Loss-of-function mutations in ndh do not confer delamanid, ethionamide, isoniazid, or pretomanid resistance in Mycobacterium tuberculosis.

Results from clinical strains and knockouts of the H37Rv and CDC1551 laboratory strains demonstrated that ndh (Rv1854c) is not a resistance-conferring gene for isoniazid, ethionamide, delamanid, or pretomanid in Mycobacterium tuberculosis. This difference in the susceptibility to NAD-adduct-forming drugs compared with other mycobacteria may be driven by differences in the absolute intrabacterial NADH concentration.

Cross-Border Movement of Highly Drug-Resistant Mycobacterium tuberculosis from Papua New Guinea to Australia through Torres Strait Protected Zone, 2010-2015.

In this retrospective study, we used whole-genome sequencing (WGS) to delineate transmission dynamics, characterize drug-resistance markers, and identify risk factors of transmission among Papua New Guinea residents of the Torres Strait Protected Zone (TSPZ) who had tuberculosis diagnoses during 2010-2015. Of 117 isolates collected, we could acquire WGS data for 100; 79 were Beijing sublineage 2.2.1.1, which was associated with active transmission (odds ratio 6.190, 95% CI 2.221-18.077). Strains were distributed widely throughout the TSPZ. Clustering occurred more often within than between villages (p = 0.0013). Including 4 multidrug-resistant tuberculosis isolates from Australia citizens epidemiologically linked to the TSPZ into the transmission network analysis revealed 2 probable cross-border transmission events. All multidrug-resistant isolates (33/104) belonged to Beijing sublineage 2.2.1.1 and had high-level isoniazid and ethionamide co-resistance; 2 isolates were extensively drug resistant. Including WGS in regional surveillance could improve tuberculosis transmission tracking and control strategies within the TSPZ.

Rapidly growing mycobacteria associated with laparoscopic gastric banding, Australia, 2005-2011.

Laparoscopic gastric banding is a common bariatric procedure worldwide. Rapidly growing mycobacteria are environmental organisms increasingly seen as pathogens,often in infected prosthetic material. We report 18 cases of infection associated with laparoscopic gastric banding caused by Mycobacterium fortuitum and M. abscessus in Australia during 2005­2011. We identified cases by reviewing positive cultures at the Queensland state reference laboratory or through correspondence with clinicians, and we obtained clinical and epidemiologic data. Eleven cases of M. fortuitum and 7 cases of M. abscessus infection were identified. The port was thought to be the primary site of infection in 10 of these cases. Complications included peritonitis,band erosion, and chronic ulceration at the port site.Rapidly growing mycobacteria can infect both port and band and can occur as either an early perioperative or late infection.Combination antimicrobial therapy is used on the basis of in vitro susceptibilities. Device removal seems to be vital to successful therapy.

Diversity of Mycobacterium tuberculosis and drug resistance in different provinces of Papua New Guinea.

BACKGROUND: Papua New Guinea (PNG) is a high tuberculosis (TB) burden country of the WHO Western Pacific Region, but so far research on drug resistance (DR) and genotypes of Mycobacterium tuberculosis (M. tuberculosis) was only conducted in few provinces in the country. The aim of the present study was to obtain baseline data on the level of drug resistance and the genotypic diversity of circulating M. tuberculosis in additional provinces and to investigate the differences between three selected sites across PNG. RESULTS: Genotyping of 147 M. tuberculosis clinical isolates collected in Goroka, Eastern Highlands Province, in Alotau, Milne Bay Province and in Madang, Madang Province revealed three main lineages of M. tuberculosis: Lineage 4 (European-American lineage), Lineage 2 (East-Asian lineage) and Lineage 1 (Indo-Oceanic lineage). All three lineages were detected in all three sites, but the individual lineage compositions varied significantly between sites. In Madang Lineage 4 was the most prevalent lineage (76.6%), whereas in Goroka and Alotau Lineage 2 was dominating (60.5% and 84.4%, respectively) (p < 0.001). Overall, phenotypic drug susceptibility testing showed 10.8% resistance to at least one of the first-line drugs tested. Of all resistant strains (23/212) 30.4% were Streptomycin mono-resistant, 17.4% were Isoniazid mono-resistant and 13% were Rifampicin mono-resistant. Multi-drug resistant (MDR) TB was found in 2.8% of all tested cases (6/212). The highest amount of MDR TB was found in Alotau in Milne Bay Province (4.6%). CONCLUSION: A large number of drug resistant TB infections are present in the country and MDR TB has already been detected in all three surveyed regions of PNG, highlighting the importance of monitoring drug resistance and making it a high priority for the National Control Program. Due to the high prevalence of Lineage 2 in Milne Bay Province and given the frequent association of this lineage with drug resistance, monitoring of the latter should especially be scaled up in that province.

Surgical site infections following coronary artery bypass graft procedures: 10 years of surveillance data.

BACKGROUND: Surgical site infections following coronary artery bypass graft (CABG) procedures pose substantial burden on patients and healthcare systems. This study aims to describe the incidence of surgical site infections and causative pathogens following CABG surgery over the period 2003-2012, and to identify risk factors for complex sternal site infections. METHODS: Routine computerised surveillance data were collected from three public hospitals in Queensland, Australia in which CABG surgery was performed between 2003 and 2012. Surgical site infection rates were calculated by types of infection (superficial/complex) and incision sites (sternal/harvest sites). Patient and procedural characteristics were evaluated as risk factors for complex sternal site infections using a logistic regression model. RESULTS: There were 1,702 surgical site infections (518 at sternal sites and 1,184 at harvest sites) following 14,546 CABG procedures performed. Among 732 pathogens isolated, Methicillin-sensitive Staphylococcus aureus accounted for 28.3% of the isolates, Pseudomonas aeruginosa 18.3%, methicillin-resistant Staphylococcus aureus 14.6%, and Enterobacter species 6.7%. Proportions of Gram-negative bacteria elevated from 37.8% in 2003 to 61.8% in 2009, followed by a reduction to 42.4% in 2012. Crude rates of complex sternal site infections increased over the reporting period, ranging from 0.7% in 2004 to 2.6% in 2011. Two factors associated with increased risk of complex sternal site infections were identified: patients with an ASA (American Society of Anaesthesiologists) score of 4 or 5 (reference score of 3, OR 1.83, 95% CI 1.36-2.47) and absence of documentation of antibiotic prophylaxis (OR 2.03, 95% CI 1.12-3.69). CONCLUSIONS: Compared with previous studies, our data indicate the importance of Gram-negative organisms as causative agents for surgical site infections following CABG surgery. An increase in complex sternal site infection rates can be partially explained by the increasing proportion of patients with more severe underlying disease.

TB incidence and characteristics in the remote gulf province of Papua New Guinea: a prospective study.

BACKGROUND: The incidence and characteristics of tuberculosis (TB) in remote areas of Papua New Guinea (PNG) are largely unknown. The purpose of our study was to determine the incidence of TB in the Gulf Province of PNG and describe disease characteristics, co-morbidities and drug resistance profiles that could impact on disease outcomes and transmission. METHODS: Between March 2012 and June 2012, we prospectively collected data on 274 patients presenting to Kikori Hospital with a presumptive diagnosis of TB, and on hospital inpatients receiving TB treatment during the study period. Sputum was collected for microscopy, GeneXpert analysis, culture and genotyping of isolates. RESULTS: We estimate the incidence of TB in Kikori to be 1290 per 100,000 people (95% CI 1140 to 1460) in 2012. The proportion of TB patients co-infected with HIV was 1.9%. Three of 32 TB cases tested were rifampicin resistant. Typing of nine isolates demonstrated allelic diversity and most were related to Beijing strains. CONCLUSIONS: The incidence of TB in Kikori is one of the highest in the world and it is not driven by HIV co-infection. The high incidence and the presence of rifampicin resistant warrant urgent attention to mitigate substantial morbidity in the region.

Sustained transmission over two decades of a previously unrecognised MPT64 negative Mycobacterium tuberculosis strain in Queensland, Australia: a whole genome sequencing study.

Background: MPT64 is a key protein used for Mycobacterium tuberculosis (MTB) complex strain identification. We describe protracted transmission of an MPT64 negative MTB strain in Queensland, Australia, and explore genomic factors related to its successful spread. Methods: All MPT64 negative strains identified between 2002 and 2022 by the Queensland Mycobacteria Reference Laboratory, and an additional 2 isolates from New South Wales (NSW), were whole genome sequenced. Bayesian modelling and phylogeographical analyses were used to assess their evolutionary history and transmission dynamics. Protein structural modelling to understand the putative functional effects of the mutated gene coding for MPT64 protein was performed. Findings: Forty-three MPT64 negative isolates were sequenced, belonging to a single MTB cluster of Lineage 4.1.1.1 strains. Combined with a UK dataset of the same lineage, molecular dating estimated 1990 (95% HPD 1987-1993) as the likely time of strain introduction into Australia. Although the strain has spread over a wide geographic area and new cases linked to the cluster continue to arise, phylodynamic analysis suggest the outbreak peaked around 2003. All MPT64 negative strains had a frame shift mutation (delAT, p.Val216fs) within the MPT64 gene, which confers two major structural rearrangements at the C-terminus of the protein. Interpretation: This study uncovered the origins of an MPT64 negative MTB outbreak in Australia, providing a richer understanding of its biology and transmission dynamics, as well as guidance for clinical diagnosis and public health action. The potential spread of MPT64 negative strains undermines the diagnostic utility of the MPT64 immunochromatographic test. Funding: This study was funded from an operational budget provided to the Queensland Mycobacterium Reference Laboratory by Pathology Queensland, Queensland Department of Health.

Management of the infant born to a mother with tuberculosis: a systematic review and consensus practice guideline.

Infants born to mothers with tuberculosis disease are at increased risk of developing tuberculosis disease themselves. We reviewed published studies and guidelines on the management of these infants to inform the development of a consensus practice guideline. We searched MEDLINE, CINAHL, and Cochrane Library from database inception to Dec 1, 2022, for original studies reporting the management and outcome of infants born to mothers with tuberculosis. Of the 521 published papers identified, only three met inclusion criteria and no evidence-based conclusions could be drawn from these studies, given their narrow scope, variable aims, descriptive nature, inconsistent data collection, and high attrition rates. We also assessed a collection of national and international guidelines to inform a consensus practice guideline developed by an international panel of experts from different epidemiological contexts. The 16 guidelines reviewed had consistent features to inform the expert consultation process. Two management algorithms were developed-one for infants born to mothers considered potentially infectious at the time of delivery and another for mothers not considered infectious at the time of delivery-with different guidance for high and low tuberculosis incidence settings. This systematic review and consensus practice guideline should facilitate more consistent clinical management, support the collection of better data, and encourage the development of more studies to improve evidence-based care.

A case series of medically managed infective endocarditis after transcatheter aortic valve replacement.

We report a series of 4 cases of transcatheter aortic valve replacement infective endocarditis from our institution. Atypical diagnostic features were noted echocardiographically. Infection with enterococcal species was prevalent and a coexisting mitral valve vegetation was found in 2 patients. All 4 patients had excellent responses to intravenous antibiotics.

A mariner's tale: Invasive endotracheal Mycobacterium marinum infection.

Mycobacterium marinum is a ubiquitous water-borne non-tuberculous mycobacterial (NTM) pathogen. In humans, M. marinum infections are acquired through direct inoculation of skin wounds and are almost exclusively localized to skin and soft tissues. Pulmonary infection with M. marinum is extremely rare, and to our knowledge, invasive endobronchial disease has not been reported. Here, we present a case of a 71-year-old immunocompetent male surfer with invasive endotracheal M. marinum granulomatous disease. The patient was successfully cured with a regimen of azithromycin 250 mg daily, ethambutol 900 mg (15 mg/kg) daily and rifampicin 600 mg daily for 12 months following culture conversion. This case highlights several important concepts: Firstly, M. marinum infection, including invasive endobronchial infection, should be considered a rare cause of NTM pulmonary disease. Secondly, endotracheal infection can be successfully eradicated with this selected therapeutic regimen. Finally, the absence of M. marinum skin or soft-tissue infection in this patient, raises the possibility that human disease might also be acquired via inhalation of M. marinum contaminated water in rare circumstances.

Drug resistance-conferring mutations in Mycobacterium tuberculosis from Madang, Papua New Guinea.

BACKGROUND: Monitoring drug resistance in Mycobacterium tuberculosis is essential to curb the spread of tuberculosis (TB). Unfortunately, drug susceptibility testing is currently not available in Papua New Guinea (PNG) and that impairs TB control in this country. We report for the first time M. tuberculosis mutations associated with resistance to first and second-line anti-TB drugs in Madang, PNG. A molecular cluster analysis was performed to identify M. tuberculosis transmission in that region. RESULTS: Phenotypic drug susceptibility tests showed 15.7% resistance to at least one drug and 5.2% multidrug resistant (MDR) TB. Rifampicin resistant strains had the rpoB mutations D516F, D516Y or S531L; Isoniazid resistant strains had the mutations katG S315T or inhA promoter C15T; Streptomycin resistant strains had the mutations rpsL K43R, K88Q, K88R), rrs A514C or gidB V77G. The molecular cluster analysis indicated evidence for transmission of resistant strain. CONCLUSIONS: We observed a substantial rate of MDR-TB in the Madang area of PNG associated with mutations in specific genes. A close monitoring of drug resistance is therefore urgently required, particularly in the presence of drug-resistant M. tuberculosis transmission. In the absence of phenotypic drug susceptibility testing in PNG, molecular assays for drug resistance monitoring would be of advantage.

Community-associated methicillin-resistant Staphylococcus aureus endocarditis 'down under': case series and literature review.

Infective endocarditis is a common complication of Staphylococcus aureus bacteraemia, but literature reports of community-associated methicillin-resistant S. aureus (CA-MRSA) endocarditis are relatively uncommon and mostly comprise intravenous drug users (IVDUs) with the USA300 strain. We report 5 cases of CA-MRSA endocarditis in previously healthy young Australian adults, 4 in IVDUs. Morbidity was high with frequent septic emboli; 3 patients required cardiac surgery and 1 patient died. Typing revealed the 2 most common Australian strains, the Panton-Valentine leukocidin (PVL)-positive ST93 (Queensland) strain and the PVL-negative ST1 (WA-MRSA-1) strain.

Blood cultures for assessment of postoperative fever in arthroplasty patients.

Postoperative fever in arthroplasty patients is common. The value of diagnostic workup of fever in this instance is of questionable utility. Studies have shown that blood cultures in this scenario add little to clinical management, but sample sizes have been small and the use of blood cultures in this setting continues. This study aimed to examine the value of blood cultures in the assessment of postoperative fever in a large arthroplasty population. The medical records of 101 patients who had 141 blood culture sets taken during a 2-year period were retrospectively analyzed. Of the 141 blood culture sets, only 2 returned positive results. These were both thought to be as a result of skin contamination at the time of venipuncture. No infectious sequelae occurred in either patient. We conclude that blood cultures have no role to play in the assessment of the febrile, otherwise asymptomatic arthroplasty patient in the early postoperative period.

Rifampin stability in 7H9 broth and Löwenstein-Jensen medium.

The objectives of this study were to monitor the stability of rifampin (RIF) in Löwenstein-Jensen medium (L-J medium) and 7H9 broth, which are the media commonly used for drug susceptibility testing (DST) of Mycobacterium tuberculosis. Rifampin degradation in stock solution, 7H9 broth, and L-J medium and during the inspissation process for L-J medium preparation was serially monitored by high-performance liquid chromatography (HPLC). L-J medium-based DST was conducted to examine the effect of L-J medium storage on the DST outcome. The RIF stock solution was stable for at least 3 months when kept at either 4°C or -20°C; RIF in 7H9 broth and L-J medium was almost 50% decayed after 1 week of storage at 37°C, and rifampin could not be detected in 7H9 or L-J medium after 3 weeks or 6 weeks of storage at 37°C. Approximately half of the drug was decomposed after 4 months of storage at 4°C for both media, and after 6 months of storage at 4°C, RIF in L-J medium was undetectable, while 38% of RIF remained in 7H9 medium. Approximately 21, 24, 29, and 35% RIF degradations were detected when the L-J medium was coagulated at 75°C, 80°C, 85°C, and 90°C, respectively. The DST outcomes when using L-J medium stored for different periods of time were consistent with the RIF concentration monitoring data. Rifampin in stock solution is stable for at least 3 months at a reduced storage temperature. Media containing RIF should be prepared strictly according to validated standard operating procedures. RIF degradation is a possible reason for false resistance categorizations of M. tuberculosis isolates in the clinical laboratory.

Characteristics of embB mutations in multidrug-resistant Mycobacterium tuberculosis isolates in Henan, China.

OBJECTIVES: To determine the association between embB mutations and drug resistance, and to further investigate the mechanism of embB mutations involved in the development of ethambutol and multidrug resistance in Mycobacterium tuberculosis. METHODS: One hundred and thirty-eight multidrug-resistant clinical M. tuberculosis isolates, including 86 ethambutol-resistant and 52 ethambutol-susceptible strains, were analysed to characterize mutations within the entire coding region of the embB gene. Moreover, a two-step genotyping was performed to identify the genetic lineage. RESULTS: In total, 27 embB mutation types were detected in 19 distinct codons. Though a strong association was observed between embB mutations and ethambutol resistance, 19.2% of embB306 mutants and 11.5% of embB406 or embB497 mutants were ethambutol susceptible. Among 39 ethambutol-resistant strains without embB306 mutations, 51.3% harboured mutations at codons 406 or 497. Particularly, three pairs of isolates with identical embB mutations and genotyping features were identified with variant ethambutol susceptibility. Among 77 isoniazid, rifampicin, streptomycin and ethambutol quadruple drug-resistant isolates, 89.6% carried embB mutations and 83.1% could be identified by detecting 10 embB mutations. CONCLUSIONS: Our results suggest embB mutations alone are not sufficient for the development of full resistance to ethambutol in M. tuberculosis and mutations other than embB are also needed. Our study confirms the importance of mutations at embB406 and embB497 as hotspots, in addition to embB306, for detecting ethambutol resistance. Ten selected mutations of embB, covered by a short PCR product, can be used as candidate markers for the prediction of quadruple resistance to isoniazid, rifampicin, streptomycin and ethambutol.

Genomic epidemiology of tuberculosis in eastern Malaysia: insights for strengthening public health responses.

Tuberculosis is a leading public health priority in eastern Malaysia. Knowledge of the genomic epidemiology of tuberculosis can help tailor public health interventions. Our aims were to determine tuberculosis genomic epidemiology and characterize resistance mutations in the ethnically diverse city of Kota Kinabalu, Sabah, located at the nexus of Malaysia, Indonesia, Philippines and Brunei. We used an archive of prospectively collected Mycobacterium tuberculosis samples paired with epidemiological data. We collected sputum and demographic data from consecutive consenting outpatients with pulmonary tuberculosis at the largest tuberculosis clinic from 2012 to 2014, and selected samples from tuberculosis inpatients from the tertiary referral centre during 2012-2014 and 2016-2017. Two hundred and eight M. tuberculosis sequences were available for analysis, representing 8 % of cases notified during the study periods. Whole-genome phylogenetic analysis demonstrated that most strains were lineage 1 (195/208, 93.8 %), with the remainder being lineages 2 (8/208, 3.8 %) or 4 (5/208, 2.4 %). Lineages or sub-lineages were not associated with patient ethnicity. The lineage 1 strains were diverse, with sub-lineage 1.2.1 being dominant (192, 98 %). Lineage 1.2.1.3 isolates were geographically most widely distributed. The greatest diversity occurred in a border town sub-district. The time to the most recent common ancestor for the three major lineage 1.2.1 clades was estimated to be the year 1966 (95 % HPD 1948-1976). An association was found between failure of culture conversion by week 8 of treatment and infection with lineage 2 (4/6, 67 %) compared with lineage 1 strains (4/83, 5 %) (P<0.001), supporting evidence of greater virulence of lineage 2 strains. Eleven potential transmission clusters (SNP difference ≤12) were identified; at least five included people living in different sub-districts. Some linked cases spanned the whole 4-year study period. One cluster involved a multidrug-resistant tuberculosis strain matching a drug-susceptible strain from 3 years earlier. Drug resistance mutations were uncommon, but revealed one phenotype-genotype mismatch in a genotypically multidrug-resistant isolate, and rare nonsense mutations within the katG gene in two isolates. Consistent with the regionally mobile population, M. tuberculosis strains in Kota Kinabalu were diverse, although several lineage 1 strains dominated and were locally well established. Transmission clusters - uncommonly identified, likely attributable to incomplete sampling - showed clustering occurring across the community, not confined to households or sub-districts. The findings indicate that public health priorities should include active case finding and early institution of tuberculosis management in mobile populations, while there is a need to upscale effective contact investigation beyond households to include other contacts within social networks.

Rapid macrolide and amikacin resistance testing for Mycobacterium abscessus in people with cystic fibrosis.

Introduction. Mycobacterium abscessus complex (MABSC) is an environmental organism and opportunistic pathogen. MABSC pulmonary infections in people with cystic fibrosis are of growing clinical concern. Resistance data guide the use of macrolides and amikacin in MABSC pulmonary disease treatment. MABSC can acquire resistance against macrolides or amikacin via 23S or 16S rRNA gene mutations, respectively.Gap Statement. Current culture-based methods for MABSC detection and antibiotic resistance characterization are typically prolonged, limiting their utility to directly inform treatment or clinical trials. Culture-independent molecular methods may help address this limitation.Aim. To develop real-time PCR assays for characterization of key 23S or 16S rRNA gene mutations associated with constitutive resistance in MABSC.Methodology. We designed two real-time PCR assays to detect the key 23S and 16S rRNA gene mutations. The highly conserved nature of rRNA genes was a major design challenge. To reduce potential cross-reactivity, primers included non-template bases and targeted single-nucleotide polymorphisms unique to MABSC. We applied these assays, as well as a previously developed real-time PCR assay for MABSC detection, to 968 respiratory samples from people with cystic fibrosis. The results from the molecular methods were compared to those for gold standard culture methods and 23S and 16S rRNA gene sequencing.Results.The real-time PCR MABSC detection assay provided a sensitivity of 83.8 % and a specificity of 97.8 % compared to culture. The results from the real-time PCR resistance detection assays were mostly concordant (>77.4 %) with cultured isolate sequencing. The real-time PCR resistance detection assays identified several samples harbouring both resistant and susceptible MABSC, while culture-dependent methods only identified susceptible MABSC in these samples.Conclusion. Using the molecular methods described here, results for health care providers or researchers could be available days or weeks earlier than is currently possible via culture-based antibiotic susceptibility testing.

Characterization of pncA mutations in multi-drug and pyrazinamide resistant Mycobacterium tuberculosis isolates cultured from Queensland migrants and Papua New Guinea residents.

Outbreak of drug resistant tuberculosis in the Western province, Papua New Guinea is a concern to Queensland, Australia due to migration. We performed pncA mutation analysis and genotyping of multi-drug/pyrazinamide (MDR/PZA) resistant isolates from 18 Queensland (Qld) migrants and 81 Papua New Guinea (PNG) residents, to compare with phenotypic evidence of PZA resistance and to evaluate the genotypes obtained from the two countries. Seven different mutations were seen from Qld isolates of which 2 have not been described previously. A cluster of mutations were found between amino acids L35 and S65. Amongst the PNG isolates, 10 mutations were identified, of which 6 were unique and have not been described previously. Majority of the mutations formed 2 clusters, between amino acids Q10 to A20 and W68 to W119. Mutations identified at nucleotide (nt) position 202 and 307 were found to be the most common types, occurring in 25% and 51% of the PNG isolates respectively. The majority of the mutations were seen in MDR/PZA resistant isolates. These mutations could be utilized for direct screening of PZA resistance from PNG patient samples. Genotypic analysis of the isolates showed strong clustering amongst the PNG isolates as opposed to Qld isolates. A diversity of mutations and genotypes were seen amongst the Qld migrant isolates. Majority of PNG isolates had one genotype with two distinct pncA mutation patterns (T202C and T307G) which highlight on-going transmission. pncA mutation analysis provided a satisfactory alternative to PZA culture DST with high positive predictive value and an improved result turnaround time.