There is an urgent need to develop simple and fast antimicrobial susceptibility tests (ASTs) that allow informed prescribing of antibiotics. Here, we describe a label-free AST that can deliver results within an hour, using an actively dividing culture as starting material. The bacteria are incubated in the presence of an antibiotic for 30 min, and then approximately 10⁵ cells are analysed one-by-one with microfluidic impedance cytometry for 2–3 min. The measured electrical characteristics reflect the phenotypic response of the bacteria to the mode of action of a particular antibiotic, in a 30-minute incubation window. The results are consistent with those obtained by classical broth microdilution assays for a range of antibiotics and bacterial species.

Burkholderia pseudomallei Traced to Water Treatment Plant in Australia

Limited experience and a lack of validated diagnostic reagents make Burkholderia pseudomallei , the cause of melioidosis, difficult to recognize in the diagnostic microbiology laboratory. We compared three methods of confirming the identity of presumptive B. pseudomallei strains using a collection of Burkholderia species drawn from diverse geographic, clinical, and environmental sources. The 95 isolates studied included 71 B. pseudomallei and 3 B. thailandensis isolates. The API 20NE method identified only 37% of the B. pseudomallei isolates. The agglutinating antibody test identified 82% at first the attempt and 90% including results of a repeat test with previously negative isolates. Gas-liquid chromatography analysis of bacterial fatty acid methyl esters (GLC-FAME) identified 98% of the B. pseudomallei isolates. The agglutination test produced four false positive results, one B. cepacia , one B. multivorans , and two B. thailandensis . API produced three false positive results, one positive B. cepacia and two positive B. thailandensis . GLC-FAME analysis was positive for one B. cepacia isolate. On the basis of these results, the most robust B. pseudomallei discovery pathway combines the previously recommended isolate screening tests (Gram stain, oxidase test, gentamicin and polymyxin susceptibility) with monoclonal antibody agglutination on primary culture, followed by a repeat after 24 h incubation on agglutination-negative isolates and GLC-FAME analysis. Incorporation of PCR-based identification within this schema may improve percentages of recognition further but requires more detailed evaluation.

Purpose of Review Melioidosis epidemiology is susceptible to climate change through direct and indirect effects on human encounter with the causative agent, Burkholderia pseudomallei. This review describes the current depth of knowledge and recent advances in the understanding of this relationship and applies it to observations of melioidosis in Western Australia. Recent Findings High maximum rainfall and dense cloud cover have been shown to predict environmental presence of B. pseudomallei and cases of melioidosis, probably through correspondingly high moisture levels in B. pseudomallei-receptive soils. Increased melioidosis cases have been observed following storms in Taiwan and cyclones in the Australian Northern Territory and strengthen the association between melioidosis and extreme weather events. Indirect weather effects contribute to bacterial exposure through mechanisms such as increasing B. pseudomallei output from water seeps after heavy rain or localised flooding. Climate and weather have been directly implicated in dissemination of B. pseudomallei and cases of melioidosis in several notable events in Western Australia. Over a 10-year surveillance period, the cases that lay in the path of a tropical cyclone co-located with cyclone systems that repeatedly crossed the Western Australian coast. Cyclone-associated cases were caused by different B. pseudomallei MLST genotypes, arguing against airborne dissemination from a common source. Summary Predicted increases in temperature, changes in global precipitation patterns and an increased incidence of extreme weather events are expected to change melioidosis epidemiology. Further studies of the physical geographic drivers of melioidosis will deepen understanding of the impact of climate on melioidosis.

In this study we investigated the role of the bacterial flagellum inBurkholderia pseudomalleientry toAcanthamoeba astronyxistrophozoites.B. pseudomalleicells were tethered to the external amoebic surface via their flagella. MM35, the flagellum-lackingfliCknockout derivative ofB. pseudomalleiNCTC 1026b did not demonstrate flagellum-mediated endocytosis in timed coculture, confirming that an intact flagellar apparatus assistsB. pseudomalleientry intoA. astronyxis.

The expanding global distribution of multi-resistant Klebsiella pneumoniae demands faster antimicrobial susceptibility testing (AST) to guide antibiotic treatment. Current ASTs rely on time-consuming differentiation of resistance and susceptibility after initial isolation of bacteria from a clinical specimen. Here we describe a flow cytometry workflow to determine carbapenem susceptibility from bacterial cell characteristics in an international K. pneumoniae isolate collection (n = 48), with a range of carbapenemases. Our flow cytometry-assisted susceptibility test (FAST) method combines rapid qualitative susceptible/non-susceptible classification and quantitative MIC measurement in a single process completed shortly after receipt of a primary isolate (54 and 158 minutes respectively). The qualitative FAST results and FAST-derived MIC (MIC_FAST) correspond closely with broth microdilution MIC (MIC_BMD, Matthew’s correlation coefficient 0.887), align with the international AST standard (ISO 200776-1; 2006) and could be used for rapid determination of antimicrobial susceptibility in a wider range of Gram negative and Gram positive bacteria.