Influenza A virus infections in swine: pathogenesis and diagnosis.

Influenza has been recognized as a respiratory disease in swine since its first appearance concurrent with the 1918 "Spanish flu" human pandemic. All influenza viruses of significance in swine are type A, subtype H1N1, H1N2, or H3N2 viruses. Influenza viruses infect epithelial cells lining the surface of the respiratory tract, inducing prominent necrotizing bronchitis and bronchiolitis and variable interstitial pneumonia. Cell death is due to direct virus infection and to insult directed by leukocytes and cytokines of the innate immune system. The most virulent viruses consistently express the following characteristics of infection: (1) higher or more prolonged virus replication, (2) excessive cytokine induction, and (3) replication in the lower respiratory tract. Nearly all the viral proteins contribute to virulence. Pigs are susceptible to infection with both human and avian viruses, which often results in gene reassortment between these viruses and endemic swine viruses. The receptors on the epithelial cells lining the respiratory tract are major determinants of infection by influenza viruses from other hosts. The polymerases, especially PB2, also influence cross-species infection. Methods of diagnosis and characterization of influenza viruses that infect swine have improved over the years, driven both by the availability of new technologies and by the necessity of keeping up with changes in the virus. Testing of oral fluids from pigs for virus and antibody is a recent development that allows efficient sampling of large numbers of animals.

Clinicopathological features of Swine influenza.

In this chapter, the clinical presentations, the development of infection and the macroscopic and microscopic lesions of swine influenza virus (SIV) infection are described. Both natural and experimental infections are discussed.

Kinetics of lung lesion development and pro-inflammatory cytokine response in pigs with vaccine-associated enhanced respiratory disease induced by challenge with pandemic (2009) A/H1N1 influenza virus.

The objective of this report was to characterize the enhanced clinical disease and lung lesions observed in pigs vaccinated with inactivated H1N2 swine δ-cluster influenza A virus and challenged with pandemic 2009 A/H1N1 human influenza virus. Eighty-four, 6-week-old, cross-bred pigs were randomly allocated into 3 groups of 28 pigs to represent vaccinated/challenged (V/C), non-vaccinated/challenged (NV/C), and non-vaccinated/non-challenged (NV/NC) control groups. Pigs were intratracheally inoculated with pH1N1 and euthanized at 1, 2, 5, and 21 days post inoculation (dpi). Macroscopically, V/C pigs demonstrated greater percentages of pneumonia compared to NV/C pigs. Histologically, V/C pigs demonstrated severe bronchointerstitial pneumonia with necrotizing bronchiolitis accompanied by interlobular and alveolar edema and hemorrhage at 1 and 2 dpi. The magnitude of peribronchiolar lymphocytic cuffing was greater in V/C pigs by 5 dpi. Microscopic lung lesion scores were significantly higher in the V/C pigs at 2 and 5 dpi compared to NV/C and NV/NC pigs. Elevated TNF-α, IL-1ß, IL-6, and IL-8 were detected in bronchoalveolar lavage fluid at all time points in V/C pigs compared to NV/C pigs. These data suggest H1 inactivated vaccines followed by heterologous challenge resulted in potentiated clinical signs and enhanced pulmonary lesions and correlated with an elevated proinflammatory cytokine response in the lung. The lung alterations and host immune response are consistent with the vaccine-associated enhanced respiratory disease (VAERD) clinical outcome observed reproducibly in this swine model.

The role of swine in the generation of novel influenza viruses.

The ecology of influenza A viruses is very complicated involving multiple host species and viral genes. Avian species have variable susceptibility to influenza A viruses with wild aquatic birds being the reservoir for this group of pathogens. Occasionally, influenza A viruses are transmitted to mammals from avian species, which can lead to the development of human pandemic strains by direct or indirect transmission to man. Because swine are also susceptible to infection with avian and human influenza viruses, genetic reassortment between these viruses and/or swine influenza viruses can occur. The potential to generate novel influenza viruses has resulted in swine being labelled 'mixing vessels'. The mixing vessel theory is one mechanism by which unique viruses can be transmitted from an avian reservoir to man. Although swine can generate novel influenza viruses capable of infecting man, at present, it is difficult to predict which viruses, if any, will cause a human pandemic. Clearly, the ecology of influenza A viruses is dynamic and can impact human health, companion animals, as well as the health of livestock and poultry for production of valuable protein commodities. For these reasons, influenza is, and will continue to be, a serious threat to the wellbeing of mankind.

Cardiovascular lesions in pigs naturally or experimentally infected with porcine circovirus type 2.

Abundant intracytoplasmic porcine circovirus type 2 (PCV2) was associated with myocardiocyte swelling or necrosis, or myocardial fibrosis (or both) in three naturally infected pigs aged 4-7 weeks from three different farms. One 6 week old pig from a fourth farm had severe diffuse segmental to circumferential lymphohistiocytic and plasmacytic periarteritis and endarteritis in several organs, PCV2 antigen was demonstrated in endothelial cells, and inflammatory cells in the arterial walls. In three pigs experimentally infected with PCV2, viral antigen was also associated with obliterated blood vessels in areas of granulomatous and necrotizing lymphadenitis. Together these findings suggest that the cardiovascular system in general and endothelial cells in particular play an important role in the pathogenesis of PCV2-associated diseases.

Porcine circovirus type 2 (PCV-2) coinfections in US field cases of postweaning multisystemic wasting syndrome (PMWS).

The prevalence of different pathogens detected in combination with porcine circovirus type 2 (PCV-2) was studied retrospectively in field cases of postweaning multisystemic wasting syndrome (PMWS) diagnosed at the Iowa State University Veterinary Diagnostic Laboratory, Ames, Iowa, between January 2000, and September 2001. The presence of PCV-2 antigen in lymphoid tissues and/or lung, demonstrated by immunohistochemistry, together with moderate to severe lymphoid depletion and/or granulomatous lymphadenitis, was used as the criteria for the diagnosis of PMWS. A total of 484 cases fulfilled these criteria. Most of the cases (294/369) of PMWS occurred in pigs between the ages of 8 and 18 weeks, with a peak at 10 weeks of age. Porcine reproductive and respiratory syndrome virus was detected in 51.9% of the cases, Mycoplasma hyopneumoniae in 35.5%, bacterial septicemia in 14.0%, bacterial pneumonia in 7.6%, swine influenza virus in 5.4%, and PCV-2 alone in 1.9%. In cases with bacterial septicemia the most frequently isolated pathogen was Streptococcus suis. In cases with bacterial pneumonia, Pasteurella multocida was the most prevalent.

Paramyxovirus infection in pigs with interstitial pneumonia and encephalitis in the United States.

In the last few years, newly recognized paramyxoviruses have been associated with severe disease in several animal species, including swine, as well as in human beings. Recently, a paramyxovirus was isolated from a swine herd in the northcentral United States that experienced an epizootic of respiratory and central nervous system disease. Affected pigs had interstitial pneumonia with necrotizing bronchiolitis and encephalitis characterized by lymphocytic perivasculitis and diffuse gliosis. Germ-free pigs inoculated with this isolate developed mild clinical illness and similar but less severe histopathologic lesions in lungs and brain.

Interaction between Mycoplasma hyopneumoniae and swine influenza virus.

An experimental respiratory model was used to investigate the interaction between Mycoplasma hyopneumoniae and swine influenza virus (SIV) in the induction of pneumonia in susceptible swine. Previous studies demonstrated that M. hyopneumoniae, which produces a chronic bronchopneumonia in swine, potentiates a viral pneumonia induced by the porcine reproductive and respiratory syndrome virus (PRRSV). In this study, pigs were inoculated with M. hyopneumoniae 21 days prior to inoculation with SIV. Clinical disease as characterized by the severity of cough and fever was evaluated daily. Percentages of lung tissue with visual lesions and microscopic lesions were assessed upon necropsy at 3, 7, 14, and 21 days following SIV inoculation. Clinical observations revealed that pigs infected with both SIV and M. hyopneumoniae coughed significantly more than pigs inoculated with a single agent. Macroscopic pneumonia on necropsy at days 3 and 7 was greatest in both SIV-infected groups, with minimal levels of pneumonia in the M. hyopneumoniae-only-infected pigs. At 14 days post-SIV inoculation, pneumonia was significantly more severe in pigs infected with both pathogens. However, by 21 days postinoculation, the level of pneumonia in the dual-infected pigs was similar to that of the M. hyopneumoniae-only-infected group, and the pneumonia in the pigs inoculated with only SIV was nearly resolved. Microscopically, there was no apparent increase in the severity of pneumonia in pigs infected with both agents compared to that of single-agent-challenged pigs. The results of this study found that while pigs infected with both agents exhibited more severe clinical disease, the relationship between the two pathogens lacked the profound potentiation found with dual infection with M. hyopneumoniae and PRRSV. These findings demonstrate that the relationship between mycoplasmas and viruses varies with the individual agent.

A comparison of diagnostic assays for the detection of type A swine influenza virus from nasal swabs and lungs.

Nasal swabs and lung samples from pigs experimentally infected with H1N1 swine influenza virus (SIV) were examined for the presence of SIV by the indirect fluorescent antibody assay, immunohistochemistry, cell culture virus isolation, egg inoculation, and 2 human enzyme immunoassays (membrane enzyme immunoassay, microwell enzyme immunoassay). Egg inoculation was considered to be the gold standard for assay evaluation. The 2 human enzyme immunoassays (EIA) and egg inoculation agreed 100% for the prechallenge nasal swabs. Agreement on SIV identification in nasal swabs with egg inoculation following challenge was considered to be good to excellent for membrane EIA (kappa = 0.85) and microwell EIA (kappa = 0.86). Agreement on SIV identification in lung tissue with egg inoculation following challenge was good to excellent for membrane EIA (kappa = 0.75), fair for microwell EIA, fluorescent antibody, and cell culture virus isolation (kappa = 0.48, 0.64, 0.62, respectively), and poor for immunohistochemistry (kappa = 0.36). No assay was 100% accurate, including the "gold standard," egg inoculation. In light of this information, it is important to consider clinical signs of disease and a thorough herd history in conjunction with diagnostic results to make a diagnosis of SIV infection.

Detection and isolation of coronavirus from feces of three herds of feedlot cattle during outbreaks of winter dysentery-like disease.

Clinical signs of a winter dysentery-like syndrome in 6- to 9-month-old cattle in 3 feedlots included acute onset of diarrhea with high morbidity and low mortality, respiratory tract problems that included dyspnea, coughing, and nasal discharge, and high rectal temperatures. Bovine coronavirus was detected by use of an ELISA and immune electron microscopy in fecal and nasal swab samples and by immunohistochemical analysis of intestinal sections collected from calves during necropsy. Bovine coronavirus should be considered in the differential diagnoses for diseases that cause acute onset of bloody diarrhea in feedlot cattle.

Evaluation of antemortem polymerase chain reaction and serologic methods for detection of Lawsonia intracellularis-exposed pigs.

OBJECTIVE: To evaluate polymerase chain reaction (PCR) for detection of Lawsonia intracellularis DNA in feces and an indirect fluorescent antibody test (IFAT) for detecting serum IgG antibodies in pigs exposed to L intracellularis. ANIMALS: 15 seven-week-old pigs and 42 three-week-old pigs. PROCEDURE: During 3 experiments, 23 pigs were inoculated with a pure culture of L intracellularis, 31 pigs served as noninoculated controls, and 3 pigs were used as sentinels. Fecal shedding of L intracellularis was monitored by use of PCR analysis at 7-day intervals. At euthanasia, the ileum was obtained for PCR and histologic analyses. Serum was obtained at 7-day intervals for use in the IFAT. RESULTS: Polymerase chain reaction analysis detected L intracellularis DNA in the feces of 39% of the inoculated pigs; by postinoculation days 21 to 28, 90% of inoculated pigs developed IgG antibodies detected by IFAT. Neither L intracellularis DNA nor IgG antibodies were detected in any of the noninoculated control pigs at euthanasia. Sera from pigs inoculated with enteric pathogens other than L intracellularis did not contain detectable antibodies that reacted with L intracellularis by use of the IFAT. CONCLUSION: The IFAT for L intracellularis IgG antibody detection appeared to be a more sensitive antemortem test for detecting pigs experimentally infected with L intracellularis than was a PCR method for direct detection of the organism in the feces. CLINICAL RELEVANCE: Not all animals that are infected with L intracellularis shed the organism in feces at detectable amounts.

Evaluation of rotavirus infection and diarrhea in Iowa commercial pigs based on an epidemiologic study of a population represented by diagnostic laboratory cases.

Group A, B, and C rotaviruses were identified in 9% (96/1,048) of pig fecal specimens submitted to the Iowa State University Veterinary Diagnostic Laboratory during 1987 and 1988. Six of the rotaviruses were group B, 5 were group C, and the remaining 89% were group A. Of the rotavirus cases with more than 1 serotype, 5 were multiple group A serotypes, 1 involved a group A and B serotype, and 1 included 2 group C serotypes. A retrospective epidemiologic evaluation of pig diarrhea in herds of origin was done using data obtained from the accession records of the rotavirus and 88 matched nonrotavirus pig diarrhea control cases. Herds from which rotavirus cases were derived experienced lower morbidity, mortality, and case fatality rates than matched control herds. The incidence of diarrhea decreased rapidly among all pigs from birth to 3 weeks of age. The peak incidence for piglet diarrhea occurred in February, and a moderate rise occurred in August-September. Definitive evidence for transmissible gastroenteritis virus was found in 12% of nonrotavirus cases but none of the rotavirus cases in which it was sought. Other pathogenic microorganisms were identified less frequently and inconsistently.

An indirect microimmunofluorescence test for detection of chlamydial antibodies in ovine fetal fluids.

The objective of this study was to evaluate an indirect microimmunofluorescence test (IMIF) for detection of chlamydial antibodies in serum and/or thoracic fluids of aborted ovine fetuses. One hundred eighty-two ovine fetuses, including 64 fetuses from 40 ewes that were experimentally infected with an ovine abortion strain of Chlamydia psittaci at gestation days 90-100, 10 fetuses from 6 normal ewes, and 108 fetuses selected from those received at the Iowa Veterinary Diagnostic Laboratory, were evaluated in this study. Fetuses from experimentally infected ewes were examined 4-60 days after inoculation. The IMIF findings were compared with the results of complement fixation serology for chlamydiae and concentrations of immunoglobulin (IgG). Chlamydiae-specific antibodies were detected by IMIF in 28 of 38 fetuses infected with C. psittaci. Elevated levels of IgG and IMIF titers > or = 1:8 were consistent findings in ovine fetuses infected with chlamydiae for more than 24 days. IgG levels and titers of chlamydial antibodies increased with maturity of the fetus and duration of chlamydial infection. Chlamydial antibodies were not detected with the complement fixation test. Fluids from ovine fetuses aborted as a result of other causes also were examined, and IMIF results were negative. The results of this study indicate that the IMIF is a useful and relatively rapid test for identification of chlamydial antibodies in ovine fetuses.

Clinical signs and lesions in gnotobiotic pigs inoculated with Shiga-like toxin I from Escherichia coli.

Gnotobiotic pigs were used as a model to study the contribution of Shiga-like toxin I to natural disease caused by enterohemorrhagic Escherichia coli in calves and human beings. Eleven 2- to 7-day-old gnotobiotic pigs of either sex, obtained by closed hysterotomy, were injected intramuscularly with graded doses of partially purified Shiga-like toxin I derived from a lysogenized Escherichia coli strain. Four other gnotobiotic pigs were injected with a mock toxin preparation obtained from a nonlysogenized culture of the same E. coli strain. All toxin-injected pigs developed diarrhea, and three displayed signs of neurologic disease. Pigs either died or were euthanatized 2 to 4 days post-inoculation. Necrosis of muscle was grossly evident at the site of injection in all toxin-inoculated pigs. Hemorrhage in the lumen of the small and large intestines and blood in the feces were also evident in two toxin-inoculated pigs. Microscopically, severe necrotizing myositis at the injection site, multifocal encephalomalacia, and mucosal infarcts and hemorrhage in the small and large intestines were seen. In small vessels at lesion sites, endothelial cells were frequently swollen or necrotic. Pigs inoculated with mock toxin did not develop diarrhea or exhibit signs of neurologic disease, and the only apparent lesion was mild microscopic myositis at the injection site in 1/4 pigs. The results of this study indicate that Shiga-like toxin I causes vascular damage and ischemic necrosis in the intestines and brains of gnotobiotic pigs. These lesions are similar to those seen in the intestines of calves and human beings with hemorrhagic colitis and in the brains of human beings with thrombotic thrombocytopenic purpura.

Relative prevalence of typical and atypical strains among rotaviruses from diarrheic pigs in conventional swine herds.

Polyacrylamide gel electrophoresis was conducted on genomic RNA extracted from rotaviruses detected in diarrheic pigs from conventional swine herds. Ninety samples contained sufficient virus for RNA band visualization and genome classification. Genome profiles were characteristic of typical group A rotaviruses in 67.8% of the 90 samples, of group B rotaviruses in 10.0%, and of group C rotaviruses in 11.1%. In 11.1% of the samples, the presence of more than 11 bands suggested concurrent infection with more than 1 strain of rotavirus. In infections among nursing pigs, 76.4% were group A rotaviruses, 7.4% were group B, 7.4% were group C, and 8.8% were coinfections. In infections among weaned pigs, 40.9% were group A, 18.2% were group B, 22.7% were group C, and 18.2% were coinfections. Coelectrophoresis with prototype OSU and Gottfried strains revealed a great diversity in electropherotype among field strains of rotavirus.

Attaching and effacing Escherichia coli infection as a cause of diarrhea in young calves.

A form of enteric Escherichia coli infection was identified in 60 calves from 59 farming operations. The E coli responsible for these infections principally colonized the colon, inducing a distinctive lesion described as attaching and effacing. Hemorrhagic enterocolitis or blood in the feces was observed on 40% of the farms. Of affected calves, 86.6% were dairy calves (average age, 11.8 days). Forty-four calves were infected concurrently with other enteropathogens (cryptosporidia, rotavirus, coronavirus, enterotoxigenic E coli, bovine viral diarrhea virus, coccidia). Verotoxin-producing E coli was recovered from 31 calves; 8 were serotype O111:NM isolates, 3 were serotype O5:NM, and 1 was serotype O26:NM.

Acute selenium toxicosis in a dog.

A toxic dose of selenium administered by IM injection to a 3-year-old Chihuahua resulted in pulmonary edema and death. The compound had been dispensed to the owner inadvertently in combination with a vitamin E preparation. Vitamin and mineral products often are considered safe for use in megadoses by the uninformed public. The potential danger of selenium overdosage should not be underestimated.