Artificial Insemination and Its Role in Transmission of Swine Viruses

1. Introduction Artificial insemination (AI) in swine is not a new technique and reports as early as the 1930s (Lush, 1925) describe collecting semen for AI. However, because of farm structure changes, increasing farm sizes and separation of production stages, interest in intensive pig production is growing and AI has become a critical component in modern pig production. In 2001, nearly 60% of North American swine herds utilized AI (Singleton, 2001), a drastic increase from the estimated 5% in the 1990’s (Flowers & Esbenshade, 1993). This is still relatively low compared to the 90% or greater use of AI in Western Europe (Madsen, 2005; Maes et al., 2008). The extensive use of AI in pig reproduction in the last decade has facilitated the exchange of desirable genetic characteristics at an international level, allowing producers to make greater use of superior genetics at a lower cost than some natural-service systems (Gerrits et al., 2005). However, the growth in use of AI has increased the risk of quick and widespread transmission of venereally transmissible pathogens (Thacker et al., 1984). It has been reported that the porcine male reproductive tract is highly susceptible to viral infections (Phillips et al., 1972; Spradbrow, 1968). This, coupled with the ability of boars to produce tens to thousands of insemination doses per week and the widespread distribution of the processed semen (both nationally and internationally), further increases the risk of wide transmission of viral pathogens by semen.This book chapter is published as Opriessnig T, Giménez-Lirola LG, Halbur PG. (2012). Artificial insemination and its role in transmission of swine viruses. In: Carlos C Perez-Marin (Ed), A Bird’s-Eye View of Veterinary Medicine, pp. 255-280. InTech, Croatia. ISBN 978-953-51-0031-7. DOI: 10.5772/17961. Copyright 2012 InTech. Attribution 3.0 International (CC BY 3.0). Posted with permission

Characterization of a novel porcine parvovirus tentatively designated PPV5

A new porcine parvovirus (PPV), provisionally designated as PPV5, was identified in U.S. pigs. Cloning and sequencing from a circular or head-to-tail concatemeric array revealed that the PPV5 possesses the typical genomic organization of parvoviruses with two major predicted open reading frames (ORF1 and ORF2), and is most closely related to PPV4 with overall genomic identities of 64.1-67.3%. The amino acid identities between PPV5 and PPV4 were 84.6%-85.1% for ORF1 and 54.0%-54.3% for ORF2. Unlike PPV4, but similar to bovine parvovirus 2 (BPV2), PPV5 lacks the additional ORF3 and has a much longer ORF2. Moreover, the amino acid sequences of ORF1 and ORF2 of BPV2 showed higher homologies to PPV5 than to PPV4. The conserved motifs of the Ca(2+) binding loop (YXGXG) and the catalytic center (HDXXY) of phospholipase A2 (PLA2) were identified in VP1 (ORF2) of PPV5, as well as in BPV2, but were not present in PPV4. Phylogenetic analyses revealed that PPV5, PPV4 and BPV2 form a separate clade different from the genera Parvovirus and Bocavirus. Further epidemiologic investigations of PPV4 and PPV5 in U.S. pigs of different ages indicated a slightly higher prevalence for PPV5 (6.6%; 32/483) compared to PPV4 (4.1%; 20/483), with detection of concurrent PPV4 and PPV5 in 15.6% (7/45) of lungs of infected pigs. Evidence for potential vertical transmission or association with reproductive failure was minimal for both PPV4 and PPV5. The high similarity to PPV4 and the lack of ORF3 may suggest PPV5 is an intermediate of PPV4 during the evolution of parvoviruses in pigs

Detection of porcine reproductive and respiratory syndrome virus (PRRSV)-specific IgM-IgA in oral fluid samples reveals PRRSV infection in the presence of maternal antibody

The ontogeny of PRRSV antibody in oral fluids has been described using isotype-specific ELISAs. Mirroring the serum response, IgM appears in oral fluid by 7 days post inoculation (DPI), IgA after 7 DPI, and IgG by 9 to 10 DPI. Commercial PRRSV ELISAs target the detection of IgG because the higher concentration of IgG relative to other isotypes provides the best diagnostic discrimination. Oral fluids are increasingly used for PRRSV surveillance in commercial herds, but in younger pigs, a positive ELISA result may be due either to maternal antibody or to antibody produced by the pigs in response to infection. To address this issue, a combined IgM-IgA PRRSV oral fluid ELISA was developed and evaluated for its capacity to detect pig-derived PRRSV antibody in the presence of maternal antibody. Two longitudinal studies were conducted. In Study 1 (modified-live PRRS vaccinated pigs), testing of individual pig oral fluid samples by isotype-specific ELISAs demonstrated that the combined IgM-IgA PRRSV ELISA provided better discrimination than individual IgM or IgA ELISAs. In Study 2 (field data), testing of pen-based oral fluid samples confirmed the findings in Study 1 and established that the IgM-IgA ELISA was able to detect antibody produced by pigs in response to wild-type PRRSV infection, despite the presence of maternal IgG. Overall, the combined PRRSV IgM-IgA oral fluid ELISA described in this study is a potential tool for PRRSV surveillance, particularly in populations of growing pigs originating from PRRSV-positive or vaccinated breeding herds

Detection of African Swine Fever Virus Antibodies in Serum and Oral Fluid Specimens Using a Recombinant Protein 30 (p30) Dual Matrix Indirect ELISA

In the absence of effective vaccine(s), control of African swine fever caused by African swine fever virus (ASFV) must be based on early, efficient, cost-effective detection and strict control and elimination strategies. For this purpose, we developed an indirect ELISA capable of detecting ASFV antibodies in either serum or oral fluid specimens. The recombinant protein used in the ELISA was selected by comparing the early serum antibody response of ASFV-infected pigs (NHV-p68 isolate) to three major recombinant polypeptides (p30, p54, p72) using a multiplex fluorescent microbead-based immunoassay (FMIA). Non-hazardous (non-infectious) antibody-positive serum for use as plate positive controls and for the calculation of sample-to-positive (S:P) ratios was produced by inoculating pigs with a replicon particle (RP) vaccine expressing the ASFV p30 gene. The optimized ELISA detected anti-p30 antibodies in serum and/or oral fluid samples from pigs inoculated with ASFV under experimental conditions beginning 8 to 12 days post inoculation. Tests on serum (n = 200) and oral fluid (n = 200) field samples from an ASFV-free population demonstrated that the assay was highly diagnostically specific. The convenience and diagnostic utility of oral fluid sampling combined with the flexibility to test either serum or oral fluid on the same platform suggests that this assay will be highly useful under the conditions for which OIE recommends ASFV antibody surveillance, i.e., in ASFV-endemic areas and for the detection of infections with ASFV isolates of low virulence

Evaluation of serological cross-reactivity and cross-neutralization between the United States porcine epidemic diarrhea virus prototype and S-INDEL-variant strains

BACKGROUND: At least two genetically different porcine epidemic diarrhea virus (PEDV) strains have been identified in the United States (U.S. PEDV prototype and S-INDEL-variant strains). The current serological assays offered at veterinary diagnostic laboratories for detection of PEDV-specific antibody are based on the U.S. PEDV prototype strain. The objectives of this study were: 1) isolate the U.S. PEDV S-INDEL-variant strain in cell culture; 2) generate antisera against the U.S. PEDV prototype and S-INDEL-variant strains by experimentally infecting weaned pigs; 3) determine if the various PEDV serological assays could detect antibodies against the U.S. PEDV S-INDEL-variant strain and vice versa. RESULTS: A U.S. PEDV S-INDEL-variant strain was isolated in cell culture in this study. Three groups of PEDV-negative, 3-week-old pigs (five pigs per group) were inoculated orally with a U.S. PEDV prototype isolate (previously isolated in our lab), an S-INDEL-variant isolate or virus-negative culture medium. Serum samples collected at 0, 7, 14, 21 and 28 days post inoculation were evaluated by the following PEDV serological assays: 1) indirect fluorescent antibody (IFA) assays using the prototype and S-INDEL-variant strains as indicator viruses; 2) virus neutralization (VN) tests against the prototype and S-INDEL-variant viruses; 3) PEDV prototype strain whole virus based ELISA; 4) PEDV prototype strain S1-based ELISA; and 5) PEDV S-INDEL-variant strain S1-based ELISA. The positive antisera against the prototype strain reacted to and neutralized both prototype and S-INDEL-variant viruses, and the positive antisera against the S-INDEL-variant strain also reacted to and neutralized both prototype and S-INDEL-variant viruses, as examined by IFA antibody assays and VN tests. Antibodies against the two PEDV strains could be detected by all three ELISAs although detection rates varied to some degree. CONCLUSIONS: These data indicate that the antibodies against U.S. PEDV prototype and S-INDEL-variant strains cross-reacted and cross-neutralized both strains in vitro. The current serological assays based on U.S. PEDV prototype strain can detect antibodies against both U.S. PEDV strains

Detection of Actinobacillus pleuropneumoniae ApxIV toxin antibody in serum and oral fluid specimens from pigs inoculated under experimental conditions

Introduction: The prevention and control of Actinobacillus pleuropneumoniae in commercial production settings is based on serological monitoring. Enzyme-linked immunosorbent assays (ELISAs) have been developed to detect specific antibodies against a variety of A. pleuropneumoniae antigens, including long-chain lipopolysaccharides (LPS) and the ApxIV toxin, a repeats-in-toxin (RTX) exotoxin unique to A. pleuropneumoniae and produced by all serovars. The objective of this study was to describe ApxIV antibody responses in serum and oral fluid of pigs

Summary of the history (age, states, farms) and prevalence of PPV4, PPV5 or concurrent PPV4/5 DNA in 185 fecal samples from pigs in the U.S.

<p>Summary of the history (age, states, farms) and prevalence of PPV4, PPV5 or concurrent PPV4/5 DNA in 185 fecal samples from pigs in the U.S.</p

The phylogenetic trees were constructed by using the Maximum Likelihood method based on the Poisson correction model, with the amino acid sequences of the nonstructural protein (ORF1, NS1) and the capsid protein (ORF2, VP1).

<p>The percentage of the tree in which the associated sequences clustered together is shown above the branches (only >70% are shown). The trees are drawn to scale, with branch lengths measured in the number of substitutions per site. Evolutionary analyses were conducted in MEGA5 <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0065312#pone.0065312-Tamura1" target="_blank">[34]</a>. The analyses involved 39 sequences of vertebrate parvoviruses with their GenBank accession numbers marked in the tree. PPV5 sequences characterized in the present study are in bold and marked with “♦”. The sequence of <i>Mythimna loreyi</i> densovirus (AY461507) was used as outgroup to root the tree. The asterisk indicates the new genus proposed by ICTV. (A) Tree constructed based on the amino acid sequences of NS1. All positions containing gaps and missing data except ambiguous positions were included. There were a total of 900 positions in the final dataset. (B) Tree constructed based on the amino acid sequences of VP1. All positions containing gaps and missing data except ambiguous positions were included. There were a total of 1,177 positions in the final dataset.</p