Kurt Rossow

Porcine Reproductive and Respiratory Syndrome

In 1987, porcine reproductive and respiratory syndrome (PRRS) was recognized in the USA as a new disease of swine causing late-term reproductive failure and severe pneumonia in neonatal pigs. The syndrome is caused by an RNA virus referred to as PRRS virus (PRRSV), which is classified in the family Arteriviridae. Swine macrophages are the only indigenous cell type known to support PRRSV replication. Direct contact between infected and naive pigs is the predominant route of PRRSV transmission. Exposure of a mucosal surface to PRRSV leads to virus replication in regional macrophages, a prolonged viremia and systemic distribution of virus to other macrophage populations. Reproductive failure induced by PRRSV infection in late-gestation sows is characterized by premature farrowing of stillborn, partially autolyzed, and mummified fetuses. Pneumonia caused by PRRSV infection is more severe in young pigs compared to adults and may be complicated by concurrent bacterial infections. Gross lung lesions associated with PRRSV infection vary from none to diffuse consolidation. In addition, multiple lymph nodes may be markedly enlarged. Microscopically, PRRSV-pneumonia is characterized by multifocal, interstitial thickening by macrophages and necrotic cell debris in alveoli. Other less common microscopic lesions of PRRSV infection include myocarditis, vasculitis, encephalitis, and lymphoid hypertrophy and hyperplasia. In acute or subacute PRRSV infections, serum and lung are the best specimens for diagnosis. Persistent PRRSV infections can be produced by transplacental or intranasal infection. Persistent PRRSV infections are an important factor for virus survival and transmission within a swine herd and will complicate control programs.

Experimental porcine reproductive and respiratory syndrome virus infection in one-, four-, and 10-week-old pigs.

One-, 4-, and lo-week-old pigs were exposed to porcine reproductive and respiratory syndrome virus (PRRSV) to determine the effect of age on clinical signs, hematologic alterations, the onset and duration of viremia, routes of virus shedding, antibody production, and microscopic lesions produced by PRRSV isolate ATCC VR-2332. The response to PRRSV infection was similar among age groups. Fever, usually prolonged, and a marked dyspnea with cutaneous erythema when restrained for sample collection were the most consistent clinical signs. Prolonged periocular edema was unique to the 1-week-old pigs. The white blood cell count was decreased on day 4 postexposure (PE) due to decreases in neutrophils and lymphocytes. The virus was isolated from buffy coats at day 1 PE and was isolated from serum, buffy coat, or plasma at each sample collection period through the end of the trial (day 28 PE). Virus was most consistently isolated from lung, lymph node, spleen, and tonsil on day 7 PE and exclusively from lymph node, spleen, and tonsil on day 28 PE. Virus was infrequently isolated from urine and fecal and nasal swabs. Consistent microscopic changes in all age groups included interstitial pneumonia and lymph node hypertrophy and hyperplasia on days 7 and 28 PE, lymph node necrosis on day 7 PE, and subacute mononuclear myocarditis on day 28 PE. Findings presented here indicate that interstitial pneumonia, lymphoid necrosis, and mononuclear myocarditis are characteristic lesions of PRRSV isolate ATCC VR-2332 infection in 1-, 4-, and lo-week-old pigs.

Pathogenesis of Porcine Reproductive and Respiratory Syndrome Virus Infection in Gnotobiotic Pigs

The pathogenesis of porcine reproductive and respiratory syndrome virus (PRRSV) was determined in gnotobiotic pigs by studying the sequential development of microscopic lesions and sites of virus distribution and replication. Thirty-two pigs (three pigs/infected group and one pig/control group) were inoculated by nasal instillation of either PRRSV isolate ATCC VR-2332 (total dose 102,6 TCID50) or uninfected cell culture supernatant. Infected and control pigs were euthanized at 12 hours, and 1, 2, 3, 5, 7, 14, and 21 days postexposure (PE). Gnotobiotic pigs experimentally infected with PRRSV were viremic by 12 hours PE and subsequently developed pneumonia, lymphadenopathy, vasculitis, myocarditis, and encephalitis. Lung lesions developed by day 3 PE, persisted through day 21 PE and were characterized by alveolar septa thickened by macrophages, alveolar proteinaceous and karyorrhectic debris, alveolar syncytial cells, and multifocal type II pneumocyte hypertrophy. Lymph node lesions varied in distribution and severity and were characterized by germinal center hypertrophy and hyperplasia, lymphocyte necrosis, multiple cystic spaces, and polykaryocytes within the cystic spaces. Heart lesions were a late feature of infection and all infected pigs had heart lesions on day 21 PE characterized by subendocardial, myocardial, and perivascular foci of lymphocytes. Vasculitis also varied in distribution and severity and affected all sizes of vessels. Results of this experiment indicate that PRRSV is a multisystem disease characterized initially by viremia with subsequent virus distribution and replication in multiple organs causing interstitial pneumonia, vasculitis, lymphadenopathy, myocarditis, and encephalitis.

Characterization of Emerging European-Like Porcine Reproductive and Respiratory Syndrome Virus Isolates in the United States

ABSTRACT European-like field isolates of porcine reproductive and respiratory syndrome virus (PRRSV) have recently emerged in North America. The full-length genomic sequence of an index isolate characterized in 1999, strain EuroPRRSV, served as the reference strain for further studies of the evolution and epidemiology of European-like isolates (type 1) in the United States. Strain EuroPRRSV shared 90.1 to 100% amino acid identity with the prototype European strain, Lelystad, within the structural and nonstructural open reading frames (ORFs) and 95.3% overall nucleotide identity. The 5′ untranslated region and two nonstructural regions within ORF 1 were closely examined due to significant divergence from strain Lelystad. A 51-bp deletion in a region within ORF 1a, coding for nonstructural protein 2 (NSP2), was observed. Sequence analysis of the structural ORFs 2 to 7 of additional European-like isolates indicated that these isolates share 93% nucleotide identity with one another and 95 to 96% identity with the Lelystad strain but only 70% identity with the North American reference strain VR-2332. Phylogenetic analysis with published PRRSV ORF 3, 5, and 7 nucleotide sequences indicated that these newly emerging isolates form a clade with the Lelystad and United Kingdom PRRSV isolates. Detailed analysis of four of these isolates with a panel of 60 monoclonal antibodies directed against the structural proteins confirmed a recognition pattern that was more consistent with strain Lelystad than with other North American isolates.

Distinct characteristics and complex evolution of pedv strains, North america, May 2013-february 2014

Sequence analysis showed heterogeneity among 74 strains and distinct molecular characteristics of highly virulent strains and variants.

Chronological Immunohistochemical Detection and Localization of Porcine Reproductive and Respiratory Syndrome Virus in Gnotobiotic Pigs

An immunogold-silver immunohistochemical technique was used to determine the chronological distribution and localization of porcine reproductive and respiratory syndrome virus (PRRSV) in experimentally infected gnotobiotic pigs. Thirty-two pigs were randomly allocated to infected (n = 24) or control (n = 8) groups. Pigs in infected groups were inoculated at 3 days of age by nasal instillation of PRRSV isolate ATCC VR-2332 (total dose = 102.64 TCID50), and control pigs were exposed in the same manner to uninfected cell culture supernatant. Three infected and one control pigs were euthanatized at 12 hours and at 1, 2, 3, 5, 7, 14, and 21 days postexposure (DPE). Bronchiolar epithelial cells, arteriolar endothelial cells, monocytes, and interstitial, alveolar, and intravascular macrophages stained for PRRSV antigen at 12 hours postexposure. Staining for PRRSV antigen in endothelial cells, monocytes, and alveolar, interstitial, and intravascular macrophages was most intense and widespread in lung sections from 14 and 21 DPE. In the heart, macrophages in the interstitial and subendocardial spaces and endothelial cells in a few arterioles stained for PRRSV antigen at 14 and 21 DPE. Tonsillar macrophages and mucosal epithelium stained for PRRSV antigen at 12 hours postexposure and sporadically with less intensity in subsequent sampling periods. In the nasal turbinate, PRRSV antigen was identified in macrophages within the mucosal epithelium at 12 hours postexposure and again at 14 and 21 DPE. There was focal staining for PRRSV antigen in the choroid plexus in one pig at 14 DPE. Based on the results of this experiment, the pathogenesis of PRRSV infection in gnotobiotic pigs can be described as initial virus entry through nasal epithelial, tonsillar, and pulmonary macrophages, with viremia occurring by 12 hours postexposure followed by the development of pneumonia, myocarditis, encephalitis, rhinitis, vasculitis, and lymphoid necrosis. Although PRRSV can infect macrophages in heart, tonsil, turbinate, and choroid plexus, pulmonary macrophages are predominantly and consistently infected and are the predominant cells for virus replication in gnotobiotic pigs.

Isolation and Genetic Characterization of New Reassortant H3N1 Swine Influenza Virus from Pigs in the Midwestern United States

ABSTRACT Since the introduction of H3N2 swine influenza viruses (SIVs) into U.S. swine in 1998, H1N2 and H1N1 reassortant viruses have emerged from reassortment between classical H1N1 and H3N2 viruses. In 2004, a new reassortant H3N1 virus (A/Swine/Minnesota/00395/2004) was identified from coughing pigs. Phylogenetic analyses revealed a hemagglutinin segment similar to those of contemporary cluster III H3N2 SIVs and a neuraminidase sequence of contemporary H1N1 origin. The internal genes were of swine, human, and avian influenza virus origin, similar to those of contemporary U.S. cluster III H3N2 SIVs. The recovery of H3N1 is further evidence of reassortment among SIVs and justifies continuous surveillance.

Emergence of H3N2 reassortant influenza A viruses in North American pigs.

In late summer through early winter of 1998, there were several outbreaks of respiratory disease in the swine herds of North Carolina, Texas, Minnesota and Iowa. Four viral isolates from outbreaks in different states were analyzed, both antigenically and genetically. All of the isolates were identified as H3N2 influenza viruses with antigenic profiles similar to those of recent human H3 strains. Genotyping and phylogenetic analysis demonstrated that the four swine viruses had emerged through two different pathways. The North Carolina isolate is the product of genetic reassortment between human and swine influenza viruses, while the others arose from reassortment of human, swine and avian viral genes. The hemagglutinin genes of the four isolates were all derived from the human H3N2 virus circulating in 1995. It remains to be determined if either of these recently emerged viruses will become established in the pigs in North America and whether they will become an economic burden.

Lymphoid tissue tropism of porcine reproductive and respiratory syndrome virus replication during persistent infection of pigs originally exposed to virus in utero.

Abstract The ability of porcine reproductive and respiratory syndrome virus (PRRSV) to establish a persistent infection is the principal contributing factor to the world-wide spread of the disease. Several studies have documented the course of viral infection in postnatally infected pigs; however, very little is known regarding sites of virus replication during persistent infection of pigs exposed to PRRSV in utero. In this study, virus replication and PRRSV-specific antibody were followed for several hundred days in a group of pigs derived from three sows infected at 90 days of gestation with PRRSV isolate VR-2332. Eighty-four percent of pigs were born viremic with a mortality of 54% within 21 days after birth. At approximately 60 days sera from pigs were negative for virus by virus isolation. Analysis of virus replication in the tissues of pigs randomly sacrificed between 63 and 132 days showed no evidence of virus in lung and other non-lymphoid organs. However, virus was easily recovered from tonsil and lymph nodes and in situ hybridization identified these tissues as sites of virus replication. Even though replication was at a low level, virus was easily transmitted to sentinel pigs. By 260 days pigs became seronegative and did not transmit virus to sentinel pigs. Sacrifice of remaining pigs after 300 days showed no evidence of virus in blood and tissues. This study shows that congenital PRRSV-infected pigs can support virus replication for an extended period during which virus replication is primarily restricted to tonsil and lymph nodes.

Porcine reproductive and respiratory syndrome virus infection of gnotobiotic pigs: sites of virus replication and co-localization with MAC-387 staining at 21 days post-infection

The organ distribution of PRRSV-infected cells in gnotobiotic piglets at 21 days after infection with PRRSV isolate VR-2332 was examined by in situ hybridization. Cells that expressed PRRSV RNA were identified in all tissues examined, including organs not usually characterized as sites of PRRSV infection. PRRSV-infected cells frequently appeared in clusters and were not always associated with microscopic lesions. The expression of PRRSV RNA co-localized with a macrophage monoclonal antibody, MAC-387, in lymph nodes. Some, but not all infected cells stained with MAC-387. The wide distribution of PRRSV-infected cells and co-localization with MAC-387 staining is consistent with the macrophage-tropism of PRRSV and is similar to observations made during persistent infection with other arteriviruses.