Emergence of Canonical and Noncanonical Genomic Variants following In Vitro Exposure of Clinical Mycobacterium tuberculosis Strains to Bedaquiline or Clofazimine.

In Mycobacterium tuberculosis, bedaquiline and clofazimine resistance occurs primarily through Rv0678 variants, a gene encoding a repressor protein that regulates mmpS5/mmpL5 efflux pump gene expression. Despite the shared effect of both drugs on efflux, little else is known about other pathways affected. We hypothesized that in vitro generation of bedaquiline- or clofazimine-resistant mutants could provide insight into additional mechanisms of action. We performed whole-genome sequencing and determined phenotypic MICs for both drugs on progenitor and mutant progenies. Mutants were induced through serial passage on increasing concentrations of bedaquiline or clofazimine. Rv0678 variants were identified in both clofazimine- and bedaquiline-resistant mutants, with concurrent atpE SNPs occurring in the latter. Of concern was the acquisition of variants in the F420 biosynthesis pathway in clofazimine-resistant mutants obtained from either a fully susceptible (fbiD: del555GCT) or rifampicin mono-resistant (fbiA: 283delTG and T862C) progenitor. The acquisition of these variants possibly implicates a shared pathway between clofazimine and nitroimidazoles. Pathways associated with drug tolerance and persistence, F420 biosynthesis, glycerol uptake and metabolism, efflux, and NADH homeostasis appear to be affected following exposure to these drugs. Shared genes affected by both drugs include Rv0678, glpK, nuoG, and uvrD1. Genes with variants in the bedaquiline resistant mutants included atpE, fadE28, truA, mmpL5, glnH, and pks8, while clofazimine-resistant mutants displayed ppsD, fbiA, fbiD, mutT3, fadE18, Rv0988, and Rv2082 variants. These results show the importance of epistatic mechanisms as a means of responding to drug pressure and highlight the complexity of resistance acquisition in M. tuberculosis.

Diagnostic performance of dobutamine stress echocardiography: A South African experience.

BACKGROUND: Dobutamine stress echocardiography (DSE) is a well-established modality for the diagnosis of coronary artery disease, but there are no reported diagnostic data in southern Africa. Objectives. To compare the safety, sensitivity and specificity of a South African (SA) DSE programme with larger, international series. Methods. All patients undergoing DSE from 2019 to 2021 at a single SA centre were included. A new wall motion abnormality (≥2 segments) signified inducible ischaemia. Results. A total of 106 patients (mean (standard deviation) age 61 (11) years, 68% male) were analysed. Six patients (6%) experienced chest pain during DSE and 4 (4%) developed an atrial arrhythmia. The sensitivity and specificity for epicardial coronary stenosis were 77% and 74%, respectively, changing to 82% and 72% when excluding those who had previous coronary artery bypass surgery. Conclusion. The sensitivity, specificity and safety of an SA DSE programme were comparable to international series. A DSE programme is feasible in a resource-constrained environment.

Percutaneous left atrial appendage occlusion: A South African experience.

BACKGROUND: Atrial fibrillation (AF) is associated with all-cause mortality, heart failure and non-fatal stroke, and thromboprophylaxis is traditionally provided with oral anticoagulants (OACs). Percutaneous left atrial appendage occlusion (LAAO) with a dedicated device is an alternative approach to thromboprophylaxis in patients with AF who are: (i) intolerant to OACs (e.g. life-threatening haemorrhage); (ii) non-adherent to OACs; or (iii) at a high bleeding risk with OACs. Non-inferiority of LAAO compared with OACs was demonstrated in e.g. the WATCHMAN Left Atrial Appendage System for Embolic Protection in Patients With Atrial Fibrillation (PROTECT AF) trial. Only very limited data are available on percutaneous LAAO in South Africa (SA), and no local outcome data have been reported. OBJECTIVES: To compare the safety and efficacy outcomes of an SA percutaneous LAAO programme with larger international series. METHODS: All patients undergoing percutaneous LAAO from 2013 to 2020 at a single centre (SAEndovascular, Kuils River Netcare Hospital, SA) were included from an ongoing registry. Survival analysis was performed with the Kaplan-Meier method. RESULTS: Of 101 LAAO recipients (mean (standard deviation) age 77 (10) years, 64% male) analysed, 90 (90%) had permanent AF, 1 (1%) persistent AF and 9 (9%) paroxysmal AF. The most common indication for LAAO was previous severe bleeding (n=23; 23%). The mean device size was 23 (3) mm and the procedural success rate was 98%. After a median (interquartile range) follow-up of 21 (5 - 41) months, 6 patients (6%) experienced stroke or all-cause mortality. Four patients (4%) had a life-threatening procedural complication (tamponade n=2 (2%) and device embolisation n=2 (2%)). These outcomes are comparable to large international series, e.g. PROTECT AF. CONCLUSIONS: The safety and efficacy outcomes of an SA percutaneous LAAO programme were comparable to large international series. A successful percutaneous LAAO programme is feasible in a southern African context.

Mechanically compliant single crystals of a stable organic radical.

Mechanically compliant organic crystals are the foundation of the development of future flexible, light-weight single-crystal electronics, and this requires reversibly deformable crystalline organic materials with permanent magnetism. Here, we report and characterize the first instance of a plastically bendable single crystal of a permanent organic radical, 4-(4'-cyano-2',3',4',5'-tetrafluorophenyl)-1,2,3,5-dithiadiazolyl. The weak interactions between the radicals render single crystals of the ß phase of this material exceedingly soft, and the S-N interactions facilitate plastic bending. EPR imaging of a bent single crystal reveals the effect of deformation on the three-dimensional spin density of the crystal. The unusual mechanical compliance of this material opens prospects for exploration into flexible crystals of other stable organic radicals towards the development of flexible light-weight organic magnetoresistance devices based on weak, non-hydrogen-bonded interactions in molecular crystals.

Whole genome sequencing reveals genomic heterogeneity and antibiotic purification in Mycobacterium tuberculosis isolates.

BACKGROUND: Whole genome sequencing has revolutionised the interrogation of mycobacterial genomes. Recent studies have reported conflicting findings on the genomic stability of Mycobacterium tuberculosis during the evolution of drug resistance. In an age where whole genome sequencing is increasingly relied upon for defining the structure of bacterial genomes, it is important to investigate the reliability of next generation sequencing to identify clonal variants present in a minor percentage of the population. This study aimed to define a reliable cut-off for identification of low frequency sequence variants and to subsequently investigate genetic heterogeneity and the evolution of drug resistance in M. tuberculosis. METHODS: Genomic DNA was isolated from single colonies from 14 rifampicin mono-resistant M. tuberculosis isolates, as well as the primary cultures and follow up MDR cultures from two of these patients. The whole genomes of the M. tuberculosis isolates were sequenced using either the Illumina MiSeq or Illumina HiSeq platforms. Sequences were analysed with an in-house pipeline. RESULTS: Using next-generation sequencing in combination with Sanger sequencing and statistical analysis we defined a read frequency cut-off of 30% to identify low frequency M. tuberculosis variants with high confidence. Using this cut-off we demonstrated a high rate of genetic diversity between single colonies isolated from one population, showing that by using the current sequencing technology, single colonies are not a true reflection of the genetic diversity within a whole population and vice versa. We further showed that numerous heterogeneous variants emerge and then disappear during the evolution of isoniazid resistance within individual patients. Our findings allowed us to formulate a model for the selective bottleneck which occurs during the course of infection, acting as a genomic purification event. CONCLUSIONS: Our study demonstrated true levels of genetic diversity within an M. tuberculosis population and showed that genetic diversity may be re-defined when a selective pressure, such as drug exposure, is imposed on M. tuberculosis populations during the course of infection. This suggests that the genome of M. tuberculosis is more dynamic than previously thought, suggesting preparedness to respond to a changing environment.

Interferon-gamma release assays for the detection of Mycobacterium tuberculosis infection in children with cancer.

SETTING: The paediatric oncology unit at Tygerberg Children's Hospital, South Africa. OBJECTIVES: To assess the use of the tuberculin skin test (TST) and two commercial interferon-gamma release assays (IGRAs) for the detection of Mycobacterium tuberculosis infection in children with cancer before initiating chemotherapy treatment. DESIGN: Prospective hospital-based study, including children newly diagnosed with cancer; all underwent TST and IGRA testing. RESULTS: Of the 34 children enrolled, seven (17.6%) tested positive with either test: TST (3/7, 8.8%), T-SPOT.TB (n = 6, 17.6%) and QuantiFERON-TB Gold In-Tube (QFT-G; n = 3, 8.8%). T-SPOT.TB assay results were negative in 17 (50.0%) and indeterminate in four (11.8%) children. Six T-SPOT.TB tests could not be completed due to low cell counts (<100,000 per well), and one clotted. QFT-G results were negative in 26 (76.5%) and indeterminate in five (14.7%). CONCLUSIONS: TST and IGRAs were frequently discordant, with fewer positive results than expected. T-SPOT.TB produced more positive results, but inadequate cell counts were a particular problem. The sample size was too small to comment with confidence on test accuracy. All latent TB infection tests appear to perform sub-optimally in this group of children, and therefore none of them can be used in isolation to confirm or disprove TB infection.

Cancer nutrition and rehabilitation-its time has come!

Cancer is a systemic disease that can affect nearly every organ in the body, resulting in a progressive loss of organ function. That loss of function may be initially slow, having minimal effect, or it may be rapid, resulting in more dramatic changes.The usual medical management of patients with cancer has focused more specifically on the administration of cytotoxic treatments. These treatments can potentially eradicate or minimize the tumour, but they may also have toxic side effects that in turn can also affect the patient.Cancer rehabilitation is a process that assists the individual with a cancer diagnosis to obtain optimal physical, social, psychological, and vocational functioning within the limits created by the disease and its treatment. The McGill Cancer Nutrition and Rehabilitation (CNR) program developed as a result of the ever-increasing demand for a focus on addressing individual cancer patients and their needs, as well as on achieving optimal tumour-related outcomes. Using an interdisciplinary approach, the CNR's global objective is to empower individuals who are experiencing loss of function, fatigue, malnutrition, psychological distress, and other symptoms as a result of cancer or its treatment to improve their own quality of life. All team members-experts in their respective fields-assess all patients. At a subsequent team discussion and planning meeting, a specific 8-week program is designed for each patient. The hoped-for outcome for the CNR program is primarily to empower patients to "take control" or to enable them to improve their own quality of life. This article reviews the philosophy of the CNR's approach and the roles played by the various members of the team.

Improved outcome in South African children of mixed ethnicity treated for all.

A historical cohort study with an analytical component was conducted to determine whether risk-appropriate chemotherapy can improve survival in children of mixed ethnicity with ALL. Eighty-one coloured children treated for ALL in South Africa were divided into 2 groups: group A (n = 39), treated prior to 1992, and group B (n = 42), treated after 1992. A comparison was made of survival, treatment complications, and supportive measures. The two groups were comparable. The mean nadirs of the white cell count (p < .01), platelet count (p = .01), and hemoglobin value (p < .01) were significantly lower in group B. The survival rate of 37% in group A improved to 66% in group B (p = .025). The results show that a risk-adapted regimen increased survival in children of mixed ethnicity in the Western Cape, despite increased hematological toxicity and episodes of febrile neutropenia.