Association of vaccination, international travel, public health and social measures with lineage dynamics of SARS-CoV-2.

Continually emerging SARS-CoV-2 variants of concern that can evade immune defenses are driving recurrent epidemic waves of COVID-19 globally. However, the impact of measures to contain the virus and their effect on lineage diversity dynamics are poorly understood. Here, we jointly analyzed international travel, public health and social measures (PHSM), COVID-19 vaccine rollout, SARS-CoV-2 lineage diversity, and the case growth rate (GR) from March 2020 to September 2022 across 63 countries. We showed that despite worldwide vaccine rollout, PHSM are effective in mitigating epidemic waves and lineage diversity. An increase of 10,000 monthly travelers in a single country-to-country route between endemic countries corresponds to a 5.5% (95% CI: 2.9 to 8.2%) rise in local lineage diversity. After accounting for PHSM, natural immunity from previous infections, and waning immunity, we discovered a negative association between the GR of cases and adjusted vaccine coverage (AVC). We also observed a complex relationship between lineage diversity and vaccine rollout. Specifically, we found a significant negative association between lineage diversity and AVC at both low and high levels but not significant at the medium level. Our study deepens the understanding of population immunity and lineage dynamics for future pandemic preparedness and responsiveness.

No evidence for persistent natural plague reservoirs in historical and modern Europe.

Caused by Yersinia pestis, plague ravaged the world through three known pandemics: the First or the Justinianic (6th-8th century); the Second (beginning with the Black Death during c.1338-1353 and lasting until the 19th century); and the Third (which became global in 1894). It is debatable whether Y. pestis persisted in European wildlife reservoirs or was repeatedly introduced from outside Europe (as covered by European Union and the British Isles). Here, we analyze environmental data (soil characteristics and climate) from active Chinese plague reservoirs to assess whether such environmental conditions in Europe had ever supported "natural plague reservoirs". We have used new statistical methods which are validated through predicting the presence of modern plague reservoirs in the western United States. We find no support for persistent natural plague reservoirs in either historical or modern Europe. Two factors make Europe unfavorable for long-term plague reservoirs: 1) Soil texture and biochemistry and 2) low rodent diversity. By comparing rodent communities in Europe with those in China and the United States, we conclude that a lack of suitable host species might be the main reason for the absence of plague reservoirs in Europe today. These findings support the hypothesis that long-term plague reservoirs did not exist in Europe and therefore question the importance of wildlife rodent species as the primary plague hosts in Europe.

Quantitative and dynamic profiling of human gut core microbiota by real-time PCR.

The human gut microbiota refers to a diverse community of microorganisms that symbiotically exist in the human intestinal system. Altered microbial communities have been linked to many human pathologies. However, there is a lack of rapid and efficient methods to assess gut microbiota signatures in practice. To address this, we established an appraisal system containing 45 quantitative real-time polymerase chain reaction (qPCR) assays targeting gut core microbes with high prevalence and/or abundance in the population. Through comparative genomic analysis, we selected novel species-specific genetic markers and primers for 31 of the 45 core microbes with no previously reported specific primers or whose primers needed improvement in specificity. We comprehensively evaluated the performance of the qPCR assays and demonstrated that they showed good sensitivity, selectivity, and quantitative linearity for each target. The limit of detection ranged from 0.1 to 1.0 pg/µL for the genomic DNA of these targets. We also demonstrated the high consistency (Pearson's r = 0.8688, P < 0.0001) between the qPCR method and metagenomics next-generation sequencing (mNGS) method in analyzing the abundance of selected bacteria in 22 human fecal samples. Moreover, we quantified the dynamic changes (over 8 weeks) of these core microbes in 14 individuals using qPCR, and considerable stability was demonstrated in most participants, albeit with significant individual differences. Overall, this study enables the simple and rapid quantification of 45 core microbes in the human gut, providing a promising tool to understand the role of gut core microbiota in human health and disease. KEY POINTS: • A panel of original qPCR assays was developed to quantify human gut core microbes. • The qPCR assays were evaluated and compared with mNGS using real fecal samples. • This method was used to dynamically profile the gut core microbiota in individuals.

Assessing the origins of the European Plagues following the Black Death: A synthesis of genomic, historical, and ecological information.

The second plague pandemic started in Europe with the Black Death in 1346 and lasted until the 19th century. Based on ancient DNA studies, there is a scientific disagreement over whether the bacterium, Yersinia pestis, came into Europe once (Hypothesis 1) or repeatedly over the following four centuries (Hypothesis 2). Here, we synthesize the most updated phylogeny together with historical, archeological, evolutionary, and ecological information. On the basis of this holistic view, we conclude that Hypothesis 2 is the most plausible. We also suggest that Y. pestis lineages might have developed attenuated virulence during transmission, which can explain the convergent evolutionary signals, including pla decay, that appeared at the end of the pandemics.

Progress on the quality control technology of next generation sequencing library.

As a key supporting technology in the fields of life sciences and medicine, high-throughput sequencing has developed rapidly and become increasingly mature. The workflow of this technology can be divided into nucleic acid extraction, library construction, sequencing, and data analysis. Among these, library construction is a pivotal step that bridges the previous and subsequent stages. The effectiveness of library construction is contingent on the quality of upstream samples and also impacts the data analysis following sequence data output. The selection and implementation of library construction quality control techniques are crucial for enhancing the reliability of results and reducing errors in sequencing data. This review provides an in-depth discussion of library construction quality control techniques, summarizing and evaluating their principles, advantages and disadvantages, and applicability. It also discusses the selection of relevant technologies in practical application scenarios. The aim is to offer theoretical foundations and references for researchers, disease prevention and control personnel, and others when choosing library quality control techniques, thereby promoting the quality and efficiency of high-throughput sequencing work.

Competitive fitness and homologous recombination of SARS-CoV-2 variants of concern.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants continue to emerge and cocirculate in humans and wild animals. The factors driving the emergence and replacement of novel variants and recombinants remain incompletely understood. Herein, we comprehensively characterized the competitive fitness of SARS-CoV-2 wild type (WT) and three variants of concern (VOCs), Alpha, Beta and Delta, by coinfection and serial passaging assays in different susceptible cells. Deep sequencing analyses revealed cell-specific competitive fitness: the Beta variant showed enhanced replication fitness during serial passage in Caco-2 cells, whereas the WT and Alpha variant showed elevated fitness in Vero E6 cells. Interestingly, a high level of neutralizing antibody sped up competition and completely reshaped the fitness advantages of different variants. More importantly, single clone purification identified a significant proportion of homologous recombinants that emerged during the passage history, and immune pressure reduced the frequency of recombination. Interestingly, a recombination hot region located between nucleotide sites 22,995 and 28,866 of the viral genomes could be identified in most of the detected recombinants. Our study not only profiled the variable competitive fitness of SARS-CoV-2 under different conditions, but also provided direct experimental evidence of homologous recombination between SARS-CoV-2 viruses, as well as a model for investigating SARS-CoV-2 recombination.

Oxaliplatin derived monofunctional triazole-containing platinum(II) complex counteracts oxaliplatin-induced drug resistance in colorectal cancer.

Oxaliplatin-based chemotherapy is the current standard of care in adjuvant therapy for advanced colorectal cancer (CRC). But acquired resistance to oxaliplatin eventually occurs and becoming a major cause of treatment failure. Thus, there is an unmet need for developing new chemical entities (NCE) as new therapeutic candidates to target chemotherapy-resistant CRC. Novel Pt(II) complexes were designed and synthesized as cationic monofunctional oxaliplatin derivatives for DNA platination-mediated tumor targeting. The complex Ph-glu-Oxa sharing the same chelating ligand of diaminocyclohexane (DACH) with oxaliplatin but is equally potent in inhibiting the proliferation of HT29 colon cancer cells and its oxaliplatin-resistant phenotype of HT29/Oxa. The in vivo therapeutic potential of Ph-glu-Oxa was confirmed in oxaliplatin-resistant xenograft model demonstrating the reversibility of the drug resistance by the new complex and the efficacy was associated with the unimpaired high intracellular drug accumulation in HT29/Oxa. Guanosine-5'-monophosphate (5'-GMP) reactivity, double-strand plasmid DNA cleavage, DNA-intercalated ethidium bromide (EB) fluorescence quenching and atomic force microscopy (AFM)-mediated DNA denaturing studies revealed that Ph-glu-Oxa was intrinsically active as DNA-targeting agent. The diminished susceptibility of the complex to glutathione (GSH)-mediated detoxification, which confers high intracellular accumulation of the drug molecule may play a key role in maintaining cytotoxicity and counteracting oxaliplatin drug resistance.

Exploring the industrial potential of Lactobacillus delbrueckii ssp. bulgaricus by population genomics and genome-wide association study analysis.

Lactobacillus delbrueckii ssp. bulgaricus is one of the most commonly used starter cultures for yogurt production. However, how its genetic background affects acid production capacity is unclear. This study investigated the industrial potential of L. delbrueckii ssp. bulgaricus using population genomics and GWAS analysis. To meet our goal, population genetics and functional genomics analyses were performed on 188 newly sequenced L. delbrueckii ssp. bulgaricus strains isolated from naturally fermented dairy products together with 19 genome sequences retrieved from the NCBI database. Four distinct clusters were identified, and they were correlated with the geographical sites where the samples were collected. The GWAS analysis about acidification fermentation results with sucrose-fortified reconstituted skim milk revealed a significant association between l-lactate dehydrogenase (lldD; Ldb2036) and the bacterial acid production rate. Our study has broadened the understanding of the population structure and genetic diversity of L. delbrueckii ssp. bulgaricus by identifying potential targets for further research, development, and use of lactic acid bacteria in the dairy industry.

Integrated Whole-Genome Sequencing Infrastructure for Outbreak Detection and Source Tracing of Salmonella enterica Serotype Enteritidis.

As an important foodborne pathogen, Salmonella enterica serotype Enteritidis is recognized as one of the most common causes of human salmonellosis globally. Outbreak detection for this highly homogenous serotype, however, has remained challenging. Rapid advances in sequencing technologies have presented whole-genome sequencing (WGS) as a significant advancement for source tracing and molecular typing of foodborne pathogens. A retrospective analysis was conducted using Salmonella Enteritidis isolates (n = 65) from 11 epidemiologically confirmed outbreaks and a collection of contemporaneous sporadic isolates (n = 258) during 2007-2017 to evaluate the performance of WGS in delineating outbreak-associated isolates. Whole-genome single-nucleotide polymorphism (SNP)-based phylogenetic analysis revealed well-supported clades in concordance with epidemiological evidence and pairwise distances of ≤3 SNPs for all outbreaks. WGS-based framework of outbreak detection was thus proposed and applied prospectively to investigate isolates (n = 66) from nine outbreaks during 2018-2019. We further demonstrated the superior discriminatory power and accuracy of WGS to resolve and delineate outbreaks for pragmatic food source tracing. The proposed integrated WGS framework is the first in China for Salmonella Enteritidis and has the potential to serve as a paradigm for outbreak detection and source tracing of Salmonella throughout the stages of food production, as well as expanded to other foodborne pathogens.

Whole-Genome Analysis of Salmonella enterica Serovar Enteritidis Isolates in Outbreak Linked to Online Food Delivery, Shenzhen, China, 2018.

In July 2018, an outbreak of 10 cases of Salmonella enterica serovar Enteritidis infection occurred in Shenzhen, China. Outbreak investigation complemented by whole-genome sequencing traced the source to food ordered online. Our investigation highlights the role of online food delivery platforms as a new mode of foodborne disease transmission.

Vertical Flow Assay for Inflammatory Biomarkers Based on Nanofluidic Channel Array and SERS Nanotags.

There is a great demand for the development of detection assays for inflammation infection diagnosis with high throughput and ultrasensitivity. Herein, a vertical flow assay system with functionalized nanoporous anodic aluminum oxide (AAO) as sensing membrane, and encoded core-shell surface enhanced Raman scattering (SERS) nanotags as labels for multiple inflammatory biomarkers detection is presented. A 2 × 2 test array on the porous AAO is developed and modified with multiple capture antibodies to capture inflammatory biomarkers from samples. Due to the high surface area to volume ratio of the AAO membrane, and its influence on plasmonic coupling, the electromagnetic field of the encoded core-shell SERS nanotags is enhanced. Detection limits of 53.4, 4.72, 48.3, and 7.53 fg mL-1 are realized for C reactive protein, interleukin-6, serum amyloid A, and procalcitonin, respectively, with a linear dynamic range spanning at least five orders of magnitude. In addition, the proposed method also shows acceptable accuracy and repeatability for the detection of clinical samples. Therefore, this approach is expected to be a powerful point of care testing tool for disease diagnosis in facility limited areas.

Complete genome sequence of a novel bovine hepacivirus from Yunnan, China.

We detected a novel bovine hepacivirus N (HNV) subtype, IME_BovHep_01, in the serum of cattle in Tengchong, Yunnan, China, by high-throughput sequencing. The complete genome of IME_BovHep_01, was sequenced using an Illumina MiSeq sequencer and found to be 8850 nt in length, encoding one hypothetical protein. BLASTn analysis showed that the genome sequence shared similarity with the bovine hepacivirus isolate BovHepV_209/Ger/2014, with 88% query coverage and 70.8% identity. However, the highest similarity was to bovine hepacivirus N strain BRBovHep_RS963, for which only a partial genome sequence is available, with 68% query coverage and 81.5% identity. Sequence comparisons and phylogenetic analysis suggested that IME_BovHep_01 is a novel HNV subtype. Importantly, IME_BovHep_01 is the first member of this new genotype for which the complete genome sequence was determined.

Anthropogenically driven environmental changes shift the ecological dynamics of hemorrhagic fever with renal syndrome.

Zoonoses are increasingly recognized as an important burden on global public health in the 21st century. High-resolution, long-term field studies are critical for assessing both the baseline and future risk scenarios in a world of rapid changes. We have used a three-decade-long field study on hantavirus, a rodent-borne zoonotic pathogen distributed worldwide, coupled with epidemiological data from an endemic area of China, and show that the shift in the ecological dynamics of Hantaan virus was closely linked to environmental fluctuations at the human-wildlife interface. We reveal that environmental forcing, especially rainfall and resource availability, exert important cascading effects on intra-annual variability in the wildlife reservoir dynamics, leading to epidemics that shift between stable and chaotic regimes. Our models demonstrate that bimodal seasonal epidemics result from a powerful seasonality in transmission, generated from interlocking cycles of agricultural phenology and rodent behavior driven by the rainy seasons.

Genetic diversity, virulence factors and farm-to-table spread pattern of Vibrio parahaemolyticus food-associated isolates.

Vibrio parahaemolyticus is the leading bacterial cause of seafood-associated gastroenteritis worldwide. Moreover, infections and outbreaks caused by V. parahaemolyticus has kept increasing over the last two decades. In this study, we investigated the genetic diversity, virulence factors and farm-to-table spread pattern of V. parahaemolyticus by analyzing 383 genomes of food-associated isolates. These strains were isolated from diverse sample types from six provinces of China in 2014, being classified into three tiers of the farm-to-table spread process: food production, circulation and consumption. The genetic diversity of V. parahaemolyticus in different classifications, including geographical location, sample type, source and spread tier, was similar, as the median number of pairwise SNPs within each classification was between 33,013 and 33,659. Specifically, there was no clear boundaries in genetic diversity of the isolates from inland vs. coastal provinces, as well as of those from freshwater vs. seawater products. Moreover, the virulence genes and genomic islands were only found in a small number of isolates, indicating a low disease risk of the food-associated isolates in this study. By further exploring 28 recently emerged clonal groups, we identified seven farm-to-table spread events, showing a common pattern of single-source radial spread accompanied with occasional gene gain/loss events. Generally speaking, our work highlighted the colonization of V. parahaemolyticus in inland provinces and freshwater environment, and provided a snapshot of the farm-to-table spread pattern of V. parahaemolyticus food-associated isolates. Our results showed the feasibility of tracking the farm-to-table spread of foodborne pathogen, which would help construct the whole genome sequencing-based molecular tracking network in the future.

Avian influenza H5N1 viral and bird migration networks in Asia.

The spatial spread of the highly pathogenic avian influenza virus H5N1 and its long-term persistence in Asia have resulted in avian influenza panzootics and enormous economic losses in the poultry sector. However, an understanding of the regional long-distance transmission and seasonal patterns of the virus is still lacking. In this study, we present a phylogeographic approach to reconstruct the viral migration network. We show that within each wild fowl migratory flyway, the timing of H5N1 outbreaks and viral migrations are closely associated, but little viral transmission was observed between the flyways. The bird migration network is shown to better reflect the observed viral gene sequence data than other networks and contributes to seasonal H5N1 epidemics in local regions and its large-scale transmission along flyways. These findings have potentially far-reaching consequences, improving our understanding of how bird migration drives the periodic reemergence of H5N1 in Asia.

Bacterial genome-wide association study: methodologies and applications.

With the development of genome sequencing and the accumulation of whole genome sequences, genome-wide association study (GWAS) has achieved remarkable advances in understanding of human complex disease, and tens of thousands of disease risk factors have been found. Meanwhile, GWAS provides a new tool for exploring the genetic mechanism of bacterial phenotypes. Since the publication of the first bacterial GWAS (BGWAS) work in 2013, there have been more than 10 reports, which reveal the genetic basis of host adaption, drug resistance and virulence, etc. These findings greatly enhance our understanding on genetics, evolution and spread of bacteria. In this review, we summarize the current methodologies, applications and problems of BGWAS and highlight its potential in future research, which aims to provide helps for the applications of BGWAS in the field of microbiology.

Features of Ebola Virus Disease at the Late Outbreak Stage in Sierra Leone: Clinical, Virological, Immunological, and Evolutionary Analyses.

We performed Ebola virus disease diagnosis and viral load estimation for Ebola cases in Sierra Leone during the late stage of the 2014-2015 outbreak (January-March 2015) and analyzed antibody and cytokine levels and the viral genome sequences. Ebola virus disease was confirmed in 86 of 1001 (9.7%) patients, with an overall case fatality rate of 46.8%. Fatal cases exhibited significantly higher levels of viral loads, cytokines, and chemokines at late stages of infection versus early stage compared with survivors. The viruses converged in a new clade within sublineage 3.2.4, which had a significantly lower case fatality rate.

Host transcriptomic responses to pneumonic plague reveal that Yersinia pestis inhibits both the initial adaptive and innate immune responses in mice.

Pneumonic plague is the most deadly form of infection caused by Yersinia pestis and can progress extremely fast. However, our understanding on the host transcriptomic response to pneumonic plague is insufficient. Here, we used RNA-sequencing technology to analyze transcriptomic responses in mice infected with fully virulent strain 201 or EV76, a live attenuated vaccine strain lacking the pigmentation locus. Approximately 600 differentially expressed genes (DEGs) were detected in lungs from both 201- and EV76-infected mice at 12h post-infection (hpi). DEGs in lungs of 201-infected mice exceeded 2000 at 48hpi, accompanied by sustained large numbers of DEGs in the liver and spleen; however, limited numbers of DEGs were detected in those organs of EV-infected mice. Remarkably, DEGs in lungs were significantly enriched in critical immune responses pathways in EV76-infected but not 201-infected mice, including antigen processing and presentation, T cell receptor signaling among others. Pathological and bacterial load analyses confirmed the rapid systemic dissemination of 201-infection and the confined EV76-infection in lungs. Our results suggest that fully virulent Y. pestis inhibits both the innate and adaptive immune responses that are substantially stimulated in a self-limited infection, which update our holistic views on the transcriptomic response to pneumonic plague.

Epidemic Clones, Oceanic Gene Pools, and Eco-LD in the Free Living Marine Pathogen Vibrio parahaemolyticus.

We investigated global patterns of variation in 157 whole-genome sequences of Vibrio parahaemolyticus, a free-living and seafood associated marine bacterium. Pandemic clones, responsible for recent outbreaks of gastroenteritis in humans, have spread globally. However, there are oceanic gene pools, one located in the oceans surrounding Asia and another in the Mexican Gulf. Frequent recombination means that most isolates have acquired the genetic profile of their current location. We investigated the genetic structure in the Asian gene pool by calculating the effective population size in two different ways. Under standard neutral models, the two estimates should give similar answers but we found a 27-fold difference. We propose that this discrepancy is caused by the subdivision of the species into a hundred or more ecotypes which are maintained stably in the population. To investigate the genetic factors involved, we used 51 unrelated isolates to conduct a genome-wide scan for epistatically interacting loci. We found a single example of strong epistasis between distant genome regions. A majority of strains had a type VI secretion system associated with bacterial killing. The remaining strains had genes associated with biofilm formation and regulated by cyclic dimeric GMP signaling. All strains had one or other of the two systems and none of isolate had complete complements of both systems, although several strains had remnants. Further "top down" analysis of patterns of linkage disequilibrium within frequently recombining species will allow a detailed understanding of how selection acts to structure the pattern of variation within natural bacterial populations.

Yersinia protein kinase A phosphorylates vasodilator-stimulated phosphoprotein to modify the host cytoskeleton.

Pathogenic Yersinia species evolved a type III secretion system that injects a set of effectors into the host cell cytosol to promote infection. One of these effectors, Yersinia protein kinase A (YpkA), is a multidomain effector that harbours a Ser/Thr kinase domain and a guanine dissociation inhibitor (GDI) domain. The intercellular targets of the kinase and GDI domains of YpkA were identified to be Gαq and the small GTPases RhoA and Rac1, respectively, which synergistically induce cytotoxic effects on infected cells. In this study, we demonstrate that vasodilator-stimulated phosphoprotein (VASP), which is critical for regulation of actin assembly, cell adhesion and motility, is a direct substrate of YpkA kinase activity. Ectopic co-expression of YpkA and VASP in HEK293T cells leads to the phosphorylation of VASP at S157, and YpkA kinase activity is essential for VASP phosphorylation at this site. Moreover, YpkA directly phosphorylates VASP in in vitro kinase assay. YpkA-mediated VASP phosphorylation significantly inhibits actin polymerization and promotes the disruption of actin cytoskeleton, which inhibits the phagocytosis. Taken together, our study found a novel molecular mechanism used by YpkA to disrupt cytoskeleton dynamics, thereby promoting the anti-phagocytosis ability of pathogenic Yersiniae.