Neelsen-positive bacilli that were immunoreactive by immunohistochemistry using a polyclonal primary antibody that recognizes tuberculous and nontuberculous Mycobacterium species. DNA extracted from formalin-fixed, paraffin-embedded skin sections was tested by a Mycobacterium genus-specific nested PCR assay targeting the 16S rRNA gene. BLAST sequence analysis of 214-bp and 178-bp amplicons showed 99.5% identity with members of the MTBC; however, the agent could not be identified at the species level. Although CLG has been associated traditionally with nontuberculous mycobacterial infections, the role of Mycobacterium spp. within the MTBC as a cause of this condition, and the role of dogs with CLG as possible sources of MTBC to other animals and humans, should not be disregarded given its zoonotic potential.