Great genetic diversity of vector-borne bacteria and protozoan in wild rodents from Guangxi, China.

BACKGROUND: Rodents are recognized as the hosts of many vector-borne bacteria and protozoan parasites and play an important role in their transmission and maintenance. Intensive studies have focused on their infections in vectors, especially in ticks, however, vector-borne bacterial and protozoan infections in rodents are poorly understood although human cases presenting with fever may due to their infection have been found. METHODS: From May to October 2019, 192 wild rodents were trapped in wild environment of Guangxi Province, and the spleen samples were collected to reveal the presence of vector-borne bacterial and protozoan infections in them. The microorganisms in rodents were identified by detecting their DNA using (semi-)nested PCR. All the PCR products of the expected size were subjected to sequencing, and then analyzed by BLASTn. Furthermore, all the recovered sequences were subjected to nucleotide identity and phylogenetic analyses. RESULTS: As a result, 192 rodents representing seven species were captured, and Bandicota indica were the dominant species, followed by Rattus andamanensis. Based on the (semi-)nested PCR, our results suggested that Anaplasma bovis, Anaplasma capra, Anaplasma ovis, Anaplasma phagocytophilum, "Candidatus Neoehrlichia mikurensis", "Candidatus E. hainanensis", "Candidatus E. zunyiensis", three uncultured Ehrlichia spp., Bartonella coopersplainsensis, Bartonella tribocorum, Bartonella rattimassiliensis, Bartonella silvatica, two uncultured Bartonella spp., Babesia microti and diverse Hepatozoon were identified in six rodent species. More importantly, six species (including two Anaplasma, two Bartonella, "Ca. N. mikurensis" and Bab. microti) are zoonotic pathogens except Anaplasma bovis and Anaplasma ovis with zoonotic potential. Furthermore, dual infection was observed between different microorganisms, and the most common type of co-infection is between "Ca. N. mikurensis" and other microorganisms. Additionally, potential novel Bartonella species and Hepatozoon species demonstrated the presence of more diverse rodent-associated Bartonella and Hepatozoon. CONCLUSIONS: The results in this work indicated great genetic diversity of vector-borne infections in wild rodents, and highlighted the potential risk of human pathogens transmitted from rodents to humans through vectors.

Anaplasma, Bartonella, and Rickettsia infections in Daurian ground squirrels (Spermophilus dauricus), Hebei, China.

Rodents have been confirmed as hosts of various vector-borne zoonotic pathogens and are important for the maintenance of these microbes in nature. However, surveillance for zoonotic pathogens is limited for many wild rodent species in China, so our knowledge of pathogen ecology, genetic diversity, and the risk of cross-species transmission to humans is limited. In this study, 165 spleen samples of Daurian ground squirrels (Spermophilus dauricus) were collected from Weichang Manchu and the Mongolian Autonomous County of Hebei Province, China, and Rickettsia, Bartonella, and Anaplasma were identified by DNA detection using polymerase chain reaction (PCR). Sequence analysis identified eight bacterial pathogens: R. raoultii, R. sibirica, Candidatus R. longicornii, B. washoensis, B. grahamii, B. jaculi, A. capra, and Candidatus Anaplasma cinensis. Co-infection of B. grahamii and R. raoultii in one sample was observed. Our results demonstrated the genetic diversity of bacteria in Daurian ground squirrels and contributed to the distribution of these pathogens. Six species, A. capra, R. raoultii, R. sibirica, Candidatus R. longicornii, B. washoensis, and B. grahamii, are known to be pathogenic to humans, indicating a potential public health risk to the local human population, especially to herders who frequently have close contact with Daurian ground squirrels and are thus exposed to their ectoparasites.

Genetic diversity of vector-borne zoonotic pathogens in companion dogs and cats, Tianjin, China.

Background: Dogs and cats are the hosts of many vector-borne human pathogens that can be transmitted to humans. Given their direct and intimate contact with humans, companion dogs and cats are considered direct sentinels of vector-borne human pathogens. However, limited information is currently available regarding canine and feline zoonotic pathogens in China. This study detected canine and feline vector-borne human pathogens to better understand the potential risk to humans. Methods: Blood samples were collected from 275 domestic companion animals (117 dogs and 158 cats) living in Tianjin city, China, and the presence of DNA from Anaplasma, Babesia, Bartonella, and Rickettsia was detected by semi-nested polymerase chain reaction (PCR). The PCR products of the expected size were sequenced, and these newly generated sequences were subjected to BLASTN, nucleotide identity, and phylogenetic analyses. Results: A total of 24 blood samples tested positive for vector-borne pathogens in companion dogs and cats in Tianjin city, China, with a relatively low positive rate of 8.7%. Specifically, seven human pathogens, including Rickettsia raoultii, Candidatus Rickettsia jingxinensis, Rickettsia sibirica, Rickettsia felis, Babesia venatorum, Bartonella tribocorum, and Bartonella Henselae, were identified. In addition, Anaplasma ovis with zoonotic potential and Candidatus A. cinensis were detected. Conclusion: Our results indicate substantial genetic diversity in the vector-borne human pathogens circulating in companion dogs and cats. Interventions based on "One Health" should be taken to reduce the potential risks of contracting infection from companion dogs and cats in Tianjin, China.

Molecular evidence of Rickettsia canadensis in ticks, Hebei, China.

China was affected severely by tick-borne rickettsiosis, and more than 10 Rickettsia species pathogenic to humans have been identified. In recent years, several Rickettsia members, with unknown pathogenicity, firstly identified abroad have been found in China. In this study, parasitic and questing ticks were recovered from two sampling sites in Hebei, China. Specific primers targeting outer membrane protein B (ompB) gene were designed to test the presence of Rickettsia canadensis by nested Polymerase Chain Reaction (PCR). As a result, a total of 428 ticks, including 232 ticks (including 230 Haemaphysalis longicornis and two H. japonica) from Laiyuan County and 196 (H. longicornis) from Luanping County, were collected. Sequencing of PCR products with the expected size and subsequently BLAST showed that 38H. longicornis ticks tested positive for R. canadensis, with an overall positive rate of 8.8%. In addition, 800-bp ompB gene and nearly complete citrate synthase (gltA) gene were recovered from six randomly selected positive samples to better understand their genetic characteristics. Nucleotide identity and phylogenetic analyses showed that R. canadensis presented geographical clustering with evidence that variants identified in the current study presented closer genetic relationship with others identified in Asian than those found in North America. In addition, epidemiological data suggested that H. longicornis may be the competent vector, and more attention should be paid to R. canadensis due to its zoonotic potential. In sum, R. canadensis was confirmed to be present in Hebei Province, China, and its surveillance in ticks should be strengthened due to potential pathogenicity, higher positive rate in ticks and wide distribution of possible vector tick species.

Transcriptome profiling of A549 non-small cell lung cancer cells in response to Trichinella spiralis muscle larvae excretory/secretory products.

Trichinella spiralis (T. spiralis) muscle-larva excretory/secretory products (ML-ESPs) is a complex array of proteins with antitumor activity. We previously demonstrated that ML-ESPs inhibit the proliferation of A549 non-small cell lung cancer (NSCLC) cell line. However, the mechanism of ML-ESPs against A549 cells, especially on the transcriptional level, remains unknow. In this study, we systematically investigated a global profile bioinformatics analysis of transcriptional response of A549 cells treated with ML-ESPs. And then, we further explored the transcriptional regulation of genes related to glucose metabolism in A549 cells by ML-ESPs. The results showed that ML-ESPs altered the expression of 2,860 genes (1,634 upregulated and 1,226 downregulated). GO and KEGG analysis demonstrated that differentially expressed genes (DEGs) were mainly associated with pathway in cancer and metabolic process. The downregulated genes interaction network of metabolic process is mainly associated with glucose metabolism. Furthermore, the expression of phosphofructokinase muscle (PFKM), phosphofructokinase liver (PFKL), enolase 2 (ENO2), lactate dehydrogenase B (LDHB), 6-phosphogluconolactonase (6PGL), ribulose-phosphate-3-epimerase (PRE), transketolase (TKT), transaldolase 1 (TALDO1), which genes mainly regulate glycolysis and pentose phosphate pathway (PPP), were suppressed by ML-ESPs. Interestingly, tricarboxylic acid cycle (TCA)-related genes, such as pyruvate dehydrogenase phosphatase 1 (PDP1), PDP2, aconitate hydratase 1 (ACO1) and oxoglutarate dehydrogenase (OGDH) were upregulated by ML-ESPs. In summary, the transcriptome profiling of A549 cells were significantly altered by ML-ESPs. And we also provide new insight into how ML-ESPs induced a transcriptional reprogramming of glucose metabolism-related genes in A549 cells.

Cell membrane-bound toll-like receptor-1/2/4/6 monomers and -2 heterodimer inhibit enterovirus 71 replication by activating the antiviral innate response.

Host immune activation is critical for enterovirus 71 (EV71) clearance and immunopathogenesis. However, the mechanism of innate immune activation, especially of cell membrane-bound toll-like receptors (TLRs), against EV71 remains unknown. We previously demonstrated that TLR2 and its heterodimer inhibit EV71 replication. In this study, we systematically investigated the effects of TLR1/2/4/6 monomers and TLR2 heterodimer (TLR2/TLR1, TLR2/TLR6, and TLR2/TLR4) on EV71 replication and innate immune activation. We found that the overexpression of human- or mouse-derived TLR1/2/4/6 monomers and TLR2 heterodimer significantly inhibited EV71 replication and induced the production of interleukin (IL)-8 via activation of the phosphoinositide 3-kinase/protein kinase B (PI3K/AKT) and mitogen-activated protein kinase (MAPK) pathways. Furthermore,human-mouse chimeric TLR2 heterodimer inhibited EV71 replication and activated innate immunity. Dominant-negative TIR-less (DN)-TLR1/2/4/6 did not exert any inhibitory effects, whereas DN-TLR2 heterodimer inhibited EV71 replication. Prokaryotic expression of purified recombinant EV71 capsid proteins (VP1, VP2, VP3, and VP4) or overexpression of EV71 capsid proteins induced the production of IL-6 and IL-8 via activation of the PI3K/AKT and MAPK pathways. Notably, two types of EV71 capsid proteins served as pathogen-associated molecular patterns for TLR monomers (TLR2 and TLR4) and TLR2 heterodimer (TLR2/TLR1, TLR2/TLR6, and TLR2/TLR4) and activated innate immunity. Collectively, our results revealed that membrane TLRs inhibited EV71 replication via activation of the antiviral innate response, providing insights into the EV71 innate immune activation mechanism.

Human pathogens in ticks removed from humans in Hebei, China.

Ticks are the hosts or vectors of many human pathogens, including viruses, bacteria and protozoa, and can transmit these causative agents to humans when feeding on human bodies. In this study, 26 ticks removed from humans in Hebei, China were tested for the presence of human-pathogenic microorganisms by Polymerase Chain Reaction (PCR) or Reversed Transcript PCR (RT-PCR). As a result, 11 ticks tested positive for at least one human pathogen. Specifically, four validated human pathogens, including Rickettsia raoultii, Candidatus Rickettsia tarasevichiae, Babesia venatorum, and Borrelia garinii, as well as Anaplasma ovis with zoonotic potential, were identified in Ixodes persulcatus, Dermacentor silvarum and Haemaphysalis concinna. Importantly, this is the first report of Anaplasma and Babesia species pathogenic to humans in Hebei province. Moreover, the co-infections, including double infection and quadruple infection were observed. In addition, Candidatus R. principis with unknown pathogenicity was identified in one tick, which may be the same species as Candidatus R. hongyuanensis based on the nucleotide identity and phylogenetic analysis. Concluding, four validated tick-borne pathogens and one with zoonotic potential were identified in ticks parasitizing humans, suggesting the potential high public health risk in the local human population.

Genetic diversity of Bartonella infection in residential and field rodents in Hebei, China.

Rodents are the primary natural reservoirs of Bartonella spp., and some of which are zoonotic causative agents. Hence, surveillance of Bartonella sp. infection in rodents is very important for the prevention of human bartonellosis caused by them. In this study, rodents were captured, and their spleen samples were collected for Bartonella sp. DNA detection and identification by amplifying the 16S rRNA, gltA, and ftsz genes using semi-nested polymerase chain reaction (PCR). The results indicated that Bartonella sp. DNA was detected in seven Rattus norvegicus individuals with a detection rate of 6.7% in Chengde City and bacterial DNA in 31 Apodemus agrarius individuals with a detection rate of 28.4% in Handan City. The DNA detection rate across the genders and ages of rodents was not found to be statistically significant. Furthermore, sequence analysis of the above-mentioned three genes demonstrated that at least eight Bartonella species were circulating in Hebei Province, of which three, including Bartonella rattimassiliensis, Bartonella grahamii, and Bartonella tribocorum, are human pathogens, thus suggesting the existence of a major public health risk. Overall, these results revealed the detection rate and genetic diversity of Bartonella species infection in rodents in Hebei Province, which could be potentially helpful for the prevention of bartonellosis caused by rodent-associated Bartonella species. This study highlights the urgent need for the surveillance of Bartonella infections in rodents and ectoparasites that affect both rodents and humans and can cause fever of unknown origin or endocarditis.

Molecular detection of "Candidatus Rickettsia tarasevichiae" by Loop-mediated Isothermal Amplification (LAMP) of the ompA gene.

"Candidatus Rickettsia tarasevichiae" (CRT) is increasingly being recognized as a disease causative agent in China and poses a great challenge to public health. Rapid and accurate detection is indispensable for laboratory diagnosis of infection caused by CRT and its surveillance in ticks. In the present study, a novel DNA-based loop-mediated isothermal amplification (LAMP) assay targeting the ompA gene was developed for the detection of CRT in tick samples. A set of universal primers specific to CRT were designed using PrimerExplorer V5 software. The analytical sensitivity, evaluated using recombinant plasmids containing the ompA gene, reached up to 1 copy per reaction, greater than that of the PCR assay targeting the same gene. This LAMP assay specifically detected CRT and showed no cross-reaction with other species common in China within the genus Rickettsia. In addition, this newly developed LAMP assay presented high diagnostic sensitivity of CRT detection validated by known positive DNA samples from ticks and simulated clinical samples. The applicability of the LAMP assay was evaluated by screening CRT from ticks, and the result showed that CRT circulation in Weichang County, China, was confirmed. Our findings indicate that this LAMP method is sensitive and specific for the detection of CRT and may have a potential application in the detection of CRT infection in patients and ticks.

Prevalence and clinical characteristics of hospitalized children with community-acquired Mycoplasma pneumoniae pneumonia during 2017/2018, Chengde, China.

ABSTRACT: Community acquired-pneumonia (CAP) has varying causative pathogens and clinical characteristics. This study investigated the prevalence of Mycoplasma pneumoniae (M pneumoniae) and evaluated the clinical characteristics in infected hospitalized children by disease severity.From throat swabs of hospitalized children (5 months to 14 years) with CAP collected between November 2017 and May 2018, M pneumoniae and other CAP pathogens were identified using polymerase chain reaction (PCR). Differences in clinical and laboratory test data were compared between severe and mild case groups.Of 333 hospitalized children enrolled, 221/333 (66.4%) tested positive for M pneumoniae and 24/221 (10.9%) patients were (n = 9, aged <5 years vs n = 15, ≥5 years) single infection by PCR, however, only 170/333 (51.1%) patients were presented with M pneumoniae IgM-positive. M pneumoniae detection rate by PCR was higher than by immunoglobulin (IgM) serology. In 123/221 (55.7%) M pneumoniae infected patients, coinfection with bacterial pathogens (n = 61, <5 years vs n = 62, ≥5 years) occurred. Children (aged 3-8 years) had most M pneumoniae infection. Severe M pneumoniae pneumonia (MPP) in children occurred mostly in older age (7 [interquartile ranges {IQR}, 6-8] years; P < .0001), with longer cough days (14 [IQR, 10-19.5] days; P = .002) and hospitalization duration (9.5 [IQR, 7-12.3] days; P < .0001), lower lymphocyte ratio (24.1, [IQR, 20.0-31.1] %; P = .001), higher neutrophils ratio (66.0, [IQR, 60.2-70.3]%; P < .0001), and serum C-reactive protein (CRP) level (3.8, [IQR, 1.3-10.9] mg/L; P = .027).M pneumoniae is the most commonly detected pathogen in CAP. High coinfection prevalence increases diagnosis difficulty by clinically nonspecific characteristics. M pneumoniae detection by PCR with IgM may improve precise and reliable diagnosis of community-acquired MPP.

Orthohantaviruses infections in humans and rodents in Baoji, China.

In recent years, hemorrhagic fever with renal syndrome (HFRS) incidence has been becoming a severe public health problem again due to its significant increase in Shaanxi Province, China. Baoji, located in the Guanzhong Plain in the central part of Shaanxi Province, has been severely affected by HFRS since its first emergence in 1955. To better understand the epidemiology of orthohantaviruses infection in humans and the causative agents carried by the rodents, the long-term incidence patterns were analyzed and a molecular epidemiological investigation of orthohantaviruses infection in humans and rodents was performed. During 1984-2019, 13,042 HFRS cases were registered in Baoji, including 275 death cases. Except the first high prevalence of HFRS in 1988-1993, another two epidemic peaks were observed in 1998-2003 and 2012, respectively, although vaccination project was started since 1996. During the same period, HFRS cases in Baoji mainly were recorded in winter suggesting they may be caused by Hantaan orthohantavirus (HTNV), while a small peak of HFRS was also found in summer with unknown reason. Nucleotide identity and phylogenetic analyses demonstrated that a novel clade of HTNV sequences recovered from HFRS cases were closely related to those from rodents, including species close contact with humans, suggesting a direct viral transmission from rodents to humans and the important role in the HTNV transmission the nontraditional rodent hosts may play. Moreover, two distant related Dabieshan orthohantavirus (DBSV) lineages were also identified in Niviventer niviventer in this area demonstrating its considerable genetic diversity. Our data indicated that continual spillover of HTNV from rodents to humans, contributing to the high prevalence of HFRS in humans in Baoji.

Global profiling of the alternative splicing landscape reveals transcriptomic diversity during the early phase of enterovirus 71 infection.

The alteration of host cell splicing is a major strategy favouring viral replication; however, the interaction between human tonsillar epithelial cells (HTECs) and enterovirus 71 (EV71) has not been fully elucidated. Here, a total of 201 differentially expressed genes (DEGs) and 3266 novel genes with coding potential were identified. A total of 3479 skipped exons (SEs), 515 alternative 3' splice sites (A3SSs), 391 alternative 5' splice sites (A5SSs), 531 mutually exclusive exons (MXEs) and 825 retained introns (RIs) were identified as significantly altered alternative splicing (AS) events. The enriched DEGs were mainly related to the cell cycle, spliceosome, and Toll-like receptor (TLR) signalling pathways. Finally, the replication of EV71 was significantly inhibited by TLR2 heterodimers. Our findings suggest that AS events induced by EV71 increase the transcriptomic diversity of HTECs in response to EV71 infection. Additionally, TLR2 heterodimers have the potential to protect HTECs against EV71.

An adult co-presented with varicella and herpes zoster caused by varicella zoster virus genotype J, China: a case report.

BACKGROUND: Varicella zoster virus (VZV) causes varicella primarily in childhood, and some rare adults also report varicella. Herpes zoster mainly occurs in adults by endogenous reactivation of latent VZV. Until now, varicella and herpes zoster have seldom been reported simultaneously in one patient. Here, we report a rare case co-presenting with varicella and herpes zoster in a Chinese adult. CASE PRESENTATION: A 44-year-old Chinese man suffered papules and vesicles with pain on the left ear. Five days after onset, he was admitted to the Department of Dermatology of The Third Hospital of Xiamen. Physical examination revealed that small vesicles surrounded by erythema had developed on his trunk, back and neck, and unilateral papules and vesicles in ribbons had also developed on the left ear. This patient was excluded from human immunodeficiency virus and Treponema pallidum infections by ELISA antibody tests. Laboratory tests revealed that the ratio of eosinophils (0.1%) and eosinophil count (0.0 × 109/L) were significantly downregulated. Treatment with valacyclovir, ebastine, mecobalamine, pregabalin and calamine lotion for 5 days was effective therapy for varicella and herpes zoster. Polymerase chain reaction for vesicular fluids from varicella and herpes zoster was positive for VZV, and further phylogenetic analysis and single nucleotide polymorphism variations confirmed that the VZV genotype was type J (clade 2). CONCLUSIONS: This rare case highlights awareness of varicella and herpes zoster caused by VZV infection in adults. Our report provides novel insight into the rare clinical presentation of VZV genotype J.

Molecular detection of Hepatozoon canis in dogs and ticks in Shaanxi province, China.

Hepatozoon canis, transmitted by Rhipicephalus sanguineus, is a tick-borne pathogen and causes canine hepatozoonosis. Until now, only limited previous studies were conducted on the molecular detection and characterization of Hepatozoon sp. in dogs in China. Blood samples were collected from 93 sick dogs that were clinically diagnosed as babesiosis but tested negative for Babesia, and 103 apparently healthy dogs, as well as their infesting ticks in Xi'an and Hanzhong cities, Shaanxi province of China. PCR amplifying partial 18S rRNA gene was used to detect the DNA of Hepatozoon sp. Genetic and phylogenetic analysis were performed to determine the Hepatozoon species. Our results demonstrated that H. canis was identified from the sick dogs and the infested ticks in Hanzhong, with no significant differences of prevalence between both genders and ages. No positive blood or tick samples were found in Xi'an. Moreover, all the 18S rRNA gene sequences recovered from both dogs and the infested ticks showed a high genetic similarity with each other, and also presented a close relationship with other known sequences in and outside China. In conclusion, H. canis was identified in babesiosis-suspected dogs and ticks infesting them in Shaanxi, China, although the association between clinical signs and H. canis need further study.

Molecular detection and genetic characteristics of Babesia gibsoni in dogs in Shaanxi Province, China.

BACKGROUND: Several members of genus Babesia are important pathogens causing babesiosis in dogs. In China, at least five Babesia species have been described in dogs or ticks. This study sought to determine the prevalence and molecular characteristics of various Babesia spp. in dogs in cities in Shaanxi Province in China, including Xi'an and Hanzhong. METHODS: A total of 371 blood samples were collected from pet dogs presenting to veterinary clinics in the cities of Xi'an and Hanzhong in Shaanxi, China. Babesia spp. DNA was detected via amplification of partial 18S rRNA genes by semi-nested PCR. Almost full-length 18S rRNA, ITS, partial TRAP and complete cytb genes were recovered for analysis of the genetic characteristics and relationships with known isolates. RESULTS: A single species, Babesia gibsoni, was identified in dogs in Xi'an and Hanzhong. Consistently, B. gibsoni was also detected in 14 ticks collected from positive dogs. Sequence similarities and phylogenetic analysis suggested that the isolates identified herein showed a closer genetic relationship with isolates from East Asian countries rather than India, Bangladesh, or the USA. Sequence analysis based on tandem repeat analysis of the TRAP gene further revealed that specific haplotypes were circulating in both Xi'an and Hanzhong, with no specific regionality. In addition, 10.9% of all isolates with atovaquone (ATV)-resistance were identified because of M121I mutation in the deduced cytb protein. CONCLUSIONS: This study revealed a high prevalence rate of Babesia infection. Babesia gibsoni was the only Babesia species identified in cases of canine babesiosis in the cities of Xi'an and Hanzhong cities in Shaanxi, China. In addition, the TRAP gene presented high genetic diversity across isolates. Such information is useful for elucidating the epidemiological characteristics of canine babesiosis, as well as the overall genetic diversity of Babesia spp. circulating in dog populations in Shaanxi Province.

Extensive genetic diversity of Anaplasma bovis in ruminants in Xi'an, China.

Anaplasma bovis is an organism significant to cattle and buffalo since it is one of the causative agents of bovine anaplasmosis. Previous studies have shown the worldwide distribution of A. bovis. However, most of these studies about its genetic diversity only focused on the rrs gene. In this study, DNA of A. bovis was detected in blood samples of cattle and goats in Xi'an city, China by nested-PCR. Near full-length rrs, groEL, and gltA genes were amplified successfully from the positive samples. Genetic analysis showed that specific genetic marker (an insertion and a deletion) was found in the rrs sequences in some strains, as well as clone 88 from monkeys in previous study. Phylogenetic analysis based on the rrs, groEL, and gltA genes revealed that A. bovis circulating in Xi'an exhibited great genetic diversity. Our results also indicated that variants outside China presented geographic clustering, and all A. bovis isolates based on the groEL or gltA gene also showed a host origin clustering. Also of note was that the phylogenetic analyses of the groEL and gltA genes suggested that both frequent dispersals over long distances in recent years and local adaptation over long evolutionary timescales played important roles in the distribution and evolution of A. bovis in China. Finally, a potential recombination event in the genome of Zhouzhi-cattle-10 based on inconsistent positions in the groEL and gltA trees was also observed. These results also reinforce the need for assessing the pathogenicity to humans of A. bovis variants with specific marker in the rrs gene.

Mosaic genome of Human Coxsackievirus A4 associated with herpangina and HFMD in Yancheng, China, 2016 and 2018.

OBJECTIVES: To better understand the spectrums of pathogens causing herpangina and circulation of Coxsackievirus A4 in Yancheng, China. METHODS: Stool samples from herpangina and HFMD cases were collected. Real Time PCR Kits was used to identify Enterovirus 71, CV-A16 and CV-A6, and nested reverse transcription PCR (nRT-PCR) to detect the other enterovirus types. Complete VP1 and genome sequence of CV-A4 were amplified by using nRT-PCR. Genetic, phylogenetic and recombination analysis were performed. RESULTS: Co-circulation of three recombinant CV-A4 groups, including one novel (C2 lineage), was identified in Yancheng, China, 2016 and 2018. One was the major causative agent of herpangina, and another two were responsible for HFMD. Phylogenetic and recombination analysis indicated that the non-structural region of their genome originated from the same ancestry and subsequently adaptation. C2 lineage of CV-A4 group may be introduced from countries outside China and its genome occurred recombination in China. CONCLUSION: Novel recombinant CV-A4 was mainly associated with herpanginain in Yancheng, 2018, China. C2 lineage of CV-A4 group with recombinant non-structural region was also identified in HFMD patients.

Experimental infection of Marmota monax with a novel hepatitis A virus.

To establish an animal model for the newly identified Marmota Himalayana hepatovirus, MHHAV, so as to develop a better understanding of the infection of hepatitis A viruses. Five experimental woodchucks (Marmota monax) were inoculated intravenously with the purified MHHAV from wild woodchuck feces. One animal injected with PBS was defined as a control. Feces and blood were routinely collected. After the animals were subjected to necropsy, different tissues were collected. The presence of viral RNA and negative sense viral RNA was analyzed in all the samples and histopathological and in situ hybridization analysis was performed for the tissues. MHHAV infection caused fever but no severe symptoms or death. Virus was shed in feces beginning at 2 dpi, and MHHAV RNA persisted in feces for ~2 months, with a biphasic increase, and in blood for ~30 days. Viral RNA was detected in all the tissues, with high levels in the liver and spleen. Negative-strand viral RNA was detected only in the liver. Furthermore, the animals showed histological signs of hepatitis at 45 dpi. MHHAV can infect M. monax and is associated with hepatic disease. Therefore, this animal can be used as a model of HAV pathogenesis and to evaluate antiviral and anticancer therapeutics.

The suppression of innate immune response by human rhinovirus C.

Rhinovirus C (RV-C), a newly identified group of human rhinoviruses (RVs), is associated with exacerbation of severe asthma. The type I interferon (IFN) response induced by this virus and the mechanisms of evasion of IFN-mediated innate immunity for RV-C remain unclear. In this study, we constructed a full-length cDNA clone of RV-C (LZ651) from a clinical sample. IFN-ß mRNA and protein levels were not elevated in differentiated Human bronchial epithelial (HBE) cells at the air-liquid interface infected with RV-C, except in the early stage of infection. The ability to attenuate IFN-ß activation was ascribed to 3Cpro of RV-C, and the 40-His site of 3Cpro played an important role. Furthermore, RIG-I was degraded by 3Cpro in a caspase-dependent manner and 3Cpro cleaved MAVS at 148 Q/A, which inhibited IFN signaling. Taken together, our results demonstrate the mechanism by which RV-C circumvents the production of type I IFN in infected cells.

Study on the mitochondrial apoptosis pathways of small cell lung cancer H446 cells induced by Trichinella spiralis muscle larvae ESPs.

Trichinella spiralis (T.spiralis) muscle-larva (ML) excretory-secretory proteins (ESPs) contain antitumour-active substances. ESPs have been shown to inhibit tumour growth. To explore the effects of these proteins on small cell lung cancer cells and the possible mechanisms of their antineoplastic action, H446 SCLC cells were co-cultured with different concentrations of T. spiralis ML ESPs for 12, 24 and 48 h. Our results showed that T. spiralis ML ESPs significantly inhibited H446 cell proliferation, which was dose-and time-dependent. The results of flow cytometry testing indicate a clear apoptosis trend in H446 cells co-cultured with ESPs for 24 h. Reverse transcription polymerase chain reaction and Western blotting results showed increased expression of pro-apoptosis genes Bax, Cyt-C, Apaf-1, caspase-9 and caspase-3, compared with the negative control group, and decreased the expression of anti-apoptosis genes Bcl-2 and Livin. Our results suggest that T. spiralis ML ESPs can induce apoptosis in H446 cells through a mitochondrial pathway, which may be a mechanism of antineoplastic action in T. spiralis ML ESPs.