Colostral transmission of maedi visna virus: sites of viral entry in lambs born from experimentally infected ewes

Maedi visna virus (MVV) vertical transmission in sheep via infected colostrums is a very important route of infection in lambs. To verify colostral transmission and to study early viral entry in lambs, colostrum samples, and small intestine and mesenteric lymph nodes of lambs born from experimentally infected ewes were examined by histopathology, immunohistochemistry (IHC) and in situ hybridisation (ISH) studies. In particular, newborn lambs were naturally fed maternal colostrum and humanely killed at 10, 24, 48, 72, 96 h and 7 and 10 days after birth; two caesarian-derived lambs served as uninfected controls. No lesions suggestive of MVV infection were found, but marked immunoreactions for MVV capsid antigen (CA, p28) were detected in lambs fed maternal colostrum and in macrophages cultured from colostrum. IHC results in lambs suggest an initial viral absorption by intestinal epithelial cells at the tip of the villi, passage to mononuclear cells in the lamina propria and involvement of ileum Peyers' patches and mesenteric lymph nodes, with different staining patterns depending on infection times. ISH on intestinal sections of the 72 h lamb revealed the presence of proviral DNA in epithelial cells at the tip of the villi, suggesting a role for these cells in early MVV replication. The results contribute to knowledge about the pathogenesis of ovine lentivirus infection suggesting that the small intestine and mesenteric nodes are the sites of entry and propagation of MVV in lambs fed colostrums from infected ewes.     

Four-layer enzyme immunoassay (EIA) detection of differences in IgG, IgM and IgA antibody response to Aujeszky's disease virus in infected and vaccinated pigs

The use of the four-layer enzyme immunoassay (EIA) for the detection of IgG, IgM and IgA antibodies against Aujeszky's disease virus in blood and oropharyngeal swabs of infected and vaccinated pigs is described. Mean antibody titres obtained using the four-layer EIA were 6.1 and 3829 times higher compared with the indirect enzyme-linked immunosorbent assay (ELISA) and virus neutralization (VN) test, respectively. The VN test detected mainly IgG antibodies, while the IgM antibodies did not react. Using the EIA, the first antiviral antibodies in sera were demonstrated on Days 5-7 after infection or vaccination. Up to the 7th day, demonstrable antibodies were almost exclusively of the IgM class. In infected pigs high titres of IgM antibodies were still detected on Day 18, while in vaccinated animals they were absent by this time. Antibodies of the IgG class appeared in infected pigs sooner (Day 7) than in vaccinated pigs (Day 10) and reached higher mean titres. Antibodies of the IgA class were demonstrable from Day 10 only in samples from infected pigs. Similar antibody dynamics and distribution were detected in oropharyngeal swabs, except that the IgG and IgM titres were roughly 100 times lower than in sera. However, titres of IgA antibodies in oropharyngeal swabs were two times higher than in sera. The greatest differences between both groups of animals were recorded on Day 18; in the infected pigs, IgG, IgM and IgA antibodies were present in sera and oropharyngeal swabs at that time, while in vaccinated pigs only IgG antibodies were demonstrable. The effect of infection and vaccination on the pattern of the immune response as well as the importance of the detection of individual immunoglobulin classes for the specificity of the enzyme immunoassay are discussed.     

Cross-protection studies between feline infectious peritonitis and porcine transmissible gastroenteritis viruses

Cross-protection studies between the feline infectious peritonitis (FIP) and the porcine transmissible gastroenteritis (TGE) viruses were conducted in cats, pigs and pregnant gilts. Cats vaccinated with TGE virus developed neutralizing antibodies against TGE virus and low titer antibody against FIP virus detected by an indirect fluorescent antibody technique but were not protected against a virulent FIP virus challenge. Baby pigs and pregnant gilts vaccinated with FIP virus did not develop detectable antibodies to TGE virus. Nevertheless, it appeared that vaccination of swine with FIP virus conferred some immunity against TGE virus infection. Seventeen-day-old pigs vaccinated with two doses of FIP virus had a 67% survival rate following a virulent TGE virus challenge, and 75% of the 3-day-old pigs suckling either FIP or TGE-virus-vaccinated gilts survived virulent TGE virus infection in contrast to 0% survival of baby pigs suckling unvaccinated gilts.     

Age-dependent resistance to transmissible gastroenteritis of swine. III. Effects of epithelial cell kinetics on coronavirus production and on atrophy of intestinal villi.

Coronavirus titers in small intestine, degree of villous atrophy and apparent rates of regeneration of intestinal villi were compared in newborn, 3-week-old and adult pigs for 1 week after they were exposed to the transmissible gastroenteritis virus of swine. The response within the newborn group was homogeneous, resulting in high virus titers, maximal villous atrophy and comparatively slow regeneration. In general, virus titers were lower, villous atrophy was less severe and regeneration more rapid in both older groups than in the newborn pigs. However, the response varied greatly in the older groups. The 3-week-old group was divided into two populations. The major population had low virus titers and developed partial villous atrophy, whereas the minor population had marked villous atrophy and virus titers comparable to those of the newborn pigs. These observations support the hyposthesis that the accelerated replacement of villous epithelium in the small intestine of pigs during the first 3 weeks contributes to the innate age-dependent resistance to transmissible gastroenteritis. The accelerated replacement of villous epithelial cells in older pigs contributes to resistance in two ways. The increased proliferative capacity of crypt epithelium results in a more rapid regeneration of atrophic villi; and the comparatively young villous absorptive cells resulting from accelerated replacement produce less virus per cell than the older ones of the newborn pig.     

Immunological parameters in the intestinal lymph of pigs including changes during experimentally induced diarrhoea

Experiments were undertaken to provide basic immunological data on the intestinal lymph of young pigs. For this purpose indwelling cannulae were established in the main efferent intestinal lymphatic ducts of 12 animals and measurements were made of lymph flow rate, and concentrations of IgG, IgA and IgM. Measurements were also made of cell numbers, differential counts and immunoglobulin specificity of lymphoid cells in lymph. Similar measurements were also made on six pigs in which experimental diarrhoea was induced. The mean number of leucocytes in intestinal lymph was extremely low (0.66 X 10(5)/ml). However a high proportion of lymphocytes contained cytoplasmic IgA (19.65 per cent) and IgM (12.53 per cent), with few containing IgG (1.35 per cent). The concentrations of IgM and IgA in intestinal lymph were 0.51 mg/ml and 1.64 mg/ml respectively, values which suggest that the intestine is an important organ for synthesis of these two classes of immunoglobulin in young pigs. Following induction of diarrhoea and consequent dehydration of the intestine, the lymph: serum concentration ratios for immunoglobulins increased but subsequently declined when the water balance in the intestine returned to normal.     

Analysis of the quality of protection induced by a porcine influenza A vaccine to challenge with an H3N2 virus

Antigenic drift of swine influenza A (H3N2) viruses away from the human A/Port Chalmers/1/73 (H3N2) strain, used in current commercial swine influenza vaccines, has been demonstrated in The Netherlands and Belgium. Therefore, replacement of this human strain by a more recent swine H3N2 isolate has to be considered. In this study, the efficacy of a current commercial swine influenza vaccine to protect pigs against a recent Dutch field strain (A/Sw/Oedenrode/96) was assessed. To evaluate the level of protection induced by the vaccine it was compared with the optimal protection induced by a previous homologous infection. Development of fever, virus excretion, and viral transmission to unchallenged group mates were determined to evaluate protection. The vaccine appeared efficacious in the experiment because it was able to prevent fever and virus transmission to the unchallenged group mates. Nevertheless, the protection conferred by the vaccine was sub-optimal because vaccinated pigs excreted influenza virus for a short period of time after challenge, whereas naturally immune pigs appeared completely protected. The immune response was monitored, to investigate why the vaccine conferred a sub-optimal protection. The haemagglutination inhibiting and virus neutralising antibody responses in sera, the nucleoprotein-specific IgM, IgG, and IgA antibody responses in sera and nasal secretions and the influenza-specific lymphoproliferation responses in the blood were studied. Vaccinated pigs developed the same or higher serum haemagglutination inhibiting, virus neutralising, and nucleoprotein-specific IgG antibody titres as infected pigs but lower nasal IgA titres and lymphoproliferation responses. The lower mucosal and cell-mediated immune responses may explain why protection after vaccination was sub-optimal.     

Recovery of transmissible gastroenteritis virus from chronically infected experimental pigs.

Transmissible gastroenteritis (TGE) virus was reisolated from pulmonary and intestinal tissues from 6 of 9 chronically infected experimental pigs (principals) necropsied 30 to 104 days after inoculation. Tissue homogenates (lung and small intestine) from the principals were prepared and inoculated into 3- to 5-day-old gnotobiotic pigs. The virus reisolated from the tissue homogenates produced a milder disease on 1st passage and a more severe disease on 2nd passage. The chronically infected experimental pigs (principals) developed serum-neutralization titers to TGE of 1:30 to 1:525. There appeared to be no relationship between serum titers and reisolation of TGE virus from the 9 principals. The persistence of virus in lung or intestine to 104 days indicates the recovered (or carrier) pig may be considered the primary source of TGE virus infection.     

Pathogenicity of an attenuated strain of transmissible gastroenteritis virus for newborn pigs.

The pathogenicity of a cell culture-attenuated strain of transmissible gastroenteritis virus for newborn pigs was investigated. Newborn (1- to 2-day-old) pigs were orally given 2 x 10(6) plaque-forming units of attenuated virus. All pigs developed mild diarrhea, but deaths did not occur. As determined by immunofluorescence and villous atropy, infection of the small intestine was limited to the caudal 50 to 66%. Fluorescing cells and atrophic villi were seen from 2 to 3 days until 6 to 7 days after exposure. Attenuated virus-exposed pigs produced circulating virus-neutralizing antibodies detectable as early as 5 days after exposure. By contrast, all pigs orally given 1 x 10(2) pig infective doses of virulent transmissible gastroenteritis virus developed severe diarrhea, and almost all of those not killed died within 2 to 5 days after exposure. In the latter pigs, the entire length of the small intestine, except for the first 4 to 5 cm, was infected with virus by 24 to 36 hours after exposure.     

Overcoming maternal antibody interference by vaccination with human adenovirus 5 recombinant viruses expressing the hemagglutinin and the nucleoprotein of swine influenza virus

Sows and gilts lack immunity to human adenovirus 5 (Ad-5) vectored vaccines so immunogens of swine pathogens can be expressed with these vaccines in order to immunize suckling piglets that have interfering, maternally derived antibodies. In this study 7-day-old piglets, that had suckled H3N2 infected gilts, were sham-inoculated with a non-expressing Ad-5 vector or given a primary vaccination with replication-defective Ad-5 viruses expressed the H3 hemagglutinin and the nucleoprotein of swine influenza virus (SIV) subtype H3N2. The hemagglutination inhibition (HI) titer of the sham-inoculated group (n = 12) showed continued antibody decay whereas piglets vaccinated with Ad-5 SIV (n = 23) developed an active immune response by the second week post-vaccination. At 4 weeks-of-age when the HI titer of the sham-inoculated group had decayed to 45, the sham-inoculated group and half of the Ad-5 SIV vaccinated pigs were boosted with a commercial inactivated SIV vaccine. The boosted pigs that had been primed in the presence of maternal interfering antibodies had a strong anamnestic response while sham-inoculated pigs did not respond to the commercial vaccine. Two weeks after the booster vaccination the pigs were challenged with a non-homologous H3N2 virulent SIV. The efficacy of the vaccination protocol was demonstrated by abrogation of clinical signs, by clearance of challenge virus from pulmonary lavage fluids, by markedly reduced virus shedding in nasal secretions, and by the absence of moderate or severe SIV-induced lung lesions. These recombinant Ad-5 SIV vaccines are useful for priming the immune system to override the effects of maternally derived antibodies which interfere with conventional SIV vaccines.     

Local humoral and cellular responses in Aujeszky's disease virus infection in pigs

Mucosal and tracheal washings from pigs vaccinated parenterally and intranasally with Aujeszky's disease virus were tested for specific anti-Aujeszky's disease virus responses. Antibody tests included complement dependent antibody lysis, antibody dependent cellular cytotoxicity, virus neutralisation, and anti-Aujeszky's disease virus IgA and IgG levels. There was no correlation between the levels of these antibodies and protection from subsequent challenge. Direct lymphocyte cytotoxicity against cells infected with Aujeszky's disease virus was found in lymph nodes draining the tonsillar area.     

Effect of recombinant DNA-derived bovine alpha-1 interferon on transmissible gastroenteritis virus infection in swine

Recombinant DNA-derived bovine interferon alpha 1-1 (BoIFN) inhibited replication of both vesicular stomatitis virus and transmissible gastroenteritis virus in cultures of swine testicular cells. Newborn pigs were orally inoculated with BoIFN and subsequently had interferon in their gastric contents and serum; however, interferon was found only occasionally in intestinal washings. Incubation of BoIFN with gastric contents from a newborn suckling pig did not affect antiviral activity, whereas intestinal (small intestine) contents from the same animal inactivated BoIFN within 1 minute. Beginning at 6 hours of age, newborn, colostrum-deprived pigs were given 1 mg of BoIFN orally every 12 hours. These pigs were not protected against challenge exposure to virulent transmissible gastroenteritis virus at 48 hours of age; disease and mortality were similar for these pigs and for control pigs not given BoIFN prior to challenge exposure. The BoIFN did not impair growth rate of pigs and did not cause obvious disease or lesions.     

Antibody response of pigs to inactivated monovalent and bivalent vaccines for porcine parvovirus and pseudorabies virus

Groups of pigs vaccinated with an inactivated bivalent vaccine containing porcine parvovirus (PPV) and pseudorabies virus (PRV) developed geometric mean titers (GMT) of humoral antibody for each of the viruses as high or slightly higher than those of other groups of pigs that were vaccinated with inactivated monovalent vaccines containing one or the other of the same viruses. An increase in GMT after challenge exposure of vaccinated pigs to live virus indicated that vaccination did not prevent virus replication. However, an indication that replication was less extensive in vaccinated pigs was provided by the following. Although neither vaccinated nor nonvaccinated (control) pigs had clinical signs after exposure to the live PPV, the effect of vaccination was evident by the fact that GMT were higher in nonvaccinated pigs after exposure than they were in vaccinated pigs. Conversely, all pigs exposed to live PRV had clinical signs, but these signs varied between mild-to-moderate and transient for vaccinated pigs to severe and fatal for nonvaccinated pigs.     

Protection against progressive atrophic rhinitis by vaccination with Pasteurella multocida toxin purified by monoclonal antibodies

Pasteurella multocida toxin was purified by affinity chromatography and inactivated by treatment with formaldehyde before use as a single component vaccine against progressive atrophic rhinitis in pigs. Twenty pregnant gilts which were vaccinated twice before farrowing with either low or high doses of the purified toxoid, developed dose-dependent positive serum and colostrum titres to the toxin and, unlike the progeny of 10 untreated control gilts, the offspring of the vaccinated gilts also had serum titres. These titres could be measured in blood samples taken for more than eight weeks from birth for most pigs born to gilts vaccinated with low doses and more than 12 weeks for pigs born to gilts vaccinated with high doses of the vaccine. All the piglets were inoculated intranasally with Bordetella bronchiseptica and toxigenic P multocida. The clinical and post mortem examinations of snouts revealed a significant reduction in the frequency and degree of conchal atrophy in the two groups of pigs from the vaccinated gilts compared with the pigs from control gilts. Clinically 90 per cent of the snouts of pigs born to vaccinated gilts appeared normal whereas only 28 per cent of the snouts of control pigs were not shortened or deviated at eight weeks of age. At slaughter 11 per cent of the pigs born to vaccinated gilts and 81 per cent of the control pigs had severe turbinate atrophy.(ABSTRACT TRUNCATED AT 250 WORDS)     

Mucosal vaccination against influenza: protection of pigs immunized with inactivated virus and ether-split vaccine

Effective vaccinations against swine influenza reduce the economic loss of pig industries, and also may minimize the possibility of emergence of new pandemic viruses, since pigs are intermediate hosts to generate reassortant viruses among avian and mammalian influenza viruses. In this study, we showed that intranasal immunization of pigs with formalin-inactivated or ether-split influenza vaccine (A/Aichi/2/68) induced virus-specific IgG, IgM, and IgA antibodies in their nasal secretions and sera, resulting in complete protection from virus challenge. Antibody response to the challenge virus was not observed in the immunized pigs, suggesting that the replication of the virus in the primary targets, respiratory epithelial cells, was inhibited. The present results indicate that intranasal immunization of pigs with inactivated vaccines is effective to control swine influenza, and also provide a good model, as well as a mouse model, to evaluate an intranasal application of influenza vaccine for humans.     

Colostral IgA, IgG, and IgM-IgA fractions as fluorescent antibody for the detection of the coronavirus of transmissible gastroenteritis.

Colostrum from sows and gilts inoculated with virulent transmissible gastroenteritis virus was fractionated into the 3 major immunoglobulin classes, IgA, IgG, and IgM-IgA fractions, by chromatographic and gel-filtration procedures. Each fraction was assayed for purity with rabbit anti-porcine serum and rabbit monospecific anti-porcine IgG, anti-porcine IgA, and anti-porcine IgM. These analyses showed that the IgG and IgA fractions were pure. The IgM fraction contained some IgA in the polymeric form and was designated the IgM-IgA fraction. Each Ig was assayed for virus-neutralizing activity on swine testes cells by the plaque-reduction method before and after conjugation with fluorescein isothiocyanate. On the basis of activity per milligram of protein, the virus-neutralizing titers were 1:641, 1:44, and 1:6.8 for the IgA, IgG, and IgM-IgA fractions respectively; the fluorescent antibody titers were 1:31.3, 1:0.1, and 1:15.6, respectively, for the same Ig.     

Randomised, placebo-controlled trial of a live vaccine against porcine reproductive and respiratory syndrome virus in sows on infected farms

A randomised, double-blinded, placebo-controlled study into the effectiveness and safety of a vaccine against porcine reproductive and respiratory syndrome virus (PRRSV) was carried out on three farms with a history compatible with chronic PRRSV infection; representative groups of sows and gilts were injected with a live vaccine against PRRSV, and during the next six weeks no side effects were observed. The remaining sows and gilts on the three farms were then vaccinated with the same vaccine. Again, no side effects were observed. There were significant reductions in abortion, reproductive disease, returns to oestrus and the numbers of stillborn pigs per sow, and significant increases in the numbers of liveborn pigs per sow and weaned pigs per sow among the vaccinated animals.     

Protection against enteric colibacillosis in pigs suckling orally vaccinated dams: evidence for pili as protective antigens

Pregnant gilts were vaccinated orally with Escherichia coli that produced pilus antigens K99 or 987P. The vaccines were live or dead enterotoxigenic E coli (ETEC) or a liver rough non-ETEC strain which has little ability to colonize pig intestine. Pigs born to the gilts were challenge exposed orally with K99+ or 987P+ ETEC, which did not produce heat-labile enterotoxin or flagella and which produced somatic and capsular antigens different from those of the vaccine strains. Control gilts had low titers of serum and colostral antibodies against pilus antigens, and their suckling pigs frequently had fatal diarrhea after challenge exposure. Serum antibody titers against pilus antigens of the vaccine strains increased in the gilts after vaccination with liver ETEC, and the colostral antibody titers of these gilts were higher than those of controls. Pigs suckling such vaccinated gilts were more resistant than controls to challenge strains were of different pilus types, and it could not be attributed to enterotoxin neutralization by colostrum. In contrast to the live ETEC vaccines given to the pregnant gilts, the liver rough non-ETEC and dead ETEC vaccines stimulated little or no production of antibody against pilu, and the pigs born of these vaccinated gilts remained highly susceptible to challenge exposure. The results support the hypothesis that pilu can be protective antigens in oral ETEC vaccines. It was indicated that in the system reported, protection depended on living bacteria for the production of pilus antigens in vivo or for the transport of pilus antigens across intestinal epithelium.     

Immunoglobulins in porcine umbilical cord blood and maternal placenta.

Studies were made of the immunoglobulin (Ig) in serums from umbilical cord of newborn pigs and maternal placenta. The neutralization test for porcine parvovirus and Japanese encephalitis virus was carried out with the serum of the sow and that of the umbilical cord of the newborn pig. Comparative studies of the serums from the dam and the umbilical cord were also done with gel filtration. Of 20 umbilical cord serum samples, IgG was seen in 5 samples (25%), IgA in 1 sample (5%), and IgM in 9 samples (45%). The amount of any 1 of the 3 classes of Ig in the serums was between 13.5 and 28.0 mg/dl. Among the samples examined, 1 had both IgG and IgA and 1 had IgG and IgM, but none had both IgA and IgM and none had 3 classes of immunoglobulins (i.e., IgG, IgA, and IgM). Only 7 samples (35%) did not have any class of Ig. The IgG disappeared from the blood of hysterectomy-produced colostrum-deprived pigs at 3 days of age, and IgM disappeared when pigs were 7 days of age. Neutralization antibodies of porcine parvovirus and Japanese encephalitis virus in maternal serum were not transferred to the fetus through the placenta. Results of immunohistologic surveys indicated that the sow's Ig were not transferred to the fetus through the placenta. Therefore, it is believed that the Ig in the porcine fetus might be synthesized in certain cells in the placental tissue, and the degree of production of the Ig in the placental tissues may differ in each case. The component, which seems to be Ig, was observed as the obscure band of the beta- to gamma-globulin area in serum of the umbilical cord. Comparison was made, with gel filtration, of maternal serum and serum from the umbilical cord of the newborn pig originating from the same sow. Seemingly, the IgG in the umbilical cord serum is mainly in the lower molecular weight fraction, whereas IgG in the sow's serum was distributed in the high to low molecular weight fractions.     

寡果糖对人源菌群仔猪肠道中IgA和IgG分泌细胞的影响

本文主要研究寡果糖对人源菌群仔猪肠道中IgA和IgG分泌细胞的影响。通过无菌剖腹产获取15头无特定病原菌(specific pathogen free,SPF)仔猪,随机分为三组。第一组为SPF组,经口灌服磷酸钠缓冲液(内含10%甘油)以示对照;第二组为人源菌群(human flora-associated,HFA)组,经口服途径接种人源菌群;第三组为寡果糖(fructo-oligosaccharides,FOS)组,口服途径接种人源菌群且灌喂寡果糖。仔猪饲养于屏障系统内,无菌条件下人工哺育45天。应用免疫组织化学方法进行研究。结果表明:(1)所有仔猪小肠和结肠的固有层中均分布有IgA和IgG分泌细胞。(2)IgA和IgG分泌细胞在十二指肠中分布最多,随着肠段的向后推移IgA和IgG分泌细胞数量有逐渐下降趋势。(3)HFA组和FOS组IgA分泌细胞数量在回肠显著高于SPF组(P<0.01);十二指肠中HFA组IgG分泌细胞数量显著高于SPF组(P<0.01)。(4)FOS组IgA分泌细胞数量在空肠显著高于HFA组外(P<0.05),其他肠段总体上低于HFA组,但差异不显著。本结果提示给新生仔猪接种人源菌群能促进仔猪肠道中IgA和IgG分泌细胞的发育,而寡果糖使肠道IgA和IgG分泌细胞数量呈现下降的趋势。     

Enzymatic and acidic sensitivity profiles of selected virulent and attenuated transmissible gastroenteritis viruses of swine

This study identifies the in vitro differences (markers) between virulent and attenuated transmissible gastroenteritis (TGE) viruses. Exposure of virulent Miller strain and attenuated Purdue strain TGE viruses to a spectrum of acidities indicated that the Miller strain was more stable at pH 2. Acidities at or above pH 3 did not reduce viral infectivity of either strain. When virulent and attenuated viruses were exposed to gastric fluids of either fed or fasted swine, there was a similar degree of sensitivity. Carboxypeptidase B, alpha-amylase, and alkaline phosphatase present in porcine small intestinal fluids did not cause a significant difference in sensitivity between virulent and attenuated virus isolates. The digestive enzymes: trypsin, alpha-chymotrypsin, pancreatin, peptidase, and carboxypeptidase A did not (or only slightly) inactivate virulent Miller strain TGE virus, but greatly reduced infectivity of attenuated viruses (Purdue strain and TGE vaccine virus isolates). The attenuated strains were significantly more sensitive to small intestinal fluids from both fasted and fed adult swine. Differential sensitivities between virulent and attenuated TGE viruses to digestive fluids from stomach and small intestine further substantiate the notion of differential susceptibility to small intestinal proteases as a correlate of viral virulence.