On the pathogenesis of spontaneous parvovirus infection

The internal organs of 21 beagles spontaneously infected with parvovirus were examined histologically and in 10 of the dogs immunofluorescence examination was also performed. The study showed that the pathological process had started in the small intestine and from there the viral agent had spread through the regional lymph nodes into the other lymphatic and the haemopoietic organs causing there depletion predominantly of lymphocytes and arrest of haemopoiesis. Four dogs displayed conspicuous oedema of the media of the arterioli in the liver. In the cytoplasm of probably Kupffer's cells specific fluorescence was present in all the 10 dogs examined. On the other hand neither viral antigen nor histological changes were found in kidneys. Our observations suggest that spontaneous infection of dogs with parvovirus takes place per os. Parvovirus is not eliminated from the organism with urine.     

Isolation and Genomic Characterization of a Duck-Origin GPV-Related Parvovirus from Cherry Valley Ducklings in China

A newly emerged duck parvovirus, which causes beak atrophy and dwarfism syndrome (BADS) in Cherry Valley ducks, has appeared in Northern China since March 2015. To explore the genetic diversity among waterfowl parvovirus isolates, the complete genome of an identified isolate designated SDLC01 was sequenced and analyzed in the present study. Genomic sequence analysis showed that SDLC01 shared 90.8%–94.6% of nucleotide identity with goose parvovirus (GPV) isolates and 78.6%–81.6% of nucleotide identity with classical Muscovy duck parvovirus (MDPV) isolates. Phylogenetic analysis of 443 nucleotides (nt) of the fragment A showed that SDLC01 was highly similar to a mule duck isolate (strain D146/02) and close to European GPV isolates but separate from Asian GPV isolates. Analysis of the left inverted terminal repeat regions revealed that SDLC01 had two major segments deleted between positions 160–176 and 306–322 nt compared with field GPV and MDPV isolates. Phylogenetic analysis of Rep and VP1 encoded by two major open reading frames of parvoviruses revealed that SDLC01 was distinct from all GPV and MDPV isolates. The viral pathogenicity and genome characterization of SDLC01 suggest that the novel GPV (N-GPV) is the causative agent of BADS and belongs to a distinct GPV-related subgroup. Furthermore, N-GPV sequences were detected in diseased ducks by polymerase chain reaction and viral proliferation was demonstrated in duck embryos and duck embryo fibroblast cells.     

人类博卡病毒HBoV1基因组克隆及启动子活性分析

人类博卡病毒 (Human bocavirus,HBoV) 是继细小病毒B19之后,第2个被发现可引起人类疾病的细小病毒。通过PCR扩增方法从患有下呼吸道感染的患儿痰液中鉴定HBoV,以鉴定的阳性样本为模板,利用分子生物学方法构建病毒基因组克隆并进行序列分析。2007年10月?2009年3月从湖北省妇幼保健院共收集941例下呼吸道感染患儿的痰液标本,检测到33份HBoV阳性样品,阳性率为3.51％ (33/941);其中1岁以下婴幼儿患者占阳性样72.7％;构建了含有HBoV中间大片段基因克隆WHL-1,     

Experimental infection of cynomolgus monkeys with simian parvovirus.

Simian parvovirus is a recently discovered parvovirus that was first isolated from cynomolgus monkeys. It is similar to human B19 parvovirus in terms of virus genome, tropism for erythroid cells, and characteristic pathology in natural infections. Cynomolgus monkeys were infected with simian parvovirus to investigate their potential usefulness as an animal model of human B19 parvovirus. Six adult female cynomolgus monkeys were inoculated with purified simian parvovirus by the intravenous or intranasal route and monitored for evidence of clinical abnormalities; this included the preparation of complete hematological profiles. Viremia and simian parvovirus-specific antibody were determined in infected monkeys by dot blot and Western blot assays, respectively. Bone marrow was examined at necropsy 6, 10, or 15 days postinfection. All of the monkeys developed a smoldering, low-grade viremia that peaked approximately 10 to 12 days after inoculation. Peak viremia coincided with the appearance of specific antibody and was followed by sudden clearance of the virus and complete, but transient, absence of reticulocytes from the peripheral blood. Clinical signs were mild and involved mainly anorexia and slight weight loss. Infection was associated with a mild decrease in hemoglobin, hematocrit, and erythrocyte numbers. Bone marrow showed marked destruction of erythroid cells coincident with peak viremia. Our findings indicate that infection of healthy monkeys by simian parvovirus is self-limited and mild, with transient cessation of erythropoiesis. Our study has reproduced Koch's postulates and further shown that simian parvovirus infection of monkeys is almost identical to human B19 parvovirus infection of humans. Accordingly, this animal model may prove valuable in the study of the pathogenesis of B19 virus infection.     

Impaired gamma interferon responses against parvovirus B19 by recently infected children

Parvovirus B19 is the causative agent of "fifth disease" of childhood. It has been implicated in a variety of conditions, including unsuccessful pregnancy and rheumatoid arthritis, and is a potential contaminant of blood products. There has been little study of immunity to parvovirus B19, and the exact nature of the protective humoral and cell-mediated immune response is unclear. Immune responses to purified virus capsid proteins, VP1 and VP2, were examined from a cohort of recently infected children and compared with responses from long-term convalescent volunteers. The results demonstrate that antibody reactivity is primarily maintained against conformational epitopes in VP1 and VP2. The unique region of VP1 appears to be a major target for cell-mediated immune responses, particularly in recently infected individuals. We confirm that antibody reactivity against linear epitopes of VP2 is lost shortly after infection but find no evidence of the proposed phenotypic switch in either the subclass of parvovirus B19-specific antibody or the pattern of cytokine production by antigen-specific T cells. The dominant subclass of specific antibody detected from both children and adults was immunoglobulin G1. No evidence was found for interleukin 4 (IL-4) or IL-5 production by isolated lymphocytes from children or adults. In contrast, lymphocytes from convalescent adults produced a typical type 1 response associated with high levels of IL-2 and gamma interferon (IFN-gamma). However, we observed a significant (P<0.001) deficit in the production of IFN-gamma in response to VP1 or VP2 from lymphocytes isolated from children. Taken together, these results imply that future parvovirus B19 vaccines designed for children will require the use of conformationally preserved capsid proteins incorporating Th1 driving adjuvants. Furthermore, these data suggest novel mechanisms whereby parvovirus B19 infection may contribute to rheumatoid arthritis and unsuccessful pregnancy.     

Novel B19-Like Parvovirus in the Brain of a Harbor Seal

Using random PCR in combination with next-generation sequencing, a novel parvovirus was detected in the brain of a young harbor seal (Phoca vitulina) with chronic non-suppurative meningo-encephalitis that was rehabilitated at the Seal Rehabilitation and Research Centre (SRRC) in the Netherlands. In addition, two novel viruses belonging to the family Anelloviridae were detected in the lungs of this animal. Phylogenetic analysis of the coding sequence of the novel parvovirus, tentatively called Seal parvovirus, indicated that this virus belonged to the genus Erythrovirus, to which human parvovirus B19 also belongs. Although no other seals with similar signs were rehabilitated in SRRC in recent years, a prevalence study of tissues of seals from the same area collected in the period 2008-2012 indicated that the Seal parvovirus has circulated in the harbor seal population at least since 2008. The presence of the Seal parvovirus in the brain was confirmed by real-time PCR and in vitro replication. Using in situ hybridization, we showed for the first time that a parvovirus of the genus Erythrovirus was present in the Virchow-Robin space and in cerebral parenchyma adjacent to the meninges. These findings showed that a parvovirus of the genus Erythrovirus can be involved in central nervous system infection and inflammation, as has also been suspected but not proven for human parvovirus B19 infection.     

Comparison of canine parvovirus with mink enteritis virus by restriction site mapping.

The genomes of canine parvovirus and mink enteritis virus were compared by restriction enzyme analysis of their replicative-form DNAs. Of 79 mapped sites, 68, or 86%, were found to be common for both types of DNA, indicating that canine parvovirus and mink enteritis virus are closely related viruses. Whether they evolved from a common precursor or whether canine parvovirus is derived from mink enteritis virus, however, cannot be deduced from our present data.     

Generation of an adenovirus-parvovirus chimera with enhanced oncolytic potential

In this study, our goal was to generate a chimeric adenovirus-parvovirus (Ad-PV) vector that combines the high-titer and efficient gene transfer of adenovirus with the anticancer potential of rodent parvovirus. To this end, the entire oncolytic PV genome was inserted into a replication-defective E1- and E3-deleted Ad5 vector genome. As we found that parvoviral NS expression inhibited Ad-PV chimera production, we engineered the parvoviral P4 early promoter, which governs NS expression, by inserting into its sequence tetracycline operator elements. As a result of these modifications, P4-driven expression was blocked in the packaging T-REx-293 cells, which constitutively express the tetracycline repressor, allowing high-yield chimera production. The chimera effectively delivered the PV genome into cancer cells, from which fully infectious replication-competent parvovirus particles were generated. Remarkably, the Ad-PV chimera exerted stronger cytotoxic activities against various cancer cell lines, compared with the PV and Ad parental viruses, while being still innocuous to a panel of tested healthy primary human cells. This Ad-PV chimera represents a novel versatile anticancer agent which can be subjected to further genetic manipulations in order to reinforce its enhanced oncolytic capacity through arming with transgenes or retargeting into tumor cells.     

Assembly of empty capsids by using baculovirus recombinants expressing human parvovirus B19 structural proteins.

Empty parvovirus B19 capsids were isolated from insect cells infected with a recombinant baculovirus expressing parvovirus B19 VP2 alone and also with a double-recombinant baculovirus expressing both VP1 and VP2. That VP2 alone can assemble to form capsids is a phenomenon not previously observed in parvoviruses. The stoichiometry of the capsids containing both VP1 and VP2 was similar to that previously observed in parvovirus B19-infected cells. The capsids were similar to native capsids in size and appearance, and their antigenicity was demonstrated by immunoprecipitation and enzyme-linked immunosorbent assay with B19-specific antibodies.     

The evolution of defective and autonomous parvoviruses.

Because of the small size and genetic simplicity of small DNA viruses, parvoviruses would appear to be excellent models for studying viral evolution and adaptation. In an earlier publication we hypothesized the evolution of sequences of cellular "junk" DNA into protective interfering transposons. These transposons would interfere with invading pathogenic viruses by competing with the pathogen DNA for replicative enzymes. We speculated that a small, defective parvovirus, the adeno-associated virus (AAV), which usually requires the presence of a pathogenic helper virus to replicate, may have evolved from such a piece of cellular "junk" DNA. Our theory predicted that AAVs, as a consequence of their defective nature, developed under pressures favoring maintenance of their transposon like qualities. In contrast, disease-causing, autonomous, non-defective parvoviruses such as the B19 agent of humans and the canine parvovirus, even though their origins may have been in cellular DNA, would appear to have developed under totally different evolutionary pressures. In this paper we will present evidence for a common ancestry for the defective and autonomous parvoviruses and discuss the divergent paths this evolution may have taken in establishing the two genera.     

Cellular adaptive immune response against porcine circovirus type 2 in subclinically infected pigs

Background  

Porcine circovirus type 2 (PCV2) is a dominant causative agent of postweaning multisystemic wasting syndrome (PMWS), a multifactorial disease complex with putative immunosuppressive characteristics. Little is known about adaptive PCV2-specific immune responses in infected pigs. Therefore, the T and B cell responses following PCV2 infection in 3-week old SPF piglets infected with PCV2 or PCV2 plus porcine parvovirus (PPV) were studied.     

犬科、猫科动物细小病毒血清抗体调查

肉食兽细小病毒属于细小病毒科、细小病毒属中的一类病毒,能够感染多种动物,导致犬的出血性肠炎、幼龄犬的心肌炎、猫的白细胞减少、出血性肠炎、幼龄猫的共济失调症以及水貂的肠炎等多种疾病.血清学调查发现,由肉食兽细小病毒引起的疾病存在于世界各地.在我国,无论是家养还是野生动物均有细小病毒相关疫病的流行,对我国犬科和猫科动物的生存和健康构成巨大威胁(许树林等,1996;宋桂强等,2007).     

Detection of rodent parvoviruses by use of fluorogenic nuclease polymerase chain reaction assays

Polymerase chain reaction (PCR) assays have proven useful for detection of rodent parvoviruses in animals and contaminated biological materials. Fluorogenic nuclease PCR assays combine PCR with an internal fluorogenic hybridization probe, eliminating post-PCR processing and potentially enhancing specificity. Consequently, three fluorogenic nuclease PCR assays were developed, one that detects all rodent parvoviruses, one that specifically detects minute virus of mice (MVM), and one that specifically detects mouse parvovirus 1 (MPV) and hamster parvovirus (HaPV). When rodent parvoviruses and other rodent DNA viruses were evaluated, the rodent parvovirus assay detected only rodent parvovirus isolates, whereas the MVM and MPV/HaPV assays detected only the MVM or MPV/ HaPV isolates, respectively. Each assay detected the equivalent of 10 or fewer copies of target template, and all fluorogenic nuclease PCR assays exceeded the sensitivities associated with previously reported PCR assays and mouse antibody production testing. In addition, each fluorogenic nuclease PCR assay detected the targeted parvovirus DNA in tissues obtained from mice experimentally infected with MVM or MPV. Results of these studies indicate that fluorogenic nuclease PCR assays provide a potentially high-throughput, PCR-based method to detect rodent parvoviruses in infected mice and contaminated biological materials.     

Management strategies for controlling endemic and seasonal mouse parvovirus infection in a barrier facility

Despite improved diagnostic and rederivation capabilities, research facilities still struggle to manage parvovirus infections (e.g., mouse parvovirus (MPV) and minute virus of mice) in mouse colonies. Multi-faceted approaches are needed to prevent adventitious organisms such as MPV from breaching a barrier facility. In this article, the authors document recent changes to the Salk Institute's animal care program that were intended to help manage mouse parvovirus in the barrier facility. Specifically, the Institute started to use a new disinfectant and to give mice irradiated feed. The authors found an association between these modifications and a reduction in MPV incidence and prevalence in endemically infected colonies. These data suggest that using irradiated feed and appropriate disinfectants with contemporary management practices can be an effective plan for eradicating or controlling MPV infection in a research facility. The authors recommend further study of the environmental risk factors for parvovirus infection and of potential biological interactions associated with the use of irradiated feed.     

Parvovirus host range,cell tropism and evolution

The past few years have seen major advances in our understanding of the controls of evolution, host range and cell tropism of parvoviruses. Notable findings have included the identification of the transferrin receptor TfR as the cell surface receptor for canine parvovirus and feline panleukopenia virus, and also the finding that specific binding to the canine TfR led to the emergence of canine parvovirus as a new pathogen in dogs. The structures of the adeno-associated virus-2 and porcine parvovirus capsids, along with those of the minute virus of mice, have also advanced our understanding of parvovirus biology. Structure-function studies have shown that in several different parvoviruses the threefold spikes or peaks of the capsid control several aspects of cell tropism and host range, and that those are subject to selective pressures leading to viral evolution. The cell and tissue tropisms of different adeno-associated virus serotypes were demonstrated to be due, in part, to specific receptor binding.     

鸭细小病毒04Nb株的分离鉴定与rep基因测序与分析

采集浙江宁波地区以腹泻、呼吸困难为主要症状的病鸭肝组织,接种正常鸭胚尿囊腔增殖病毒。雏鸭感染试验显示发病症状及病理变化明显,死亡率为75%。电镜下可见纯化病毒直径约20nm左右的球形病毒粒子。免疫琼脂扩散实验结果显示与鸭细小病毒(duckparvovirusDPV)标准株阳性血清有明显沉淀线。经SDS-PAGE呈现3条结构蛋白带,与DPV标准株一致;参照GenBankDPV非结构蛋白基因序列设计引物,PCR扩增反应获得目的条带,克隆测序后,与DPV代表株序列同源性达98%。根据上述实验结果,确定引起本次鸭场疫病的病原为DPV。为进一步研究该分离株rep基因的序列特征,对其rep基因克隆测序,与GenBank中两株DPV、两株鹅细小病毒(GPV)进行序列比对,结果显示rep基因核苷酸序列与DPV参考毒株同源性为98%以上,与GPV同源性为80%左右。     

Human parvovirus B19 can infect cynomolgus monkey marrow cells in tissue culture.

The human pathogenic parvovirus B19 cannot be grown in standard tissue culture but propagates in human bone marrow, where it is cytotoxic to erythroid progenitor cells. We now show that parvovirus B19 can replicate in cynomolgus bone marrow. Cynomolgus monkeys may be a suitable animal model for pathogenesis studies of parvovirus B19.     

Parvovirus infection-induced cell death and cell cycle arrest

The cytopathic effects induced during parvovirus infection have been widely documented. Parvovirus infection-induced cell death is often directly associated with disease outcomes (e.g., anemia resulting from loss of erythroid progenitors during parvovirus B19 infection). Apoptosis is the major form of cell death induced by parvovirus infection. However, nonapoptotic cell death, namely necrosis, has also been reported during infection of the minute virus of mice, parvovirus H-1 and bovine parvovirus. Recent studies have revealed multiple mechanisms underlying the cell death during parvovirus infection. These mechanisms vary in different parvoviruses, although the large nonstructural protein (NS)1 and the small NS proteins (e.g., the 11 kDa of parvovirus B19), as well as replication of the viral genome, are responsible for causing infection-induced cell death. Cell cycle arrest is also common, and contributes to the cytopathic effects induced during parvovirus infection. While viral NS proteins have been indicated to induce cell cycle arrest, increasing evidence suggests that a cellular DNA damage response triggered by an invading single-stranded parvoviral genome is the major inducer of cell cycle arrest in parvovirus-infected cells. Apparently, in response to infection, cell death and cell cycle arrest of parvovirus-infected cells are beneficial to the viral cell lifecycle (e.g., viral DNA replication and virus egress). In this article, we will discuss recent advances in the understanding of the mechanisms underlying parvovirus infection-induced cell death and cell cycle arrest.     

Efficacy of three microbiological monitoring methods in a ventilated cage rack

The use of individually ventilated caging (IVC) to house mice presents new challenges for effective microbiological monitoring. Methods that exploit the characteristics of IVC have been developed, but to the authors' knowledge, their efficacy has not been systematically investigated. Air exhausted from the IVC rack can be monitored, using sentinels housed in cages that receive rack exhaust air as their supply air, or using filters placed on the exhaust air port. To aid laboratory animal personnel in making informed decisions about effective methods for microbiological monitoring of mice in IVC, the efficacy of air monitoring methods was compared with that of contact and soiled bedding sentinel monitoring. Mice were infected with mouse hepatitis virus (MHV), mouse parvovirus (MPV), murine rotavirus (agent of epizootic diarrhea of mice [EDIM]), Sendai virus (SV), or Helicobacter spp. All agents were detected using contact sentinels. Mouse hepatitis virus was effectively detected in air and soiled bedding sentinels, and SV was detected in air sentinels only. Mouse parvovirus and Helicobacter spp. were transmitted in soiled bedding, but the efficacy of transfer was dependent on the frequency and dilution of soiled bedding transferred. Results were similar when the IVC rack was operated under positive or negative air pressure. Filters were more effective at detecting MHV and SV than they were at detecting MPV. Exposure of sentinels or filters to exhaust air was effective at detecting several infectious agents, and use of these methods could increase the efficacy of microbiological monitoring programs, especially if used with soiled bedding sentinels. In contemporary mouse colonies, a multi-faceted approach to microbiological monitoring is recommended.