Development of a highly sensitive point-of-care test for African swine fever that combines EZ-Fast DNA extraction with LAMP detection: Evaluation using naturally infected swine whole blood samples from Vietnam.

BACKGROUND: While early detection and early containment are key to controlling the African swine fever (ASF) pandemic, the lack of practical testing methods for use in the field are a major barrier to achieving this feat. OBJECTIVES: To describe the development of a rapid and sensitive point-of-care test (POCT) for ASF, and its evaluation using swine whole blood samples for field settings. METHODS: In total, 89 swine whole blood samples were collected from Vietnamese swine farms and were performed the POCT using a combination of crude DNA extraction and LAMP (loop-mediated isothermal amplification) amplification. RESULTS: The POCT enabled crude DNA to be extracted from swine whole blood samples within 10 min at extremely low cost and with relative ease. The entire POCT required a maximum of 50 min from the beginning of DNA extraction to final judgment. Compared to a conventional real-time PCR detection, the POCT showed a 1 log reduction in detection sensitivity, but comparable diagnostic sensitivity of 100% (56/56) and diagnostic specificity of 100% (33/33). The POCT was quicker and easier to perform and did not require special equipment. CONCLUSIONS: This POCT is expected to facilitate early diagnosis and containment of ASF invasion into both regions in which it is endemic and eradicated.

Identification of Escherichia coli and Related Enterobacteriaceae and Examination of Their Phenotypic Antimicrobial Resistance Patterns: A Pilot Study at A Wildlife-Livestock Interface in Lusaka, Zambia.

A cross-sectional study was used to identify and assess prevalence and phenotypic antimicrobial resistance (AMR) profiles of Escherichia coli and other enterobacteria isolated from healthy wildlife and livestock cohabiting at a 10,000 acres game ranch near Lusaka, Zambia. Purposive sampling was used to select wildlife and livestock based on similarities in behavior, grazing habits and close interactions with humans. Isolates (n = 66) from fecal samples collected between April and August 2018 (n = 84) were examined following modified protocols for bacteria isolation, biochemical identification, molecular detection, phylogenetic analysis, and antimicrobial susceptibility testing by disc diffusion method. Data were analyzed using R software, Genetyx ver.12 and Mega 6. Using Applied Profile Index 20E kit for biochemical identification, polymerase chain reaction assay and sequencing, sixty-six isolates were identified to species level, of which Escherichia coli (72.7%, 48/66), E. fergusonii (1.5%, 1/66), Shigella sonnei (22.7%, 14/66), Sh. flexinerri (1.5%, 1/66) and Enterobacteriaceae bacterium (1.5%, 1/66), and their relationships were illustrated in a phylogenetic tree. Phenotypic antimicrobial resistance or intermediate sensitivity expression to at least one antimicrobial agent was detected in 89.6% of the E. coli, and 73.3% of the Shigella isolates. The E. coli isolates exhibited the highest resistance rates to ampicillin (27%), ceftazidime (14.3%), cefotaxime (9.5%), and kanamycin (9.5%). Multidrug resistance (MDR) was detected in 18.8% of E. coli isolates while only 13.3% Shigella isolates showed MDR. The MDR was detected among isolates from impala and ostrich (wild animals in which no antimicrobial treatment was used), and in isolates from cattle, pigs, and goats (domesticated animals). This study indicates the possible transmission of drug-resistant microorganisms between animals cohabiting at the wildlife-livestock interface. It emphasizes the need for further investigation of the role of wildlife in the development and transmission of AMR, which is an issue of global concern.

Optimum fabrication parameters for preparing high performance SERS substrates based on Si pyramid structure and silver nanoparticles.

In this work, we propose simple and inexpensive methods to prepare micro/nano hierarchical Surface-Enhanced Raman Scattering (SERS) substrates, in which pyramid structure is created by using anisotropic wet etching of a silicon wafer and a silver thin film is deposited on these pyramid arrays by thermal evaporation. The ensemble is then annealed at 450 °C for 2 hours to form silver nanoparticles (AgNPs). The sizes and density of the pyramids and AgNPs are optimized mainly by changing the etching temperature (60-80 °C), the thickness of the Ag-film (15-45 nm) and etching time (3-10 min). The ultraviolet visible (UV-Vis) absorbance spectra show that the AgNPs formed with the 30 nm-thick film exhibit the strongest plasmonic effect. Under these conditions, the spherical AgNPs with sizes of 42-48 nm are densely distributed on the silicon micro-pyramid array. The obtained SERS signal is the strongest at the pyramid base-edge size of 7-10 µm. The enhancement factor obtained from the abamectin probe molecules is as high as 1 × 106 and the SERS substrates enable the detection of abamectin concentrations as low as 5.7 × 10-9 M. Therefore, this work provides a novel SERS substrate structure that has a high potential for use in medicine and biotechnology or as a food security sensor.