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Summary: Background: Genetically distinct viral variants of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have been recorded since January 2020. The introduction of global vaccine programs has contributed to lower COVID-19 hospitalisation and mortality rates, particularly in developed countries. In late 2021, Omicron BA.1 emerged, with substantially altered genetic differences and clinical effects from other variants of concern. Shortly after dominating global spread in early 2022, BA.1 was supplanted by the genetically distinct Omicron lineage BA.2. A sub-lineage of BA.2, designated BA.5, presently has an outgrowth advantage over BA.2 and other BA.2 sub-lineages. Here we study the neutralisation of Omicron BA.1, BA.2 and BA.5 and pre-Omicron variants using a range of vaccine and convalescent sera and therapeutic monoclonal antibodies using a live virus neutralisation assay. Using primary nasopharyngeal swabs, we also tested the relative fitness of BA.5 compared to pre-Omicron and Omicron viral lineages in their ability to use the ACE2-TMPRSS2 pathway. Methods: Using low passage clinical isolates of Clade A.2.2, Beta, Delta, BA.1, BA.2 and BA.5, we determined humoral neutralisation in vitro in vaccinated and convalescent cohorts, using concentrated human IgG pooled from thousands of plasma donors, and licensed monoclonal antibody therapies. We then determined infectivity to particle ratios in primary nasopharyngeal samples and expanded low passage isolates in a genetically engineered ACE2/TMPRSS2 cell line in the presence and absence of the TMPRSS2 inhibitor Nafamostat. Findings: Peak responses to 3 doses of BNT162b2 vaccine were associated with a 9-fold reduction in neutralisation for Omicron lineages BA.1, BA.2 and BA.5. Concentrated pooled human IgG from convalescent and vaccinated donors and BNT162b2 vaccination with BA.1 breakthrough infections were associated with greater breadth of neutralisation, although the potency was still reduced 7-fold across all Omicron lineages. Testing of clinical grade antibodies revealed a 14.3-fold reduction using Evusheld and 16.8-fold reduction using Sotrovimab for the BA.5. Whilst the infectivity of BA.1 and BA.2 was attenuated in ACE2/TMPRSS2 entry, BA.5 was observed to be equivalent to that of an early 2020 circulating clade and had greater sensitivity to the TMPRSS2 inhibitor Nafamostat. Interpretation: Observations support all Omicron variants to significantly escape neutralising antibodies across a range of vaccination and/or convalescent responses. Potency of therapeutic monoclonal antibodies is also reduced and differs across Omicron lineages. The key difference of BA.5 from other Omicron sub-variants is the reversion in tropism back to using the well-known ACE2-TMPRSS2 pathway, utilised efficiently by pre-Omicron lineages. Monitoring if these changes influence transmission and/or disease severity will be key for ongoing tracking and management of Omicron waves globally. Funding: This work was primarily supported by Australian Medical Foundation research grants MRF2005760 (ST, GM & WDR), MRF2001684 (ADK and ST) and Medical Research Future Fund Antiviral Development Call grant (WDR), Medical Research Future Fund COVID-19 grant (MRFF2001684, ADK & SGT) and the New South Wales Health COVID-19 Research Grants Round 2 (SGT).

Introduction: The emergence of multidrug resistance in Bacteroides fragilis, especially the phylogenetic lineage carrying the carbapenemase gene cfiA, represents an increasing threat to human health. However, knowledge on the diversity of the multidrug-resistant strains and the genetic elements carrying the antibiotic resistance genes (ARGs) remains limited. Aim: The objective of the study was to describe the resistome in cfiA-positive B. fragilis. Methods: A collection of cfiA-positive B. fragilis from diverse human (8 bacteremias, 15 wound infections) and animal (2 chickens, 2 pigs, 6 dogs, 3 cats) sources in Hong Kong, 2015–2017 was analysed by whole genome sequencing. Results: In the 36 isolates, 13 distinct ARGs (total number 83, median 2, range 0–7 per isolate) other than cfiA were detected. ARGs encoding resistance to aminoglycosides, β-lactams, macrolides, sulphonamides and tetracyclines were carried by CTn341-like, CTnHyb-like, Tn5220-like, Tn4555-like and Tn613-like transposons and were detected in phylogenetically diverse isolates of different host sources. Only few ARGs encoding resistance to metronidazole and tetracyclines were localized on plasmids. In two chicken isolates, a novel transposon (designated as Tn6994) was found to be involved in the dissemination of multiple ARGs mediating resistance to multiple antibiotics, including metronidazole and linezolid that are critically important for treatment of anaerobic infections. In mating experiments, Tn6994 and the associated phenotypic resistance could be transferred to Bacteroides nordii recipient. Conclusion: This study illustrates the importance of transposons in the dissemination of ARGs in the cfiA-positive division of B. fragilis. One Health approach is necessary to track the dissemination of ARGs.

Objectives: This study evaluated disc diffusion tests and agar screening for detecting mecA-mediated oxacillin resistance in Staphylococcus lugdunensis (S. lugdunensis). Methods: Staphylococcus lugdunensis isolates (n = 179) from diverse sources in Hong Kong during 1998–2018 were investigated by disc diffusion tests (cefoxitin and oxacillin) and inoculation onto oxacillin (1 μg/mL and 2 μg/mL) and chromID methicillin-resistant Staphylococcus aureus (MRSA) agars. The results were compared with mecA PCR as the reference. Isolates with discordant results were further tested by MIC and penicillin-binding protein 2a (PBP2a) assays. Results: Cefoxitin and oxacillin zone diameters were not distributed in ways that allowed reliable division of the mecA-positive (n = 52) and mecA-negative (n = 127) isolates. On applying the 2019 Clinical Laboratory Standards Institute (CLSI) M100 breakpoints for cefoxitin disc results, there was 88% categorical agreement (CA) and 40% very major error (VME). Screening using 2 μg/mL oxacillin agar reliably differentiated mecA-positive and mecA-negative isolates (100% CA) without any major error (ME) or VME results. The performance of screening using 1 μg/mL oxacillin agar or ChromID MRSA agar was variable (74–89% CA, 0–38% ME and 0–37% VME). The mecA-positive isolates (n = 21) that could not be detected by the cefoxitin disc test were further characterised. The cefoxitin MIC for all 21 isolates was ≤4 μg/mL. Twenty isolates had an oxacillin MIC of 1–2 μg/mL and one had an oxacillin MIC of 4 μg/mL. All had positive PBP2a results and were typed as clonal cluster 27/SCCmec V. Conclusions: These findings highlight the need to evaluate phenotypic methods using mecA-positive S. lugdunensis with different oxacillin resistance phenotypes.

Background: Subject C135 is one of the members of the Sydney Blood Bank Cohort, infected in 1981 through transfusion with attenuated nef/3′ long terminal repeat (LTR)-deleted HIV-1, and has maintained undetectable plasma viral load and steady CD4 cell count, in the absence of therapy. Uniquely, C135 combines five factors separately associated with control of viraemia: nef/LTR-deleted HIV-1, HLA-B57, HLA-DR13, heterozygous CCR5 Δ32 genotype and vigorous p24-stimulated peripheral blood mononuclear cell (PBMC) proliferation. Therefore, we studied in detail viral burden and immunological responses in this individual. Methods: PBMC and gut and lymph node biopsy samples were analysed for proviral HIV-1 DNA by real-time and nested PCRs, and nef/LTR alleles by nested PCR. HIV-specific antibodies were studied by Western blotting, and CD4+ and CD8+ T lymphocyte responses were measured by proliferation and cytokine production in vitro. Results: PBMC samples from 1996, but not since, showed amplification of nef alleles with gross deletions. Infectious HIV-1 was never recovered. Proviral HIV-1 DNA was not detected in recent PBMC or gut or lymph node biopsy samples. C135 has a consistently weak antibody response and a substantial CD4+ T cell proliferative response to a previously described HLA-DR13-restricted epitope of HIV-1 p24 in vitro, which augmented a CD8+ T cell response to an immunodominant HLA-B57-restricted epitope of p24, while his T cells show reduced levels of CCR5. Conclusions: Subject C135's early PCR and weak antibody results are consistent with limited infection with a poorly replicating nef/LTR-deleted strain of HIV-1. With his HLA-B57-restricted gag-specific CD8 and helper HLA-DR13-restricted CD4 T cell proliferative responses, C135 appears to have cleared his HIV-1 infection 37 years after transfusion.