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Mukawa virus (MKWV), a novel tick-borne virus (TBV) of the genus Phlebovirus of family Phenuiviridae , has been firstly reported in Ixodes persulcatus in Japan. In this study, we made an epidemiological investigation in China to obtain the geographic distribution and genetic features of this virus outside Japan. We screened 1,815 adult ticks (665 I. persulcatus , 336 Dermacentor silvarum , 599 Haemaphysalis longicornis , 170 Rhipicephalus microplus , 45 Haemaphysalis concinna ) and 805 wild small mammals collected from eight provinces. The positive rate of 6.77% (45/665, including 18 female and 27 male I. persulcatus ) and 2.22% (1/45, 1 male H. concinna ) were obtained from I. persulcatus and H. concinna in Heilongjiang province, respectively. No evidence of MKWV infection was found in other three tick species or any of the mammalian species. The virus can infect the Vero cells successfully, indicating the ability of MKWV to replicate in mammalian cells. A phylogenetic tree based on the nucleotide sequences of L, M, and S segments demonstrated that the Japanese MKWV variant, our two MKWV variants, and KURV were clustered with the members of the mosquito/sandfly-borne phleboviruses and distant from other tick-borne phenuiviruses. A phylogenetic analysis based on 895 bp partial L gene sequences ( n = 46) showed that all MKWV sequences were separated into three lineages. Our results showed the presence of MKWV in I. persulcatus and H. concinna in northeast of China, highlighting the necessity of epidemiological study in wider regions. Due to the ability of MKWV to replicate in mammalian cells, the potential for zoonosis, and wide distribution of I. persulcatus and H. concinna in China, the important vectors of MKWV, further screening to more tick species, wild animals, domestic animals, and humans raises up practical significance., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Wang, Jiang, Ding, Zhang, Wang, Zhang, Li, Chen, Zhang, Li, Jiang, Liu, Yu, Wang, Zhang and Liu.)

Background: Severe fever with thrombocytopenia (SFTS) caused by SFTS virus (SFTSV) was a tick-borne hemorrhagic fever that posed significant threat to human health in Eastern Asia. The study was designed to measure the seroprevalence of SFTSV antibody in healthy population residing in a high endemic region., Methods: A cohort study was performed on healthy residents in Shangcheng County in Xinyang City from April to December in 2018, where the highest SFTS incidence in China was reported. Anti-SFTSV IgG was measured by indirect enzyme-linked immunosorbent assay and neutralizing antibody (NAb) was detected by using PRNT50. The logistic regression models were performed to analyze the variables that were associated with seropositive rates., Results: Totally 886 individuals were recruited. The baseline seroprevalence that was tested before the epidemic season was 11.9% (70/587) for IgG and 6.8% (40/587) for NAb, which was increased to 13.4% (47/350) and 7.7% (27/350) during the epidemic season, and further to 15.8% (80/508) and 9.8% (50/508) post epidemic. The IgG antibody-based seropositivity was significantly related to the patients aged ≥ 70 years old [adjusted odds ratio (OR) = 2.440, 95% confidence interval (CI): 1.334-4.461 compared to the group of < 50 years old, P = 0.004], recent contact with cats (adjusted OR = 2.195, 95% CI: 1.261-3.818, P = 0.005), and working in tea garden (adjusted OR = 1.698, 95% CI: 1.002-2.880, P = 0.049) by applying multivariate logistic regression model. The NAb based seropositivity was similarly related to the patients aged ≥ 70 years old (adjusted OR = 2.691, 95% CI: 1.271-5.695 compared to the group of < 50 years old, P = 0.010), and recent contact with cats (OR = 2.648, 95% CI: 1.419-4.941, P = 0.002). For a cohort of individuals continually sampled with 1-year apart, the anti-SFTSV IgG were maintained at a stable level, while the NAb level reduced., Conclusions: Subclinical infection might not provide adequate immunity to protect reinfection of SFTSV, thus highlighting the ongoing threats of SFTS in endemic regions, which called for an imperative need for vaccine development. Identification of risk factors might help to target high-risk population for public health education and vaccination in the future., (© 2021. The Author(s).)

Human parechoviruses (HPeVs) are important causes of infection in children. However, without a comprehensive and persistent surveillance, the epidemiology and clinical features of HPeV infection remain ambiguous. We performed a hospital-based surveillance study among three groups of pediatric patients with acute respiratory infection (Group 1), acute diarrhea (Group 2), and hand, foot and mouth disease (Group 3) in Chongqing, China, from 2009 to 2015. Among 10,212 tested patients, 707 (6.92%) were positive for HPeV, with the positive rates differing significantly among three groups (Group 1, 3.43%; Group 2, 14.94%; Group 3, 3.55%; P < 0.001). The co-infection with other pathogens was detected in 75.2% (531/707) of HPeV-positive patients. Significant negative interaction between HPeV and Parainfluenza virus (PIV) ( P = 0.046, OR = 0.59, 95% CI = 0.34-0.98) and positive interactions between HPeV and Enterovirus (EV) ( P = 0.015, OR = 2.28, 95% CI = 1.23-4.73) were identified. Among 707 HPeV-positive patients, 592 (83.73%) were successfully sequenced, and 10 genotypes were identified, with HPeV1 ( n = 396), HPeV4 ( n = 86), and HPeV3 ( n = 46) as the most frequently seen. The proportion of genotypes differed among three groups ( P < 0.001), with HPeV1 and HPeV4 overrepresented in Group 2 and HPeV6 overrepresented in Group 3. The spatial patterns of HPeV genotypes disclosed more close clustering of the currently sequenced strains than those from other countries/regions, although they were indeed mixed. Three main genotypes (HPeV1, HPeV3, and HPeV4) had shown distinct seasonal peaks, highlighting a bi-annual cycle of all HpeV and two genotypes (HPeV 1 and HPeV 4) with peaks in odd-numbered years and with peaks in even-numbered years HPeV3. Significantly higher HPeV1 viral loads were associated with severe diarrhea in Group 2 ( P = 0.044), while associated with HPeV single infection than HPeV-EV coinfection among HFMD patients ( P = 0.001). It's concluded that HPeV infection was correlated with wide clinical spectrum in pediatric patients with a high variety of genotypes determined. Still no clinical significance can be confirmed, which warranted more molecular surveillance in the future., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Zhang, Zhao, Chen, Yuan, Tang, Zhou, Ren, Lu, Wang, Zhang, Zhang, Fang, Zhou, Liu, Xu and Liu.)

Beilong virus (BeiV), a member of the newly recognized genus Jeilongvirus of family Paramyxoviridae, has been reported with limited geographic and host scopes, only in Hongkong, China and from two rat species. Here, by next-generation sequencing (NGS) on dominant wild small animal species in 4 provinces in China, we obtained a complete sequence of BeiV strain from Rattus norvegicus in Guangdong, neighboring HongKong, China. We then made an expanded epidemiological investigation in 11 provinces to obtain the geographic distribution and genetic features of this virus. Altogether 7168 samples from 2005 animals (1903 rodents, 100 shrews, 2 mustelidaes) that belonged to 33 species of Cricetidae, Muridae, Sciuridae and Dipodidae family of Rodentia, 3 species of Soricidae family of Soricomorpha, 2 species of Mustelidae family of Carnivora were examined by RT-PCR and sequencing. A positive rate of 3.7% (266/7168) was obtained that was detected from 22 animal species, including 5 species of Cricetidae family, 12 species of Muridae family, 2 species of Sciuridae family and 3 species of Soricidae family. Phylogenetic analyses based on 154 partial Large gene sequences grouped the current BeiV into two lineages, that were related to their geographic regions and animal hosts. Our study showed the wide distribution of BeiV in common species of wild rodents and shrews in China, highlighting the necessity of epidemiological study in wider regions., (Copyright © 2020 The Authors. Published by Elsevier B.V. All rights reserved.)

Severe Fever with Thrombocytopenia Syndrome (SFTS) is an emerging infectious disease caused by a novel bunyavirus, SFTS virus (SFTSV), with fatal outcome developed in approximately 17% of the cases. Thrombocytopenia is a hallmark feature of SFTS, and associated with a higher risk of fatal outcome, however, the pathophysiological involvement of platelet in the clinical outcome of SFTS remained under-investigated. In the current study, by retrospectively analyzing 1538 confirmed SFTS patients, we observed that thrombocytopenia was associated with enhanced activation of the cytokine network and the vascular endothelium, also with a disturbed coagulation response. The platelet phenotypes were also extensively altered in the process of thrombocytopenia development of SFTS patients. More importantly, all these disturbed host responses were related to the severity of thrombocytopenia, thus were considered to play in a synergistic way to influence the disease outcome. Moreover, the clinical effect of platelet transfusion was assessed by comparing two groups of patients with or without receiving this therapy. As a result, we observed no therapy effect in altering frequencies of fatal outcome, clinical bleeding development, or dynamic change of platelet count during the hospitalization. It's suggested that platelet supplementation alone acted a minor role in improving disease outcome, therefore new therapeutic intervention to regulate host response should be proposed. The current results revealed some evidence of interrelationship between platelet count and clinical outcome of SFTS disease from the perspective of activation of the cytokine network, the vascular endothelium, and the coagulation/fibrinolysis system. These evaluations might help to attain a better understanding of the pathogenesis and therapy choice in SFTS., Competing Interests: The authors have declared that no competing interests exist.

Severe fever with thrombocytopenia syndrome virus (SFTSV) is a novel phlebovirus in the Bunyaviridae family, causing SFTS with high mortality rate. Haemaphysalis longicornis ticks has been demonstrated as a competent vector of SFTSV by experimental transmission study and field study. However, there has been query whether other tick species that infest human beings in the SFTS endemic regions are capable of transmitting the pathogen. Here by performing experimental transmission study, we compared the capable of transmitting SFTSV among Ixodes sinensis, Ixodes persulcatus and Dermacentor silvarum ticks. The transovarial transmission was seen in the I. sinensis ticks with a rate of 40%, but neither in I. persulcatus nor in D. silvarum ticks. I. sinensis ticks also have the ability to transmit SFTSV horizontally to uninfected mice at 7 days after feeding, but not for I. persalcatus or D. silvarum ticks. In the transstadial transmission of I. persulcatus and D. silvarum ticks, I. persulcatus ticks were tested negative from larvae to adults. But the D. silvarum ticks were tested positive from larvae to nymphs, with the positive rate of 100% (10/10) for engorged larval ticks and 81.25% (13/16) for molted nymphs. However, the mice bitten by SFTSV-infected D. silvarum nymphs were negative for SFTSV detection. Therefore, there is not enough evidence to prove the transstadial transmission of SFTSV in I. persalcatus and D. silvarum ticks., Competing Interests: The authors have declared that no competing interests exist.

In 2015, we evaluated 221 patients with undifferentiated fever and tick bite or animal exposure in Xinyang, China, for Rickettsia infection. Three with mild disease were infected with Candidatus R. xinyangensis, which clustered with R. fournieri and R. vini in phylogenetic analyses. Field investigations suggest Haemaphysalis longicornis ticks might be involved in transmission.

An effective differentiation between severe fever with thrombocytopenia syndrome and hemorrhagic fever with renal syndrome was attained by a model considering patients' age, mouse/tick contact, presence of blush, low back pain, diarrhea, enlarged lymph nodes, and white blood cell count., (© The Author(s) 2019. Published by Oxford University Press on behalf of Infectious Diseases Society of America.)

Severe fever with thrombocytopenia syndrome (SFTS), an emerging tick-borne infectious disease caused by a novel phlebovirus (SFTS virus, SFTSV), was listed among the top 10 priority infectious diseases by the World Health Organization due to its high fatality of 12%-50% and possibility of pandemic transmission. Currently, effective anti-SFTSV intervention remains unavailable. Here, by screening a library of FDA-approved drugs, we found that benidipine hydrochloride, a calcium channel blocker (CCB), inhibited SFTSV replication in vitro. Benidipine hydrochloride was revealed to inhibit virus infection through impairing virus internalization and genome replication. Further experiments showed that a broad panel of CCBs, including nifedipine, inhibited SFTSV infection. The anti-SFTSV effect of these two CCBs was further analyzed in a humanized mouse model in which CCB treatment resulted in reduced viral load and decreased fatality rate. Importantly, by performing a retrospective clinical investigation on a large cohort of 2087 SFTS patients, we revealed that nifedipine administration enhanced virus clearance, improved clinical recovery, and remarkably reduced the case fatality rate by >5-fold. These findings are highly valuable for developing potential host-oriented therapeutics for SFTS and other lethal acute viral infections known to be inhibited by CCBs in vitro.

During 2014-2017, we screened for Rickettsia japonica infection in Xinyang, China, and identified 20 cases. The major clinical manifestations of monoinfection were fever, asthenia, myalgia, rash, and anorexia; laboratory findings included thrombocytopenia and elevated hepatic aminotransferase concentrations. Physicians in China should consider R. japonica infection in at-risk patients.

Severe fever with thrombocytopenia syndrome (SFTS) is an emerging infectious disease that is caused by a novel bunyavirus SFTSV. Currently our knowledge of the host-related factors that influence the pathogenesis of disease is inadequate to allow prediction of fatal outcome. Here we conducted a prospective study of the largest database on the SFTS patients, to identify the presence of comorbidities in SFTS, and estimate their effect on the fatal outcome. Among 2096 patients eligible for inclusion, we identified nine kinds of comorbidities, from which hyperlipidemia (12.2%; 95% CI: 10.8%-13.6%), hypertension (11.0%; 95% CI: 9.6%-12.3%), chronic viral hepatitis (CVH) (9.3%; 95% CI: 8.1%-10.5%), and diabetes mellitus (DM) (6.8%; 95% CI: 5.7%-7.9%) were prevalent. Higher risk of death was found in patients with DM (adjusted OR = 2.304; 95% CI: 1.520-3.492; P<0.001), CVH (adjusted OR = 1.551; 95% CI: 1.053-2.285; P = 0.026) and chronic obstructive pulmonary diseases (COPD) (adjusted OR = 2.170; 95% CI: 1.215-3.872; P = 0.009) after adjusting for age, sex, delay from disease onset to admission and treatment regimens. When analyzing the comorbidities separately, we found that the high serum glucose could augment diseases severity. Compared to the group with max glucose < 7.0 mmol/L, patients with glucose between 7.0-11.1 mmol/L and glucose ≥11.1 mmol/L conferred higher death risk, with the adjusted OR to be 1.467 (95% CI: 1.081-1.989; P = 0.014) and 3.443 (95% CI: 2.427-4.884; P<0.001). Insulin therapy could effectively reduce the risk of severe outcome in DM patients with the adjusted OR 0.146 (95% CI: 0.058-0.365; P<0.001). For CVH patients, severe damage of liver and prolongation of blood coagulation time, as well as high prevalence of bleeding phenotype were observed. These data supported the provocative hypothesis that treating SFTS related complications can attain potentially beneficial effects on SFTS., Competing Interests: The authors have declared that no competing interests exist.

In this study, human enterovirus C117 (EV-C117) was detected in a 3-month-old boy diagnosed with pneumonia in China. A phylogenetic analysis showed that this strain was genetically closer to the Lithuanian strain than to the USA strain.

Severe fever with thrombocytopenia syndrome (SFTS) caused by a recently identified bunyavirus, SFTSV, is an emerging infectious disease with extensive geographical distribution and high mortality. Progressive viral replication and severe thrombocytopenia are key features of SFTSV infection and fatal outcome, whereas the underlying mechanisms are unknown. We revealed arginine deficiency in SFTS cases by performing metabolomics analysis on two independent patient cohorts, suggesting that arginine metabolism by nitric oxide synthase and arginase is a key pathway in SFTSV infection and consequential death. Arginine deficiency was associated with decreased intraplatelet nitric oxide (Plt-NO) concentration, platelet activation, and thrombocytopenia. An expansion of arginase-expressing granulocytic myeloid-derived suppressor cells was observed, which was related to T cell CD3-ζ chain down-regulation and virus clearance disturbance, implicating a role of arginase activity and arginine depletion in the impaired anti-SFTSV T cell function. Moreover, a comprehensive measurement of arginine bioavailability, global arginine bioavailability ratio, was shown to be a good prognostic marker for fatal prediction in early infection. A randomized controlled trial demonstrated that arginine administration was correlated with enhanced Plt-NO concentration, suppressed platelet activation, and elevated CD3-ζ chain expression and eventually associated with an accelerated virus clearance and thrombocytopenia recovery. Together, our findings revealed the arginine catabolism pathway-associated regulation of platelet homeostasis and T cell dysregulation after SFTSV infection, which not only provided a functional mechanism underlying SFTS pathogenesis but also offered an alternative therapy choice for SFTS., (Copyright © 2018 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.)

Severe fever with thrombocytopenia syndrome virus (SFTSV) infection typically causes acute fever, thrombocytopenia and leucopenia, presenting with a high case fatality rate. The pathogenesis of SFTSV infection, however, is not well described. It was hypothesized that endothelial dysfunction might play part in the disease process. In current study, we retrospectively analyzed the clinical manifestations among a large group of confirmed SFTS cases and found evidence of plasma leakage and vascular endothelial injury. Then we established a SFTSV infection cell model and determined the infectivity and stimulation of SFTSV on vascular endothelial cells in vitro. The hyperpermeability of endothelial cells directly induced by SFTSV was confirmed by electrical resistance and dextran diffusion assay. The virus induced alterations of cell junctions and cytoskeleton was also revealed. It's suggested that vascular endothelial cell injury and barrier function damage were induced after SFTSV infection, which is a vital but neglected pathogenesis of SFTS., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Severe fever with thrombocytopenia syndrome (SFTS) is an emerging infectious disease that is caused by a novel bunyavirus, SFTSV. We assessed whether the single nucleotide polymorphisms (SNPs) in the tumor necrosis factor-alpha (TNF-α) were associated with risk to severity of SFTS. Five TNF-α SNPs (SNP1: T-1031C; SNP2: C-863A; SNP3: C-857T; SNP4: G-308A; SNP5: G-238A) were genotyped in 987 hospitalized SFTS patients and 633 asymptomatic/mild SFTSV-infected subjects of Chinese Han origin. Multivariate logistic regression analysis was used to calculate adjusted odds ratios (ORs) and 95% confidence intervals (95% CIs). The hospitalized SFTS patients had significantly lower frequency of G-238A A allele than those with mild/asymptomatic infection (P = 0.006). Furthermore, T-1031C C allele (P < 0.001) and G-238A A allele (P < 0.001) were significantly associated with decreased risk of death. Multiple haplotypes were significantly associated with decreased risk of SFTS hospital admission (SNP1-2, CC; SNP1-3, CCC; SNP1-4, CCCG; SNP1-5, CCCGA; SNP2-4, CCGA; SNP3-5, CGA; SNP4-5, GA) and death (SNP1-2, CA; SNP1-3, CAG; SNP1-4, CACG; SNP1-5, CACGG; SNP2-3, AC; SNP2-4, ACG; SNP2-5, ACGG) after correction for multiple comparisons. By using the ELISA assay, we observed that TNF-α concentration of hospitalized patients was significantly increased in acute phase than in convalescent phase (P < 0.001). Elevated TNF-α concentration was also revealed from fatal patients (P < 0.001). The -238A allele was associated with decreased serum TNF-α levels in SFTS patients in acute phase (P = 0.01). Our findings suggest that polymorphisms in TNF-α gene may play a role in mediating the risk to disease severity of SFTS in Chinese Han population., Competing Interests: The authors have declared that no competing interests exist.

Background: A wide variety of pathogens could be maintained and transmitted by Haemaphysalis longicornis. The aim of this study is to systematically examine the variety of pathogens carried by Haemaphysalis longicornis, an importnatn vector, in tick-borne diseases epidemic area, and to estimate the risk of human infection imposed by tick bites., Methods: Adult questing ticks were collected in Xinyang, central China. Genomic DNA and RNA were extracted from 144 H. longicornis ticks individually, and sequenced respectively as the templates for high-throughput sequencing. Clean reads were compared against the database of NCBI nucleotide collection and specific PCR was performed to confirm the presence of pathogen. Phylogenetic analysis was performed to explore the evolutionary status of pathogens., Results: The assignment of reads to taxa based on BLASTN results revealed the existence of several potential pathogens, including Anaplasma spp., Rickettsia spp., Babesia sp., as well as severe fever with thrombocytopenia syndrome bunyavirus (SFTSV). Comfirmantory PCR assays revealed the existence of Anaplasma bovis (13/144, 9.03%), Anaplasma centrale (2/144, 1.39%), Rickettsia heilongjiangensis (3/144, 2.08%), Rickettsia sp. LON-13 (1/144, 0.69%), Rickettsia raoultii (5/144, 3.47%), Babesia sp. (1/144, 0.69%). SFTSV accounted for the highest detected pathogen with a positive rate of 18.75% (27/144). Three of the ticks (2.08%) were co-infected with SFTSV and A. bovis., Conclusion: Our study provided a broadened list of microorganism that harbored by H. longicornis. In previously unrecognized endemic regions, prokaryotic and eukaryotic infection including Anaplasma spp., Rickettsiae spp., and Babesia spp. should be considered, along with the well-known SFTSV for patients with tick bites history. A novel Babesia species was identified in local natural foci, which needs further investigation in the future.

Background: Rickettsia raoultii is frequently detected in multiple tick species, whereas human infection remains scarcely studied., Methods: A surveillance study was performed at 3 sentinel hospitals in China, to recruit participants with suspected tick exposure. Rickettsia raoultii infection was identified through polymerase chain reaction, followed by sequencing, and confirmed serologically. Isolation by cell culture was performed and the isolates were genome sequenced., Results: Twenty-six subjects were determined to have R. raoultii infection, including 7 with asymptomatic infection, 15 with mild to moderate illness, and 4 with severe illness. Common nonspecific manifestations in the 19 patients with mild to moderate or severe illness included fever (100%), malaise (95%), myalgia (58%), lymphadenopathy (53%), and nausea (42%). Only 5% of them had rash, and 16% had eschar. Scalp eschar and neck lymphadenopathy after a tick bite syndrome was only seen in 2 patients. Of the 4 patients with severe complications, 3 developed pulmonary edema, and 1 developed clouding of consciousness and lethargy. Frequent abnormalities of laboratory testing included leukopenia, thrombocytopenia, lymphopenia, neutropenia, hypoproteinemia, and elevated levels of total bilirubin, hepatic aminotransferases, lactate dehydrogenase, and creatine kinase. All the 19 patients recovered without sequelae after receiving doxycycline treatment. Two R. raoultii strains were isolated, and a significantly less degraded genome was observed than other more virulent Rickettsia strains, indicating a low pathogenicity of the current strain., Conclusions: Human infection with R. raoultii has a wide clinical spectrum that ranged from subclinical infection to severe complications. Physicians need to be aware of the high potential and clinical complexity of R. raoultii infection, to ensure appropriate testing and treatment in endemic regions.

Background: Pathogenesis of severe fever with thrombocytopenia syndrome (SFTS) has not been well described yet. Recent studies indicate that SFTSV could replicate in endothelial cells. Here we performed a case-control study to determine whether endothelial activation/dysfunction occurred in SFTSV infection and to identify the biomarkers reflecting endothelial dysfunction., Methodology/principal Findings: In a case-control study of 134 SFTS patients and 68 healthy controls, serum levels of plasminogen activator inhibitor 1, tissue plasminogen activator, P-selectin, platelet endothelial cell adhesion molecular, CD40 ligand, E-selectin, vascular endothelial growth factor A, serum amyloid antigen 1 (SAA-1) and vascular cell adhesion molecular 1 were significantly enhanced in the patients than the controls (all P<0.05), indicating the occurrence of endothelial activation/dysfunction in SFTS. The intercellular adhesion molecular 1 (ICAM-1) and SAA-1 at the convalescent phase were also significantly associated with severe patients, after adjusting for the potential confounders. The odds ratio was estimated to be 3.364 (95% CI 1.074-10.534) for ICAM-1, and 1.881 (95% CI 1.166-3.034) for SAA-1, respectively. Cutoff value of 1.1×107 pg/mL SAA-1 or 1.2×106 pg/mL ICAM-1 were found to have moderate power of predicting fatal cases., Conclusions: The endothelial dysfunction may be one of the pathogenic mechanism of SFTS. The serum levels of ICAM-1 and SAA-1 might be used to predict adverse outcome.

Aim: To explore the molecular epidemiology and clinical characteristics of Washington University polyomavirus (WUPyV) infection in pediatric patients with acute respiratory tract infections in China., Materials & Methods: A laboratory surveillance was performed to recruit pediatric patients with acute respiratory tract infections. WUPyV was detected using real-time PCR and complete genome was sequenced for randomly selected positive nasopharyngeal aspirate., Results: Altogether 122 (7.5%) of 1617 children found to be infected with WUPyV and 88 (72.1%) were coinfected with other viruses during 2012-2015. The phylogenetic analysis showed that 14 strains from our study formed two new clusters (Id and IIIc) within the Branch I and Branch III, respectively., Conclusion: WUPyV is persistently circulating in China. Surveillance on WUPyV infection in wider areas and long persistence is warranted.

Only 4 species of spotted fever group rickettsiae have been detected in humans in China. However, phylogenetic analysis of samples from 5 ill patients in China indicated infection with a novel spotted fever group Rickettsia, designated Rickettsia sp. XY99. Clinical signs resembled those of severe fever with thrombocytopenia syndrome.

During 2013-2015 in central China, co-infection with spotted fever group rickettsiae was identified in 77 of 823 patients infected with severe fever with thrombocytopenia syndrome virus. Co-infection resulted in delayed recovery and increased risk for death, prompting clinical practices in the region to consider co-infection in patients with severe fever with thrombocytopenia syndrome.

Background: Severe fever with thrombocytopenia syndrome (SFTS), caused by novel bunyavirus (SFTSV) is a potentially fatal disease that was first identified in China. Person to person transmission through contact with blood or body fluids was considered as an important infection route., Objectives: The study is designed to investigate the longitudinal viral loads following SFTSV infection and to identify factors affecting viral shedding in SFTS patients., Methods: A prospective, observational study was performed on 208 laboratory-confirmed SFTSV infected patients in Xinyang, Henan Province. Sequential serum samples were collected on admission and during the hospitalization for quantification of SFTSV RNA by real-time RT-PCR., Results: The viral RNA was undetectable in 55.6% of the patients on admission into the hospital, becoming detectable in most cases until three days and attained maximum level on six days after disease onset. This was followed by an obvious decrease thereafter, but maintained detectable for over 20 days. Viral load was independently predictable of severe disease outcome throughout the hospitalization. Viral load of >10(7)copies/mL was predictable of fatal outcome. The serum levels of PLT, WBC, LDH, AST and CK were significantly associated with viral loads level., Conclusions: The diagnosis of SFTSV infection based on PCR test should be performed at least three days after disease onset. Peaking viral loads were attained around six days after disease, posing a highest risk of human-to-human transmission., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Objective: To investigate the pregnancy complications, perinatal outcomes, and congenital abnormalities (CAs) that occurred in Beijing, China, when pregnant women became infected with the 2009 pandemic influenza A (H1N1) (H1N1 pdm)., Methods: Pregnancy complications, perinatal outcomes, and CAs were compared among 3 groups of pregnant women. The 23 women in group 1 were confirmed to harbor viral RNA; the 23 in group 2 had serum levels of virus-specific antibodies against H1N1 pdm, meaning that they were suspected of being infected with the virus; and the 93 in group 3 had no detectable virus-specific antibodies., Results: Perinatal outcomes and pregnancy complications were not significantly different in groups 1 and 3. Higher percentages of stillbirths (12.0%) and placental disorders (13.0%) were observed in group 2 than in group 3. Many women in group 2 (62.5%) experienced symptoms of having a cold during pregnancy and most took no medication. Two cases of CA occurred in group 1, in the offspring of women infected in the second trimester., Conclusion: When left untreated, infection with the 2009 H1N1 pdm virus during pregnancy appears to have increased fetal mortality and morbidity. Because CAs are traumatic for all concerned, their possible association with the virus should be further evaluated., (Copyright © 2011. Published by Elsevier Ireland Ltd.)

Thirty-two strains of Borrelia burgdorferi sensu lato were isolated from Ixodes persulcatus ticks collected from northeastern China from May to June in 2004 and 2005. Restriction fragment length polymorphism (RFLP) analysis and sequence analysis of 5S-23S rRNA intergenic spacer revealed that 29 (90.6%) belonged to Borrelia garinii, demonstrating B, C, and a unique pattern. The remaining three isolates (9.4%) were Borrelia afzelii with pattern D. The phylogenetic analysis based on 5S-23S rRNA intergenic spacer showed that B. garinii and B. afzelii genospecies clustered into two separate lineages. B. garinii strains were classified into three different branches: All the strains with RFLP pattern C were in the same branch, strain VH10 with a unique RFLP pattern clustered with strains VH9 and MDH2 with pattern B, and the rest of the strains with pattern B constitute another branch. These findings demonstrate the genetic diversity of B. burgdorferi sensu lato isolates from northeastern China.

Background: Hemorrhagic fever with renal syndrome (HFRS) is highly endemic in mainland China, and has extended from rural areas to cities recently. Beijing metropolis is a novel affected region, where the HFRS incidence seems to be diverse from place to place., Methodology/principal Findings: The spatial scan analysis based on geographical information system (GIS) identified three geo-spatial "hotspots" of HFRS in Beijing when the passive surveillance data from 2004 to 2006 were used. The Relative Risk (RR) of the three "hotspots" was 5.45, 3.57 and 3.30, respectively. The Phylogenetic analysis based on entire coding region sequence of S segment and partial L segment sequence of Seoul virus (SEOV) revealed that the SEOV strains circulating in Beijing could be classified into at least three lineages regardless of their host origins. Two potential recombination events that happened in lineage #1 were detected and supported by comparative phylogenetic analysis. The SEOV strains in different lineages and strains with distinct special amino acid substitutions for N protein were partially associated with different spatial clustered areas of HFRS., Conclusion/significance: Hotspots of HFRS were found in Beijing, a novel endemic region, where intervention should be enhanced. Our data suggested that the genetic variation and recombination of SEOV strains was related to the high risk areas of HFRS, which merited further investigation.

Thirty-two strains of Borrelia burgdorferi sensu lato were isolated from Ixodes persulcatus ticks collected from northeastern China from May to June in 2004 and 2005. Restriction fragment length polymorphism (RFLP) analysis and sequence analysis of 5S-23S rRNA intergenic spacer revealed that 29 (90.6%) belonged to Borrelia garinii, demonstrating B, C, and a unique pattern. The remaining three isolates (9.4%) were Borrelia afzelii with pattern D. The phylogenetic analysis based on 5S-23S rRNA intergenic spacer showed that B. garinii and B. afzelii genospecies clustered into two separate lineages. B. garinii strains were classified into three different branches: All the strains with RFLP pattern C were in the same branch, strain VH10 with a unique RFLP pattern clustered with strains VH9 and MDH2 with pattern B, and the rest of the strains with pattern B constitute another branch. These findings demonstrate the genetic diversity of B. burgdorferi sensu lato isolates from northeastern China.

Hantavirus genome sequences were recovered from lung tissues of Chinese white-bellied rats (Niviventer confucianus) captured in Yunnan province, China. Pairwise comparison of the nucleotide and deduced amino acid sequences of the entire S and partial M and L segments indicated that the newly discovered virus strain, which was designated as strain YN509, was very different from other rodent-borne hantaviruses. Phylogenetic analysis showed that the new strain fit into a clade containing Da Bie Shan virus (DBSV) (also carried by N. confucianus), which is mainly found in Anhui Province in mainland China. Strain YN509 appears to be in a sister taxa of the DBSV group described previously. These data suggest that strain YN509 is a new subtype of DBSV, which appears to be widely distributed in China with a higher genetic diversity than expected., ((c) 2010 Elsevier B.V. All rights reserved.)

To characterize the strains of Anaplasma phagocytophilum in wild and domestic animals in China, we isolated the organism from rodents and sheep in northeastern China. We isolated 3 strains (2 from rodents and 1 from sick sheep) through propagation in BALB/c mice and then cell culture in HL60 cells. The 3 isolates were identified by Wright-Giemsa staining, immunofluorescence, and electronic microscopy and were characterized by sequence analyses of the 16S rRNA gene, partial citrate synthase gene, major surface protein 4 gene, and heat shock protein gene. The multiple sequences of the 3 isolates were identical to each other but different from all known strains from other countries. The public health and veterinary relevance of the isolates deserves further investigation.

A total of 705 rodents from 6 provinces and autonomous regions of mainland People's Republic of China were tested by PCRs for tick-borne agents (Anaplasma phagocytophilum, Borrelia burgdorferi sensu lato, spotted fever group rickettsiae, and Francisella tularensis). Infection rates were 5.5%, 6.7%, 9.1% and 5.0%, respectively. Eighteen (2.6%) rodents of 10 species were positive for 2 or 3 agents. Sequence analysis of PCR products confirmed the presence and genotypes of detected agents. These findings demonstrate that these tick-borne agents cocirculate and that a variety of rodent species may be involved in their enzootic maintenance.

Background: Before 1986, scrub typhus was only found endemic in southern China. Because human infections typically occur in the summer, it is called "summer type". During the autumn-winter period of 1986, a new type of scrub typhus was identified in Shandong and northern Jiangsu province of northern China. This newly recognized scrub typhus was subsequently reported in many areas of northern China and was then called "autumn-winter type". However, clinical characteristics of associated cases have not been reported., Methods: From 1995 to 2006, all suspected scrub typhus cases in five township hospitals of Feixian county, Shandong province were enrolled. Indirect immunofluorescent assay (IFA) was used as confirmatory serodiagnosis test. Polymerase chain reaction (PCR) connected with restriction fragment length polymorphism (RFLP) and sequence analyses were used for genotyping of O. tsutsugamushi DNAs. Clinical symptoms and demography of confirmed cases were analyzed., Results: A total of 480 scrub typhus cases were confirmed. The cases occurred every year exclusively between September and December with a peak occurrence in October. The case numbers were relatively higher in 1995, 1996, 1997, and 2000 than in other years. 57.9% of cases were in the group aged 21-50. More cases occurred in male (56%) than in female (44%). The predominant occupational group of the cases was farmers (85.0%). Farm work was reported the primary exposure to infection in 67.7% of cases. Fever, rash, and eschar were observed in 100.0%, 90.4%, and 88.5% of cases, respectively. Eschars formed frequently on or around umbilicus, abdomen areas, and front and back of waist (34.1%) in both genders. Normal results were observed in 88.7% (WBC counts), 84.5% (PLT counts), and 89.7% (RBC counts) of cases, respectively. Observations from the five hospitals were compared and no significant differences were found., Conclusion: The autumn-winter type scrub typhus in northern China occurred exclusively from September to December with a peak occurrence in October, which was different from the summer type in southern China. In comparison with the summer type, complications associated with autumn-winter type scrub typhus were less severe, and abnormalities of routine hematological parameters were less obvious.

We determined the complete nucleotide sequence of a Japanese encephalitis virus (JEV) isolate (designated SH17M-2007) from a pool of Culex tritaeniorhynchus collected in southern China in 2007. The genome consisted of 10,965 nucleotides and included a single open reading frame (10,296 nucleotides) that encodes a 3,432-amino-acid polyprotein. The SH17M-2007 had 97.3 to 98.4% nucleotide identity with two Korean strains (KV1899, K94P05) and two Japanese strains (Ishikawa, JEV/sw/Mie/40/2004), but only 88.8% identity with the Chinese vaccine strain SA14-14-2. Five unique amino acid substitutions including one in the envelope (E) protein (Glu(E-306)-Lys) were found in the SH17M-2007 strain. Phylogenetic relationships based on the full-length nucleotide sequences were similar to those based on the E gene.

The hemagglutination inhibition (HI) assay is a widely used serological method to measure the levels of protective antibody responses against influenza viruses. However, the traditional HI assay which uses chicken erythrocytes is not sufficiently sensitive for detecting HI antibodies specific to avian influenza viruses. Previously, it was demonstrated that employing an assay using horse erythrocytes was able to increase the sensitivity of HI assay. The current report describes further optimization of this modified HI assay. It was shown that this method was able to increase detection of HI activities in rabbit sera immunized with H5 HA antigens, and proved that this increased sensitivity is useful in dissecting the strain specificity of HI antibody responses. In addition, the modified HI assay using horse erythrocytes increased the sensitivity of detecting HI antibodies specific for three major serotypes of avian influenza viruses, H5, H7 and H9, in people who may have asymptomatic infection with avian influenza viruses. Based on these results, the optimized use of horse erythrocytes should be standard practice for detecting HI activities against avian influenza viruses.

By using multilocus sequence analysis, five Borrelia valaisiana-related strains isolated from rodents and ticks in southwestern China were eventually classified as a new genospecies of B. burgdorferi sensu lato rather than B. valaisiana. The finding explained the differences in transmission cycle and phenotype between B. valaisiana strains from Europe and B. valaisiana-related strains from eastern Asia.

A molecular epidemiological survey was conducted to investigate the presence of pathogenic Borrelia burgdorferi sensu lato (s.l.) species in the forest areas of Zhejiang province, south-east China. A total of 182 ticks of 6 species and 200 rodents of 8 species were collected and individually examined for the presence of B. burgdorferi s.l. DNA by nested PCR targeting the 5S-23S rRNA intergenic spacer. Forty-one ticks of four species, Haemaphysalis concinna, Haemaphysalis longicornis, Rhipicephalus microplus and Haemaphysalis warburconi, were infected with B. burgdorferi s.l., with an overall infection rate of 23 %. Sixteen rodents of four species, Nivivener confucianus, Nivivener coxingi, Apodemus sylvaticus and Rattus losea, were positive for B. burgdorferi s.l., with an overall prevalence of 8 %. MseI RFLP analysis and sequence analysis of the positive PCR products showed that Borrelia spirochaetes in specimens consisted of Borrelia garinii, Borrelia afzelii and Borrelia valaisiana-related group. Forty (98 %) of the B. burgdorferi s.l.-positive ticks were infected with B. garinii and one (2 %) was infected with B. afzelii. Twelve (75 %) of the positive rodents were infected with B. garinii and four (25 %) were infected with the Borrelia spirochaete belonging to B. valaisiana-related group.

Anaplasma phagocytophilum was detected by polymerase chain reaction in 13 (14.1%) of 92 rodents captured from a mountainous area of Zhejiang Province in southeastern China. The nucleotide sequences of 1442-bp, nearly entire 16S rRNA gene amplified from these rodents, had 100% identity, but varied from all known corresponding sequences of A. phagocytophilum deposited in GenBank. To further identify and classify the variant, fragments of 357-bp partial citrate synthase gene (gltA), 849-bp major surface protein 4 gene (msp4), and 443-bp groESL heat-shock operon gene, were amplified and analyzed. The nucleotide sequences of the partial gltA gene amplified from the rodents were identical to each other, but distinct from previously reported A. phagocytophilum sequences,as were msp4 and groESL. These findings indicate that the newly discovered agent represents a novel A. phagocytophilum variant.

Background: Cytokines play important roles in antiviral action. We examined whether polymorphisms of interleukin (IL)-12 receptor B1 (IL-12RB1) affect the susceptibility to and outcome of severe acute respiratory syndrome (SARS)., Methods: A case-control study was carried out in Chinese SARS patients and healthy controls. The genotypes of 4SNPs on IL-12 RB1 gene, +705A/G,+1158T/C, +1196G/C and +1664 C/T, were determined by PCR-RFLP. Haplotypes were estimated from the genotype data using the expectation-maximisation algorithm., Results: Comparison between patients and close contacts showed that individuals with the +1664 C/T (CT and TT) genotype had a 2.09-fold (95% confidence interval [CI], 1.90-7.16) and 2.34-fold (95% CI, 1.79-13.37) increased risk of developing SARS, respectively. For any of the other three polymorphisms, however, no significant difference can be detected in allele or genotype frequencies between patients and controls. Additionally, estimation of the frequencies of multiple-locus haplotypes revealed potential risk haplotypes (GCCT) for SARS infection., Conclusions: Our data indicate that genetic variants of IL12RB1confer genetic susceptibility to SARS infection, but not necessary associated with the progression of the disease in Chinese population.

Hemorrhagic fever with renal syndrome (HFRS) is a significant public health problem with an increasing incidence in Beijing, China (report of disease surveillance from the Center for Disease Control and Prevention of Beijing, China). Hantaviruses were detected using RT-PCR method in blood samples of HFRS patients and lung tissues of rodents captured in Beijing. Phylogenetic analyses of 724bp partial S segment of the hantavirus gene showed that the detected Seoul virus (SEOV) fell into three different lineages, two of which circulated in Beijing. A nucleotide sequence identity of 99.7% for one of the cases of HFRS--the human- and Rattus norvegicus-originated SEOV sequences--had only two silent substitutions, suggesting genetic analysis is an essential tool for "case-investigation."

A total of 835 rodents captured in Beijing, China, were tested for hantavirus infection. Fifty-five (6.6%) were positive for viral RNA when lung tissue samples were examined by reverse transcriptase-polymerase chain reaction. Of 666 sera collected from the above rodents, 50 (7.5%) were positive for IgG antibody by ELISA. Among the 50 seropositive rodents, 37 were positive for viral RNA. In addition, five rodents were positive for viral RNA but negative for IgG antibody. The infection rates among study sites (chi(2) = 28.93, df = 8, P = 0.001) and habitats (chi(2) = 22.88,df = 7, P = 0.02) were significantly different. The sequences of partial M-segment of hantaviruses detected in 11 representative rodents had 0.1-8.2% divergence. Phylogenetic analysis showed that our hantavirus sequences fell into three different lineages regardless of geographical origin or rodent species. A strain detected from a trading center of agricultural products, which might be imported from other provinces, was genetically different from other strains of Beijing.

Lung tissue samples of 76 Korean field mice (Apodemus peninsulae) collected from northeastern China bordering with Far East Russia and Korea were detected for hantavirus partial M-segment or entire S-segment sequences by RT-PCR and 481-nt mitochondrial DNA fragment of the rodents. Four A. peninsulae mice were found positive for partial M-segment of hantavirus. Sequence analyses of partial M-segment or/and entire S-segment of the hantaviruses revealed that three were closely related to Hantaan virus (HTNV) strain 76-118. One new variant of HTNV-like virus designated as "Jilin-AP06" was much different from other rodent-borne hantavirus from China, and clustered with Amur (AMR) virus strains, which represent a distinct genetic lineage. These findings imply that hantavirus Jililn-AP06 strain from A. peninsulae is a new record of rodent-borne AMR virus in China. A. peninsulae might be a natural carrier of two distinct hantaviruses, AMR virus and HTNV in China.

Amur virus (AMRV) and Soochong virus (SOOV) were reported to be carried by Korean field mice (Apodemus peninsulae) in the Far East of Russia, China, and Korea. The distinction and demarcation between these two viruses have been a matter of debate. In order to clarify this issue and to confirm taxonomic position of AMRV and SOOV, the extensive phylogenetic analyses based on entire S segment and entire M segment sequences of AMRV, SOOV and other reference virus strains deposited in GenBank, were carried out using maximum likelihood and distant matrix methods. All inferred phylogenies revealed that all AMRV strains from China and Far East and SOOV (especially SOO-1/2 strains from Northeastern Korea) shared high identities of nucleotide sequences and were monophyletic distinct from Apodemus agrarius HTNV. Although two genetic sublineages of SOOV exist, these findings revealed that AMRV and SOOV might belong to the same entities of hantavirus., ((c) 2007 Wiley-Liss, Inc.)

Objective: To analyze the genetic differences of Orientia tsutsugamushi (Ot) Sta56 gene between Shandong isolates and other strains deposited in GenBank., Methods: PCR and restriction fragment length polymorphism (RFLP) were used to amplify the complete sequence of Ot-Sta56 gene. RFLP profiles of Ot were predicted by a computer program according to their complete sequences of Ot-Sta56 gene. PCR amplicon from XDM2 strain was sequenced and analyzed by Clustal X (1.8) and PHYLIP software., Results: The complete sequences (about 1.6 kbp) of Ot-Sta56 gene were amplified from B16 strain (isolated from patients), FXS2 strain (isolated from A. agrarius) and XDM2 strain. Four species of restriction endonucleases (Hha I, Hinf I, Hae III, Pst I) were used to digest the PCR amplicons from the 3 isolates. When comparing with the RFLP profiles of prototype Ot, the RFLP profiles of PCR amplicons from the 3 isolates were similar to those of Japan Kawasaki strain, but were quite different from the international reference strains Gilliam, Karp, Kato. Results from DNA sequence analysis showed that the complete sequence of Ot-Sta56 gene homology to Japan Kawasaki strain of XDM2 strain was 97%, and deduced amino acid sequence was 92%., Conclusion: Data from the complete sequence of Sta56 gene indicated that the genotypes of Ot isolates in Shandong province were similar, but with distinction from the Kawasaki strain.

Objective: To confirm the existence of Amur-like viruses in Apodemus peninsulae in China, and to understand the molecular characteristics of these viruses., Methods: Total RNA was extracted from lungs of A. peninsulae captured in Jilin of Northeast China with Trizol reagent. Complete S and partial M segments of Amur virus were amplified and sequenced. Phylogenetic analyses on multiple nucleotide sequences were performed with the Clustal method and DNASTAR software., Results: 383 bp cDNA of M segment and 1696 bp of S segment of Amur like virus were recovered from lung tissue of A. peninsulae, named JilinAP06. The full-length of its S gene comprised of 1696 nucleotides with ORF including 1287 nucleotides and encoding a protein which comprised 429 amino acids. The phylogenetic analysis of this sample with other hantaviruses revealed that the complete S and partial M segment sequence of JilinAP06 both were closely related to those Amur viruses such as AP63, AP61, AP1371 and AP1168 found in A. peninsulae from Far East region of Russia and B78 strain, Liu strain and H5 strain, which were all from Chinese patients. The complete S and partial M segment sequence of JilinAP06 had only 81.0% identities with the nucleotide sequences of HV prototype 76-118 strain., Conclusion: Amur-like viruses did exist in A. peninsulae from Northeasern China while A. peninsulae might be the natural reservoir of Amur-like viruses in China and was the important infectious source to HFRS patients which were caused by Amur-like viruses.

Both scrub typhus and hemorrhagic fever with renal syndrome (HFRS) are severely epidemic in northern China and often present with acute undifferentiated fever. To correctly distinguish the two diseases at an early stage, we collected and compared clinical and routine laboratory data of 46 patients with confirmed scrub typhus and 49 patients with confirmed HFRS presenting to the outpatient departments of three town hospitals in northern China. Most patients with HFRS but none of the patients with scrub typhus had hemorrhagic manifestations. Retro-orbital pain, lumbar back pain, flank tenderness, proteinuria, and occult blood in urine often occurred in patients with HFRS. However, skin eschar, regional lymphadenopathy, and maculopapular rash were more commonly found in patients with scrub typhus. In addition, platelet counts in patients with HFRS were significantly lower than in patients with scrub typhus. These findings will be useful for physicians to distinguish scrub typhus from HFRS.