Detecting horizontal gene transfer with metagenomics co-barcoding sequencing.

Horizontal gene transfer (HGT) is the process through which genetic information is transferred between different genomes and that played a crucial role in bacterial evolution. HGT can enable bacteria to rapidly acquire antibiotic resistance and bacteria that have acquired resistance is spreading within the microbiome. Conventional methods of characterizing HGT patterns include short-read metagenomic sequencing (short-reads mNGS), long-read sequencing, and single-cell sequencing. These approaches present several limitations, such as short-read fragments, high amounts of input DNA, and sequencing costs, respectively. Here, we attempt to circumvent present limitations to detect HGT by developing a metagenomics co-barcode sequencing workflow (MECOS) and applying it to the human and mouse gut microbiomes. In addition to that, we have over 10-fold increased contig length compared to short-reads mNGS; we also obtained exceeding 30 million paired reads with co-barcode information. Applying the novel bioinformatic pipeline, we integrated this co-barcoding information and the context information from long reads, and observed over 50-fold HGT events after we corrected the potential wrong HGT events. Specifically, we detected approximately 3,000 HGT blocks in individual samples, encompassing ~6,000 genes and ~100 taxonomic groups, including loci conferring tetracycline resistance through ribosomal protection. MECOS provides a valuable tool for investigating HGT and advance our understanding on the evolution of natural microbial communities within hosts.IMPORTANCEIn this study, to better identify horizontal gene transfer (HGT) in individual samples, we introduce a new co-barcoding sequencing system called metagenomics co-barcoding sequencing (MECOS), which has three significant improvements: (i) long DNA fragment extraction, (ii) a special transposome insertion, (iii) hybridization of DNA to barcode beads, and (4) an integrated bioinformatic pipeline. Using our approach, we have over 10-fold increased contig length compared to short-reads mNGS, and observed over 50-fold HGT events after we corrected the potential wrong HGT events. Our results indicate the presence of approximately 3,000 HGT blocks, involving roughly 6,000 genes and 100 taxonomic groups in individual samples. Notably, these HGT events are predominantly enriched in genes that confer tetracycline resistance via ribosomal protection. MECOS is a useful tool for investigating HGT and the evolution of natural microbial communities within hosts, thereby advancing our understanding of microbial ecology and evolution.

A chromosome-encoded T4SS independently contributes to horizontal gene transfer in Enterococcus faecalis.

Bacterial type IV secretion systems (T4SSs) are the specific devices that mediate the dissemination of antibiotic resistant genes via horizontal gene transfer (HGT). Multi-drug-resistant Enterococcus faecalis (E. faecalis) represents a clinical public health threat because of its transferable plasmid with a functional plasmid-encoded (PE)-T4SS. Here, we report a chromosome-encoded (CE)-T4SS that exists in 40% of E. faecalis isolates. Compared with the PE-T4SS, CE-T4SS displays distinct characteristics in protein architecture and is capable of mediating large and genome-wide gene transfer in an imprecise manner. Reciprocal exchange of CE-T4SS- or PE-T4SS-associated origin of transfer (oriT) could disrupt HGT function, indicating that CE-T4SS is an independent system compared with PE-T4SS. Taken together, the CE-T4SS sheds light on the knowledge of HGT in gram-positive bacteria and triggers us to explore more evolutionary mechanisms in E. faecalis.

Role of the fsr Quorum-Sensing System in Enterococcus faecalis Bloodstream Infection.

Enterococcus faecalis is an important intestinal colonizing bacteria and can cause various tissue infections, including invasive blood infection (BI). The annual incidence of E. faecalis BI has been estimated to be ~4.5 per 100,000, with a fatality rate that can reach 20%. However, whether bacterial colonization or invasive infections are tissue based has not been thoroughly studied. In this study, we analyzed 537 clinical isolates from 7 different tissues to identify the key genomic elements that facilitate the colonization and invasive infection of E. faecalis. Comparative genomic analysis revealed that the BI E. faecalis isolates had the largest genome size but the lowest GC content, fsr quorum-sensing system genes were enriched in the BI E. faecalis, and the fsr gene cluster could enhance biofilm formation and serum resistance ability. Our findings also provide deep insight into the genomic differences between different tissue isolates, and the fsr quorum-sensing systems could be a key factor promoting E. faecalis invasion into the blood. IMPORTANCE First, we conducted an advanced study on the genomic differences between colonizing and infecting E. faecalis, which provides support and evidence for early and accurate diagnoses. Second, we discovered that fsr was significantly associated with blood infections, which also provides additional information for studies exploring the invasiveness of E. faecalis. Most importantly, we found that fsr played an important role in both biofilm formation and serum resistance ability in E. faecalis.

Alteration of microbiota and immune response of mice gavaged with Klebsiella oxytoca.

Interactions between the microbiota and immune system play a vital role in the host homeostasis. Increasing studies have investigated environmental perturbations affecting the microbiota. However, studies also are needed to model how an organ-specific immune response affects the microbiota to understand the dynamic changes between the immune system and microbiota. We constructed a murine Klebsiella oxytoca infection model, in which mice were gavaged with K. oxytoca, and the microbiota and immune responses of both the digestive tract and respiratory tract were compared for 1-2 weeks after infection. Metagenomic and cytokine analysis of the samples displayed a delayed colonization of K. oxytoca, but an early immune response in the respiratory tract, as compared with that in the digestive tract, suggested niche-specific characteristics of bacterial colonization and the corresponding immune response. Furthermore, we constructed an interaction map of K. oxytoca in both the digestive tract and respiratory tract that furthers our understanding of the host-microbe biology in K. oxytoca-infected hosts.

Two-step fitness selection for intra-host variations in SARS-CoV-2.

Spontaneous mutations introduce uncertainty into coronavirus disease 2019 (COVID-19) control procedures and vaccine development. Here, we perform a spatiotemporal analysis on intra-host single-nucleotide variants (iSNVs) in 402 clinical samples from 170 affected individuals, which reveals an increase in genetic diversity over time after symptom onset in individuals. Nonsynonymous mutations are overrepresented in the pool of iSNVs but underrepresented at the single-nucleotide polymorphism (SNP) level, suggesting a two-step fitness selection process: a large number of nonsynonymous substitutions are generated in the host (positive selection), and these substitutions tend to be unfixed as SNPs in the population (negative selection). Dynamic iSNV changes in subpopulations with different gender, age, illness severity, and viral shedding time displayed a varied fitness selection process among populations. Our study highlights that iSNVs provide a mutational pool shaping the rapid global evolution of the virus.

Specific Redistribution of Severe Acute Respiratory Syndrome Coronavirus 2 Variants in the Respiratory System and Intestinal Tract.

Intrahost analysis of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) genomic sequences identified 2 viral haplotypes comprised of 3 genetically linked mutations from the respiratory and intestinal tracts of a patient with coronavirus disease 2019. Spatiotemporal data suggest that this patient initially had dual infection of 2 SARS-CoV-2 variants, which subsequently redistributed into the 2 systems.

[Analysis of particle size characteristics of road sediments in Beijing Olympic Park].

Particle size analysis of road sediment collected in October and November in Beijing Olympic Park indicates that most of the sediments are 76-830 µm; the grain size of the sediments in the area of large population flow is mainly coarse but the grain size in the area of large traffic volume is fine relatively while most of the sediments are <300 p.m. Moreover, sediments of size range <300 µm can be easily accumulated on the road with moderate traffic density. The results demonstrate that the effect of pedestrian flow on the composition of the particles is unobvious and the main influences are the traffic density, extensive construction. With the length of dry period increasing, the content of sediments of size range >300 µm decreases and the content of sediments of size range < 150 µm increases, however, the change of the content of sediments of size range 150-300 µm is not obvious. The results indicate that the effectiveness of the road sediment removal depends on the length of dry period, and the accumulation of different size particles varies differently under the different dry days. Compared with the stone road, surface particles can accumulate on the asphalt road more easily as the accumulation of particles is affected by the road material significantly. Therefore, to reduce the urban surface water pollution, it is necessary to improve the design of park road such as using the stone road, which can decrease the roughness of the road.