Investigation of the Causes of Shigatoxigenic Escherichia coli PCR Positive and Culture Negative Samples

peer-reviewedMolecular methods may reveal the presence of pathogens in samples through the detection of specific target gene(s) associated with microorganisms, but often, the subsequent cultural isolation of the pathogen is not possible. This discrepancy may be related to low concentration of the cells, presence of dead cells, competitive microflora, injured cells and cells in a viable but non-culturable state, free DNA and the presence of free bacteriophages which can carry the target gene causing the PCR-positive/culture-negative results. Shiga-toxigenic Escherichia coli (STEC) was used as a model for studying this phenomenon, based on the phage-encoded cytotoxins genes (Stx family) as the detection target in samples through real-time qPCR. Stx phages can be integrated in the STEC chromosome or can be isolated as free particles in the environment. In this study, a combination of PCR with culturing was used for investigating the presence of the stx1 and stx2 genes in 155 ovine recto-anal junction swab samples (method (a)-PCR). Samples which were PCR-positive and culture-negative were subjected to additional analyses including detection of dead STEC cells (method (b)-PCR-PMA dye assay), presence of Stx phages (method (c)-plaque assays) and inducible integrated phages (method (d)-phage induction). Method (a) showed that even though 121 samples gave a PCR-positive result (78%), only 68 samples yielded a culturable isolate (43.9%). Among the 53 (34.2%) PCR-positive/culture-negative samples, 21 (39.6%) samples were shown to have STEC dead cells only, eight (15.1%) had a combination of dead cells and inducible stx phage, while two samples (3.8%) had a combination of dead cells, inducible phage and free stx phage, and a further two samples had Stx1 free phages only (3.8%). It was thus possible to reduce the samples with no explanation to 20 (37.7% of 53 samples), representing a further step towards an improved understanding of the STEC PCR-positive/culture-negative phenomenon

Klebsiella , a Hitherto Underappreciated Zoonotic Pathogen of Importance to One Health: A Short Review

Members of the genus, Klebsiella , are becoming increasingly challenging to control due to the recent convergence of multidrug resistant (MDR) and hypervirulent (hv) phenotypes in some species of concern to One Health . This short review will provide an introduction to this bacterial genus in the hospital and other settings, update Klebsiella taxonomy, and comment on recent findings describing the prevalence of Klebsiella species in the food chain, a hitherto infrequently recognised ecologic niche. The paper will also consider this bacterium in the context of the One Health paradigm and its importance to food safety and security

Meta-analysis of cheese microbiomes highlights contributions to multiple aspects of quality

peer-reviewedA detailed understanding of the cheese microbiome is key to the optimization of flavour, appearance, quality and safety. Accordingly, we conducted a high-resolution meta-analysis of cheese microbiomes and corresponding volatilomes. Using 77 new samples from 55 artisanal cheeses from 27 Irish producers combined with 107 publicly available cheese metagenomes, we recovered 328 metagenome-assembled genomes, including 47 putative new species that could influence taste or colour through the secretion of volatiles or biosynthesis of pigments. Additionally, from a subset of samples, we found that differences in the abundances of strains corresponded with levels of volatiles. Genes encoding bacteriocins and other antimicrobials, such as pseudoalterin, were common, potentially contributing to the control of undesirable microorganisms. Although antibiotic-resistance genes were detected, evidence suggested they are not of major concern with respect to dissemination to other microbiomes. Phages, a potential cause of fermentation failure, were abundant and evidence for phage-mediated gene transfer was detected. The anti-phage defence mechanism CRISPR was widespread and analysis thereof, and of anti-CRISPR proteins, revealed a complex interaction between phages and bacteria. Overall, our results provide new and substantial technological and ecological insights into the cheese microbiome that can be applied to further improve cheese production.Science Foundation Irelan

Draft genome sequences of four Yersinia enterocolitica strains, isolated from wild ungulate carcasses

This study describes the draft genome sequences of four Yersinia enterocolitica strains, originally isolated from ungulate carcasses. These isolates were typed biochemically and two were determined to be highly virulent (biotype 1B). The draft genome sequences had a mean size of 4.77 Mb and a mean G+C content of 47.1%

Aptamer-functionalized nanopipettes : a promising approach for viral fragment detection via ion current rectification

In this report, ion current rectification, an electrochemical phenomenon observed in asymmetric nanopipettes, is used for the label-free detection of SARS-CoV-2 viral fragments in nasopharyngeal samples. Quartz nanopipettes are functionalized with aptamers targeting the spike protein S1 domain, wherein changes to the surface charge magnitude, distribution, and ion transport behavior modulate the current-voltage response upon binding. The aptamer-modified nanopipette provides a selective and sensitive method for detecting SARS-CoV-2, with a limit of detection in the laboratory of 0.05 pg/mL. The effectiveness of this low-cost platform was demonstrated by sensing SARS-CoV-2 in nasopharyngeal samples, successfully discriminating between positive and negative cases with minimal template preparation, highlighting the platform’s potential as a versatile sensing strategy for infectious disease detection in clinical diagnosis