Pseudorabies: experimental studies in raccoons with different virus strains

Raccoons (Procyon lotor) were infected by the nasal/oral route with as little as 10(2) plaque forming units (PFU) of pseudorabies virus (PrV). There was no apparent difference in the susceptibility of raccoons to infection with either of two virulent field strains or with the naturally avirulent K strain which has been used in modified live virus vaccines. Each of these three viruses was transmitted by contact to uninfected raccoons. All raccoons that were infected with virulent field strains died; however only two of 11 (18%) raccoons that were infected with the K strain died. One of four raccoons that survived infection with the K strain survived superinfection with virulent virus. This finding was significant because it could be a mechanism by which virulent PrV can be introduced and persist in the raccoon population. The possibility of this event occurring is increasing because of the widespread prevalence of PrV and the use of modified live virus vaccines for controlling clinical pseudorabies in swine. Virus neutralizing activity was found in five of 47 serums collected from raccoons that were trapped in PrV endemic areas. This observation implies that a herpesvirus, possibly PrV, may be present in the wild raccoon population.     

Serologic survey for selected infectious disease agents in raccoons from Illinois

The determination of serologic titers to infectious organisms is a valuable tool for quantitating exposure to disease organisms. Raccoons (Procyon lotor) were live-trapped from September 1989 to October 1993 and samples collected from two distinct locations in west-central Illinois (USA); a state recreational facility (Park) and privately owned farming property (Farm). Sera were submitted for testing Leptospira interrogans (serovars bratislava, canicola, grippotyphosa, hardjo, icterohemmorhagiae, and pomona), canine distemper virus (CDV), pseudorabies virus (PV), and Toxoplasma gondii. Two-hundred and twenty-two (48%) of 459 raccoons were seropositive for L. interrogans. Eighty-five (23%) out of 368 raccoons were seropositive for canine distemper virus. Eighty-two (17%) of 479 raccoons raccoons were seropositive for pseudorabies virus. One hundred and eight-four (49%) of 379 raccoons were seropositive for T. gondii. A significant difference (P < 0.05) in seroprevalence for L. interrogans between the park (43%) and farm (52%) areas was found. A correlation between increasing age and seroprevalence was found for L. interrogans, CDV, PV, and T. gondii. Furthermore, there was a significant difference in seroprevalence for T. gondii during the spring trapping seasons (73%), when compared with the fall (33%). This type of information on exposure to infectious agents is important for developing control programs to manage raccoon-human and raccoon-domestic animals interactions.     

A field trial to determine the feasibility of delivering oral vaccines to wild swine

A field study was conducted on Ossabaw Island, Georgia (USA) to determine the feasibility of delivering oral vaccines to wild swine (Sus scrofa). Baits were made of polymerbound fish meal and contained a gelatin capsule as a potential vaccine chamber. Two biomarkers, iophenoxic acid and tetracycline, were incorporated into each bait, and soured chicken mash was used as an attractant. Baits (n = 1,980) were distributed in a grid pattern on a 405-ha test site and monitored for animal disturbance. Within 72 hr, 88% of 393 monitored baits were gone, and observations of track-beds surrounding 100 baits indicated that at least 52% were taken by wild swine. Subsequent testing of 80 wild swine for the biomarkers revealed that 95% of the animals had consumed bait. Track-bed observations indicated that raccoons (Procyon lotor) were the only non-target animal that frequently took baits. Biomarker analyses indicated 44% of 16 raccoons tested had eaten bait. It was concluded that oral vaccine delivery to wild swine should be considered as a feasible method of control or eradication of pseudorabies and/or swine brucellosis in wild swine if effective vaccines become available.     

Potential sites of virus latency associated with indigenous pseudorabies viruses in feral swine

Free-ranging feral swine (Sus scrofa) are known to be present in at least 32 states of the USA and are continuously expanding their range. Infection with pseudorabies virus (PRV) occurs in feral swine and the primary route of transmission in free-living conditions seems to be venereal. Between 1995 and 1999, naturally infected feral swine and experimentally infected hybrid progeny of feral and domestic swine, were kept in isolation and evaluated for occurrence of latent PRV indigenous to feral swine in sacral and trigeminal ganglia and tonsil. Sacral ganglia were shown, by polymerase chain reaction (PCR) amplification of the thymidine kinase (TK) gene of PRV, to be the most frequent sites of latency of PRV. Nine (56%) of 16 sacral ganglia, seven (44%) of 16 trigeminal ganglia, and five (39%) of 13 tonsils from naturally infected feral swine were positive for PCR amplification of TK sequences of PRV. These tissues were negative for PRV when viral isolation was attempted in Vero cells. DNA sequencing of cloned TK fragments from the sacral ganglia of two feral swine, showed only one nucleotide difference between the two fragments and extensive sequence homology to fragment sequences from various domestic swine PRV strains from China, Northern Ireland, and the USA. The hybrid feral domestic swine, experimentally inoculated with an indigenous feral swine PRV isolate by either the genital or respiratory route, acquired the infection but showed no clinical signs of pseudorabies. Virus inoculated into either the genital or respiratory tract could, at times, be isolated from both these sites. The most common latency sites were the sacral ganglia, regardless of the route and dose of infection in these experimentally infected hybrids. Nine of 10 sacral ganglia, six of 10 trigeminal ganglia, and three of 10 tonsils were positive for PCR amplification of TK sequences. No virus was isolated from these tissues in Vero cells. The demonstration of the sacral ganglia as the most common sites of latency of pseudorabies viruses indigenous to feral swine, supports the hypothesis that these viruses are primarily transmitted venereally, and not by the respiratory route as is common in domestic swine, in which the trigeminal ganglia are the predominant sites of virus latency.     

Venereal transmission of pseudorabies viruses indigenous to feral swine

Between 1995 and 1998, we designed a series of studies in which we attempted to determine the main routes of transmission involved in the natural infection of pseudorabies virus (PRV) indigenous to free-ranging feral swine (Sus scrofa). Naturally infected feral sows transmitted the infection to uninfected feral boars, with which they had been commingled for a 6-wk period. Pseudorabies virus was isolated from boar preputial swabs, but not from nasal swabs. Three of the same PRV-infected feral sows did not transmit the infection to domestic boars during a 16 wk commingling period, despite the fact that they became pregnant. Feral boars, naturally infected with PRV transmitted the virus to domestic gilts while penned together during 6 wk. Pseudorabies virus was isolated from vaginal swabs, but not from nasal swabs of gilts, after 2 and 3 wk of commingling. When the same infected boars were commingled with either feral or domestic boars for 13 wk, PRV transmission did not occur. None of the exposed boars developed neutralizing antibodies or yielded virus from their preputial or nasal swabs. Our results indicate that PRV indigenous to feral swine is preferentially transmitted to feral or domestic swine of the opposite sex by the venereal route. This mode of transmission differs from that seen in the natural transmission of PRV prevalent in domestic swine, where contaminated secretions, excretions and aerosols are responsible for the spread of the virus. Based on these results, we feel that as long as feral swine do not come into direct contact with domestic swine, PRV-infected feral swine probably pose only a limited risk to the success of the National Pseudorabies Eradication Program. The fact that PRV is usually transmitted from feral to domestic swine at the time of mating would indicate that the isolation of domestic herds by the use of a "double fence," should be adequate protection against reinfection with PRV.     

Persistence of pseudorabies virus in feral swine populations

Serologic surveys for evidence of exposure to pseudorabies virus (PRV) in feral swine were conducted from November 2001 to April 2002 at 10 sites in the southeastern United States, where evidence of previous PRV exposure had been documented during 1979-89. Sera were tested in the field on the day of collection by latex agglutination. Maximum sample size per site was to be 30 animals, but sampling was discontinued before reaching this number when positive results were obtained. Positive results were obtained at all of the study sites, demonstrating long-term persistence of PRV in feral swine populations. Overall, 38 of 100 (38%) animals were positive for antibodies. Consistent results from latex agglutination tests conducted in the field and laboratory demonstrated that this test was useful as a rapid and reliable diagnostic tool when used in the field.     

Activation of a ribosomal protein S6 kinase in mouse fibroblasts during infection with herpesvirus

If confluent fibroblasts are infected with the swine alpha-herpes virus, pseudorabies virus, ribosomal protein S6 becomes phosphorylated after a lag of approximately 2 h. When cell-free extracts were prepared from such cells in the presence of glycerol 2-phosphate and EGTA, a ribosomal protein S6 kinase activity was found to appear at approximately the same time as the phosphorylation in vivo. This protein kinase was similar to that activated in the same cells by replenishing the nutrient medium, and in other quiescent cells by the action of growth factors and mitogens. It was distinct from the previously described pseudorabies virus protein kinase, which is unique to infected cells. When medium from cells infected with pseudorabies virus was freed of virus and added to confluent fibroblasts, rapid activation of the ribosomal protein S6 kinase activity occurred. A similar, although more limited, effect could be seen when the pH of the medium was increased. These results suggest that the phosphorylation of ribosomal protein S6 in cells infected with herpes virus is a consequence of the production of a factor which initiates the metabolic programme for cellular growth. The possible function of this effect in the infective strategy of herpes viruses is discussed in relation to requirements for the replication of viral DNA.     

猪源胰酶中特异病毒和非特异病毒安全检测简

目的：对猪源胰酶样品进行病毒检测,以评价其病毒安全性。方法：非特异性病毒检测采用致细胞病变、血凝吸附试验和形态学检测方法;特异性病毒检测包括猪繁殖与呼吸障碍综合征病毒（PRRSV）、猪瘟病毒（CSFV）、猪圆环病毒（PCV）、猪细小病毒（PPV）、猪口蹄疫病毒（FMDV）和猪伪狂犬病病毒（PRV）特异性核酸检测,以及CSFV、PPV、PRV特定性抗原蛋白直接免疫荧光检测。结果：受检样品非特异性病毒检测中,未见可观察到的细胞病变产生和病毒粒子,对豚鼠、鸡和人的0型红细胞无凝集现象。特异性病毒检测中,PRRSV、CSFV、PCV、PPV、FMDV和PRV核酸检测均为阴性,CSFV、PPV、PRV免疫荧光检测均为阴性。结论：猪源胰酶样品经非特异性病毒检测和特异性病毒检测均无可检出的病毒存在。     

Live attenuated pseudorabies virus expressing envelope glycoprotein E1 of hog cholera virus protects swine against both pseudorabies and hog cholera

To investigate whether live attenuated pseudorabies virus (PRV) can be used as a vaccine vector, PRV recombinants that expressed envelope glycoprotein E1 of hog cholera virus (HCV) were generated. Pigs inoculated with these recombinants developed high levels of neutralizing antibodies against PRV and HCV and were protected against both pseudorabies and hog cholera (classical swine fever).     

Experimental infection of European wild swine (Sus scrofa L.) with pseudorabies virus

Experimental infection with pseudorabies virus was carried out by oral exposure of four young wild swine held in contact with two unexposed controls. No disease was observed but virological procedures indicated that the virus was shed in saliva and, in one case, in the nasal discharge, with subsequent infection of the control animals. After slaughter the virus was reisolated from the tonsils but not from lungs and brain. Virus reisolation from the tonsils was obtained in two animals after the throat swabs became negative. Virus neutralizing antibodies were detected.     

Vaccination of free-ranging Pennsylvania raccoons (Procyon lotor) with inactivated rabies vaccine

Thirty-nine free-ranging raccoons (Procyon lotor) in an endemic rabies area of Pennsylvania (USA) were vaccinated with a single intramuscular inoculation of commercial inactivated rabies virus vaccine, 17 June to 23 August 1987. Paired serum samples, pre- and postvaccination, were obtained from eight raccoons and were analyzed in vitro for rabies virus neutralizing antibody using a modified rapid fluorescent focus inhibition test. Seven of eight (88%) recaptured raccoons demonstrated seroconversion within 15 to 26 days of vaccination. At 1 yr postvaccination, three vaccinated raccoons were recaptured and challenged in captivity with street rabies virus, resulting in the death of two of three vaccinates and five of five unvaccinated control raccoons.     

Detection of porcine circovirus type 1 in commercial porcine vaccines by loop-mediated isothermal amplification

A loop-mediated isothermal amplification (LAMP) method with a real-time monitoring system was developed for the detection of porcine circovirus type 1 (PCV1) in commercial swine vaccines. This method was highly specific for PCV1. No cross-reaction to porcine circovirus type 2, porcine parvovirus, pseudorabies virus, classical swine fever virus, and porcine reproductive and respiratory syndrome virus was observed. The analytical sensitivity of the LAMP for PCV1 DNA was 10 copies/μl in the case of positive recombinant plasmid comparable to that obtained from the nested polymerase chain reaction (nested PCR). Furthermore, 25 commercial swine vaccines were tested by both the LAMP and the nested PCR, and three of them were tested positive for PCV1 DNA. These results indicate that PCV1 DNA can be real-time detected by the LAMP; the method was highly specific, sensitive, and rapid for the detection of PCV1 DNA, particularly in commercial swine vaccines.     

A serosurvey for Brucella suis, classical swine fever virus, porcine circovirus type 2, and pseudorabies virus in feral swine (Sus scrofa) of eastern North Carolina

As feral swine (Sus scrofa) populations expand their range and the opportunity for feral swine hunting increases, there is increased potential for disease transmission that may impact humans, domestic swine, and wildlife. From September 2007 to March 2010, in 13 North Carolina, USA, counties and at Howell Woods Environmental Learning Center, we conducted a serosurvey of feral swine for Brucella suis, pseudorabies virus (PRV), and classical swine fever virus (CSFV); the samples obtained at Howell Woods also were tested for porcine circovirus type 2 (PCV-2). Feral swine serum was collected from trapped and hunter-harvested swine. For the first time since 2004 when screening began, we detected B. suis antibodies in 9% (9/98) of feral swine at Howell Woods and <1% (1/415) in the North Carolina counties. Also, at Howell Woods, we detected PCV-2 antibodies in 59% (53/90) of feral swine. We did not detect antibodies to PRV (n=512) or CSFV (n=307) at Howell Woods or the 13 North Carolina counties, respectively. The detection of feral swine with antibodies to B. suis for the first time in North Carolina warrants increased surveillance of the feral swine population to evaluate speed of disease spread and to establish the potential risk to commercial swine and humans.     

A toxin fromPasteurella multocida serogroup D enhances swine herpesvirus 1 replication/lethality in vitro and in vivo

In swine, the nasal turbinate epithelium is both a site of swine herpesvirus 1 (pseudorabies virus, PRV) replication and a tissue affected by toxin fromPasteurella multocida serogroup D. We examined the effects of exposure to PRV and exposure to toxin in mice, swine, and nasal turbinate cell cultures. Increased mortality in mice was observed when nonlethal doses of PRV (1000 or 100 plaque-forming units, PFU) were administered along with nonlethal doses (60–200 ng/kg) of toxin. In swine, clinical disease and death in adult pigs was observed after an intradermal injection of toxin (20 ng/kg) and intranasal exposure to 1000 PFU/kg of PRV. Nasal turbinate cell cultures incubated with toxin and PRV had increased protein synthesis, DNA synthesis, and increased recovery of virus particles. These findings indicate that a toxin fromP. multocida serogroup D enhances swine herpesvirus 1 replication and lethality in cell cultures and animal models.     

Aerosol transmission of a viable virus affecting swine: Explanation of an epizootic of pseudorabies

A Gaussian diffusion model was applied to an epizootic of pseudorabies in ten swine herds located in Decatur County, Indiana, USA to test the hypothesis that the virus can be spread via aerosol. The epizootic occurred during January to March, 1988, spreading through ten farms across an area of about 150 km². The model included a receptor component that provided an estimate of viruses received by the pig within an enclosed barn. Results show that the diffusion model can explain the spread of the virus during the epizootic for all nine farms to which the virus spread.     

Trichinella spiralis in an agricultural ecosystem. II. Evidence for natural transmission of Trichinella spiralis spiralis from domestic swine to wildlife

Epidemiological investigations of an outbreak of trichinellosis were carried out in a domestic swine herd and it was established that the parasite also occurred in rats, and in skunks, opossums, and raccoons. Because considerable uncertainty exists regarding the role of sylvatic trichinellosis as a reservoir for the synanthropic cycle, studies were conducted to determine the genetic nature of the various isolates from this ecosystem. Pig infectivity trials, isoenzyme analyses, and repetitive DNA sequence analyses were performed. The results showed that all isolates from the farm environs were genetically similar and that they are related to Trichinella spiralis isolated from domestic pigs. The implication of these findings, in contrast to studies on isolates from wildlife elsewhere, is that this parasite is transmitted from domestic swine to sylvatic hosts and that any control or eradication efforts must take into account the potential for reinfection of hogs from wild animals.     

Prevalence and transmission of pseudorabies virus in an isolated population of feral swine

Six hundred sixty-one feral swine (Sus scrofa) from Ossabaw Island, Georgia (USA) were captured, bled, and their sera tested for pseudorabies virus (PRV) antibody during a 6 yr period. Prevalence of seroconversion in females was somewhat higher than in males (10% versus 7%), but the difference was not statistically significant. Adults had a significantly higher prevalence than juveniles (29% versus 1%). An important finding in this study was that seroconversion occurred primarily in the adult feral swine.     

中国西藏部分地区猪戊型肝炎病毒流行病学调查

戊型肝炎病毒(Hepatitis E Virus,HEV)感染是一个重要的全球公共卫生问题,而猪被认为是HEV的天然宿主。HEV可以跨种间传播,且已经证实生吃感染的猪肉会导致人感染。在中国西藏许多地区仍然有生吃猪肉、猪肝等的习惯,且不同种家畜混合饲养,极易造成HEV感染和传播。然而中国西藏地区猪HEV流行情况报道甚少。文中对中国西藏5个地区市(拉萨、日喀则、山南、那曲和昌都)猪血清进行HEV Immunoglobulin-M(Ig M)和Ig G抗体检测,并通过逆转录巢氏PCR(RT-n PCR)进行HEV RNA检测和定量RT-PCR(q RT-PCR)进行病毒拷贝计算,首次报道了藏猪血清HEV RNA阳性率。结果显示,在西藏猪中HEV有较高的流行趋势。猪血清HEV Ig M抗体阳性率高达7.6%(26/340),HEV Ig G抗体阳性率为1.8%(6/340),HEV RNA阳性率高达7.6%(26/340),血清中病毒拷贝高达1.7×107 copies/m L,而且5个地区有不同的流行趋势。结果表明西藏猪HEV感染情况严重。有关部门应加强管理,以避免人与动物之间的交叉感染和暴发。     

Demonstration of rabies virus-specific antibody in the sera of free-ranging Iowa raccoons (Procyon lotor).

Between 1984 and 1988, a study was conducted to evaluate the frequency of rabies virus neutralizing antibodies in raccoons (Procyon lotor) in two counties in Iowa. Nine hundred eighty five raccoons were trapped and tagged in Guthrie and Cerro Gordo counties during the spring, summer and fall of each year. Sex, age and weight were recorded for each animal and a blood sample was collected. Serum samples were tested for the presence of serum neutralizing antibodies (SNA) by the rapid fluorescent focus inhibition test (RFFIT), mouse serum neutralization test (MSN), and an indirect fluorescent antibody (IFA) technique for detecting immunoglobulin G. Fifty-one raccoons (5%) were found to have SNA by the RFFIT. Thirty-six serum samples (24 with RFFIT antibody titer greater than 3.0, and 12 less than 3.0) were also tested by the MSN, with results correlating well with the RFFIT results (r = 0.86, P less than 0.01, Kappa = 0.93). In 35 raccoons with SNA by the RFFIT, six individuals had immunoglobulin G binding activity by the IFA test. These results provided serologic evidence of exposure of raccoons to rabies virus in an area free of enzootic raccoon rabies.     

Hosts of Lutzomyia shannoni (Diptera: Psychodidae) in relation to vesicular stomatitis virus on Ossabaw Island, Georgia, U.S.A.

Abstract. Hosts of Lutzomyia shannoni Dyar, a suspected biological vector of the New Jersey serotype of vesicular stomatitis (VSNJ) virus, were determined using an indirect enzyme-linked immunosorbent assay (ELISA) of 333 blood-fed female sandflies collected from their diurnal resting shelters on Ossabaw Island, Georgia, U.S.A. Sandflies had fed primarily on white-tailed deer ( Odocoileus virginianus ) (81%) and to a lesser extent on feral swine ( Sus scrofa ) (16%), two species of host infected annually with VSNJ. Other hosts were raccoons ( Procyon lotor ) and horses ( Equus caballus ) or donkeys ( E. asinus ), with only two (<1%) mixed bloodmeals from deer/raccoon and deer/swine. A larger proportion of feedings on feral swine was detected in maritime live oak forests than in mixed hardwood forests. These findings are consistent with the hypothesis that L. shannoni is a primary vector of VSNJ virus on Ossabaw Island.