Biology subjects

Chan, K.-G.

Publications and source records attributed to Chan, K.-G..

9 recordsLinked to original sources

bioRxiv2022

Exosome-mediated viral nucleic acid presentation in a crustacean reveals innate immunity evolution from a novel perspective

As an enduring hot topic in the field of innate immunity, apoptosis is widely considered as an effective approach to eliminate pathogenic microbes, and possesses crucial role during host-pathogen interactions. Recently, researchers have found that virus-containing host cells could transmit apoptotic signals to surrounding uninfected cells during infection, but the mechanism remains unclear. Here, we found that exosomes secreted by WSSV-infected mud crab hemocytes contain viral nucleic acid wsv277, which could be transported to the recipient cells and further expressed viral protein with phosphokinase activity. Besides, by using transcriptome, proteome, ChIP-seq and coIP techniques, the results revealed that wsv277 could activate the transcription and translation of apoptotic genes via interacting with CBF and EF-1, so as to suppress the spread of virus infection by inducing apoptosis of the surrounding cells. Therefore, for the first time, our study proved that the components of DNA virus could be encapsulated into exosomes, and elucidated the mechanism of apoptotic signal transduction between cells from the perspective of exosome. Moreover, comparing those of invertebrates, we hypothesized that the innate immune system of vertebrate was weakened or degenerated, allowing exosomes to be employed by virus as the vehicle for immune escape. Significance statementOur study revealed that the components of DNA virus could be packaged and transmitted through the exosomes of lower invertebrates, which strongly demonstrated the diversity of exosome-mediated viral immunity and its universality in animals. Furthermore, we elucidated the mechanism of apoptotic signal transduction between cells from the perspective of exosome, and revealed a novel strategy for the host to cope with viral infection. Since exosome is an important innate immune regulation approach, we hypothesize that the innate immune system of vertebrates is weakened or degenerated, allowing exosomes to be employed by virus as the vehicle for immune escape.

immunology↗

bioRxiv2022

Migratory behavior is positively associated with genetic diversity in butterflies

Migration is typically associated with risk and uncertainty at the population level, but little is known about its cost and benefit trade-offs at the species level. Migratory insects often exhibit strong demographic fluctuations due to local bottlenecks and outbreaks. Here, we use genomic data to investigate levels of heterozygosity and long-term population size dynamics in migratory insects, as an alternative to classical local and short-term approaches such as regional field monitoring. We analyze whole-genome sequences from 97 Lepidoptera species and show that migratory species have significantly higher levels of genome-wide heterozygosity, a proxy for effective population size, than non-migratory species. Also, we contribute whole-genome data for one of the most emblematic insect migratory species, the painted lady butterfly (Vanessa cardui), sampled across its worldwide distribution range. This species exhibits one of the highest levels of heterozygosity described in Lepidoptera (2.95% {+/-} 0.15). Coalescent modeling (PSMC) shows historical demographic stability in V. cardui, and high effective population size estimates of 2 to 20 million individuals 10,000 years ago. The study reveals that the high risks associated with migration and local environmental fluctuations do not seem to decrease overall genetic diversity and demographic stability in migratory Lepidoptera. We propose a "compensatory" demographic model for migratory r-strategist organisms in which local bottlenecks are counterbalanced by reproductive success elsewhere within their typically large distributional ranges. Our findings highlight that the delimitations of populations are substantially different for sedentary and migratory insects, and that, in the latter, local and even regional field monitoring results may not reflect whole population dynamics. Genomic diversity patterns may elucidate key aspects of an insects migratory nature and population dynamics at large spatiotemporal scales.

evolutionary biology↗

bioRxiv2021

Insight in the quorum sensing-driven lifestyle of the non-pathogenic Agrobacterium tumefaciens 6N2 and the interactions with the yeast Meyerozyma guilliermondii

Agrobacterium tumefaciens is considered a prominent phytopathogen, though most isolates are nonpathogenic. Agrobacteria can inhabit plant tissues interacting with other microorganisms. Yeasts are likewise part of these communities. We analyzed the quorum sensing (QS) systems of A. tumefaciens strain 6N2, and its relevance for the interaction with the yeast Meyerozyma guilliermondii, both sugarcane endophytes. We show that strain 6N2 is nonpathogenic, produces OHC8-HSL, OHC10-HSL, OC12-HSL and OHC12-HSL as QS signals, and possesses a complex QS architecture, with one truncated, two complete systems, and three additional QS-signal receptors. A proteomic approach showed differences in QS-regulated proteins between pure (64 proteins) and dual (33 proteins) cultures. Seven proteins were consistently regulated by quorum sensing in pure and dual cultures. M. guilliermondii proteins influenced by QS activity were also evaluated. Several up- and down- regulated proteins differed depending on the bacterial QS. These results show the importance of the QS regulation in the bacteria-yeast interactions. HighlightsThe avirulent A. tumefaciens 6N2 has two replicons and a complex QS architecture The profile of QS-regulated proteins is modified in dual cultures with Pa. laurentii The bacterial QS activity alters the proteome of the yeast Pa. laurentii

microbiology↗

bioRxiv2020

Exosomal miR-224 contributes to hemolymph microbiota homeostasis during bacterial infection in crustacean

The modulation of hemolymph microbiota homeostasis is vital for the marine invertebrate innate immunity, while growing evidence shows that exosomes could serves as anti-bacterial immune factors, however, the relationship between exosomes and hemolymph microbiota homeostasis during pathogenic bacteria infection has not been addressed. Here, we determined that exosomes released from Vibrio parahaemolyticus-infected Scylla paramamosain (mud crabs) could reduce the mortality of the host during the infection by maintaining the homeostasis of hemolymph microbiota. We further confirmed that miR-224 was densely packaged in these exosomes and targeting to HSP70, which resulted in disruption of the HSP70-TRAF6 complex to release TRAF6 that allows it to interact with Ecsit. The interaction of TRAF6 with Ecsit regulates the production of mitochondrial ROS (mROS) and Anti-lipopolysaccharide factors (ALFs) expression in recipient hemocytes, which affects homeostasis of hemolymph microbiota in response to the pathogenic bacteria infection in mud crab. To the best of our knowledge, this is the first document that reports the role of exosome in the homeostasis of hemolymph microbiota during pathogen infection and a novel regulatory mechanism and crosstalk between exosomal miRNAs and innate immune response in crustaceans. Author summaryExosomes are small membrane vesicles of endocytic origin which are widely involved in the regulation of a variety of pathological processes in mammals. Yet, although the antibacterial function of exosomes has been discovered for many years, the relationship between exosomes and hemolymph microbiota homeostasis remains unknown. In the present study, we identified the miRNAs packaged by exosomes that were possibly involved in Vibrio parahaemolyticus infection by modulating hemolymph microbiota homeostasis in crustacean mud crab Scylla paramamosain. Moreover, it was found that miR-224 was densely packaged in exosomes after Vibrio parahaemolyticus challenge, resulting in the suppression of HSP70 and disruption of the HSP70-TRAF6 complex in recipient hemocytes, then the released TRAF6 was further interacted with Ecsit to regulate ROS and ALFs levels, which eventually affected hemolymph microbiota homeostasis to cope with pathogenic bacteria infection. Our finding is the first to reveal the relationship between exosomes and hemolymph microbiota homeostasis in animals, which shows a novel molecular mechanism of invertebrate resistance to pathogenic microbial infection.

immunology↗

bioRxiv2020

Strain and lineage-level methylome heterogeneity in the multi-drug resistant pathogenic Escherichia coli ST101 clone

Escherichia coli Sequence Type (ST)101 is an emerging, multi-drug resistant lineage associated with carbapenem resistance. We recently completed a comprehensive genomics study on mobile genetic elements (MGEs) and their role in blaNDM-1 dissemination within the ST101 lineage. DNA methyltransferases (MTases) are also frequently associated with MGEs, with DNA methylation guiding numerous biological processes including genomic defence against foreign DNA and regulation of gene expression. The availability of Pacific Biosciences Single Molecule Real Time Sequencing data for seven ST101 strains enabled us to investigate the role of DNA methylation on a genome-wide scale (methylome). We defined the methylome of two complete (MS6192 and MS6193) and five draft (MS6194, MS6201, MS6203, MS6204, MS6207) ST101 genomes. Our analysis identified 14 putative MTases and eight N6-methyladenine DNA recognition sites, with one site that has not been described previously. Furthermore, we identified a Type I MTase encoded within a Transposon 7-like Transposon and show its acquisition leads to differences in the methylome between two almost identical isolates. Genomic comparisons with 13 previously published ST101 draft genomes identified variations in MTase distribution, consistent with MGE differences between genomes, highlighting the diversity of active MTases within strains of a single E. coli lineage. It is well established that MGEs can contribute to the evolution of E. coli due to their virulence and resistance gene repertoires. This study emphasises the potential for mobile genetic elements to also enable highly similar bacterial strains to rapidly acquire genome-wide functional differences via changes to the methylome. Impact StatementEscherichia coli ST101 is an emerging human pathogen frequently associated with carbapenem resistance. E. coli ST101 strains carry numerous mobile genetic elements that encode virulence determinants, antimicrobial resistance, and DNA methyltransferases (MTases). In this study we provide the first comprehensive analysis of the genome-wide complement of DNA methylation (methylome) in seven E. coli ST101 genomes. We identified a Transposon carrying a Type I restriction modification system that may lead to functional differences between two almost identical genomes and showed how small recombination events at a single genomic region can lead to global methylome changes across the lineage. We also showed that the distribution of MTases throughout the ST101 lineage was consistent with the presence or absence of mobile genetic elements on which they are encoded. This study shows the diversity of MTases within a single bacterial lineage and shows how strain and lineage-specific methylomes may drive host adaptation. Data SummarySequence data including reads, assemblies and motif summaries have previously been submitted to the National Center for Biotechnology Information (https://www.ncbi.nlm.nih.gov) under the BioProject Accessions: PRJNA580334, PRJNA580336, PRJNA580337, PRJNA580338, PRJNA580339, PRJNA580341 and PRJNA580340 for MS6192, MS6193, MS6194, MS6201, MS6203, MS6204 and MS6207 respectively. All supporting data, code, accessions, and protocols have been provided within the article or through supplementary data files.

genomics↗

bioRxiv2020

Oecophyllibacter saccharovorans gen. nov. sp. nov., a bacterial symbiont of the weaver ant Oecophylla smaragdina with a plasmid-borne sole rrn operon

In this study, bacterial strains Ha5T, Ta1 and Jb2 were isolated from different colonies of weaver ant Oecophylla smaragdina. They were distinguished as different strains based on matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry and distinctive random-amplified polymorphic DNA (RAPD) fingerprints. Cells of these bacterial strains were Gram-negative, rod-shaped, aerobic, non-motile, catalase-positive and oxidase-negative. They were able to grow at 15-37{degrees}C (optimum, 28-30{degrees}C) and in the presence of 0-1.5 % (w/v) NaCl (optimum 0%). Their predominant cellular fatty acids were C18:1{omega} 7c, C16:0, C19:0{omega} 8c cyclo, C14:0 and C16:0 2-OH. Strains Ha5T, Ta1 and Jb2 shared highest 16S rRNA gene sequence similarity (94.56-94.63%) with Neokomagataea tanensis NBRC106556T but were phylogenetically closer to Bombella spp. and Saccharibacterfloricola DSM15669T. Both 16S rRNA gene sequence-based phylogenetic analysis and core gene-based phylogenomic analysis placed them in a distinct lineage in family Acetobacteraceae. These bacterial strains shared higher than species level thresholds in multiple overall genome-relatedness indices which indicated that they belonged to the same species. In addition, they did not belong to any of the current taxa of Acetobacteraceae as they had low pairwise average nucleotide identity ([≤]70.7%), in silico DNA-DNA hybridization ([≤]39.5%) and average amino acid identity ([≤]66.u%) values with all the type members of the family. Based on these results, bacterial strains Ha5T, Ta1 and Jb2 represent a novel species of a novel genus in family Acetobacteraceae, for which we propose the name Oecophyllibacter saccharovorans gen. nov. sp. nov., and strain Ha5T as the type strain.

microbiology↗

bioRxiv2019

Genomic characterisation and context of the blaNDM-1 carbapenemase in Escherichia coli ST101.

Carbapenems are last-resort antibiotics; however, the spread of plasmid-encoded carbapenemases such as the New Delhi metallo-{beta}-lactamase 1 (NDM-1) challenges their effectiveness. The rise of NDM-1 has coincided with the emergence of extensively multidrug resistant (MDR) lineages such as Escherichia coli ST101. Here we present a comprehensive genomic analysis of seven E. coli ST101 isolates that carry the blaNDM-1 gene. We determined the complete genomes of two isolates and the draft genomes of five isolates, enabling complete resolution of the plasmid context of blaNDM-1. Comparisons with thirteen previously published ST101 genomes revealed a monophyletic lineage within the B1 phylogroup forming two clades (designated Clade 1 and Clade 2). Most Clade 1 strains are MDR, encoding resistance to at least 9 different antimicrobial classes, including extended spectrum cephalosporins. Additionally, we characterised different pathways for blaNDM-1 carriage and persistence in the ST101 lineage. For IncC plasmids, carriage was associated with recombination and local transposition events within the antibiotic resistance island. In contrast, we revealed recent transfer of a large blaNDM-1 resistance island between F-type plasmids. The complex acquisition pathways characterised here highlight the benefits of long-read Single Molecule Real Time sequencing in revealing evolutionary events that would not be apparent by short-read sequencing alone. These high-quality E. coli ST101 genomes will provide an important reference for further analysis of the role of mobile genetic elements in this emerging multidrug resistant lineage. ImportanceCarbapenem resistant Escherichia coli are urgent priority organisms as they are resistant to our drugs of last resort. E. coli ST101 have been reported as carriers of the New Delhi Metallo-beta-Lactamase 1 gene (blaNDM-1), conferring resistance to carbapenems, however there is limited genomic information available for this lineage. In this study we used long-read genome sequencing to characterise the complete genomes of two E. coli ST101 strains and determine the carriage of blaNDM-1 in a collection of E. coli ST101 strains. We showed that carriage of blaNDM-1 and resistance determinants to eight other antimicrobial classes was confined to a single clade. We also showed two different pathways for the carriage of blaNDM-1, which was dependent on the type of plasmid. Long-read sequencing allowed us to show the full complexities of these resistance regions and highlighted how strains from an emerging E. coli lineage have become resistant to nearly all available antimicrobials.

genomics↗

bioRxiv2019

Characterisation of Quorum Sensing System and Its Role in Global Regulation in Hafnia alvei

Quorum sensing (QS) is a regulatory process achieved via cell-to-cell communication that involves release and detection of autoinducers (AIs), and which occurs in a wide range of bacteria. To date, QS has been associated to events of pathogenesis, biofilm formation, and antibiotic resistance in clinical, industrial, and agricultural contexts. The main objective of this study was to characterise the role of N-Acyl homoserine lactone (AHL) type QS in Hafnia alvei FB1, a bacterial strain isolated from frozen vacuum-packed fish paste meatballs, via identification of QS core genes using a genomic approach, followed by comparative transcriptomic profiling between QS-deficient mutants and wild-type strains. H. alvei FB1 is known to produce two types of AHLs, namely, N-(3-oxohexanoyl) homoserine lactone (3OC6-HSL) and N-(3-oxooctanoyl) homoserine lactone (3OC8-HSL). The complete genome sequence of strain FB1 was obtained and a single gene for AHL synthase (halI) and its cognate receptor (halR) were identified. QS-deficient mutants of FB1 were constructed via the {lambda}-Red recombineering method. Removal of the QS genes in strain FB1 affected mainly mechanisms in cell division and nutrient uptake, as well as resistance to a number of antibiotics, which are crucial for survival, adaptation and colonisation of both food and the host gut environment. Impact statementThe Hafnia genus is known as opportunistic pathogen in both nosocomial and community-acquired infections, however, involvement and mechanism of pathogenesis of Hafnia in infection diseases is uncertain. We investigate the role of the signalling molecule, N-acyl homoserine lactones (AHLs), in a Hafnia alvei strain, since AHLs play important roles in pathogenicity, survival or adaptation in other pathogen. This comparative transciptomic study has revealed that AHLs are involved in mechanisms in cell division and nutrient uptake, as well as resistance to a number of antibiotics, which are crucial for survival, adaptation and colonisation of both food and the host gut environment. This finding provides insight and possible strategy to combat this opportunistic pathogen. Data summaryGenome sequence is deposited in NCBI GenBank under accession number CP009706. The transcriptomic data, have been deposited in the Gene Expression Omnibus (GEO) database (https://www.ncbi.nlm.nih.gov/gds) under accession number GSE93000.

genomics↗

bioRxiv2017

Genomic investigation of an outbreak of carbapenemase-producing Enterobacter cloacae: long-read sequencing reveals the context of blaIMP4 on a widely distributed IncHI2 plasmid

Carbapenem-resistant Enterobacteriaceae (CRE) represent one of the most urgent threats to human health posed by antibiotic resistant bacteria. Enterobacter hormaechei and other members of the Enterobacter cloacae complex are the most commonly encountered Enterobacter spp. within clinical settings, responsible for numerous outbreaks and ultimately poorer patient outcomes. Here we applied three complementary whole genome sequencing (WGS) technologies to characterise a hospital cluster of blaIMP-4 carbapenemase-producing E. hormaechei.\n\nIn response to a suspected CRE outbreak in 2015 within an Intensive Care Unit (ICU)/Burns Unit in a Brisbane tertiary referral hospital we used Illumina sequencing to determine that all outbreak isolates were sequence type (ST)90 and near-identical at the core genome level. Comparison to publicly available data unequivocally linked all 10 isolates to a 2013 isolate from the same ward, confirming the hospital environment as the most likely original source of infection in the 2015 cases. No clonal relationship was found to IMP-4-producing isolates identified from other local hospitals. However, using Pacific Biosciences long-read sequencing we were able to resolve the complete context of the blaIMP-4 gene, which was found to be on a large IncHI2 plasmid carried by all IMP-4-producing isolates. Continued surveillance of the hospital environment was carried out using Oxford Nanopore long-read sequencing, which was able to rapidly resolve the true relationship of subsequent isolates to the initial outbreak. Shotgun metagenomic sequencing of environmental samples also found evidence of ST90 E. hormaechei and the IncHI2 plasmid within the hospital plumbing.\n\nOverall, our strategic application of three WGS technologies provided an in-depth analysis of the outbreak, including the transmission dynamics of a carbapenemase-producing E. hormaechei cluster, identification of possible hospital reservoirs and the full context of blaIMP-4 on a multidrug resistant IncHI2 plasmid that appears to be widely distributed in Australia.

genomics↗