A Novel Pathogenic Mammalian Orthoreovirus from Diarrheic Pigs and Swine Blood Meal in the United States

Abstract

Since May 2013, outbreaks of porcine epidemic diarrhea have devastated the U.S. swine industry, causing immense economic losses. Two different swine enteric coronaviruses (porcine epidemic diarrhea virus and Delta coronavirus) have been isolated from the affected swine population. The disease has been reported from at least 32 states of the United States and other countries, including Mexico, Peru, Dominican Republic, Canada, Columbia, Ecuador, and Ukraine, with repeated outbreaks in previously infected herds. Here we report the isolation and characterization of a novel mammalian orthoreovirus 3 (MRV3) from diarrheic feces of piglets from these outbreaks in three states and ring-dried swine blood meal from multiple sources. MRV3 could not be isolated from healthy or pigs that had recovered from epidemic diarrhea from four states. Several MRV3 isolates were obtained from chloroform-extracted pig feces or blood meal in cell cultures or developing chicken embryos. Biological characterization of two representative isolates revealed trypsin resistance and thermostability at 90°C. NextGen sequencing of ultrapurified viruses indicated a strong homology of the S1 segment to mammalian and bat MRV3. Neonatal piglets experimentally infected with these viruses or a chloroform extract of swine blood meal developed severe diarrhea and acute gastroenteritis with 100% mortality within 3 days postinfection. Therefore, the novel porcine MRV3 may contribute to enteric disease along with other swine enteric viruses. The role of MRV3 in the current outbreaks of porcine epidemic diarrhea in the United States remains to be determined, but the pathogenic nature of the virus warrants further investigations on its epidemiology and prevalence. IMPORTANCE Porcine orthoreoviruses causing diarrhea have been reported in China and Korea but not in the UnitedStates. We have isolated and characterized two pathogenic reassortant MRV3 isolates from swine fecal samples from porcine epidemic diarrhea outbreaks and ring-dried swine blood meal in the United States. These fecal and blood meal isolates or a chloroform extract of blood meal induced severe diarrhea and mortality in experimentally infected neonatal pigs. Genetic and phylogenetic analyses of two MRV3 isolates revealed that they are identical but differed significantly from nonpathogenic mammalian orthoreoviruses circulating in the United States. The present study provides a platform for immediate development of suitable vaccines and diagnostics to prevent and control porcine orthoreovirus diarrhea.

Nanopore sequencing as a revolutionary diagnostic tool for porcine viral enteric disease complexes identifies porcine kobuvirus as an important enteric virus

“…Within each litter, one piglet was identified for longitudinal follow-up during the entire suckling period. A dry cotton rectal swab (Copan) was collected from each individual piglet at days 1,5,8,11,14,17,20 and 22 after birth. The swab was placed immediately in 2 ml of viral transport medium (phosphate buffered saline containing 1000 U/ml penicillin (Continental Pharma, Puurs, Belgium), 1 mg/ml streptomycin (Certa, Braine l′Alleud, Belgium), 1 mg/ml gentamicin (Life Technologies) and 0.01% v/v Fungizone (Bristol-Myers Squibb, Braine l′Alleud, Belgium)) in a sterile 15 ml falcon tube (Sarstedt) and stored at −20 °C.…”

Genetic and pathogenic characterization of a novel reassortant mammalian orthoreovirus 3 (MRV3) from a diarrheic piglet and seroepidemiological survey of MRV3 in diarrheic pigs from east China

“…KU558701) at 1 × 10 5 plaque-forming unit (PFU)/ml (4 ml per pig) and MRV-ZJ2013 at 6 × 10 6 PFU/ml (4 ml per pig), respectively. The challenge dosage of MRV-ZJ2013 in group C referred to a recent study (Thimmasandra Narayanappa et al, 2015). All pigs were monitored daily for rectal temperature and clinical signs.…”

“…However, in the last decade, increasing numbers of studies in humans and other mammals have shown that they can cause severe illness in humans and other mammals, including upper respiratory tract infections, encephalitis, and diarrhea (Chua et al, 2008;Ouattara et al, 2011a;Steyer et al, 2013a). MRVs have been isolated from a broad range of mammalian species, including bats (Hu et al, 2014;Kohl et al, 2012;Lelli et al, 2013;Lorusso et al, 2014;Thalmann et al, 2010;Wang et al, 2015a;Yang et al, 2015), civet cats , cows (Anbalagan et al, 2014), mink (Lian et al, 2013), pigs (Dai et al, 2012b;Kwon et al, 2012b;Thimmasandra Narayanappa et al, 2015;Zhang et al, 2011a), dogs (Decaro et al, 2005) and humans (Chua et al, 2008;Ouattara et al, 2011b). However, porcine MRV failed to arouse the concern of researchers until 2007, when it was first reported as the cause of diarrhea in newborn piglets in China (Zhang et al, 2011a).…”

“…They found that 19.0% of diarrheic fecal samples collected from 78 pig farms around the country tested positive for porcine MRV (Kwon et al, 2012b). In 2015, a novel type 3 MRV (MRV3) was isolated and characterized in the USA, having caused severe diarrhea and acute gastroenteritis in neonatal piglets with 100% mortality by 3 days post-infection (Thimmasandra Narayanappa et al, 2015). Therefore, porcine MRV may contribute to enteric disease alone or in combination with other swine pathogens.…”

Detection and Characterization of a Novel Reassortant Mammalian Orthoreovirus in Bats in Europe

“…Novel MRVs have recently been identified in several hosts, including bats in Italy and Germany [ 16 , 17 ], and a novel Orthoreovirus , with a high similarity to the MRVs found in bats in Europe, was detected in Slovenia from a child requiring hospitalization due to acute gastroenteritis [ 15 ]. Recently, a virulent MRV containing an S1 segment that was strongly homologous to bat MRV3 was detected from diarrheic pigs in the United States [ 18 ]. In addition, a bat-borne fusogenic Orthoreovirus with zoonotic potential was detected from healthy flying foxes ( Pteropus vampyrus ) legally imported from Indonesia to Europe [ 19 ].…”