OBJECTIVE: To investigate the origin of Biomphalaria straminea in China, so as to provide insights into assessment of schistosomiasis mansoni transmission risk and B. straminea control.
METHODS: Guanlan River, Dasha River, Shenzhen Reservoir, upper and lower reaches of Kuiyong River, and Xinzhen River in Shenzhen, China, were selected as sampling sites. Ten Biomphalaria samples were collected from each site, and genomic DNA was extracted from Biomphalaria samples. DNA samples were obtained from 15 B. straminea sampled from 5 sampling sites in Minas Gerais State, Pará State, Federal District, Pernambuco State, and Sao Paulo State in Brazil, South America. Cytochrome c oxidase I (COI) and mitochondrial 16S ribosomal RNA (16S rRNA) genes were sampled using the above DNA templates, and the amplified products were sequenced. The COI and 16S rRNA gene sequences were downloaded from GenBank, and the sampling sites were acquired. All COI and 16S rRNA gene sequences were aligned and evolutionary trees of B. straminea were created based on COI and 16S rRNA gene sequences to identify the genetic similarity and evolutionary relationship between B. straminea samples from China and South America.
RESULTS: A total of 60 COI gene sequences with a length of 529 bp and 3 haplotypes were obtained from B. straminea sampled from China. There were 165 COI gene sequences of B. straminea retrieved from GenBank, and following alignment with the above 60 gene sequences, a total of 33 haplotypes were obtained. Phylogenetic analysis showed that the three haplotypes of B. straminea from China were clustered into one clade, among which the haplotype China11 and three B. straminea samples from Brazil retrieved from GenBank belonged to the same haplotype. Geographical evolution analysis showed that the B. straminea samples from three sampling sites along eastern coasts of Brazil had the same haplotype with China11, and B. straminea samples from other two sampling sites were closely, genetically related to China11. A total of 60 16S rDNA gene sequences with approximately 322 bp in length were amplified from B. straminea in China, with 2 haplotypes identified. A total of 70 16S rDNA gene sequences of B. straminea were captured from GenBank. Phylogenetic analysis showed that Biomphalaria snails collected from China were clustered into a clade, and the haplotype China64 and the haplotype 229BS from Brazil shared the same haplotype. The 49 16S rDNA gene sequences of B. straminea from 25 sampling sites in southern Brazil, which were captured from GenBank, were included in the present analysis, and the B. straminea from 3 sampling sites shared the same haplotype with China64 in China. Geographical evolution analysis based on COI and 16S rRNA gene sequences showed that B. straminea sampled from eastern coastal areas of Brazil shared the same haplotypes in two gene fragment sequences with Biomphalaria snails collected from China.
CONCLUSIONS: The Biomphalaria snails in China are characterized as B. straminea, which have a low genetic diversity. The Biomphalaria snails in China have a high genetic similarity with B. straminea sampled from eastern coastal areas of Brazil, which may have originated from the eastern coastal areas of Brazil.
［摘要］ 目的 了解我国藁杆双脐螺来源, 为我国曼氏血吸虫病流行风险评估和双脐螺控制提供依据。方法 选择我 国深圳市观澜河, 大沙河, 深圳水库, 葵涌河上、下游, 新圳河作为采样点, 每个采样点采集10个双脐螺样本, 提取螺样本 基因组DNA。自南美洲巴西米纳斯吉拉斯州、帕拉州、联邦区、伯南布哥州、圣保罗州的5个采样点获得15个藁杆双脐 螺DNA样本。对上述 DNA 样本的细胞色素c 氧化酶亚基 I (cytochrome c oxidase I, COI) 和线粒体16S核糖体RNA (16S ribosomal RNA, 16S rRNA) 基因进行扩增和测序。同时, 从GenBank中下载藁杆双脐螺COI 和16S rRNA 基因序列, 并获取 其采样点信息。将所有基因序列进行比对并构建进化树, 分析我国和南美洲双脐螺样本的遗传相似度和谱系进化关系。结果 从我国双脐螺样本中共获得60个长度为529 bp的COI序列, 其中3个为单倍型。从GenBank中获得165条藁杆双 脐螺COI 序列, 与上述60条序列比对后, 共获得33个单倍型。进化树分析显示, 采自我国的双脐螺3个单倍型聚在一支, 其中单倍型China11与GenBank中获得的来自巴西的3个样本属于同一单倍型。地理进化分析结果显示, 巴西东部沿海 3个采样点的样本有与我国 China11 相同的单倍型, 另2个采样点的样本与 China11 亲缘关系较近。扩增我国双脐螺样本 16S rDNA 基因, 共获得60条长度约为322 bp的序列和2个单倍型。从GenBank中获得70条藁杆双脐螺 16S rDNA 序列。 系统进化树分析显示, 我国双脐螺样本聚为一支, 其中单倍型China64与来自巴西的 229BS 为同一单倍型。将GenBank 中获取的来自巴西南部 25 个采样点的49个藁杆双脐螺 16S rDNA 序列纳入分析, 发现其中3个采样点的藁杆双脐螺与我 国China64有相同的单倍型。综合分析藁杆双脐螺 COI 和 16S rRNA 的地理系统进化关系, 发现仅巴西东部沿海地带样本 与我国双脐螺样本在两个基因片段序列上具有相同单倍型。结论 我国双脐螺为藁杆双脐螺, 遗传多样性较低, 与来 自巴西东部沿海地区的样本遗传学相似度很高, 可能来源于巴西东部沿海地区。.

The white-bellied pangolin is subject to intense trafficking, feeding both local and international trade networks. In order to assess its population genetics and trace its domestic trade, we genotyped 562 pangolins from local to large bushmeat markets in western central Africa. We show that the two lineages described from the study region (WCA and Gab) were overlapping in ranges, with limited introgression in southern Cameroon. There was a lack of genetic differentiation across WCA and a significant signature of isolation-by-distance possibly due to unsuspected dispersal capacities involving a Wahlund effect. We detected a c. 74.1-82.5% decline in the effective population size of WCA during the Middle Holocene. Private allele frequency tracing approach indicated up to 600 km sourcing distance by large urban markets from Cameroon, including Equatorial Guinea. The 20 species-specific microsatellite loci provided individual-level genotyping resolution and should be considered as valuable resources for future forensic applications. Because admixture was detected between lineages, we recommend a multi-locus approach for tracing the pangolin trade. The Yaoundé market was the main hub of the trade in the region, and thus should receive specific monitoring to mitigate pangolins\' domestic trafficking. Our study also highlighted the weak implementation of CITES regulations at European borders.

The emergence of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) evolved into a worldwide outbreak, with the first Polish cases in February/March 2020. This study aimed to investigate the molecular epidemiology of the circulating virus lineages between March 2020 and February 2021. We performed variant identification, spike mutation pattern analysis, and phylogenetic and evolutionary analyses for 1106 high-coverage whole-genome sequences, implementing maximum likelihood, multiple continuous-time Markov chain, and Bayesian birth-death skyline models. For time trends, logistic regression was used. In the dataset, virus B.1.221 lineage was predominant (15.37%), followed by B.1.258 (15.01%) and B.1.1.29 (11.48%) strains. Three clades were identified, being responsible for 74.41% of infections over the analyzed period. Expansion in variant diversity was observed since September 2020 with increasing frequency of the number in spike substitutions, mainly H69V70 deletion, P681H, N439K, and S98F. In population dynamics inferences, three periods with exponential increase in infection were observed, beginning in March, July, and September 2020, respectively, and were driven by different virus clades. Additionally, a notable increase in infections caused by the B.1.1.7 lineage since February 2021 was noted. Over time, the virus accumulated mutations related to optimized transmissibility; therefore, faster dissemination is reflected by the second wave of epidemics in Poland.

The aim of this study is the characterization and genomic tracing by phylogenetic analyses of 59 new SARS-CoV-2 Italian isolates obtained from patients attending clinical centres in North and Central Italy until the end of April 2020. All but one of the newly-characterized genomes belonged to the lineage B.1, the most frequently identified in European countries, including Italy. Only a single sequence was found to belong to lineage B. A mean of 6 nucleotide substitutions per viral genome was observed, without significant differences between synonymous and non-synonymous mutations, indicating genetic drift as a major source for virus evolution. tMRCA estimation confirmed the probable origin of the epidemic between the end of January and the beginning of February with a rapid increase in the number of infections between the end of February and mid-March. Since early February, an effective reproduction number (Re) greater than 1 was estimated, which then increased reaching the peak of 2.3 in early March, confirming the circulation of the virus before the first COVID-19 cases were documented. Continuous use of state-of-the-art methods for molecular surveillance is warranted to trace virus circulation and evolution and inform effective prevention and containment of future SARS-CoV-2 outbreaks.

This study aimed to trace the transmission source of Salmonella enterica serovar Typhimurium and Salmonella enterica serovar Enteritidis strains associated with enteric infections in Shanghainese children, and understand the molecular mechanism of resistance to third-generation cephalosporins and ciprofloxacin.
The profiles of pulsed-field gel electrophoresis (PFGE) were compared among the isolates from children, animal, and environment. Antimicrobial susceptibility was determined using the minimal inhibitory concentrations and Kirby-Bauer disk diffusion method. Extended-spectrum β-lactamase (ESBL) producing isolates mediated by resistance genes were identified using polymerase chain reaction and sequencing.
Based on PFGE patterns, 49 (33.1%) of 148 human Salmonella Typhimurium isolates located in the dominant PFGE clusters were genetically related to the isolates from poultry source, environment water, aquatic products, and reptiles, whereas 97 (97.0%) of 100 human Salmonella Enteritidis isolates were genetically related to isolates from poultry and water. The rates of resistance to ceftriaxone among clinical Salmonella Typhimurium and Salmonella Enteritidis isolates were 42.0% and 14.2%, respectively. Besides, 35.1% of Salmonella Typhimurium isolates displayed resistance to ciprofloxacin; 64.9% of Salmonella Typhimurium isolates and 97.0% of Salmonella Enteritidis isolates displayed reduced susceptibility to ciprofloxacin. Of 64 ESBL/AmpC-producing strains, CTX-M, TEM, DHA, and CMY were detected at frequencies of 86.0%, 62.5%, 7.8%, 3.1%, and 3.1%, respectively.
The transmission sources of Salmonella Typhimurium and Salmonella Enteritidis infections in Shanghainese children were diverse. The high prevalence of resistance to third-generation cephalosporins and ciprofloxacin mediated by multiple molecular mechanisms needs continuous monitoring and intervention.

Objective: To confirm the laboratory diagnosis of dengue bordline cases reported in Henan Province and trace its origin from molecular level in 2017. Methods: The study samples were blood samples (3-5 ml), which came from 8 suspected cases of dengue fever reported in the 2017 direct reporting system of Henan provincial infectious disease monitoring network. Meanwhile, case investigation was conducted according to National dengue fever surveillance programme. Serum were separated from blood samples and tested for Dengue NS1 antigen, IgM & IgG antibodies, and dengue RNA. According to dengue diagnosis criteria, confirmed cases were identified by testing results. Samples carried dengue RNA performed for real-time PCR genotyping and amplification of E gene. Then, the amplicons were sequenced and homological and phylogenetic analyses were constructed. Results: 8 serum samples of suspected dengue cases were collected in Henan Province, 2017. Six of them were diagnosed as dengue confirmed cases. All the dengue confirmed cases belonged to outside imported cases, 5 of them were positive by dengue RNA testing. Genotyping results showed there were 1 DENV1 case, 2 DENV2 cases and 2 DENV3 cases. A DENV2 case and a DENV3 case of this study were traced its origin successfully. The sequence of Pakistan imported DENV2 case belongs to cosmopolitan genotype, which was the most consistent with Pakistan\'s DENV2 KJ010186 in 2013 (identity 99.0%). The sequence of Malaysia imported DENV3 case belongs to genotype I, which was the most consistent with Singapore\'s DENV3 KX224276 in 2014(identity 99.0%). Conclusion: The laboratory diagnosis and molecular traceability of dengue cases in Henan Province in 2017 confirmed that all cases were imported and did not cause local epidemics.
目的： 对2017年河南省报告的登革热疑似病例进行实验室确诊，从分子水平追踪其来源。 方法： 血液标本（3~5 ml）来源于2017年河南省传染病监测网络直报系统中上报的8例登革热疑似病例，按照《全国登革热监测方案》对病例进行个案调查。血液样本分离血清后，通过检测血清中登革病毒非结构蛋白1抗原（NS1）、IgM和IgG抗体以及病毒RNA进行实验室确诊；对于登革病毒RNA检测阳性的样本进行荧光PCR分型诊断和E基因序列扩增，阳性扩增产物测序后进行同源性比对。 结果： 8个登革热疑似病例中6例确诊为登革热实验室诊断病例，均为境外输入；其中5例病例登革病毒RNA检测阳性，经分型检测发现，登革病毒1型为1例，2型和3型各2例；对1例登革病毒2型和1例3型病例成功进行了分子溯源，其中由巴基斯坦输入的登革病毒2型病例核酸序列属于cosmopolitan基因型，与巴基斯坦2013年登革病毒2型KJ010186一致性最高，为99.0%；由马来西亚输入的登革病毒3型病例核酸序列属于Ⅰ型基因型，与2014年新加坡登革病毒3型KX224276一致性最高，为99.0%。 结论： 2017年河南省登革热病例经实验室诊断和分子溯源证实均为输入性病例，未引起本地流行。.

Infectious pancreatic necrosis (IPN) is an important restraint to production of salmonids in aquaculture globally. In order to implement efficacious mitigation strategies for control of this disease, it is important to understand infection routes under current production systems. IPN virus has been shown to be transmitted vertically in Rainbow trout, from broodstock to fingerlings in hatcheries, and there is circumstantial evidence suggesting that vertical transmission can also occur in Atlantic salmon, in addition to horizontal transmission between grow-out fish in farms. In this study, we show that the smolt carries infection with IPN from hatchery to the marine farm. We do this by comparing sequences from fish groups taken both in hatcheries and on corresponding marine grow-out farms. We use statistical analysis to prove that sequences obtained from the same fish group in both hatchery and marine farm are more similar than sequences obtained from random fish groups on hatcheries and marine farms.

We investigated 543 Listeria monocytogenes isolates from food having a temporal and spatial distribution compatible with that of the invasive listeriosis outbreak occurring 2012-2016 in southern Germany. Using forensic microbiology, we identified several products from 1 manufacturer contaminated with the outbreak genotype. Continuous molecular surveillance of food isolates could prevent such outbreaks.

Worldwide koi herpesvirus (KHV) causes high mortalities in Cyprinus carpio L. aquaculture. So far, it is unknown how the different variants of KHV have developed and how they spread in the fish, but also in the environmental water bodies. Therefore, a phylogenetic method based on variable number of tandem repeats (VNTR) was improved to gain deeper insights into the phylogeny of KHV and its possible worldwide distribution. Moreover, a VNTR-3 qPCR was designed which allows fast virus typing. This study presents a useful method for molecular tracing of diverse KHV types, variants, and lineages.

Knowledge on faunal diversification in African rainforests remains scarce. We used phylogeography to assess (i) the role of Pleistocene climatic oscillations in the diversification of the African common pangolin (Manis tricuspis) and (ii) the utility of our multilocus approach for taxonomic delineation and trade tracing of this heavily poached species. We sequenced 101 individuals for two mitochondrial DNA (mtDNA), two nuclear DNA and one Y-borne gene fragments (totalizing 2602 bp). We used a time-calibrated, Bayesian inference phylogenetic framework and conducted character-based, genetic and phylogenetic delineation of species hypotheses within African common pangolins. We identified six geographic lineages partitioned into western Africa, Ghana, the Dahomey Gap, western central Africa, Gabon and central Africa, all diverging during the Middle to Late Pleistocene. MtDNA (cytochrome b + control region) was the sole locus to provide diagnostic characters for each of the six lineages. Tree-based Bayesian delimitation methods using single- and multilocus approaches gave high support for \'species\' level recognition of the six African common pangolin lineages. Although the diversification of African common pangolins occurred during Pleistocene cyclical glaciations, causative correlation with traditional rainforest refugia and riverine barriers in Africa was not straightforward. We conclude on the existence of six cryptic lineages within African common pangolins, which might be of major relevance for future conservation strategies. The high discriminative power of the mtDNA markers used in this study should allow an efficient molecular tracing of the regional origin of African common pangolin seizures.