Mycobacterium Colombiense Research Articles

Absence of Tuberculosis-Causing Mycobacteria from Slaughtered Livestock Tissues and Environmental Samples, Gauteng Province, South Africa.

Mycobacterium tuberculosis complex (MTBC) is a group of bacteria responsible for causing tuberculosis in animals and humans. In South Africa (S.A), slaughterhouses are registered by the government and closely inspected and audited for hygienic slaughter practices. Meat inspection to detect lesions has been used for passive surveillance, monitoring, and diagnosis of the disease status. Information on the current status of bovine tuberculosis (bTB) in livestock in the country is limited. Hence, we investigated the occurrence of Mycobacterium spp. in the tissues of slaughtered livestock and environmental samples in abattoirs in Gauteng province of South Africa (S.A). The cross-sectional study employing random sampling from cattle, pigs, and sheep (with the collection of liver, lung, spleen, and different lymph nodes) irrespective of lesions was carried out in 19 red meat abattoirs. Five hundred animals were sampled, comprising cattle (n = 369), pigs (n = 90), and sheep (n = 41). Additionally, 19 environmental samples were collected from feedlots, or where animals drink water while awaiting slaughter, to identify mycobacterial species using culture, acid-fast bacteria staining, and polymerase chain reaction (PCR). The Chi-square and Fisher's Exact tests were used to detect statistically significant differences in the frequency of detection of Mycobacterium spp. according to the variables investigated (types of tissues, livestock, abattoirs, etc.). The PCR assays detected no MTBC complex species DNA in the bacterial isolates from cattle (n = 32). Sequence analysis (16S rDNA) of the isolates from eight cattle confirmed only two species, namely Mycobacterium colombiense (99.81% identity) and Mycobacterium simiae (99.42% identity). The remaining isolates were identified as members of the Actinomadura species. From the environmental samples, bacterial isolation was made from three samples, and two could only be identified up to the genus level (Mycobacterium species) while the remaining isolate was identified as Mycobacterium senuense (99.22% identity). The study revealed the absence of bovine tuberculosis-causing pathogens in red meat abattoirs of the Gauteng province. Although non-tuberculous Mycobacteria have been implicated as potentially causing tuberculosis-like diseases in livestock, their occurrence in the current study was found to be low, but the potential to cause disease cannot be ignored.

Diagnostic and Therapeutic Challenges in Disseminated Mycobacterium colombiense Infection Caused by Interferon-γ Neutralizing Autoantibodies.

Adult-onset immunodeficiency due to interferon-γ-neutralizing autoantibodies (nIFNγ-autoAbs) can remain underdiagnosed. We present a case of severe Mycobacterium colombiense infection with nIFNγ-autoAbs. To ensure early diagnosis, clinicians should have a high index of suspicion in patients of Asian descent with opportunistic infections and perform QuantiFERON-TB assay for disease screening.

Clinical features of rare disseminated Mycobacterium colombiense infection in nine patients who are HIV-negative in Guangxi, China

ObjectivesLocalized or disseminated infection caused by different nontuberculous mycobacteria (NTM) species has been increasingly reported in recent years, but reports of Mycobacterium colombiense infection are extremely rare. Herein, we analyzed the clinical features of patients with disseminated M. colombiense infection. MethodsPatients diagnosed with disseminated M. colombiense infection between February 4, 2016 and August 25, 2021 at the First Affiliated Hospital of Guangxi Medical University were retrospectively analyzed. ResultsNTM infection was diagnosed in 248 HIV-negative patients. Of these, nine patients with disseminated M. colombiense infection were enrolled. Five of these patients were positive for anti-interferon-γ autoantibodies. The lung, lymph nodes, bones, and joints were the most commonly involved organs. Anemia, fever, lymphadenopathy, cough and expectoration, and ostealgia were the most common symptoms. The levels of white blood cells and neutrophils were increased in eight patients. M. colombiense was detected by both metagenomic next-generation sequencing (mNGS) and culture in four patients and only by mNGS in the remaining five patients. All patients received combination anti-NTM therapy; five underwent surgery. The condition of eight patients improved, and one died during the treatment. ConclusionPatients infected with M. colombiense can present as disseminated infections, easily involving multiple organs, such as the lung, lymph nodes, bone, and joints, with fever, lymphadenopathy, and increased white blood cell and neutrophil counts. mNGS plays a crucial role in the early diagnosis of M. colombiense infection. Once diagnosed, timely and effective anti-NTM therapy, combined with local surgery if necessary, can improve the prognosis of patients with this condition.

Disseminated nontuberculous mycobacterial infection with cryptic immunodeficiency mimicking malignancy: a case report

BackgroundNontuberculous mycobacteria (NTM) usually invades vulnerable hosts. Disseminated NTM (dNTM) infection can affect nearly all organs and be easily misdiagnosed as metastatic carcinoma or other systemic diseases, especially in seemingly immunocompetent hosts. Identification of underlying immunodeficiency is critical for the diagnosis and treatment of dNTM. Adult-onset immunodeficiency (AOID) with anti-IFN-γ autoantibodies has recently been recognized as a crucial but frequently neglected risk factor for dNTM infection. Frequent relapses of infection are common in AOID patients despite appropriate anti-infective treatment and B-cell-depleting therapy has shown some promising results. Herein, we report a case of dNTM infection mimicking malignancy in an AOID patient who was successfully treated with rituximab.Case presentationA middle-aged male presented with fever, productive cough, multifocal skin abscesses and multiple osteolytic lesions with pathological fractures. Chest CT revealed consolidation of the lingula while bronchoscopy showed a mass completely blocking the airway opening of the inferior lingual segment. Metagenomic next-generation sequencing and mycobacterial culture of skin pus and bronchoalveolar lavage fluid reported Mycobacterium Colombiense, confirming the diagnosis of dNTM infection. However, anti-NTM antibiotics alone failed to prevent disease relapse and progression. Further evaluation indicated undetectable serum IFN-γ concentration and high-titer autoantibodies against IFN-γ, suggesting that AOID was the underlying reason for dNTM. Rituximab was added to treatment and successfully controlled the infection without relapse at one-year follow-up.ConclusionWe reported a rare case of disseminated Mycobacterium Colombiense infection manifested with pulmonary mass, pathological fracture and dermapostasis in a host with AOID. Our case demonstrated that AOID should be screened when patients get the episode of disseminated NTM infection particularly when other risk factors are excluded. Besides prolonged anti-NTM therapy, AOID-associated NTM infection should be treated with B-cell-depleting therapy to prevent recurrence.

Identification and drug susceptibility testing of the subspecies of Mycobacterium avium complex clinical isolates in mainland China

ObjectivesThe Mycobacterium avium complex (MAC), comprising a series of subspecies, has a worldwide distribution, with differences in drug susceptibility among subspecies. This study aimed to assess the composition of MAC and susceptibility differences among subspecies in mainland China. MethodsA total of 287 MAC clinical strains were included in the study. Multitarget sequences were applied to accurately identify subspecies, and a microdilution method was used to evaluate minimum inhibitory concentrations (MICs) among subspecies using Sensititre SLOMYCO plates. ResultsMycobacterium intracellular (N = 169), Mycobacterium avium (N = 52), Mycobacterium chimaera (N = 22), Mycobacterium marseillense (N = 25), Mycobacterium colombiense (N = 14), Mycobacterium yongonense (N = 4), Mycobacterium vulneris (N = 3) and Mycobacterium timonense (N = 2) were isolated from MAC. Clarithromycin, amikacin and rifabutin showed lower MIC50 and MIC90 values than other drugs, and the resistance rates of clarithromycin, amikacin, linezolid and moxifloxacin were 6.3%, 10.5%, 51.9% and 46.3%, respectively. The resistance rates of clarithromycin and moxifloxacin in the initial treatment group were significantly lower than those in the retreatment group (4.09% vs. 12.94%; 30.41% vs. 75.29%; P < 0.05). Drug susceptibility differences were observed in clarithromycin and moxifloxacin among the five major subspecies (P < 0.05); however, those statistically significant differences disappeared when MACs were divided into two groups according to previous anti-tuberculosis (anti-TB) treatment history. ConclusionThis study revealed that MAC, primarily comprising M. intracellulare, was susceptible to clarithromycin, amikacin and rifabutin. Drug susceptibility among subspecies did not exhibit intrinsic differences in our study. Previous anti-TB treatment patients are more resistant to drugs; thus, attention should be given to those patients in the clinic.

Mycobacterium senriense sp. nov., a slowly growing, non-scotochromogenic species, isolated from sputum of an elderly man.

A slowly growing mycobacteria, identified as strain TY59T, was isolated from sputum of an elderly man with pneumonia. Sequencing of the 16S rRNA gene indicated that this strain was similar to members of the Mycobacterium avium complex and closely related species. Strain TY59T has highest 16S rRNA gene sequence similarities to the type strains of Mycobacterium colombiense (99.80 % sequence similarity), Mycobacterium vulneris (99.74 %), Mycobacterium timonense (99.54 %), Mycobacterium avium subsp. avium (99.54 %) and Mycobacterium avium subsp. silvaticum (99.54 %). Analysis of the internal transcribed spacer (ITS) and DNA-directed RNA polymerase subunit beta (rpoB) sequences gave similar results to the 16S rRNA gene analysis. The closest species to strain TY59T were M. colombiense and M. vulneris with 97.90-98.25 % identity in ITS and 96.4-96.6 % in rpoB. The strain's 65 kDa heat shock protein (hsp65) gene was different from those of M. vulneris, M. colombiense and M. avium subsp. silvaticum with 72.4-74.2 % identity. Average nucleotide identity results showed a 93.4 % match to M. vulneris as the maximum value. Phenotypically, the non-chromogenicity, rough colonies, growth at 42 °C, negative results for nitrate reduction, β-glucosidase and Tween 80 hydrolysis, and positive results for catalase activity set this strain apart from closely related species. We propose that Mycobacterium senriense sp. nov. is a novel species of slowly growing mycobacteria. The type strain is TY59T (RIMD 1371001T=CIP 111917T).

In silico analysis of promoter regions to identify regulatory elements in TetR family transcriptional regulatory genes of Mycobacterium colombiense CECT 3035

BackgroundMycobacterium colombiense is an acid-fast, non-motile, rod-shaped mycobacterium confirmed to cause respiratory disease and disseminated infection in immune-compromised patients, and lymphadenopathy in immune-competent children. It has virulence mechanisms that allow them to adapt, survive, replicate, and produce diseases in the host. To tackle the diseases caused by M. colombiense, understanding of the regulation mechanisms of its genes is important. This paper, therefore, analyzes transcription start sites, promoter regions, motifs, transcription factors, and CpG islands in TetR family transcriptional regulatory (TFTR) genes of M. colombiense CECT 3035 using neural network promoter prediction, MEME, TOMTOM algorithms, and evolutionary analysis with the help of MEGA-X. ResultsThe analysis of 22 protein coding TFTR genes of M. colombiense CECT 3035 showed that 86.36% and 13.64% of the gene sequences had one and two TSSs, respectively. Using MEME, we identified five motifs (MTF1, MTF2, MTF3, MTF4, and MTF5) and MTF1 was revealed as the common promoter motif for 100% TFTR genes of M. colombiense CECT 3035 which may serve as binding site for transcription factors that shared a minimum homology of 95.45%. MTF1 was compared to the registered prokaryotic motifs and found to match with 15 of them. MTF1 serves as the binding site mainly for AraC, LexA, and Bacterial histone-like protein families. Other protein families such as MATP, RR, σ-70 factor, TetR, LytTR, LuxR, and NAP also appear to be the binding candidates for MTF1. These families are known to have functions in virulence mechanisms, metabolism, quorum sensing, cell division, and antibiotic resistance. Furthermore, it was found that TFTR genes of M. colombiense CECT 3035 have many CpG islands with several fragments in their CpG islands. Molecular evolutionary genetic analysis showed close relationship among the genes. ConclusionWe believe these findings will provide a better understanding of the regulation of TFTR genes in M. colombiense CECT 3035 involved in vital processes such as cell division, pathogenesis, and drug resistance and are likely to provide insights for drug development important to tackle the diseases caused by this mycobacterium. We believe this is the first report of in silico analyses of the transcriptional regulation of M. colombiense TFTR genes.

Evidence of an environmental reservoir for emergent Mycobacterium colombiense.

Mycobacterium colombiense, which belongs to the M. avium complex, is reported to have been isolated from cases of disseminated infection in both immunocompromised and immunocompetent patients. During the isolation of protists from water samples in French Guyana, we co-isolated a flagellated green alga (Polytoma sp.) and a mycobacterium identified as M. colombiense.

Investigation of non-tuberculous mycobacteria in a primary hospital from southeastern China.

Non-tuberculous mycobacterium (NTM) can colonize the human body, leading to opportunistic infection. This study was conducted to analyze the NTM species composition in a primary hospital and investigate the potential features of the patients with different NTM species. Mycobacterial strains were collected from the patients admitted at the hospital from January 2016 to May 2019. MPB64 assay was used to screen NTM strains and confirmed by Rv0577 amplification. The species were identified by hsp65 sequencing. The clinical records of patients with NTM were retrospectively reviewed. Among the 122 identified NTM isolates, the most common strains were Mycobacterium avium complex (MAC, n = 102, 83.6%), Mycobacterium abscessus (n = 9, 7.4%) and Mycobacterium lentiflavum (n = 5, 4.1%). The predominant species among MAC were Mycobacterium chimaera (n = 57, 46.7%), followed by Mycobacterium intracellulare (n = 25, 20.5%) and Mycobacterium colombiense (n = 17, 13.9%). A significantly lower percentage of positive acid-fast assay was observed in Mycobacterium colombiense positive patients than in those with Mycobacterium intracellulare and Mycobacterium chimaera. Mycobacterium intracellulare was more frequently isolated in patients from the infectious department than in other MAC members. A predominant prevalence of Mycobacterium chimaera in Dongyang of Zhejiang Province was different from other regions in China, indicating that its prevalence has been likely underestimated. The heterogeneity in clinical features, caused by different MAC members, required an accurate species identification of the NTM isolated in the primary hospitals.

A definition of the Mycobacterium avium complex for taxonomical and clinical purposes, a review.

Nontuberculous mycobacteria, particularly the Mycobacterium avium complex (MAC) bacteria, are increasingly recognized as opportunistic pathogens of humans. As a result, studies on antibiotic treatment and taxonomy of the MAC are intensifying, but an updated definition of what constitutes the MAC, either for taxonomical studies or for clinical purposes, is lacking. On the basis of literature review and phylogenetic analyses, we propose to define the MAC as a grouping of slow-growing mycobacteria that show corresponding values in at least two of the following targets against either M. avium ATCC 25291T or Mycobacterium intracellulare ATCC 13950T: >99.4 % sequence identity for the full 16S rRNA gene, >98.7 % for the partial (5') 16S rRNA gene, >97.3 % for hsp65 and >94.4 % for rpoB region V. A >97.5 % value in concatenated analyses of >2500 bp that includes 16S rRNA, hsp65 and rpoB gene sequence data or ≥85 % average nucleotide identity to M. avium ATCC 25291T or M. intracellulare ATCC 13950T on basis of whole genome sequencing data is recommended. This molecular definition is based on the distances observed between the classical members of the MAC, M. avium and M. intracellulare. Applying this definition, the complex currently consists of 12 validly published species: Mycobacterium avium, Mycobacterium intracellulare, Mycobacterium chimaera, Mycobacterium colombiense, Mycobacterium arosiense, Mycobacterium vulneris, Mycobacterium bouchedurhonense, Mycobacterium timonense, Mycobacterium marseillense, Mycobacterium yongonense, Mycobacterium paraintracellulare and Mycobacterium lepraemurium.

Draft Genome Sequences of Mycolicibacter senuensis Isolate GF74 and Mycobacterium colombiense Isolates GF28 and GF76 from a Swine Farm in Japan.

Several nontuberculous mycobacteria (NTM) occasionally infect humans and animals. Here, we report the draft genome sequences of Mycolicibacter senuensis isolate GF74 (4,792,997 bp) and Mycobacterium colombiense isolates GF28 and GF76 (5,473,554 bp and 5,426,852 bp, respectively) isolated from a swine farm in Japan. These sequences provide further information on NTM research.

Differential drug susceptibility patterns of Mycobacterium chimaera and other members of the Mycobacterium avium-intracellulare complex

ObjectivesTo determine MIC distributions for Mycobacterium chimaera, Mycobacterium intracellulare, Mycobacterium colombiense and Mycobacterium avium, and to derive tentative epidemiological cut-off (ECOFF) values. MethodsA total of 683 bacterial isolates (M. chimaera, n = 203; M. intracellulare, n = 77; M. colombiense, n = 68; M. avium, n = 335) from 627 patients were tested by broth microdilution according to CLSI protocol M24-A2 on Sensititre RAPMYCOI plates. MICs were interpreted based on CLSI breakpoints for clarithromycin, and tentative breakpoints for amikacin, moxifloxacin and linezolid. Tentative ECOFFs were determined by visual approximation and the ECOFFinder algorithm. ResultsModal MIC, MIC50 and MIC90 values were within ± one dilution step from the respective aggregated data set for 47/48 (97.9%), 48/48 (100%) and 48/48 (100%) species–drug combinations. Clarithromycin wild-type populations were mostly classified as susceptible (MIC90 4–8 mg/L; S ≤8 mg/L). Rifabutin MICs were lower than those of rifampicin. Tentative moxifloxacin, linezolid and amikacin breakpoints split wild-type populations. No ECOFFs could be set for rifampicin, ethambutol, ciprofloxacin, isoniazid, trimethoprim/sulfamethoxazole and doxycycline because of truncation of MIC distributions. Agreement between the visually determined and the modelled 97.5% ECOFFs was 90.9%. All 99.0% ECOFFs were one titre step higher than by visual approximation. ConclusionsDrug susceptibility patterns of M. chimaera are comparable to those of closely related species. Except for clarithromycin, breakpoints for Mycobacterium avium-intracellulare complex should be re-evaluated. Statistical determination of the 99.0% ECOFF may be superior to visual approximation.

Clinical features of patients with bacteraemia caused by Mycobacterium avium complex species and antimicrobial susceptibility of the isolates at a medical centre in Taiwan, 2008–2014

Advanced molecular typing methods have greatly expanded the taxonomy of Mycobacterium avium complex (MAC) species; however, little is known about the epidemiology and clinical features of bacteraemia caused by different MAC species. In this study, the clinical characteristics of patients treated for MAC bacteraemia in a tertiary-care centre in northern Taiwan during 2008–2014 were investigated. Isolates were identified to species level by rpoB gene and 16S–23S rRNA internal transcribed spacer region sequencing. Among 30 patients with bacteraemia due to MAC, the majority (n = 26) had concomitant human immunodeficiency virus (HIV) infection. Of the 30 blood isolates obtained from patients, 24 were M. avium subsp. hominissuis, 4 were Mycobacterium colombiense and 2 were Mycobacterium intracellulare. All four M. colombiense isolates were from HIV-infected patients. Bacteraemia due to M. colombiense was associated with higher 30-day mortality than bacteraemia due to M. avium subsp. hominissuis [2/4 (50%) vs. 1/24 (4%); P = 0.045, Fisher's exact test]. All four M. colombiense isolates were susceptible to clarithromycin, moxifloxacin and linezolid. Among the five patients who received ethambutol treatment and four patients who received fluoroquinolone treatment for various durations between positive MAC cultures, two and three patients, respectively, had isolates with significantly increased (≥4-fold) ethambutol and fluoroquinolone minimum inhibitory concentrations. M. colombiense was the second leading causative pathogen of MAC bacteraemia, comprising 15% of all MAC isolates obtained from HIV-positive patients. Monitoring the susceptibility of MAC isolates to ethambutol and fluoroquinolones is warranted in patients with persistent MAC bacteraemia.

Isolation and characterization of nontuberculous mycobacteria from patients with pulmonary tuberculosis in Ghana.

Nontuberculous mycobacterial (NTM) species are assuming public health importance in pulmonary diseases; they are increasingly being isolated, and importantly, most NTMs do not respond to routine tuberculosis (TB) drugs. This study aimed to identify NTMs isolated from pulmonary TB cases and also determine their susceptibility to streptomycin (STR), isoniazid (INH), and rifampicin (RIF). A total of 1755 mycobacterial isolates, obtained between August 2012 and July 2014, from 2036 smear-positive pulmonary cases were identified using polymerase chain reaction amplification of IS6110, and hsp65 gene sequencing analysis. Drug susceptibility testing (DST) was then performed for the identified NTMs against STR, INH, and RIF using microplate Alamar blue assay. The results were analyzed against patients' biodata for statistical associations. Of the 1755 analyzed isolates, we identified 43 (2.5%) NTMs, which included 18 (41.9%) Mycobacterium intracellulare, 13 (30.2%) Mycobacterium avium subs. paratuberculosis, 5 (11.3%) Mycobacterium abscessus, 3 (7.0%) each of Mycobacterium mucogenicum and Mycobacterium colombiense, and 1 (2.3%) Mycobacterium simiae. Patients infected with NTMs (52.0%) were more likely to be human immunodeficiency virus-positive (P = 0.001, odds ratio = 6.6, 95% confidence interval = 2.7-16.2) than those infected with M. tuberculosis complex (5.8%). All the 43 (100%) NTMs were resistant to INH, whereas 32 (74%) and 19 (44%) were resistant to RIF and STR, respectively. Furthermore, 16 (37.2%) NTMs were resistant to all three drugs, 20 were resistant to INH and RIF, and 3 were resistant to STR and INH. All the M. abscessus isolates were resistant to all the three drugs, whereas all the M. avium isolates were resistant to INH and RIF, but only three were resistant to STR. Among the M. intracellulare isolates, 8, 18, and 15 isolates were resistant to STR, INH, and RIF, respectively. The observed high-resistance level to INH and RIF supports the need for rapid species identification and DST of nonresponding TB cases before retreatment.

Deciphering the virulence factors of the opportunistic pathogen Mycobacterium colombiense.

Mycobacterium avium complex (MAC) contains clinically important nontuberculous mycobacteria worldwide and is the second largest medical complex in the Mycobacterium genus after the Mycobacterium tuberculosis complex. MAC comprises several species that are closely phylogenetically related but diverse regarding their host preference, course of disease, virulence and immune response. In this study we provided immunologic and virulence-related insights into the M. colombiense genome as a model of an opportunistic pathogen in the MAC. By using bioinformatic tools we found that M. colombiense has deletions in the genes involved in p-HBA/PDIM/PGL, PLC, SL-1 and HspX production, and loss of the ESX-1 locus. This information not only sheds light on our understanding the virulence mechanisms used by opportunistic MAC pathogens but also has great potential for the designing of species-specific diagnostic tools.

A fatal case of pulmonary infection by Mycobacterium colombiense in Para State, Amazon Region, Brazil

Mycobacterium avium complex (MAC) is a heterogeneous group of species found in several environmental sources and that exhibit variable degrees of pathogenicity. Among the MAC members, Mycobacterium colombiense has been related to pulmonary disease and disseminated infection in HIV-infected patients in Colombia. Lymphadenopathy cases have also been reported. We have described a fatal case of M. colombiense pulmonary disease in a Brazilian patient without evidence of HIV infection or other known causes of immunosuppression.

P1-34-3 - Disseminated Cutaneous Infection of Mycobacterium Colombiense with Myelodyspalstic Syndrome: a Case Report

Mycobacterium colombiense(M.colomboense) belongs to the Mycobacterium avium complex(MAC). It was reported to cause the opportunistic infection of immunocompromised hosts including hematological malignancy. Only a few cases of M. colombiense infection were reported all over the world. This is the first case report of a M. colombiense infection in the immunocompromised host with myelodysplastic syndrome (MDS) in Japan.Case: An 80-years-old man treated for Hypertension and Diabetes mellitus. He developed skin rash with hard exfoliation and eruption on his face and all over his head. The skin biopsy was performed, but initially mycobacterium was not detected. He repeatedly developed systemic inflammation derived from opportunistic infection including pleurapneumonia, so we considered the possibility of immunodeficiency and performed bone marrow aspiration and diagnosed as MDS(RA) with dysfunction of neutrophil. He developed the disseminated impetigos in his limbs and body, and MAC was detected in his pus. The MAC was specified as M.colombiense through Broad-range polymerase chain reaction(PCR). We prescribed the combination of antibiotics, consisting of rifampicin (450mg), ethambutol (750mg), and clarithromycin (800mg) in accordance with nontuberculous mycobacteriosis, and the skin lesions gradually resolved.In conclusion, we reported the first case of M.colombiense infection for immunocomplomised host in Japan and we could diagnose it through Broad-range PCR. The molecular diagnostic methods like PCR can improve the process of identification of the less common bacterial species that may cause persistent opportunistic infections.

Disseminated Cutaneous Infection Caused by Mycobacterium colombiense

We wish to report a case of an apparently immunocompetent 17-year-old boy with disseminated cutaneous infection caused by M. colombiense. The patient presented with deep asymptomatic erythematous nodules on his face and extremities for 4 months without any preceding history of trauma or surgery. Some of the nodules on his extremities developed into discharging ulcers, others featured slightly scaly erythematous infiltrated plaques (Fig. 1). Two months earlier, he was admitted to a local hospital on account of irregular bowel movements and an occasional slight pain in the right lower quadrant. He was diagnosed with intestinal tuberculosis according to clinical features and biopsy of the intestinal mucosa, which indicated infectious granuloma. The patient received anti-tuberculosis chemotherapy for 2 weeks but ceased taking the drugs because of severe drug-induced liver hepatitis. The past medical history of the patient only revealed chronic hepatitis B with normal liver function. Routine blood test showed mild anaemia with haemoglobin of 101 g/l. Blood culture, purified protein derivative and human immunodeficiency virus antibody tests were negative. Systemic examinations of chest X-ray, type-B ultrasonography and CT scan indicated no abnormal lesions. A skin biopsy of a nodule on the patient’s left leg displayed deep dermis showing nodular lesions with clear boundaries, collagen oedema, mucinous degeneration, blood vessel hyperplasia, infiltrated histiocytes and the absence of caseation necrosis, without special manifestation. Moderate growth of smooth, creamy, yolk yellow colonies was observed after 3 weeks of incubation only at 32 and 37°C on Lowenstein–Jensen (L–J) medium (Fig. 2A). Ziehl–Neelsen staining confirmed that the cultured organisms were acid-fast bacilli (AFB) (Fig. 2B). Pigment production testing showed that the isolated AFB was non-chromogenic. Fungal and other standard bacterial cultures were negative. Sequence analysis of 16S rDNA genes showed 99% similarity with M. colombiense strain CIP108962, and hsp65 genes indicated 99% homology with M. colombiense strain InDRE 9m. Gene sequences were analysed using the BLAST V2.0 software available at http://www.ncbi.nlm.nih.gov/ BLAST/. Gene sequencing results indicated that the strain was most similar to M. colombiense. As indicated by drug susceptibility analysis, we administered an oral regimen of clarithromycin (500 mg/ day), rifabutin (750 mg/day) and moxifloxacin (300 mg/day). After 3 months of therapy, the patient’s skin lesions greatly improved. However, his bowel symptoms did not completely improve and he continues to receive antibiotics. Disseminated Cutaneous Infection Caused by Mycobacterium colombiense

Pushing the equilibrium of regio-complementary carboxylation of phenols and hydroxystyrene derivatives

The enzymatic carboxylation of electron-rich aromatics, which represents a promising ‘green’ equivalent to the chemical Kolbe–Schmitt reaction, is thermodynamically disfavored and is therefore impeded by incomplete conversions. Optimization of the reaction conditions, such as pH, temperature, substrate concentration and the use of organic co-solvents and/or ionic liquids allowed to push the conversion in favor of carboxylation by a factor of up to 50%. Careful selection of the type of bicarbonate salt used as CO2 source was crucial to ensure optimal activities. Among two types of carboxylases tested with their natural substrates, benzoic acid decarboxylase from Rhizobium sp. proved to be significantly more stable than phenolic acid decarboxylase from Mycobacterium colombiense; it tolerated reaction temperatures of up to 50°C and substrate concentrations of up to 100mM and allowed efficient biocatalyst recycling.

Identification of potential biomarkers to distinguish Mycobacterium colombiense from other mycobacterial species

Mycobacterium avium complex (MAC) consists of 9 species of slow-growing mycobacteria with differing degrees of pathogenicity, host preference and environmental distribution. Mycobacterium colombiense is a novel member of MAC that is responsible for disseminated infections in HIV-infected patients in Colombia and lymphadenopathy cases in Europe. At present, methods to easily differentiate novel members of MAC are lacking. In this study, we identified possible biomarkers that are potentially useful for the detection of M. colombiense by PCR or chromatography. The Randomly Amplified Polymorphic DNA (RAPD) technique was used to amplify genomic fragments of M. colombiense CECT 3035 that were subsequently used in the development of a direct colony-specific PCR assay using specific primers. The designed primers amplified a 634-bp fragment of DNA from M. colombiense, which included a 450-bp genomic region that encodes a hypothetical protein of 149 amino acids that is exclusive to M. colombiense. Bioinformatic analyses revealed that this hypothetical protein had no signal peptide, active sites or functional domains to aid its identification or classification. In addition, using thin-layer chromatography, we identified a different profile of mycolates for M. colombiense strains. The test developed in this study has potential applications in the routine identification of M. colombiense and in molecular assays designed for the surveillance of MAC strains.