OBJECTIVES:To investigate the etiological characteristics of acute conjunctivitis in Shenzhen, China. METHODS:A total of 1234 conjunctival swabs collected between 2018 and 2024 were examined for coxsackievirus A24 variant (CVA24v), enterovirus D70 (EV-D70) and human adenovirus (HAdV). Complete VP1 sequences of CVA24v strains were determined and analyzed. HAdV was genotyped by PCR methods targeting the three genes (penton base, hexon and fiber) and sequencing. SPSS 22.0 software was used for statistical analysis. RESULTS:CVA24v was first detected in 2023, with a detection rate of 33.3 % (63/189). No EV-D70 was detected in 2018-2024. The annual distributions of HAdV-infected patients were 52.6 % (101/192), 62.7 % (111/177), 21.1 % (37/175), 9.0 % (14/155), 10.5 % (17/162), 15.9 % (30/189) and 21.7 % (40/184), respectively. CVA24v strains from this study clustered in a clade with significant temporal aggregation characteristic within the genotype GIV. Eight amino acid variation sites (T11A, I16L, K20I, L32P, K105R, A146T, R277G and P280S) were observed in VP1 sequences of CVA24v strains from this study when compared to the close strains. Conjunctivitis patients with one of the three symptoms (weakness, fever and blurred vision) had a higher detection rate of HAdV. Eleven known HAdV genotypes (HAdV-B3, -B14, -B21, HAdV-D8, -D37, -D42, -D53, -D64, -D85, -D115 and HAdV-E4) and 7 unknown genotypes were detected in 2020-2024, with HAdV-D37 (30.9 %), HAdV-D115 (21.1 %) and HAdV-B3 (17.1 %) being the three predominant genotypes. CONCLUSIONS:The unique genetic characteristics were observed in Shenzhen CVA24v strains. HAdVs associated with conjunctivitis exhibited a high degree of genotypic diversity in Shenzhen, and HAdV-D115 related to conjunctivitis was first reported in this study.

The study aimed to investigate epidemiological profile and molecular characteristics of coxsackievirus A10 (CVA10) associated with hand, foot and mouth disease (HFMD) in Shenzhen, China and comparatively analyze genomes of CVA10 strains related to differential clinical phenotypes. A total of 3170 clinical specimens collected between 2021 and 2024 were examined for CVA10 using real-time RT-PCR. Complete VP1 sequences and near-complete genome sequences of CVA10 were determined by RT-PCR methods and sequencing. Sequences were analyzed using a series of bioinformatics programs. Two (33.33%) out of 6 severe cases were infected with CVA10. The detection rate of CVA10 associated with mild HFMD ranged from 1.21% to 6.11% in 2021-2024, with an overall detection rate of 3.73%. There was no significant difference in the infection rate of CVA10 between males and females or different age groups. The CVA10 infections mainly occurred in Spring (March to May) and Summer (June to August) in Shenzhen. Of the 74 VP1 sequences determined, 71 (95.95%) of them were detected in the sub-genotype C2, 3 (4.05%) were assigned to the genotype D. Genomic sequence analysis indicated that the genotype D of CVA10 of this study derived from genetic recombination between CVA10 and CVA16 in 3A-3D coding region (nucleotide position: 5075-6896). Different variable sites were observed in the two CVA10 strains associated with different severe complications when compared to CVA10 strains associated with mild diseases. In conclusion, CVA10 associated with HFMD circulated at a low level in Shenzhen in 2021-2024, with C2 as the predominant genotype. Recombinant genotype D of CVA10 was introduced first to Shenzhen in 2024. The study emphasizes the importance of continuous molecular surveillance of CVA10.

Maximum Entropy model (MaxEnt), as a machine learning algorithm, is widely used to identify potential risk areas for emerging infectious diseases. However, MaxEnt usually overlooks the influence of the optimal selection of spatial grid scale and the optimal combination of factor information on identification accuracy. Furthermore, the internal level information of factors is closely related to the potential risk of disease occurrence but is rarely applied to enhance MaxEnt's accuracy. In this study, the Optimal Parameters-based Geographical Detectors-Information Value-MaxEnt (OPGD-IV-MaxEnt) was first proposed to identify the potential risk areas of hand, foot, and mouth disease (HFMD) in Shenzhen and compared its identification accuracy with that of OPGD-MaxEnt and MaxEnt. Firstly, the optimal grid scale and optimal combination of factor information were determined by OPGD. Secondly, the contributions of factors' internal level information to the potential risk of HFMD occurrence were quantified and incorporated by IV. Lastly, the spatial patterns of potential risk areas and their main driving factors were elucidated. Results showed that: (i) Area under the curve (AUC) of single MaxEnt were 0.638, 0.688, 0.763, 0.796, and 0.757 at 100 m, 250 m, 500 m, 750 m, and 1000 m scale, respectively, and 750 m were deemed the optimal scale. (ii) At the optimal scale, OPGD-IV-MaxEnt (AUC = 0.868) identified potential risk areas more accurately than MaxEnt (AUC = 0.796) and OPGD-MaxEnt (AUC = 0.827). (iii) Resident (r = 0.61, q = 0.39) and Market (r = 0.61, q = 0.36) were the primary factors affecting the identification of potential risk areas. (iv) Potential high-risk areas of HFMD were mainly distributed in northwestern, southwestern, and central Shenzhen, with dense resident and market distribution. Such insights are instrumental in devising targeted infection prevention and control measures for emerging infectious diseases and provide references for improving the identification accuracy of similar machine learning algorithms.

目的 了解广东省深圳市疱疹性咽峡炎(HA)病原组成与病原体的分子特征,为HA的预防与控制提供科学依据.方法 2017-2018年收集了 157例HA患者的314份临床样本,其中粪便样本和咽拭子各157份.使用荧光定量逆转录-聚合酶链反应(RT-PCR)和基于半巢式RT-PCR扩增的测序方法对肠道病毒(EV)进行检测与分型.使用生物信息学软件对病毒VP1基因进行序列分析.结果 在126例(80.30％,126/157)EV阳性的患者中,共检出10种EV,检出率最高的是柯萨奇病毒A 组 10 型(CVA10)(19.70％,31/157),其次是 CVA4(17.80％,28/157)、CVA6(15.30％,24/157)和 CVA2(10.80％,17/157),其他病原体包括 CVA5(4.50％,7/157)、CVA 16(3.20％,5/157)、EV-A71(1.30％,2/157)、CVA8(0.60％,1/157)、CVB5(0.60％,1/157)和埃可病毒11(E11)(0.60％,1/157),1例为CVA4与CVA10的混合感染(0.60％,1/157).2017年检出率最高的两种病原体是CVA2和CVA6,而2018年却是CVA10和CVA4.粪便样本与咽拭子的病原检出率之间的差异没有统计学意义(x2=0.019,P=0.892).基于VP1序列的分子系统发育分析表明,本研究CVA10毒株均为C2基因型.CVA4除了 1株为C5基因型外,其余均为C2基因型.1株CVA6毒株位于先前未描述的进化分支,其他CVA6毒株均为D3a基因亚型.CVA2毒株均为D2基因型.结论 粪便样本与咽拭子都适用于HA的病原体核酸检测.CVA10、CVA4、CVA6和CVA2是2017-2018年深圳市HA的优势病原体,大部分毒株是我国主流基因型,个别毒株处于不常见的基因型或进化分支.

Type 2 diabetes (T2DM) is characterized by insulin secretion deficiencies and systemic insulin resistance (IR) in adipose tissue, skeletal muscle, and the liver. Although the mechanism of T2DM is not yet fully known, inflammation and insulin resistance play a central role in the pathogenesis of T2DM. G protein‐coupled receptors (GPCRs) are involved in endocrine and metabolic processes as well as many other physiological processes. GPR50 (G protein‐coupled receptor 50) is an orphan GPCR that shares the highest sequence homology with melatonin receptors. The aim of this study was to investigate the effect of GPR50 on inflammation and insulin resistance in 3T3‐L1 preadipocytes. GPR50 expression was observed to be significantly increased in the adipose tissue of obese T2DM mice, while GPR50 deficiency increased inflammation in 3T3‐L1 cells and induced the phosphorylation of AKT and insulin receptor substrate (IRS) 1. Furthermore, GPR50 knockout in the 3T3‐L1 cell line suppressed PPAR‐γ expression. These data suggest that GPR50 can attenuate inflammatory levels and regulate insulin signaling in adipocytes. Furthermore, the effects are mediated through the regulation of the IRS1/AKT signaling pathway and PPAR‐γ expression.

Background Acquired immunodeficiency syndrome (AIDS) is a malignant infectious disease with high mortality caused by HIV (human immunodeficiency virus, and up to now there are no curable drugs or effective vaccines. In order to understand AIDS's development trend, we establish hybrid EMD-BPNN (empirical modal decomposition and Back-propagation artificial neural network model) model to forecast new HIV infection in Dalian and to evaluate model's performance. Methods The monthly HIV data series are decomposed by EMD method, and then all decomposition results are used as training and testing data to establish BPNN model, namely BPNN was fitted to each IMF (intrinsic mode function) and residue separately, and the predicted value is the sum of the predicted values from the models. Meanwhile, using yearly HIV data to established ARIMA and using monthly HIV data to established BPNN, and SARIMA (seasonal autoregressive integrated moving average) model to compare the predictive ability with EMD-BPNN model. Results From 2004 to 2017, 3310 cases of HIV were reported in Dalian, including 101 fatal cases. The monthly HIV data series are decomposed into four relatively stable IMFs and one residue item by EMD, and the residue item showed that the incidence of HIV increases firstly after declining. The mean absolute percentage error value for the EMD-BPNN, BPNN, SARIMA (1,1,2) (0,1,1)(12) in 2018 is 7.80%, 10.79%, 9.48% respectively, and the mean absolute percentage error value for the ARIMA (3,1,0) model in 2017 and 2018 is 8.91%. Conclusions The EMD-BPNN model was effective and reliable in predicting the incidence of HIV for annual incidence, and the results could furnish a scientific reference for policy makers and health agencies in Dalian.

新型冠状病毒肺炎是一种由新型冠状病毒引起的传染性极强,影响范围极大的新发突发传染病.自病原体明确以来,了解其感染性和传播途径已成为关注重点.本文总结了患者多器官系统中病毒排毒相关的文献,讨论了其在呼吸道、消化道及尿液、眼泪等体液和组织中排毒的规律,发现新型冠状病毒可以经呼吸道、消化道排出感染性病毒颗粒,还有感染眼部、肾脏、睾丸、胎盘等肺外组织的潜能,且病毒载量高的患者更容易发展为重症,为病毒致病性的研究以及疫情的防控提供了一定的思路.

On January 22, 2021, a 34-year-old male, who served as a security guard at the isolation point in Shenzhen, tested positive for coronavirus disease 2019 (COVID-19) during the every-three-day routine test After the nasopharyngeal swab was further confirmed as positive for COVID-19 by Shenzhen CDC, the patient was transferred to the Third People’s Hospital of Shenzhen and was diagnosed as a COVID-19 asymptomatic infection On January 25, 2021, Shenzhen CDC identified the 20H/501 Y V2 (B 1 351) variant, which was a variant of COVID-19 virus first emerging in the South Africa The epidemiological investigation indicated that the patient (ACC-XG00731) began working as a security guard at the isolation point on July 25, 2020 in Shenzhen On January 19, his test result was negative According to his statement, in the last 14 days, he had worked on the isolation floor occasionally but had not been exposed to the individuals who were in quarantine, had no exposure to the household or medical waste generated from the isolation personnel, and did not participate in the sewage treatment of the isolation point On January 22, 2021, 1 case (ACC-XG00741) of COVID-19 infection in this isolation point was detected in entry isolation personnel from South Africa and was diagnosed on January 22 It was considered possible that case ACC-XG00731 is a descendant of the case ACC-XG00741 On January 25, 2021, the sample of the 2 cases were sequenced by Shenzhen CDC using the second-generation (MiSeq) and third-generation sequencing technology (Nanopore) Compared with the Wuhan reference (EPI_ISL_402125) (1), these 2 strains exhibited 23 to 24 nucleotide variation sites, belonging to the Pangolin lineage B 1 351 (2), which were all classified as the 20H/501 Y V2 variant (Figure 1) The genomic sequence of ACC-XG00731 and ACC-XG00741 strains were highly homologous (99 997%), sharing 23 variation sites (G174T, C241T, C1059T, A2692T, C3037T, G5230T, C9808T, A10323G, C14408T, C21614T, A21801C, A22206G, G22813T, A23403G, C23664T, G25477T, G25563T, C25904T, C26456T, C26645T, C28253T, C28887T, and G29737T) The 2021A-XG00731 strain only had one specific variation site (C16428T) As is characteristic of 501 Y V2 variant, the ACC-XG00731 strains carried several amino acid mutation sites, including K417N, E484K, and N501Y The 501Y V2 lineage, which was connected to a fast-growing epidemic, emerged in early August in South Africa (3) The 501Y V2 lineage includes several specific mutations (D80A, LAL242-244del, R246I, K417N, E484K, N501Y, D614G, and A701V) in the spike protein of the COVID-19 virus, also known as severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV-2) The N501Y mutation had also been identified in the B 1 1 7 variant (also known as 501Y V1) first emerging in the United Kingdom (UK)(4);recently, the B 1 1 7 variant was estimated to be approximately 56% more transmissible than existing viruses in circulation (5) The N501Y mutation may influence the function of spike receptor binding domain (RBD) that promotes the interaction to ACE2 on the host cells, making infection easier (6) In the case that the 501Y V2 variant first emerging from South Africa was detected in returnees and further spread to the security guard at the isolation point in Shenzhen, the management measures of the isolation points should be heightened The 501Y V2 and 501Y V1 variant had been detected in Guangdong (7) and Shanghai in China, respectively (8), and had been identified in tens of countries worldwide, posing a more serious challenge to the global epidemic Understanding the faster-spreading variants of COVID-19 virus may be beneficial for the policy decisions to contain their spread

What is known about this topic? Few major outbreaks of coronavirus disease 2019 (COVID-19) have occurred in China after major non-pharmaceutical interventions and vaccines have been deployed and implemented. However, sporadic outbreaks that had high possibility to be linked to cold chain products were reported in several cities of China.. What is added by this report? In July 2020, a COVID-19 outbreak occurred in Dalian, China. The investigations of this outbreak strongly suggested that the infection source was from COVID-19 virus-contaminated packaging of frozen seafood during inbound unloading personnel contact. What are the implications for public health practice? Virus contaminated paper surfaces could maintain infectivity for at least 17–24 days at -25 ℃. Exposure to COVID-19 virus-contaminated surfaces is a potential route for introducing the virus to a susceptible population. Countries with no domestic transmission of COVID-19 should consider introducing prevention strategies for both inbound travellers and imported goods. Several measures to prevent the introduction of the virus via cold-chain goods can be implemented.

Background Norovirus (NoV) is the main cause of non-bacterial acute gastroenteritis (AGE) outbreaks worldwide. From September 2015 through August 2018, 203 NoV outbreaks with 2,500 patients were reported to the Shenzhen Center for Disease Control and Prevention. Methods Fecal specimens were collected from the 203 outbreaks and epidemiological data were collected through the AGE outbreak surveillance system in Shenzhen. The genotypes were determined by sequencing analysis. To gain a better understanding of evolutionary characterization of NoV in Shenzhen, the molecular evolution was analyzed by time-scale evolutionary phylogeny and amino acid mutations. Results Most of these outbreaks were associated with NoV GII.P16/GII.2 strain (45.3%,92/203) and occurred in school settings (91.6%,186/203). The timescale phylogeny suggested that the GII.P16/GII.2 strain was recombination strain and were still stable. The amino acid mutations suggested that the nonstructural proteins of the recombination strain might play a more significant role than VP1 gene in these GII.P16/GII.2 recombination strain outbreaks. Conclusions This study illustrated the characteristics of the molecular epidemiological patterns in Shenzhen, China during September 2015 to August 2018 and provided the evidence that the GII.P16/GII.2 strain was static and the epidemic trend had fade.

目的 了解新型冠状病毒肺炎(COVID-19)患者体内新型冠状病毒(SARS-CoV-2)特异性抗体一免疫球蛋白(Ig)M、IgG和IgA的应答规律.方法 使用酶联免疫吸附试验检测SARS-CoV-2特异性IgM和IgG抗体,使用磁微粒化学发光法检测SARS-CoV-2特异性IgA抗体.结果 COVID-19患者血浆中SARS-CoV-2特异性IgM、IgG抗体水平(中位数)在发病后第4周达到峰值,IgA抗体一直处于较低水平.重型患者IgM和IgG抗体水平达到峰值的时间与非重型患者一致,抗体峰值水平高于非重型患者(OD 450中位数:IgM:2.837 vs.1.133,P=0.005;IgG:3.777 vs.3.376,P=0.031),发病5周后的抗体水平高于非重型患者(OD 450中位数:IgM:1.454 vs.0.263,P=0.021;IgG:3.136 vs.2.552,P=0.027).重型患者IgA抗体水平达到峰值的时间晚于非重型患者(第5周 vs.第3周),发病第4周和第5周的抗体水平均高于非重型患者(S/CO中位数:第4周:9.654vs.2.546,P=0.035;第5周:20.580 vs.2.861,P=0.046).结论 SARS-CoV-2感染可引起机体广泛的免疫应答,血清抗体检测在辅助诊断中可发挥重要作用.重型患者产生的抗体应答有一定延迟,但抗体水平高于非重型患者.

Purpose: The goal of this study was to investigate the effect of orphan G Protein-Coupled Receptor 50 (GPR50) receptor on inflammation and insulin signaling in 3T3-L1 preadipocyte. Subjects and Methods: A high-fat diet (HFD)-induced obesity-T2DM (Type 2 Diabetes Mellitus) mouse model was used in this research, and high expression of GPR50 in mouse adipose tissue was screened by microarray technology. Expression of GPR50 in 3T3-L1 cell line and obesity-T2DM mouse adipose tissue was confirmed. To gain more insight into the potential role of this new target in obesity-associated IR development, a GPR50 knockout cell line was constructed in 3T3-L1 cell line. Inflammatory cytokine levels and insulin signaling pathways in the GPR50 knockout 3T3-L1 cell line were determined by quantitative real-time polymerase chain reaction analysis and western blot. Results: GPR50 expression was significantly increased in adipose tissue of obesity-T2DM mice. GPR50 deficiency increased inflammation in 3T3-L1 cells. In addition, GPR50 deficiency induced the phosphorylation of AKT and insulin receptor substrate (IRS)1. Furthermore, GPR50 knockout 3T3-L1 cell line had suppressed PPAR-γ expression. Conclusions: These data demonstrated a novel target GPR50 can affect inflammation and insulin signaling in adipocytes. Furthermore, the effects are mediated through the regulation of insulin signaling and PPAR-γ expression.

为了解深圳境外输入的新型冠状病毒(SARS-CoV-2)的遗传特征,本研究对2021年2月六株境外输入的SARS-CoV-2毒株进行了高通量测序与基因组序列分析.测序获得的六株SARS-CoV-2毒株基因组长度分别为29 450 nt、28 936 nt﹑28 875 nt、29 855 nt、29 146 nt 和29 528 nt.根据"Pango lineages"分型法,三个来自肯尼亚、南非和柬埔寨的毒株属于B.1.1.7系(VOC-202012/01),一个来自美国的毒株属于B.1.2系(美国谱系),两个来自南非和肯尼亚的毒株属于B.1.351系(20H/501Y.V2).与武汉毒株Wuhan-Hu-1(NC_045512.2)比较,B.1.1.7系毒株的刺突蛋白(S)中发现了多达10个氨基酸的变异,B.1.2系毒株的S蛋白仅发现一个氨基酸的变异,B.1.351系毒株的S蛋白中发现了多达11个氨基酸的变异.来自柬埔寨的一株B.1.1.7系毒株的S蛋白中发现了三个变异(H69S,V70I与Y144V)与另外两个B.1.1.7系毒株中的变异(H69del,V70del与Y144del)不同.六个毒株在ORF1b上都表现出了 P314L的变异,在S蛋白上都表现出了 D614G的变异.2021年2月深圳输入了传染性更强的B.1.1.7英国变异株和B.1.351南非变异株.境外输入的SARS-CoV-2变异株存在引起本地暴发与流行的风险,需持续对境外输入的SARS-CoV-2毒株进行分子监测.

目的 探讨酶联免疫吸附法(ELISA)检测血清中严重急性呼吸综合征冠状病毒2(SARS-CoV-2)特异性抗体——免疫球蛋白(Ig)A、IgG在新型冠状病毒肺炎(COVID-19)诊断中的临床应用价值,以及探索IgA是否可以作为比IgG更灵敏的COVID-19诊断血清标志物.方法 选取47例COVID-19确诊患者为研究对象,收集其年龄、性别、发病时间、入院时间、出院时间及采样时间等信息,采集血清样本124份,利用ELISA试剂盒和酶标仪检测患者血清中SARS-CoV-2特异性IgA和IgG抗体含量.结果 患者中＞20～40岁组人数最多占40.43％,＞40～60岁组占36.17％,＞60岁患者仅占10.64％.IgG检测阳性率较高,灵敏度最好,以IgG1(59.26％)和IgG3(44.44％)亚型为主;IgA的敏感性次于IgG,几乎所有IgA阳性标本都伴随IgG阳性.IgG和IgA阳性组患者的年龄高于抗体阴性组,15岁以下儿童和70岁以上老年患者均为IgA阴性.抗体阳性组与阴性组患者的住院天数及病程时长比较差异均无统计学意义(P＞0.05).IgG抗体水平随着发病时间延长而持续升高;而IgA抗体则在患者发病后的前3周逐渐升高,然后随着发病时间延长而逐渐下降.结论 ELISA法检测COVID-19患者血清中SARS-CoV-2特异性IgG抗体的敏感性较高,有良好的辅助诊断价值.

目的 化学发光法和酶联免疫法(ELISA)应用于快速检测新型冠状病毒(SARS-CoV-2)免疫球蛋白M(IgM)和免疫球蛋白G(IgG)以及2种方法的结果比对情况.方法 采用化学发光和ELISA分析技术检测深圳市疾病预防控制中心97例新型冠状病毒肺炎(COVID-19)核酸确诊患者血清中SARS-CoV-2抗体IgM/IgG,并用这2种检测方法检测其他非SARS-CoV-2感染的血清样本100份.结果 化学发光分析法检测SARS-CoV-2 IgG的灵敏度(92.8％,90/97)高于SARS-CoV-2 IgM(61.9％,60/97),差异有统计学意义(x2=26.455,P<0.001).SARS-CoV-2 IgM联合IgG的灵敏度(96.9％,94/97)高于1gG检测灵敏度,差异无统计学意义(x2=1.687,P=0.194).化学发光分析法检测SARS-CoV-2 IgG的特异性与SARS-CoV-2 IgM一致,均为100％.ELISA分析法检测SARS-CoV-2 IgG的灵敏度(62.9％,61/97)高于SARS-CoV-2 IgM(3.1％,3/97),差异有统计学意义(x2=78.439,P<0.001).SARS-CoV-2 IgM联合IgG后的灵敏度(62.9％,61/97)与SARS-CoV-2 IgG一致,差异无统计学意义(x2=0.000,P=1.000).该试剂盒应用于SARS-CoV-2IgG检测的特异性与IgM一致,均为100％.结论 化学发光和ELISA检测方法用于SARS-CoV-2 IgG检测的灵敏度均高于SARS-CoV-2 IgM,并且化学发光检测灵敏度要高于ELISA方法.

目的 分析影响新型冠状病毒肺炎(COVID-19)患者病毒清除的相关因素,为COVID-19的防治提供科学依据.方法 选择2020年1月14日至3月9日深圳市出院的381例COVID-19患者.收集患者年龄、性别以及感染新型冠状病毒的时间、地点等基本情况,同时利用实时荧光定量PCR和化学发光法分别检测SARS-CoV-2核酸Ct值以及患者血清中SARS-CoV-2 IgG抗体水平.分析患者基本情况、确诊时Ct值和血清抗体水平高低对患者住院时间和病程的影响.结果 COVID-19患者年龄、发病时间与住院时间及病程呈正相关(P＜0.05);60岁以上男性患者的住院时间显著长于60岁以上女性患者(P＜0.05);1月份发病患者的住院时间、发病到入院的时间和病程均显著长于2月份发病患者(P＜0.05).不同感染地点和患者确诊时Ct值与住院时间和病程之间不存在相关性(P＞0.05);发病第2周核酸已转阴患者的抗体水平显著高于核酸阳性患者的抗体水平(P＜0.05).结论 COVID-19患者的年龄、性别、感染的时间段以及IgG抗体出现的早晚与病程有密切的关系,年龄越大、男性患者、感染时间越早以及早期抗体水平低都会使病程延长.

In this study, we investigated the epidemiology and molecular characteristics of enteroviruses associated with severe hand, foot and mouth disease (HFMD) in Shenzhen, China, during 2014-2018. A total of 137 fecal specimens from patients with severe HFMD were collected. Enterovirus (EV) types were determined using real-time reverse transcription polymerase chain reaction (RT-PCR), RT nested PCR, and sequencing. Sequences were analyzed using bioinformatics programs. Of 137 specimens tested, 97 (70.8%), 12 (8.8%), and 10 (7.3%) were positive for EV-A71, coxsackievirus A6 (CVA6), and CVA16, respectively. Other pathogens detected included CVA2 (2.9%, 4/137), CVA10 (2.9%, 4/137), CVA5 (0.7%, 1/137), echovirus 6 (E6) (0.7%, 1/137) and E18 (0.7%, 1/137). The most frequent complication in patients with proven EV infections was myoclonic jerk, followed by aseptic encephalitis, tachypnea, and vomiting. The frequencies of vomiting and abnormal eye movements were higher in EV-A71-infected patients than that in CVA6-infected or CVA16-infected patients. Molecular phylogeny based on the complete VP1 gene revealed no association between the subgenotype of the virus and disease severity. Nevertheless, 12 significant mutations that were likely to be associated with virulence or the clinical phenotype were observed in the 5'UTR, 2Apro, 2C, 3A, 3Dpol and 3'UTR of CVA6. Eight significant mutations were observed in the 5'UTR, 2B, 3A, 3Dpol and 3'UTR of CVA16, and 10 significant mutations were observed in the 5'UTR, VP1, 3A and 3Cpro of CVA10. In conclusion, EV-A71 is still the main pathogen causing severe HFMD, although other EV types can also cause severe complications. Potential virulence or phenotype-associated sites were identified in the genomes of CVA6, CVA16, and CVA10.

Parechovirus A (PeV-A), which causes a wide variety of diseases, is prevalent among young children. However, little is currently known about PeV-A infections in children with acute gastroenteritis in mainland China. In this study, we investigated the molecular epidemiology of acute gastroenteritis in Shenzhen, southern China, with an emphasis on PeV-A infections. A total of 1220 stool specimens from 1220 outpatient children under 5 years old with acute gastroenteritis were collected from January 2016 to December 2018. Viral RNA was detected by a real-time RT-PCR and PCR method. The PeV-A isolates were genotyped by sequencing the VP3/VP1 region. Of 1220 specimens, 148 (12.1%) were positive for PeV-A. The predominant genotype was PeV-A 1B (68.9%), followed by PeV-A 4 (12.2%), PeV-A 14 (6.1%), PeV-A 1A (5.4%), PeV-A 6 (2.7%), PeV-A 3 (2.7%) and PeV-A 5 (2.0%). It was found that 68.2% of PeV-A infections occurred in the summer and rainy months (June to September) in southern China. The majority of PeV-A-positive patients (97.3%) were younger than 24 months old. PeV-A coinfection with norovirus, rotavirus, astrovirus and adenovirus was found in thirty specimens (30/148, 20.3%), five specimens (5/148, 3.4%), five specimens (5/148, 3.4%), and two specimens (2/148, 1.4%), respectively. Coinfections with more than one other enteric virus were not observed in any of the PeV-A-positive specimens. Phylogenetic analysis revealed that the PeV-A isolates from Shenzhen were closely related to each other and to strains circulating in China, suggesting endemic circulation of PeV-A in China. The results of this study indicate that PeV-A is one of important pathogens of acute gastroenteritis in young children and that coinfection is a possible mode of PeV-A infection. PeV-A associated with acute gastroenteritis exhibited high genotypic diversity in Shenzhen, southern China.

Objective To investigate the genetic characteristics of VP1 genes carried by coxsack-ievirus A16 strains isolated from cases of hand foot and mouth disease ( HFMD) in Shenzhen during 2016 to 2017. Methods Fecal and anal swab specimens were collected from patients with mild HFMD in four senti-nel hospitals and the Institute of Pathogen Biology, Shenzhen Center for Disease Control and Prevention, China during 2016 to 2017. All specimens were tested for CVA16 viral RNA using real-time RT-PCR. The VP1 genes of 51 randomly selected CVA16 strains were amplified by RT-PCR and then sequenced using TaKaRa Biomedical Technology ( Dalian). Bioinformatics software, including Mega6. 02, BioEdit and DNAStar, was used for comparison and analysis of the VP1 genes. Results CVA16 strains in Shenzhen during 2016 to 2017 mainly belonged to B1a and B1b subtypes as well as an emerging subtype B3. The epi-demic of B1b subtype was found in both 2016 (28 strains) and 2017 (19 strains), while the B1a subtype ( two strains) was only detected in 2017. Two B3 subtype strains were detected in 2017. The strains of B1b subtype were closely related to the strains isolated in Shanghai ( JQ314149 ) , Wenzhou ( KP289416 ) and Beijing (KU254598), while the B1a subtype strains were closely related to the strains isolated in Kunming (JQ316639) and Tailand (GQ184139). The B3 subtype strain was an emerging CVA16 epidemic strain in mainland China. Further comparison of the CVA16 epidemic strains in Shenzhen area during 2016 to 2017 with the CVA16 strains causing severe neurological symptoms showed that two amino acid mutations ( S14N and M23L) were found in VP1 protein. Conclusions The epidemic strains of CVA16 were B1b subtype in Shenzhen area in 2016. However, B1a, B1b and the emerging B3 subtype strains were prevalent in 2017. Compared with the CVA16 strains causing severe neurological symptoms, the CVA16 strains circulating in Shenzhen during 2016 to 2017 carried two amino acid mutations inVP1 protein.