Postweaning Multisystemic Wasting Syndrome (PMWS) Surveillance Study

PMWS is characterized by a clinical history of wasting or poor performance in weaned pigs and by severe lymphoid depletion and histiocytic replacement of follicles in lymphoid tissues. The detection of porcine circovirus type 2 (PCV2) antigen or nucleic acids within characteristic microscopic lesions is required for the diagnosis of PMWS. Swine veterinarians submitted a specified set of samples from one hundred field cases that they felt fit the clinical definition of PMWS. All these cases were further analyzed for the presence or absence and scored for severity of the hallmark microscopic lesions (lymphoid depletion) of PMWS, the amount of PCV2 antigen associated with the lesions, and identification of concurrent bacterial and viral infections. Fifty-four of the 100 field cases were confirmed to be PMWS, whereas, no association with PCV2 was found in 46 of the cases. This highlights the need for further diagnostic testing, specifically histopathology and antigen detection, for confirmation of cases clinically suspected to be PMWS. This will become particularly important as vaccines for PCV2-associated diseases become approved for use

Lack of reproduction of the hallmark porcine circovirus type 2-associated lesions in a mouse model

BALB/c, C57BL6, and C3H/HeN mice were experimentally-infected with porcine circovirus type 2 (PCV2). The mice were tested for their ability to become infected with porcine circovirus type 2 (PCV2) and to develop the hallmark PCV2-associated lymphoid depletion and histiocytic replacement of lymphoid follicles characteristic of postweaning multisystemic wasting syndrome. Since immunostimulation has been shown to increase PCV2-replication in the pig, half of the mice were immunostimulated with keyhole limpet hemocyanin in incomplete Freund’s adjuvant (KLH/ICFA) at the time of PCV2-inoculation. PCV2 inoculation was done twice at 4 and 5 weeks of age by using intramuscular and intranasal routes. Necropsies were performed in 5-day-intervals at 12, 17, 22, 27, 32, and 37 days post PCV2 inoculation. None of the mice developed clinical disease and none of the mice developed PCV2-associated lymphoid lesions. Immunohistochemistry (IHC) and in-situ-hybridization (ISH) for PCV2-antigen/nucleic acids was performed on all tissues of all mice and was negative. PCR was done on pooled tissues and serum samples obtained at necropsy. The majority of the mice (101/111 PCV2 infected mice) were positive for PCV2-nucleic acids in tissue samples. Forty-one percent of the mice (46/111 PCV2 infected mice) were positive for PCV2-nucleic acids in serum samples. There was no difference between treatment groups or lines. This study confirms that mice can be infected with PCV2 and could be important in the epidemiology of PCV2; however, the mouse model may not be useful to understand the pathogenesis of PCV2- associated lesions

Effect of different adjuvants on PCV2-associated lesions

Ninety, 12-14 day old pigs were randomly assigned to five groups. Group 1 (n=19) pigs were vaccinated with a Mycoplasma hyopneumoniae (M. hyopneumoniae) vaccine with an oil-in-water adjuvant (RespiSure®; Pfizer Animal Health, Inc.). Group 2 (n=17) pigs were vaccinated with a commercial M. hyopneumoniae vaccine with an aqueous adjuvant (Carbopol) (Suvaxyn® Respifend® MH; Fort Dodge Animal Health, Inc.). Group 3 (n=18) pigs were vaccinated using an oil-in-water adjuvanted vaccine containing the same amount and type of M. hyopneumoniae antigen as in group 2. Group 4 (n=18) pigs were vaccinated using an aluminum hydroxide adjuvanted vaccine containing the same amount and type of M. hyopneumoniae antigen as in group 2. Group 5 (n=18) pigs served as the controls and were sham-vaccinated with saline. Pigs were injected with 2 mL of one of the four M. hyopneumoniae vaccines at four and again at six weeks of age. PCV2 was inoculated intranasally on the day of the second vaccination at 6 weeks of age. Half of the pigs were necropsied at 21 days post inoculation (DPI). The remaining pigs were necropsied at 35 DPI. There were no differences among groups in clinical disease scores. At 21 DPI all vaccinated groups had significantly (p\u3c0.05) more severe lymphoid depletion than the saline injected group. At 35 DPI group 1 pigs had significantly (p\u3c0.05) higher amounts of PCV2 DNA in serum than pigs in groups 2, 4, and 5 as determined by quantitative real-time PCR. There was a significant (p\u3c0.05) increase in the severity of lymphoid depletion in the lymph nodes, tonsil, and spleen in groups 1 and 3 compared to groups 2, 4, and 5. Group 3 had significantly (p\u3c0.05) higher amounts of PCV2 antigen within lymph nodes, tonsil, and spleen compared to groups 2, 4 and 5. The results confirm that all adjuvants tested enhanced PCV2-induced lesions and oil-in-water products used in this study had a more severe effect

An Experimental Model for Porcine Circovirus Type 2 and Mycoplasma hyopneumoniae Co-infection

Sixty-seven pigs were randomly assigned to four groups. Group 1 served as negative control pigs, group 2 pigs were inoculated with Mycoplasma hyopneumoniae (M. hyopneumoniae), group 3 pigs were dually-inoculated with M. hyopneumoniae and porcine circovirus type 2 (PCV2) and group 4 pigs were inoculated with PCV2. Dual-infected pigs had moderate dyspnea, lethargy, and reduced weight gain. The overall severity of PCV2- associated microscopic lesions in lung and lymphoid tissues were significantly (p\u3c0.05) higher in the dualinfected pigs compared to all other pigs. Four of 17 dual infected pigs had lesions consistent with postweaning multisystemic wasting syndrome (PMWS) whereas none of the singular PCV2-infected pigs developed PMWS. This study indicates that M. hyopneumoniae potentiates the severity of PCV2-associated lesions and increases the incidence of PMWS. This co-infection model closely mimics the field situation were co-infections with PCV2 are commonly observed. In the future this model will be very useful for testing intervention strategies for the control of PCV2-associated disease in growing pigs

Attenuation of porcine reproductive and respiratory syndrome virus by molecular breeding of virus envelope genes from genetically divergent strains

Molecular breeding via DNA shuffling can direct the evolution of viruses with desired traits. By using a positive-strand RNA virus, porcine reproductive and respiratory syndrome virus (PRRSV), as a model, rapid attenuation of the virus was achieved in this study by DNA shuffling of the viral envelope genes from multiple strains. The GP5 envelope genes of 7 genetically divergent PRRSV strains and the GP5-M genes of 6 different PRRSV strains were molecularly bred by DNA shuffling and iteration of the process, and the shuffled genes were cloned into the backbone of a DNA-launched PRRSV infectious clone. Two representative chimeric viruses, DS722 with shuffled GP5 genes and DS5M3 with shuffled GP5-M genes, were rescued and shown to replicate at a lower level and to form smaller plaques in vitro than their parental virus. An in vivo pathogenicity study revealed that pigs infected with the two chimeric viruses had significant reductions in viral-RNA loads in sera and lungs and in gross and microscopic lung lesions, indicating attenuation of the chimeric viruses. Furthermore, pigs vaccinated with the chimeric virus DS722, but not pigs vaccinated with DS5M3, still acquired protection against PRRSV challenge at a level similar to that of the parental virus. Therefore, this study reveals a unique approach through DNA shuffling of viral envelope genes to attenuate a positive-strand RNA virus. The results have important implications for future vaccine development and will generate broad general interest in the scientific community in rapidly attenuating other important human and veterinary viruses

Complete genome sequence of a newly identified porcine astrovirus genotype 3 strain US-MO123

Astrovirus (AstV) infections are among the most common causes of gastroenteritis and are also associated with extraintestinal manifestations in humans and many animals. Herein, for the first time, the complete genome sequence of newly identified porcine astrovirus genotype 3 (PAstV3) strain US-MO123 was determined. Sequence comparison and phylogenetic analysis showed that PAstV3 has the closest relationship with mink AstV and the human AstV strains VA1, VA2, and SG, indicating the same ancestral origin and zoonotic potential of the virus

Molecular Characterization of Recent and Archived Erysipelothrix rhusiopathiae Isolates

Cases of erysipelas have increased considerably in 2001–2002. Diagnosis of erysipelas is typically confirmed by culture and in a limited number of cases the isolates are serotyped. Reagents for serotyping are limited and are available only at National Veterinary Service Laboratory (NVSL). In this study, we utilize pulsed-field gel electrophoresis (PFGE) to differentiate genotypes and compare archived and recent isolates. Seventy-three erysipelas field isolates (58 recent, 15 historical) and four live vaccine strains were genetically characterized. Fortysix isolates were found to belong to genotype 1A(I), three were genotype 1A(III), each one was genotype 1A(IV), 1A(V), 1A(VI), and two isolates were designated as 1A(VII). Nine different genotypes were identified among the serotype 1b isolates [1B(I-IX)]. Within serotype 2, three genotypes were identified: 2A, 2B, and 2C. The four vaccine strains tested in this study belong to the genotype group 1A(II), closely related to genotype 1A. The vaccine strains and the most common field isolates genotype 1A(I) shared 78.6% identity based on PFGE pattern

Serological profile of torque teno sus virus species 1 (TTSuV1) in pigs and antigenic relationships between two TTSuV1 genotypes (1a and 1b), between two species (TTSuV1 and -2), and between porcine and human anelloviruses

The family Anelloviridae includes human and animal torque teno viruses (TTVs) with extensive genetic diversity. The antigenic diversity among anelloviruses has never been assessed. Using torque teno sus virus (TTSuV) as a model, we describe here the first investigation of the antigenic relationships among different anelloviruses. Using a TTSuV genotype 1a (TTSuV1a) or TTSuV1b enzyme-linked immunosorbent assay (ELISA) based on the respective putative ORF1 capsid antigen and TTSuV1-specific real-time PCR, the combined serological and virological profile of TTSuV1 infection in pigs was determined and compared with that of TTSuV2. TTSuV1 is likely not associated with porcine circovirus-associated disease (PCVAD), because both the viral loads and antibody levels were not different between affected and unaffected pigs and because there was no synergistic effect of concurrent PCV2/TTSuV1 infections. We did observe a higher correlation of IgG antibody levels between anti-TTSuV1a and -TTSuV1b than between anti-TTSuV1a or -1b and anti-TTSuV2 antibodies in these sera, implying potential antigenic cross-reactivity. To confirm this, rabbit antisera against the putative capsid proteins of TTSuV1a, TTSuV1b, or TTSuV2 were generated, and the antigenic relationships among these TTSuVs were analyzed by an ELISA and by an immunofluorescence assay (IFA) using PK-15 cells transfected with one of the three TTSuV ORF1 constructs. The results demonstrate antigenic cross-reactivity between the two genotypes TTSuV1a and TTSuV1b but not between the two species TTSuV1a or -1b and TTSuV2. Furthermore, an anti-genogroup 1 human TTV antiserum did not react with any of the three TTSuV antigens. These results have important implications for an understanding of the diversity of anelloviruses as well as for the classification and vaccine development of TTSuVs

Complete genome sequence of a novel porcine circovirus type 2b variant present in cases of vaccine failures in the United States

A new porcine parvovirus (PPV) was identified from pig lung tissues in the United States. It is most closely related to the recently identified PPV4 with overall genomic identities of 64.1 to 67.3%. Unlike PPV4, this virus lacks the additional open reading frame 3 (ORF3) and has a much longer ORF2. The name PPV5 is provisionally proposed

Experimental Model for Porcine Circovirus and Porcine Parvovirus Coinfection of Specific-Pathogen-Free Pigs

Porcine parvovirus (PPV) coinfection has been shown to increase the incidence and severity of porcine circovirus 2 (PCV2) associated disease in gnotobiotic and in colostrum-deprived pigs. PPV and PCV2 coinfection is also common in the grow-finish pigs in the field today. The objectives of this study were to determine the interactions between PCV2 and PPV in conventional SPF pigs and to determine whether PPV vaccine has an effect on the coinfection. Seventy-two, 6-week-old conventional pigs were inoculated either with PCV2, PPV, both PCV2 and PPV, or sham-inoculated. Before inoculation, 56 pigs were vaccinated twice with a PPV killed-virus vaccine. Clinical signs due to postweaning multisystemic wasting syndrome (PMWS) (fever, respiratory disease, jaundice, weight loss) were seen in both coinfected groups, vaccinated as well as nonvaccinated. The majority of pigs in the PCV2, and in the PCV2/PPV-inoculated groups had mild-to-severe lymphoid depletion with histiocytic replacement of follicles, and mild lymphohistiocytic interstitial pneumonia. The majority of pigs in the PCV2/PPV-coinfected groups also had mild-to-severe lymphoplasmacytic interstitial nephritis and hepatitis. There were no statistical differences between the two coinfected groups (vaccinated and non-vaccinated) in terms of clinical disease, and macroscopic and microscopic lesions. The results indicated that PPV and PCV2 coinfection resulted in increased severity of clinical disease and lymphoid lesions typical of PMWS and that a PPV-vaccination was not able to prevent PMWS in PCV2/PPV-coinfected pigs