Antimicrobial Resistance in the Asia Pacific region: a meeting report.

The Asia Pacific region, home to two-thirds of the world's population and ten of the least developed countries, is considered a regional hot-spot for the emergence and spread of antimicrobial resistance (AMR). Despite this, there is a dearth of high-quality regional data on the extent of AMR. Recognising the urgency to close this gap, Singapore organised a meeting to discuss the problems in the region and frame a call for action. Representatives from across the region and beyond attended the meeting on the "Antimicrobial Resistance in the Asia Pacific & its impact on Singapore" held in November 2018. This meeting report is a summary of the discussions on the challenges and progress in surveillance, drivers and levers of AMR emergence, and the promising innovations and technologies that could be used to combat the increasing threat of AMR in the region. Enhanced surveillance and research to provide improved evidence-based strategies and policies are needed. The major themes that emerged for an action plan are working towards a tailored solution for the region by harnessing the One Health approach, enhancing inter-country collaborations, and collaboratively leverage upon new emerging technologies. A regionally coordinated effort that is target-driven, sustainable and builds on a framework facilitating communication and governance will strengthen the fight against AMR in the Asia Pacific region

Draft genome sequence of multidrug resistant klebsiella pneumoniae strain C43 isolated from environmental water sample

Here, we report the genome of ESBL-producing Klebsiella pneumoniae strain C43, which was isolated from an environmental water sample. The genome is 5,614,412 bp in size with GC content of 56.86% with multidrug antimicrobial resistance genes and several metal resistance gene operons.Ministry of Education (MOE)Nanyang Technological UniversityPublished versionThis work was supported by the Singapore Ministry of Education under its Academic Research Fund Tier 1 RG87/21, Research Centre for Excellence IDMxS, and a pilot grant from the Centre for Global Health, Lee Kong Chian School of Medicine (awarded to E.P.H.Y.). S.K.W. is funded by a Nanyang Technological University Research Scholarship

Rapid direct nucleic acid amplification test without RNA extraction for SARS-CoV-2 using a portable PCR thermocycler

There is an ongoing worldwide coronavirus disease 2019 (Covid-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). At present, confirmatory diagnosis is by reverse transcription polymerase chain reaction (RT-PCR), typically taking several hours and requiring a molecular laboratory to perform. There is an urgent need for rapid, simplified, and cost-effective detection methods. We have developed and analytically validated a protocol for direct rapid extraction-free PCR (DIRECT-PCR) detection of SARS-CoV-2 without the need for nucleic acid purification. As few as six RNA copies per reaction of viral nucleocapsid (N) gene from respiratory samples such as sputum and nasal exudate can be detected directly using our one-step inhibitor-resistant assay. The performance of this assay was validated on a commercially available portable PCR thermocycler. Viral lysis, reverse transcription, amplification, and detection are achieved in a single-tube homogeneous reaction within 36 min. This minimizes hands-on time, reduces turnaround-time for sample-to-result, and obviates the need for RNA purification reagents. It could enable wider use of Covid-19 testing for diagnosis, screening, and research in countries and regions where laboratory capabilities are limiting.Nanyang Technological UniversityNational Research Foundation (NRF)Published versionThis research is supported by the National Research Foundation, Prime Minister’s Office, Singapore under its Competitive Research Programme (CRP13-2014-01, E.P.H.Y.) and Lee Kong Chian School of Medicine, Nanyang Technological University Singapore Start-Up Grant (E.P.H.Y.)

Rapid Direct Nucleic Acid Amplification Test without RNA Extraction for SARS-CoV-2 Using a Portable PCR Thermocycler

There is an ongoing worldwide coronavirus disease 2019 (Covid-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). At present, confirmatory diagnosis is by reverse transcription polymerase chain reaction (RT-PCR), typically taking several hours and requiring a molecular laboratory to perform. There is an urgent need for rapid, simplified, and cost-effective detection methods. We have developed and analytically validated a protocol for direct rapid extraction-free PCR (DIRECT-PCR) detection of SARS-CoV-2 without the need for nucleic acid purification. As few as six RNA copies per reaction of viral nucleocapsid (N) gene from respiratory samples such as sputum and nasal exudate can be detected directly using our one-step inhibitor-resistant assay. The performance of this assay was validated on a commercially available portable PCR thermocycler. Viral lysis, reverse transcription, amplification, and detection are achieved in a single-tube homogeneous reaction within 36 min. This minimizes hands-on time, reduces turnaround-time for sample-to-result, and obviates the need for RNA purification reagents. It could enable wider use of Covid-19 testing for diagnosis, screening, and research in countries and regions where laboratory capabilities are limiting

Pre-isolation molecular screening for high-throughput sampling and sequencing of bacterial microbes from the environment

Environmental studies often require culture and characterization to understand the prevalence, distribution, persistence and functions of target microorganisms in ecological habitats. Isolating pure microbiological monocultures allows the phenotypic characterization of microorganisms to study their functional properties. For efficient isolation of low-prevalence organisms, enrichment followed by PCR screening is performed to identify positive samples for subsequent culture. Molecular characterization, strain-typing, and genotyping of isolated microorganisms is best comprehensively performed using whole-genome sequencing. This article outlines end-to-end protocols for screening, isolation, and sequencing of microbes from environmental samples. We provide systematic methods for environmental study design, enrichment, screening, and isolation of target microorganisms. Species identification is performed using qPCR or MALDI-TOF MS. Genomic DNA is extracted for whole-genome sequencing using the Oxford Nanopore platform. © 2023 Wiley Periodicals LLC. Basic Protocol 1: Designing and conducting an environmental soil sampling study Basic Protocol 2: Enrichment of microbes from environmental soil samples Alternate Protocol 1: Collection and enrichment of microbes from environmental water samples Basic Protocol 3: Screening of enriched samples by direct qPCR Basic Protocol 4: Enumeration and isolation of enriched samples using selective medium Basic Protocol 5: Species confirmation using colony qPCR Alternate Protocol 2: Species identification using a MALDI-TOF MS Biotyper Alternate Protocol 3: Species identification of bacterial isolates using universal PCR primers and Sanger sequencing Basic Protocol 6: Cryostorage of bacterial isolates Basic Protocol 7: Extraction of genomic DNA Basic Protocol 8: Quality check of extracted genomic DNA Basic Protocol 9: Whole-genome sequencing using the Oxford Nanopore MinION Platform.Ministry of Education (MOE)Nanyang Technological UniversityThis research is supported by the Singapore Ministry of Education under its Academic Research Fund Tier 1 RG87/21, the Research Centre for Excellence IDMxS, and internal grants from the Centre for Global Health and NTU Clinical Diagnostic Laboratory, Lee Kong Chian School of Medicine

Draft genome sequence of Enterobacter hormaechei subsp. steigerwaltii strain BEI01

Here, we report the genome sequence of Enterobacter hormaechei subsp. steigerwaltii strain BEI01, originally deposited as a member of the Enterobacter cloacae complex. The genome is 4,900,246 bp in size with a GC content of 55.44%; it contains multidrug antimicrobial resistance genes and several metal resistance gene operons.Ministry of Education (MOE)Nanyang Technological UniversityPublished versionThis work was supported by a Nanyang Assistant Professorship from the Lee Kong Chian School of Medicine, Nanyang Technological University (awarded to X.L.G.), a startup grant from the Ministry of Education, and a pilot grant from the Centre for Global Health, Lee Kong Chian School of Medicine (awarded to E.P.H.Y.). S.K.W. is funded by a Nanyang Technological University Research Scholarship

Genetic/protein association of atopic dermatitis and tooth agenesis

Atopic dermatitis and abnormalities in tooth development (including hypomineralization, hypodontia and microdontia) have been observed to co-occur in some patients. A common pathogenesis pathway that involves genes and protein interactions has been hypothesized. This review aims to first provide a description of the key gene mutations and signaling pathways associated with atopic dermatitis and tooth agenesis (i.e., the absence of teeth due to developmental failure) and identify the possible association between the two diseases. Second, utilizing a list of genes most commonly associated with the two diseases, we conducted a protein-protein network interaction analysis using the STRING database and identified a novel association between the Wnt/β-catenin signaling pathway (major pathway responsible for TA) and desmosomal proteins (component of skin barrier that affect the pathogenesis of AD). Further investigation into the mechanisms that may drive their co-occurrence and underlie the development of the two diseases is warranted.Ministry of Education (MOE)Published versionThis project was partially supported by the Academic Research Funds of Singapore Ministry of Education via the National University of Singapore (A-0002960-00-00) and the Faculty of Dentistry, National University of Singapore (A-0002960-01-00)

Optofluidic Detection for Virus Infection Monitoring using Effective Refractive Index

International audienc

Complex Copy Number Variation of AMY1 does not Associate with Obesity in two East Asian Cohorts

The human amylase gene locus at chromosome 1p21.1 is structurally complex. This region contains two pancreatic amylase genes, AMY2B, AMY2A, and a salivary gene AMY1. The AMY1 gene harbors extensive copy number variation (CNV), and recent studies have implicated this variation in adaptation to starch-rich diets and in association to obesity for European and Asian populations. In this study, we showed that by combining quantitative PCR and digital PCR, coupled with careful experimental design and calibration, we can improve the resolution of genotyping CNV with high copy numbers (CNs). In two East Asian populations of Chinese and Malay ethnicity studied, we observed a unique non-normal distribution of AMY1 diploid CN genotypes with even:odd CNs ratio of 4.5 (3.3–4.7), and an association between the common AMY2A CN = 2 genotype and odd CNs of AMY1, that could be explained by the underlying haplotypic structure. In two further case–control cohorts (n = 932 and 145, for Chinese and Malays, respectively), we did not observe the previously reported association between AMY1 and obesity or body mass index. Improved methods for accurately genotyping multiallelic CNV loci and understanding the haplotype complexity at the AMY1 locus are necessary for population genetics and association studies.Accepted versio

Fibroblast growth factor 21 mediates the associations between exercise, aging, and glucose regulation

Aging increases the prevalence of glucose intolerance, but exercise improves glucose homeostasis. The fibroblast growth factor 21 (FGF21)-adiponectin axis helps regulate glucose metabolism. However, the role of FGF21 in mediating glucose metabolism with aging and exercise remains unknown.Ministry of Education (MOE)This study is supported by the Ministry of Education Singapore Start-up Grant for Human and Metabolic disease (L0412270)