A flavin-dependent halogenase from metagenomic analysis prefers bromination over chlorination

Neubauer P, Widmann C, Wibberg D, et al. A flavin-dependent halogenase from metagenomic analysis prefers bromination over chlorination. PLoS ONE. 2018;13(5): e0196797.Flavin-dependent halogenases catalyse halogenation of aromatic compounds. In most cases, this reaction proceeds with high regioselectivity and requires only the presence of FADH2, oxygen, and halide salts. Since marine habitats contain high concentrations of halides, organisms populating the oceans might be valuable sources of yet undiscovered halogenases. A new Hidden-Markov-Model (HMM) based on the PFAM tryptophan halogenase model was used for the analysis of marine metagenomes. Eleven metagenomes were screened leading to the identification of 254 complete or partial putative flavin-dependent halogenase genes. One predicted halogenase gene (brvH) was selected, codon optimised for E. coli, and overexpressed. Substrate screening revealed that this enzyme represents an active flavin-dependent halogenase able to convert indole to 3-bromoindole. Remarkably, bromination prevails also in a large excess of chloride. The BrvH crystal structure is very similar to that of tryptophan halogenases but reveals a substrate binding site that is open to the solvent instead of being covered by a loop

Perspectives for systems biology in the management of tuberculosis

Standardised management of tuberculosis may soon be replaced by individualised, precision medicine-guided therapies informed with knowledge provided by the field of systems biology. Systems biology is a rapidly expanding field of computational and mathematical analysis and modelling of complex biological systems that can provide insights into mechanisms underlying tuberculosis, identify novel biomarkers, and help to optimise prevention, diagnosis and treatment of disease. These advances are critically important in the context of the evolving epidemic of drug-resistant tuberculosis. Here, we review the available evidence on the role of systems biology approaches - human and mycobacterial genomics and transcriptomics, proteomics, lipidomics/metabolomics, immunophenotyping, systems pharmacology and gut microbiomes - in the management of tuberculosis including prediction of risk for disease progression, severity of mycobacterial virulence and drug resistance, adverse events, comorbidities, response to therapy and treatment outcomes. Application of the Grading of Recommendations, Assessment, Development and Evaluation (GRADE) approach demonstrated that at present most of the studies provide "very low" certainty of evidence for answering clinically relevant questions. Further studies in large prospective cohorts of patients, including randomised clinical trials, are necessary to assess the applicability of the findings in tuberculosis prevention and more efficient clinical management of patients.Publisher PDFPeer reviewe

The evolving SARS-CoV-2 epidemic in Africa: Insights from rapidly expanding genomic surveillance

INTRODUCTION Investment in Africa over the past year with regard to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) sequencing has led to a massive increase in the number of sequences, which, to date, exceeds 100,000 sequences generated to track the pandemic on the continent. These sequences have profoundly affected how public health officials in Africa have navigated the COVID-19 pandemic. RATIONALE We demonstrate how the first 100,000 SARS-CoV-2 sequences from Africa have helped monitor the epidemic on the continent, how genomic surveillance expanded over the course of the pandemic, and how we adapted our sequencing methods to deal with an evolving virus. Finally, we also examine how viral lineages have spread across the continent in a phylogeographic framework to gain insights into the underlying temporal and spatial transmission dynamics for several variants of concern (VOCs). RESULTS Our results indicate that the number of countries in Africa that can sequence the virus within their own borders is growing and that this is coupled with a shorter turnaround time from the time of sampling to sequence submission. Ongoing evolution necessitated the continual updating of primer sets, and, as a result, eight primer sets were designed in tandem with viral evolution and used to ensure effective sequencing of the virus. The pandemic unfolded through multiple waves of infection that were each driven by distinct genetic lineages, with B.1-like ancestral strains associated with the first pandemic wave of infections in 2020. Successive waves on the continent were fueled by different VOCs, with Alpha and Beta cocirculating in distinct spatial patterns during the second wave and Delta and Omicron affecting the whole continent during the third and fourth waves, respectively. Phylogeographic reconstruction points toward distinct differences in viral importation and exportation patterns associated with the Alpha, Beta, Delta, and Omicron variants and subvariants, when considering both Africa versus the rest of the world and viral dissemination within the continent. Our epidemiological and phylogenetic inferences therefore underscore the heterogeneous nature of the pandemic on the continent and highlight key insights and challenges, for instance, recognizing the limitations of low testing proportions. We also highlight the early warning capacity that genomic surveillance in Africa has had for the rest of the world with the detection of new lineages and variants, the most recent being the characterization of various Omicron subvariants. CONCLUSION Sustained investment for diagnostics and genomic surveillance in Africa is needed as the virus continues to evolve. This is important not only to help combat SARS-CoV-2 on the continent but also because it can be used as a platform to help address the many emerging and reemerging infectious disease threats in Africa. In particular, capacity building for local sequencing within countries or within the continent should be prioritized because this is generally associated with shorter turnaround times, providing the most benefit to local public health authorities tasked with pandemic response and mitigation and allowing for the fastest reaction to localized outbreaks. These investments are crucial for pandemic preparedness and response and will serve the health of the continent well into the 21st century

Adverse Events Associated with Prolonged Antibiotic Therapy for Periprosthetic Joint Infections—A Prospective Study with a Special Focus on Rifampin

Periprosthetic Joint Infection (PJI) is a significant contributor to patient morbidity and mortality, and it can be addressed through a range of surgical interventions coupled with antibiotic therapies. Following surgical intervention(s), prolonged administration of oral antibiotics is recommended to cure PJI. There is a lack of reports on the adverse events (AEs) associated with oral antibiotics, particularly rifampin. This investigation sought to elucidate the occurrence of antibiotic-related AEs after an initial regimen of intravenous antibiotic administration, supplemented by an extended course of oral antibiotics. A prospective study of patients diagnosed with PJI of the hip, knee, or shoulder who underwent single-stage exchange arthroplasty (SSE) (10%), two-stage exchange arthroplasty (TSE) (81%), or debridement, antibiotics, and implant retention (DAIR) (6%) was performed. The primary outcome of interest was the detection of AEs, the secondary outcome the detection of a correlation between rifampin use and the incidence of AEs, and the tertiary outcome was whether oral antibiotic treatment needed to be adjusted or discontinued due to AEs. In addition, subjective tolerability was monitored throughout the study. A total of 336 events were identified for 73 out of 80 patients. The most frequently used antibiotics were rifampin and co-trimoxazole. Most AEs occurred in the gastrointestinal tract (46%). The most frequent AEs were nausea, inappetence, diarrhea, and skin rash. In 6% of cases, the AEs led to antibiotic discontinuation, and in 29% of cases, a dose adjustment of the oral therapy occurred, mainly with amoxicillin or co-trimoxazole. The majority of patients (55%) rated the subjective tolerability as good. In conclusion, AEs during antibiotic treatment for PJI are common. They mainly affect the gastrointestinal tract. Rifampin use might be a reason for the higher incidence of AEs compared to non-rifampin antibiotic treatment

Generation and Characterization of an Inducible Cx43 Overexpression System in Mouse Embryonic Stem Cells

Connexins (Cx) are a large family of membrane proteins that can form intercellular connections, so-called gap junctions between adjacent cells. Cx43 is widely expressed in mammals and has a variety of different functions, such as the propagation of electrical conduction in the cardiac ventricle. Despite Cx43 knockout models, many questions regarding the biology of Cx43 in health and disease remain unanswered. Herein we report the establishment of a Cre-inducible Cx43 overexpression system in murine embryonic stem (ES) cells. This enables the investigation of the impact of Cx43 overexpression in somatic cells. We utilized a double reporter system to label Cx43-overexpressing cells via mCherry fluorescence and exogenous Cx43 via fusion with P2A peptide to visualize its distribution pattern. We proved the functionality of our systems in ES cells, HeLa cells, and 3T3-fibroblasts and demonstrated the formation of functional gap junctions based on dye diffusion and FRAP experiments. In addition, Cx43-overexpressing ES cells could be differentiated into viable cardiomyocytes, as shown by the formation of cross striation and spontaneous beating. Analysis revealed faster and more rhythmic beating of Cx43-overexpressing cell clusters. Thus, our Cx43 overexpression systems enable the investigation of Cx43 biology and function in cardiomyocytes and other somatic cells