SHIP: identifying antimicrobial resistance gene transfer between plasmids.

MOTIVATION: Plasmids are carriers for antimicrobial resistance (AMR) genes and can exchange genetic material with other structures, contributing to the spread of AMR. There is no reliable approach to identify the transfer of AMR genes across plasmids. This is mainly due to the absence of a method to assess the phylogenetic distance of plasmids, as they show large DNA sequence variability. Identifying and quantifying such transfer can provide novel insight into the role of small mobile elements and resistant plasmid regions in the spread of AMR. RESULTS: We developed SHIP, a novel method to quantify plasmid similarity based on the dynamics of plasmid evolution. This allowed us to find conserved fragments containing AMR genes in structurally different and phylogenetically distant plasmids, which is evidence for lateral transfer. Our results show that regions carrying AMR genes are highly mobilizable between plasmids through transposons, integrons, and recombination events, and contribute to the spread of AMR. Identified transferred fragments include a multi-resistant complex class 1 integron in Escherichia coli and Klebsiella pneumoniae, and a region encoding tetracycline resistance transferred through recombination in Enterococcus faecalis. AVAILABILITY AND IMPLEMENTATION: The code developed in this work is available at https://github.com/AbeelLab/plasmidHGT.

Integrated omics of Saccharomyces cerevisiae CENPK2-1C reveals pleiotropic drug resistance and lipidomic adaptations to cannabidiol.

Yeast metabolism can be engineered to produce xenobiotic compounds, such as cannabinoids, the principal isoprenoids of the plant Cannabis sativa, through heterologous metabolic pathways. However, yeast cell factories continue to have low cannabinoid production. This study employed an integrated omics approach to investigate the physiological effects of cannabidiol on S. cerevisiae CENPK2-1C yeast cultures. We treated the experimental group with 0.5 mM CBD and monitored CENPK2-1C cultures. We observed a latent-stationary phase post-diauxic shift in the experimental group and harvested samples in the inflection point of this growth phase for transcriptomic and metabolomic analysis. We compared the transcriptomes of the CBD-treated yeast and the positive control, identifying eight significantly overexpressed genes with a log fold change of at least 1.5 and a significant adjusted p-value. Three notable genes were PDR5 (an ABC-steroid and cation transporter), CIS1, and YGR035C. These genes are all regulated by pleiotropic drug resistance linked promoters. Knockout and rescue of PDR5 showed that it is a causal factor in the post-diauxic shift phenotype. Metabolomic analysis revealed 48 significant spectra associated with CBD-fed cell pellets, 20 of which were identifiable as non-CBD compounds, including fatty acids, glycerophospholipids, and phosphate-salvage indicators. Our results suggest that mitochondrial regulation and lipidomic remodeling play a role in yeast's response to CBD, which are employed in tandem with pleiotropic drug resistance (PDR). We conclude that bioengineers should account for off-target product C-flux, energy use from ABC-transport, and post-stationary phase cell growth when developing cannabinoid-biosynthetic yeast strains.

Metagenomic-based surveillance systems for antibiotic resistance in non-clinical settings.

The success of antibiotics as a therapeutic agent has led to their ineffectiveness. The continuous use and misuse in clinical and non-clinical areas have led to the emergence and spread of antibiotic-resistant bacteria and its genetic determinants. This is a multi-dimensional problem that has now become a global health crisis. Antibiotic resistance research has primarily focused on the clinical healthcare sectors while overlooking the non-clinical sectors. The increasing antibiotic usage in the environment - including animals, plants, soil, and water - are drivers of antibiotic resistance and function as a transmission route for antibiotic resistant pathogens and is a source for resistance genes. These natural compartments are interconnected with each other and humans, allowing the spread of antibiotic resistance via horizontal gene transfer between commensal and pathogenic bacteria. Identifying and understanding genetic exchange within and between natural compartments can provide insight into the transmission, dissemination, and emergence mechanisms. The development of high-throughput DNA sequencing technologies has made antibiotic resistance research more accessible and feasible. In particular, the combination of metagenomics and powerful bioinformatic tools and platforms have facilitated the identification of microbial communities and has allowed access to genomic data by bypassing the need for isolating and culturing microorganisms. This review aimed to reflect on the different sequencing techniques, metagenomic approaches, and bioinformatics tools and pipelines with their respective advantages and limitations for antibiotic resistance research. These approaches can provide insight into resistance mechanisms, the microbial population, emerging pathogens, resistance genes, and their dissemination. This information can influence policies, develop preventative measures and alleviate the burden caused by antibiotic resistance.

When uncertainty becomes the norm: The Chris Hani Baragwanath Academic Hospital's Speech Therapy and Audiology Department's response to the COVID-19 pandemic.

BACKGROUND:  In March 2020 the World Health Organization declared the coronavirus disease 2019 (COVID-19) a pandemic. Management of this pandemic had significant implications for clinical departments across the world. Healthcare systems were urgently required to reorganise and redesign patient care as well as repurpose staff. OBJECTIVES:  We will share the lived experience of our response as speech therapy and audiology (STA) clinicians to the COVID-19 pandemic. METHOD:  This study adopted an autoethnographic approach within Bronfenbrenner's bioecological model to describe STA clinicians' response to the COVID-19 pandemic. RESULTS:  Adaptations to practice were made to continue service provision whilst adhering to COVID-19 regulations. We assisted in other areas to meet the immediate needs of the hospital. Service delivery strategies consisted of a review of clinical and quality assurance protocols. We developed a telehealth service package which included a hybrid approach, within a context of digital poverty. We created resources to ensure continuity of care. Collaboration within our systems facilitated innovative solutions. Mental health and well-being of staff members were key to the response developed. CONCLUSION:  South African healthcare systems' inequalities were highlighted by the pandemic. The response showed that the needs of vulnerable populations were not accounted for when developing this public health response. Lessons learnt included the importance of adaptability, becoming comfortable with uncertainty and maintaining open and transparent communication. Consultation and collaboration within various levels of our healthcare system were critical in responding to the needs of patients. Commitment to compassionate leadership and staff well-being were crucial.

Evaluating long-read de novo assembly tools for eukaryotic genomes: insights and considerations.

BACKGROUND: Assembly algorithm choice should be a deliberate, well-justified decision when researchers create genome assemblies for eukaryotic organisms from third-generation sequencing technologies. While third-generation sequencing by Oxford Nanopore Technologies (ONT) and Pacific Biosciences (PacBio) has overcome the disadvantages of short read lengths specific to next-generation sequencing (NGS), third-generation sequencers are known to produce more error-prone reads, thereby generating a new set of challenges for assembly algorithms and pipelines. However, the introduction of HiFi reads, which offer substantially reduced error rates, has provided a promising solution for more accurate assembly outcomes. Since the introduction of third-generation sequencing technologies, many tools have been developed that aim to take advantage of the longer reads, and researchers need to choose the correct assembler for their projects. RESULTS: We benchmarked state-of-the-art long-read de novo assemblers to help readers make a balanced choice for the assembly of eukaryotes. To this end, we used 12 real and 64 simulated datasets from different eukaryotic genomes, with different read length distributions, imitating PacBio continuous long-read (CLR), PacBio high-fidelity (HiFi), and ONT sequencing to evaluate the assemblers. We include 5 commonly used long-read assemblers in our benchmark: Canu, Flye, Miniasm, Raven, and wtdbg2 for ONT and PacBio CLR reads. For PacBio HiFi reads , we include 5 state-of-the-art HiFi assemblers: HiCanu, Flye, Hifiasm, LJA, and MBG. Evaluation categories address the following metrics: reference-based metrics, assembly statistics, misassembly count, BUSCO completeness, runtime, and RAM usage. Additionally, we investigated the effect of increased read length on the quality of the assemblies and report that read length can, but does not always, positively impact assembly quality. CONCLUSIONS: Our benchmark concludes that there is no assembler that performs the best in all the evaluation categories. However, our results show that overall Flye is the best-performing assembler for PacBio CLR and ONT reads, both on real and simulated data. Meanwhile, best-performing PacBio HiFi assemblers are Hifiasm and LJA. Next, the benchmarking using longer reads shows that the increased read length improves assembly quality, but the extent to which that can be achieved depends on the size and complexity of the reference genome.

Factors affecting length of hospital stay in stroke survivors in South Africa: A call for a stroke unit.

Background: Stroke in Africa is a growing and neglected crisis with the incidence more than doubling in low- to middle-income countries in the last four decades. Despite this growing threat, implementation of stroke models of care in hospitals is lacking. Stroke units as a model of care have been shown to decrease mortality, reduce length of hospital stay (LOS) and improve outcomes in stroke survivors. Objectives: To determine the profile of stroke survivors and identify factors contributing to LOS at Chris Hani Baragwanath Academic Hospital (CHBAH) in South Africa to support stroke unit implementation. Method: This study involved a retrospective record review of stroke survivors admitted to CHBAH between September 2018 and May 2019. Factors associated with LOS were determined using linear regression models; univariate and multiple regression models were fitted. Results: A total of 567 participants' data were included. Overall, 51.85% of the participants required services from all rehabilitation disciplines. The median LOS was 9 days (interquartile ranges [IQR]: 5-11 days) with each discipline providing an average of six sessions. Participants who were referred to the rehabilitation team 3 days after admission to hospital stayed 6 days longer compared with those participants who were referred earlier (p < 0.001). Conclusion: Delayed referral to the rehabilitation team resulted in increased LOS. This study supports the need for dedicated stroke units to decrease hospital LOS and improve patients' outcomes by ensuring early, well-coordinated rehabilitation intervention and discharge planning. Contribution: The study highlights the urgency for re-evaluation of stroke care infrastructure within Gauteng to streamline and provide accessible stroke models of care.

Characterisation of Campylobacter spp. Isolated from Poultry in KwaZulu-Natal, South Africa.

This study investigated the antibiotic resistance, virulence profiles, and clonality of Campylobacter jejuni and Campylobacter coli isolated from an intensive poultry farming system in KwaZulu-Natal, South Africa. Following ethical approval, samples were collected over six weeks using the farm-to-fork approach. Campylobacter spp. were identified using culture, confirmed and differentiated to species level by PCR, and subjected to antibiotic susceptibility testing. Selected antibiotic resistance (and mutations) and virulence genes were screened by PCR and confirmed by DNA sequencing. Genetic relatedness amongst the isolates was ascertained using pulsed-field gel electrophoresis. In all, 105 isolates were confirmed as belonging to both Campylobacter coli (60; 57%) and C. jejuni (45; 43%). The highest resistance was recorded against erythromycin and clindamycin. The gyrA mutation, A20175C/A2074G point mutation, tet(O), and cmeB, all associated with antibiotic resistance, were detected. All the virulence genes (pldA, ciaB, cdtA, cdtB, cdtC, dnaJ, except for cadF) were also detected. Isolates were grouped into five pulsotypes displaying 85% similarity, irrespective of their resistance profiles. The numerous permutations of clonality, antibiotic resistance, and virulence profiles evident in Campylobacter spp. pose a challenge to food safety and necessitate a comprehensive understanding of the molecular epidemiology of this organism to decrease its spread in the food chain.