Epitope mapping and cellular localization of swine acute diarrhea syndrome coronavirus nucleocapsid protein using a novel monoclonal antibody.

A swine acute diarrhea syndrome coronavirus (SADS-CoV) that causes severe diarrhea in suckling piglets was identified in Southern China in 2017. To develop an antigen that is specific, sensitive, and easy to prepare for serological diagnosis, antigenic sites in the SADS-CoV nucleocapsid (N) protein were screened. We generated and characterized an N-reactive monoclonal antibody (mAb) 3E9 from mice immunized with recombinant N protein. Through fine epitope mapping of mAb 3E9 using a panel of eukaryotic-expressed polypeptides with GFP-tags, we identified the motif 343DAPVFTPAP351 as the minimal unit of the linear B-cell epitope recognized by mAb 3E9. Protein sequence alignment indicated that 343DAPVFTPAP351 was highly conserved in different SADS-CoV strains and SADS-related coronaviruses from bat, with one substitution in this motif in HKU2-related bat coronavirus. Using mAb 3E9, we observed that N protein was expressed in the cytoplasm and was in the nucleolus during SADS-CoV replication. N protein was immunoprecipitated from SADS-CoV-infected Vero E6 cells. Taken together, our results indicated that 3E9 mAb could be a useful tool to investigate the structure and function of N protein during viral replication.

Comparison of individual, group and environmental sampling strategies to conduct influenza surveillance in pigs.

BACKGROUND: Influenza A virus (IAV) is an important pathogen in pigs that affects productivity and has important public health implications because of its zoonotic nature. Surveillance is central to the control of influenza, however, detection of IAV infections can be challenging in endemically infected herds with low prevalence of infection. METHODS: In groups of suckling (18-21 days of age) and growing (35-45 days of age) pigs, we compared various sampling approaches to detect, isolate and sequence IAV using individual (nasal swabs, nasal wipes and oropharyngeal swabs), group (oral fluids, surface wipes and sow udder skin wipes) and environmental (airborne particles deposited on surfaces and air samples) sampling approaches. All samples were tested by IAV rRT-PCR and a subset was used for virus isolation and direct sequencing. RESULTS: In general, environmental and group samples resulted in higher odd ratios (range = 3.87-16.5, p-value < 0.05) of detecting a positive sample by rRT-PCR compared to individual pooled samples, except for oropharyngeal swabs (OR = 8.07, p-value < 0.05). In contrast, individual samples were most likely to yield a viral isolate by cell culture. Oropharyngeal swabs in suckling pigs (78.4%), and nasal swabs (47.6%) or nasal wipes (45%) in growing pigs, and udder wipes in lactating sows (75%) were the preferred samples to obtain an isolate. CONCLUSIONS: Our findings indicate that group and environmental sampling strategies should be considered in influenza surveillance programs in particular if the goal is just to detect infection. This study provides new information on sampling approaches to conduct effective influenza surveillance in pigs and identifies udder wipes from lactating sows as a novel sample type that offers a convenient, cheap and sensitive manner to monitor IAV in litters prior to weaning.

Detection and genetic characterization of porcine astroviruses in piglets with and without diarrhea in Thailand.

Porcine astrovirus (PAstV) is widely distributed and highly prevalent among pigs, nevertheless its clinical significance remains unclear as it can be detected in both diarrheic and in healthy pigs. Information about the prevalence, clinical significance and molecular characterization of PAstV in Thailand is not available. This study investigated the prevalence of PAstV in 488 fecal samples collected from piglets with and without diarrhea in 28 pig farms in northern and central parts of Thailand using RT-PCR. The overall prevalence of PAstV infection was 6.5% (32/488), of which 21/251 (8.4%) were in diarrheic and 11/237 (4.6%) were in healthy pigs. Of 32 positive samples, 46.9% were positive for PAstV alone whereas 53.1% were co-infected with porcine group A rotavirus (PRVA). A phylogenetic analysis of the partial RNA-dependent RNA polymerase/capsid genes revealed two lineages of PAstV strains detected in this study. PAstV4 was the most dominant genotype (92%), followed by PAstV2 (8%). This study revealed for the first time that PAstV4 and PAstV2 were circulating in Thailand with PAstV4 as the most dominant genotype in pig herds in northern and central parts of Thailand.

Detection of atypical porcine pestivirus in Brazil in the central nervous system of suckling piglets with congenital tremor.

Atypical porcine pestivirus (APPV) has been detected in piglets with congenital tremor (CT) from three different continents including North America, Europe and Asia. Thirteen piglets from four farms in two different states in Brazil with CT were sampled. Viral RNA was detected by quantitative real-time PCR in the cerebellum or cerebellum and spinal cord in the 100% of the piglets with CT, and APPV was not detected in any tissue sample from clinically non-affected piglets with the exception of the cerebellum of one piglet from Farm A. Piglets with CT had an odds ratio of 99.0 (95% CI 3.4, 2823.8; p = .0072) compared to piglets without CT to test positive for APPV by qRT-PCR. A subset of positive samples was selected for sequencing of the NS3 gene. Phylogenetic analysis revealed that Brazilian sequences of the NS3 formed an independent cluster and had the highest sequence identity with a sequence from the United States. This is the first identification of APPV infection in piglets with CT in South America.

Identification of a natural recombinant transmissible gastroenteritis virus between Purdue and Miller clusters in China.

Transmissible gastroenteritis virus (TGEV) is an infective coronavirus (CoV) that causes diarrhea-related morbidity and mortality in piglets. For the first time, a natural recombination strain of a TGEV Anhui Hefei (AHHF) virus between the Purdue and the Miller clusters was isolated from the small intestine content of piglets in China. A phylogenetic tree based on a complete genome sequence placed the TGEV AHHF strain between the Purdue and the Miller clusters. The results of a computational analysis of recombination showed that the TGEV AHHF strain is a natural recombinant strain between these clusters. Two breakpoints located in the open reading frame 1a (ORF1a) and spike (S) genes were identified. The pathogenicity of the TGEV AHHF strain was evaluated in piglets, and the results show that TGEV AHHF is an enteric pathogenic strain. These results provide valuable information about the recombination and evolution of CoVs and will facilitate future investigations of the molecular pathogenesis of TGEV.

Increased frequency of porcine epidemic diarrhea virus shedding and lesions in suckling pigs compared to nursery pigs and protective immunity in nursery pigs after homologous re-challenge.

Porcine epidemic diarrhea virus (PEDV) causes enteric disease in pigs and spreads rapidly after entering naïve pig populations. The objectives were to (1) compare the disease course following inoculation with PEDV isolate US/Colorado/2013 in naïve 10 day and 8 week-old pigs, and (2) contrast the naïve response to homologous challenge in 8 week-old pigs. Pigs were randomly assigned into group 1 (n = 40, no PEDV exposure), group 2 (n = 43, PEDV inoculation at 10 days of age) and group 3 (n = 48, PEDV inoculation at 8 weeks of age). Thirty-three group 2 pigs received a homologous challenge at 8 weeks of age. Following primary or secondary inoculation, 3-10 pigs were euthanized at days post-inoculation (dpi) 1, 2, 3, 7 or 14. Clinical signs were more pronounced in 10 day-old pigs compared to 8 week-old pigs at dpi 2 and 3, a higher number of 10 day-old pigs shed PEDV RNA in feces compared to 8 week-old pigs. Typical severe atrophic enteritis of PEDV infection was observed at dpi 3 in both age groups, and at dpi 4 and 14 fecal shedding patterns were also similar. While both age groups had seroconverted to PEDV by dpi 14, IgG levels were higher in 8 week-old pigs. PEDV IgA antibodies were detected in feces of approximately 50% of the pigs at dpi 44. In homologous challenged pigs, no clinical signs or lesions were found, and PEDV fecal shedding was restricted to less than 10% of the pigs indicating the existence of homologous protection 44 days after initial PEDV exposure.

Jaagsiekte Sheep Retrovirus Can Reach Peyer's Patches and Mesenteric Lymph Nodes of Lambs Nursed by Infected Mothers.

Ovine pulmonary adenocarcinoma (OPA) is a contagious lung cancer of sheep caused by jaagsiekte sheep retrovirus (JSRV). It is generally accepted that transmission by the respiratory route occurs under natural conditions. However recent studies strongly indicate that JSRV can also be transmitted to lambs perinatally via colostrum and milk (C/M). The aim of this work was to confirm that C/M can transmit JSRV infection to lambs under natural conditions and investigate the initial events associated with this transmission route. We have analyzed the presence of JSRV in C/M samples from 22 naturally infected, asymptomatic ewes throughout a lactation period, and in various tissues collected from a group of 36 of their lambs that were fed naturally. The lambs were euthanized at 12, 24, 48, and 72 hours and at 5 and 10 days after birth. We detected JSRV-provirus by PCR in the somatic C/M cells from 10/22 ewes (45.45%). The virus was also detected in 9/36 lambs (25%). JSRV-infected cells, with lymphoreticular-like morphology, were observed by immunohistochemistry (IHC) and in situ hybridization (ISH) in Peyer's patches (PP) from the small intestine of the youngest lambs and in mesenteric lymph nodes (MLN) from lambs older than 72 hours. The virus was also detected by PCR in white blood cells (WBC) in 2/36 lambs (5.5%). These results confirm colostral transmission of JSRV to lambs under natural conditions. Infected lymphoreticular cells contained in C/M appear to be involved. These cells can cross the intestinal barrier of newborn lambs, reach the MLN and enter into circulation.

Influenza Transmission in the Mother-Infant Dyad Leads to Severe Disease, Mammary Gland Infection, and Pathogenesis by Regulating Host Responses.

Seasonal influenza viruses are typically restricted to the human upper respiratory tract whereas influenza viruses with greater pathogenic potential often also target extra-pulmonary organs. Infants, pregnant women, and breastfeeding mothers are highly susceptible to severe respiratory disease following influenza virus infection but the mechanisms of disease severity in the mother-infant dyad are poorly understood. Here we investigated 2009 H1N1 influenza virus infection and transmission in breastfeeding mothers and infants utilizing our developed infant-mother ferret influenza model. Infants acquired severe disease and mortality following infection. Transmission of the virus from infants to mother ferrets led to infection in the lungs and mother mortality. Live virus was also found in mammary gland tissue and expressed milk of the mothers which eventually led to milk cessation. Histopathology showed destruction of acini glandular architecture with the absence of milk. The virus was localized in mammary epithelial cells of positive glands. To understand the molecular mechanisms of mammary gland infection, we performed global transcript analysis which showed downregulation of milk production genes such as Prolactin and increased breast involution pathways indicated by a STAT5 to STAT3 signaling shift. Genes associated with cancer development were also significantly increased including JUN, FOS and M2 macrophage markers. Immune responses within the mammary gland were characterized by decreased lymphocyte-associated genes CD3e, IL2Ra, CD4 with IL1ß upregulation. Direct inoculation of H1N1 into the mammary gland led to infant respiratory infection and infant mortality suggesting the influenza virus was able to replicate in mammary tissue and transmission is possible through breastfeeding. In vitro infection studies with human breast cells showed susceptibility to H1N1 virus infection. Together, we have shown that the host-pathogen interactions of influenza virus infection in the mother-infant dyad initiate immunological and oncogenic signaling cascades within the mammary gland. These findings suggest the mammary gland may have a greater role in infection and immunity than previously thought.

Infection of farmed pigs with porcine kobuviruses in Italy.

Two-hundred eight swine fecal samples from six Italian farms were tested using a kobuvirus-specific RT-PCR with primers that amplify a region within the 3D gene. All farms were kobuvirus positive, with prevalence rates ranging between 24 % and 84 %. Overall, 57.5 % of asymptomatic pigs and 49.7 % of animals with diarrhea were positive for kobuvirus. Sequence analysis showed a different predominant strain circulating on each farm and indicated that the strains detected were related to both European and Asiatic strains. A possible pathogenic role of kobuvirus should be investigated further, since infections with this virus occur frequently in pigs of different ages.

Porcine reproductive and respiratory syndrome virus (PRRSV) surveillance using pre-weaning oral fluid samples detects circulation of wild-type PRRSV.

Oral fluid samples collected from litters of piglets (n=600) one day prior to weaning were evaluated as a method to surveil for porcine reproductive and respiratory syndrome virus (PRRSV) infections in four sow herds of approximately 12,500 sow each. Serum samples from the litters' dam (n=600) were included for comparison. All four herds were endemically infected with PRRSV and all sows had been vaccinated ≥ 2 times with PRRSV modified-live virus vaccines. After all specimens had been collected, samples were randomized and assayed by PRRSV real-time reverse transcription polymerase chain reaction (RT-qPCR) and four PRRSV antibody ELISA assays (IgM, IgA, IgG, and Commercial Kit). All sow serum samples were negative by PRRSV RT-qPCR, but 9 of 600 oral fluid samples tested positive at two laboratories. Open reading frame 5 (ORF5) sequencing of 2 of the 9 positive oral fluid samples identified wild-type viruses as the source of the infection. A comparison of antibody responses in RT-qPCR positive vs. negative oral fluid samples showed significantly higher IgG S/P ratios in RT-qPCR-positive oral fluid samples (mean S/P 3.46 vs. 2.36; p=0.02). Likewise, sow serum samples from RT-qPCR-positive litter oral fluid samples showed significantly higher serum IgG (mean S/P 1.73 vs. 0.98; p<0.001) and Commercial Kit (mean S/P 1.97 vs. 0.98; p<0.001) S/P ratios. Overall, the study showed that pre-weaning litter oral fluid samples could provide an efficient and sensitive approach to surveil for PRRSV in infected, vaccinated, or presumed-negative pig breeding herds.

Antibody response and maternal immunity upon boosting PRRSV-immune sows with experimental farm-specific and commercial PRRSV vaccines.

The porcine reproductive and respiratory syndrome virus (PRRSV) causes reproductive failure in sows and respiratory disease in pigs of all ages. Despite the frequent use of vaccines to maintain PRRSV immunity in sows, little is known on how the currently used vaccines affect the immunity against currently circulating and genetically divergent PRRSV variants in PRRSV-immune sows, i.e. sows that have a pre-existing PRRSV-specific immunity due to previous infection with or vaccination against the virus. Therefore, this study aimed to assess the capacity of commercially available attenuated/inactivated PRRSV vaccines and autogenous inactivated PRRSV vaccines - prepared according to a previously optimized in-house protocol - to boost the antibody immunity against currently circulating PRRSV variants in PRRSV-immune sows. PRRSV isolates were obtained from 3 different swine herds experiencing PRRSV-related problems, despite regular vaccination of gilts and sows against the virus. In a first part of the study, the PRRSV-specific antibody response upon booster vaccination with commercial PRRSV vaccines and inactivated farm-specific PRRSV vaccines was evaluated in PRRSV-immune, non-pregnant replacement sows from the 3 herds. A boost in virus-neutralizing antibodies against the farm-specific isolate was observed in all sow groups vaccinated with the corresponding farm-specific inactivated vaccines. Use of the commercial attenuated EU type vaccine boosted neutralizing antibodies against the farm-specific isolate in sows derived from 2 farms, while use of the commercial attenuated NA type vaccine did not boost farm-specific virus-neutralizing antibodies in any of the sow groups. Interestingly, the commercial inactivated EU type vaccine boosted farm-specific virus-neutralizing antibodies in sows from 1 farm. In the second part of the study, a field trial was performed at one of the farms to evaluate the booster effect of an inactivated farm-specific vaccine and a commercial attenuated EU-type vaccine in immune sows at 60 days of gestation. The impact of this vaccination on maternal immunity and on the PRRSV infection pattern in piglets during their first weeks of life was evaluated. Upon vaccination with the farm-specific inactivated vaccine, a significant increase in farm-specific virus-neutralizing antibodies was detected in all sows. Virus-neutralizing antibodies were also transferred to the piglets via colostrum and were detectable in the serum of these animals until 5 weeks after parturition. In contrast, not all sows vaccinated with the commercial attenuated vaccine showed an increase in farm-specific virus-neutralizing antibodies and the piglets of this group generally had lower virus-neutralizing antibody titers. Interestingly, the number of viremic animals (i.e. animals that have infectious virus in their bloodstream) was significantly lower among piglets of both vaccinated groups than among piglets of mock-vaccinated sows and this at least until 9 weeks after parturition. The results of this study indicate that inactivated farm-specific PRRSV vaccines and commercial attenuated vaccines can be useful tools to boost PRRSV-specific (humoral) immunity in sows and reduce viremia in weaned piglets.

Implication of Hsc70, PDI and integrin αvß3 involvement during entry of the murine rotavirus ECwt into small-intestinal villi of suckling mice.

In the present study, a homologous rotavirus, ECwt, infecting small intestinal villi isolated from ICR and BALB/c mice were used as a model for identifying cell-surface molecules involved in rotavirus entry. Small-intestinal villi were treated with anti-Hsc70, anti-PDI, anti-integrin ß3 or anti-ERp57 antibodies or their corresponding F(ab')2 fragments before inoculation with rotavirus ECwt, RRV or Wa. Pretreatment of villi decreased virus infectivity by about 50-100 % depending of the rotavirus strain, antibody structure and detection assay used. Similar results were obtained by treating viral inocula with purified proteins Hsc70, PDI or integrin ß3 before inoculation of untreated villi. Rotavirus infection of villi proved to be sensitive to membrane-impermeant thiol/disulfide inhibitors such as DTNB and bacitracin, suggesting the involvement of a redox reaction in infection. The present results suggest that PDI, Hsc70 and integrin ß3 are used by both homologous and heterologous rotaviruses during infection of isolated mouse villi.

Prevalence of antibodies against transmissible gastroenteritis virus and porcine respiratory coronavirus among pigs in six regions in Japan.

A total of 2,703 pig sera from 171 farms in six regions in Japan were screened for virus-neutralizing (VN) antibody against transmissible gastroenteritis virus (TGEV). Although none of the farms had clinical signs of transmissible gastroenteritis (TGE) or vaccination against TGEV, VN antibody was detected in 14.4% of sera at 30 farms (17.5%) across all six regions. On testing of 263 VN antibody-positive sera from 27 farms with a commercial blocking ELISA to distinguish TGEV and porcine respiratory coronavirus (PRCV) antibodies, 78.3% were positive for PRCV antibody only, while 12.5% were positive for TGEV antibody only or both TGEV and PRCV antibodies. Seven of the eight TGEV antibody-positive farms were also positive for PRCV antibody. Five months after the antibody examination, a TGE outbreak occurred at one of these seven farms. These results suggest that most of the detected VN antibodies were to PRCV, and that TGEV infection might be present at some PRCV-positive farms in Japan.

Efficacy of arbidol on lethal hantaan virus infections in suckling mice and in vitro.

AIM: Arbidol is an immunomodulator that was first developed in Russia. In this study, we report the antiviral activity of arbidol against Hantaan virus (HTNV) in vitro and in vivo. METHODS: The antiviral activity of arbidol in vitro was determined by plaque-forming assay, ranging from 0.5 to 8 microg/mL. To investigate whether arbidol has an antiviral effect in vivo, suckling BALB/c mice infected with HTNV were treated with arbidol at 24 h before infection with a 5, 10 or 20 mg.kg(-1).d(-1), once per day, for 10 days. On day 12 and 28 post infection (pi), histopathological changes and viral antigen were detected. On days 4, 8, 12, and 16 pi, the viral load of target organs and serum TNF-alpha levels of arbidol-treated animals were determined. RESULTS: Arbidol was found to have potent inhibitory activity against HTNV when added in vitro before or after viral infection, with a 50% inhibitory concentration (IC(50)) of 0.9 and 1.2 microg/mL, respectively. The 50% lethal dose (LD(50)) of arbidol for suckling mice was 78.42 mg.kg(-1).d(-1). Oral administration of arbidol increased both survival rate and mean time to death (MTD). Treatment with arbidol reduced histopathological changes, decreased viral load and viral antigen levels, and modulated the level of serum TNF-alpha. CONCLUSION: Arbidol has the ability to elicit protective antiviral activity against HTNV in vivo and in vitro.Acta Pharmacologica Sinica (2009) 30: 1015-1024; doi: 10.1038/aps.2009.53; published online 8 June 2009.

Excretion of hepatitis E virus by pigs of different ages and its presence in slurry stores in the United Kingdom.

Five faecal samples were collected from four different stages of production at each of 10 pig farms in the Yorkshire Humberside area of the UK, and samples of slurry were collected from nine of the farms. All the samples were tested for hepatitis E virus (HEV) RNA by a nested reverse transcriptase PCR. At least one sample from the pigs on each of the farms tested positive for hev; its prevalence in the 10 herds varied from 5 per cent to 35 per cent and its mean prevalence was 21.5 per cent. The mean prevalence in pigs aged three to five weeks was 26.0 per cent, in pigs aged 10 to 12 weeks 44.0 per cent, in pigs aged 22 to 24 weeks 8.9 per cent, and in adult dry sows 6.0 per cent. Two of the nine slurry lagoons tested positive for HEV RNA. Phylogenetic analysis of the sequence data indicated that the strains of the virus were of genotype 3 and closely related to strains detected in other pigs and in human beings in the UK.

Identification of bovine viral diarrhea virus type 2 in Korean native goat (Capra hircus).

In the genus Pestivirus, four genetically distinct viral species are currently recognized: bovine viral diarrhea viruses type 1 and 2 (BVDV-1, BVDV-2), classical swine fever virus (CSFV) and border disease virus (BDV). BVDV-1 and BDV infections have been described in goat species. Since 1998, border disease (BD) like symptoms in goats have been reported repeatedly in two southern-most provinces of Korea, which until then had been regarded as being free from BD. As a result of retrospective investigations of BD-like syndrome in goat reported between 1998 and 2004, a pestivirus was identified from intestinal content of an affected kid submitted in 1999. Both sequences of 5'-non-coding region and complete N(pro) gene from the isolate were analyzed to identify the genotype. Interestingly, the results revealed that the isolate belonged to BVDV-2 that is rarely reported even in cattle. The isolate showed close relationship to North American and European strains rather than the geographically closer Japanese strains. To authors' knowledge, this is the first identification of BVDV-2 in goat species.

Passive protection to bovine rotavirus (BRV) infection induced by a BRV VP8* produced in plants using a TMV-based vector.

We have previously reported on the use of a tobacco mosaic virus (TMV) vector TMV-30B to express foreign viral antigens for use as experimental immunogens. Here we describe the development of an improved TMV-30B vector that adds a sequence of 7 histidine residues to the C-terminus of recombinant proteins expressed in the vector. We used this TMV-30B-HISc vector to express the VP8* fragment of the VP4 protein from bovine rotavirus (BRV) strain C-486 in plants. Recombinant VP8* protein was purified from N. benthamiana leaves at 7 days post-inoculation by immobilized metal affinity chromatography. The plant-produced VP8* was initially detected using anti-His tag mAb and its antigenic nature was confirmed using both monoclonal and polyclonal specific antisera directed against BRV. Adult female mice, inoculated by the intraperinoteal route with an immunogen containing 4 microg of recombinant VP8*, developed a specific and sustained response to the native VP8* from the homologous BRV. Eighty five percent of suckling mice from immunized dams that were challenged with the homologous virus at the fifth day of age were protected from virus as compared to 35% of the pups from mothers immunized with a control protein. These results demonstrate that the plant-produced VP8* was able to induce passive protection in the new born through the immunization of dams. This suggests that the technology presented here provides a simple method for using plants as an inexpensive alternative source for production of recombinant anti-rotavirus antigens.

Virulence of variola viruses for suckling mice.

We report work done in 1971 to determine the quantitative virulence for suckling mice of 26 variola virus isolates from different countries and from cases of differing severity. Strains of recognized variola major and variola minor viruses differed up to 100-fold (expressed as the harmonic mean dose of inoculum which killed mice 2-4 d old, inoculated intracranially, in 5 d). Isolates from Indonesia and from East and West Africa gave intermediate values. Unlike tests on chick embryos, this test distinguished between African and Indonesian isolates.

Feline immunodeficiency virus is concentrated in milk early in lactation.

We studied mother-to-offspring transmission of feline immunodeficiency virus (FIV), focusing on milk-borne virus transmission in order to assess its similarities to perinatal HIV transmission. We also attempted to evaluate the influence of intragestational treatment with 9-[2-(phosphono-methoxy)-propyl]adenine (PMPA) on virus transmission to offspring. Eleven female cats (queens), chronically infected with FIV-B-2542 and bred to an FIV-negative male, produced a total of 25 viable and 18 nonviable term kittens. Overall, the vertical transmission rate by untreated queens was 22%, similar to that for HIV, which unfortunately precluded adequate assessment of PMPA efficacy. However, at delivery 9 of 10 queens (90%) had higher viral RNA loads in milk (4 x 10(4) to 4 x 10(8) viral copies/ml) than in plasma (5 x 10(3) to 2.5 x 10(6) viral copies/ml). Conversely, 10 of 11 queens (91%) had lower proviral loads in milk cells (0 to 10(2) proviral copies/microg DNA) than blood cells (10(2) to 10(4) proviral copies/microg DNA). Thus, FIV is concentrated in early milk despite relatively low proviral loads in milk cells, suggesting that virus may be actively secreted by the mammary gland for dissemination to offspring. FIV provides a model for the study of milk-borne lentivirus transmission and assessment of strategies to reduce postnatal HIV vertical transmission.

Passage of equine herpesvirus-1 in suckling mouse brain enhances extraneural virus growth and subsequent hematogenous neuroinvasion.

Intracerebral inoculation of field-isolates as well as established strains of equine herpesvirus-1 (EHV-1) in suckling mice results in viral replication in neurons and glial cells and induces encephalitis. By intraperitoneal (i.p.) inoculation, no histological lesion was observed in the central nervous system (CNS) in suckling mice with the EHV-1 HH1 strain (HH1), whereas a neuroadapted variant (NHH1) produced by serial passage of HH1 in the mouse brain caused severe encephalomyelitis after i.p. inoculation. The purpose of this study was to determine the route of neuroinvasion after i.p. inoculation of NHH1 and to clarify the effects of the brain passage on viral neuroinvasion. NHH1, but not HH1, targeted splenic and pulmonary macrophages and omental fat cells on days 1 and 2 post-inoculation (p.i.). From days 1 to 3 p.i., cell-associated viremia was occurred in NHH1-infected mice, but not in HH1-infected mice. On day 4 p.i., viral antigen was detected in a few endothelial cells, perivascular glial cells and neurons in the CNS in NHH1-infected mice. The number of viral antigen-positive cells increased markedly after day 5 p.i. In contrast, no viral antigen-positive cell was detected in the CNS in HH1-infected mice, except for a few nerve cells in the thoracic cord on day 4 p.i. These results suggest that NHH1 neuroinvasion is hematogenous and is correlated with enhanced extraneural virus growth.