Showing posts with label porcine. Show all posts
Showing posts with label porcine. Show all posts
Monday, December 19, 2016
Porcine Bocavirus Infection Associated with Encephalomyelitis in a Pig Germany1 Volume 22 Number 7—July 2016 Emerging Infectious Disease journal CDC
Porcine Bocavirus Infection Associated with Encephalomyelitis in a Pig Germany1 Volume 22 Number 7—July 2016 Emerging Infectious Disease journal CDC
Porcine Bocavirus Infection Associated with Encephalomyelitis in a Pig, Germany1 - Volume 22, Number 7—July 2016 - Emerging Infectious Disease journal - CDC
Volume 22, Number 7—July 2016
Letter
Porcine Bocavirus Infection Associated with Encephalomyelitis in a Pig, Germany1
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To the Editor: In 2013, a 6-week-old female piglet kept in a flatdeck cage had coughing, growth retardation, and diarrhea and was taken to a local veterinarian in Hannover, Germany; the piglet was euthanized. After necropsy at the University of Veterinary Medicine in Hannover, histologic investigation found interstitial pneumonia; a mild, multifocal, lymphohistiocytic panencephalitis that affected the cerebrum and cerebellum, including brain stem and medulla oblongata; and a mild, multifocal, lymphohistiocytic panmyelitis. Results from screening for typical neurotropic viruses (classical swine fever virus, suid herpesvirus 1, rabies virus, teschovirus, porcine enterovirus 8, 9, and 10) were negative; Mycoplasma hyorhinis was detected by multiplex PCR (Institute of Virology, University of Veterinary Medicine Hannover) within the lung and pulmonary lymph nodes. Cerebral tissue from the pig was processed for viral metagenomics by random RNA and DNA virus screening and next-generation sequencing (NGS) with the 454 sequencing platform (GS Junior; Roche, Basel, Switzerland), as described (1), and 21,359 reads were obtained. Analysis by using blastn and blastx (2) showed 10 reads had >97% nt identity with porcine bocavirus (PBoV) KU14. No other viral sequences were detected.
By using primers based on sequence data of the PBoV, partially overlapping PCR amplicons were obtained to confirm and extend the NGS data of the isolate, which was named PBoV S1142/13 (1; GenBank accession no. KU311698). A total of 2,176 nt of PBoV S1142/13 were obtained, consisting of the partial nucleoprotein (NP) 1 and the nearly complete viral protein (VP) 1 gene. By using MAFFT version 7 (http://mafft.cbrc.jp/alignment/server/), we aligned the nearly complete VP1 gene of PBoV S1142/13 with various closely related members of the genusBocaparvovirus and built a maximum-likelihood tree by using the general time reversible plus invariable sites plus gamma distribution method, as determined by jModelTest 2.0 (3) and default parameters in MEGA6.06 (4). Results confirmed that PBoV S1142/13 was most closely related to PBoV KU14 (Figure, panel A). The partial genome of PBoV S1142/13 differed at 8 nt positions from PBoV KU14, resulting in 99.6% nt identity. Of these nucleotide differences, 4 resulted in an amino acid difference, including position 2733 (T?C on the basis of PBoV KU14 as a reference genome), which is part of the NP1 stopcodon of PBoV KU14. These results indicate that the stopcodon was located 39 nt farther downstream than for PBoV KU14. The other 3 aa differences were present in the VP1 protein; each of these differences was within the same group of amino acids as those detected in PBoV KU14.
For further substantiation of a potential cause-effect relationship of histologic (Figure, panel B) and NGS results, we performed fluorescent in situ hybridization (FISH) on formalin-fixed, paraffin-embedded central nervous system (CNS) sections of the diseased animal and of a control pig with no CNS lesions. We used an RNA probe specific for the obtained NP1 and VP1 sequences covering 1,153 nt (Affymetrix, Santa Clara, CA, USA) according to the manufacturer’s protocol, with minor variations (ViewRNA ISH Tissue 1-Plex Assay Kit and ViewRNA Chromogenic Signal Amplification Kit, Affymetrix). A probe specific for porcine ubiquitin (Sus scrofa ubiquitin C; GenBank accession no. XM_005657305; nt 2–890) served as a positive control.
Figure. Phylogenetic analysis and staining of porcine bocavirus (PBoV) from the spinal cord of a diseased pig, Hannover, Germany. A) Phylogenetic relationship of PBoV isolate S1142/13 (bold) with other bocaviruses. The nucleotide...
The spinal cord of the diseased pig showed diffuse intracytoplasmic and intranuclear PBoV-specific signals within scattered neurons adjacent to the histologically detected inflammatory lesions (Figure, panel C). The negative control and the nonprobe incubation lacked PBoV-specific signals. The porcine ubiquitin probe provided a strong intracellular and extracellular staining within the CNS of both pigs.
PBoV (genus Bocaparvovirus, family Parvoviridae) was first described in 2009 as porcine boca-like virus in pigs in Sweden with postweaning multisystemic wasting syndrome (5). PBoV is usually involved in respiratory and intestinal diseases in pigs (5) but has not been detected in the CNS. In the pig in our study, the lack of detection of other viral sequences by using NGS indicates the potential role of PBoV as a pathogen that triggers encephalomyelitis. FISH substantiated the NGS results and revealed neuronal intracytoplasmic and intranuclear PBoV-specific signals adjacent to the lesion, indicating intraneuronal transcription and replication (6). Nevertheless, a potential synergistic effect of M. hyorhinis on the PBoV pathogenesis cannot be ruled out. Similarly, co-infection of M. hyorhinis and porcine circovirus type 2 has been associated with enhanced inflammatory lesions in the lungs of pigs (7).
The CNS tropism of PBoV S1142/13 could result from various factors, including specific amino acid changes that enable the virus to pass the blood–brain barrier and infect neurons. Additional studies are necessary to elucidate a possible role of the amino acid differences between PBoV S1142/13 and PBoV KU14 in the tropism of these viruses.
Human bocavirus has recently been found in the cerebrospinal fluid of patients having encephalitis (8), and related human parvovirus 4 (9) and human parvovirus B19 (10) have been reported in human encephalitis. The correlation of PBoV-specific signals by using FISH for histologic detection of encephalomyelitis assigns PBoV a potential role in provoking CNS lesions. PBoV should be considered as a cause of encephalomyelitis but needs further investigation.
Vanessa M. Pfankuche, Rogier Bodewes, Kerstin Hahn, Christina Puff, Andreas Beineke, André Habierski, Albert D.M.E. Osterhaus, and Wolfgang Baumgärtner
Acknowledgments
The authors thank the Lower Saxony State Office for Consumer Protection and Food Safety, the Institute of Virology at the University of Veterinary Medicine Hannover, and the Friedrich-Loeffler-Institute in Jena for performing the routine diagnostic analyses of common porcine pathogens. The authors also thank Danuta Waschke, Kerstin Rohn, Bettina Buck, Caroline Schütz, and Kerstin Schöne for excellent technical assistance.
This study was in part supported by the Niedersachsen-Research Network on Neuroinfectiology of the Ministry of Science and Culture of Lower Saxony and by the COMPARE project and received funding from the European Union’s Horizon 2020 research and innovation program COMPARE (grant agreement no. 643476). V.M.P. received a scholarship from the Akademie für Tiergesundheit e.V. in Bonn, Germany.
References
- van Leeuwen M, Williams MM, Koraka P, Simon JH, Smits SL, Osterhaus AD. Human picobirnaviruses identified by molecular screening of diarrhea samples. J Clin Microbiol. 2010;48:1787–94. DOIPubMed
- Schürch AC, Schipper D, Bijl MA, Dau J, Beckmen KB, Schapendonk CM, Metagenomic survey for viruses in Western Arctic caribou, Alaska, through iterative assembly of taxonomic units. PLoS One. 2014;9:e105227. DOIPubMed
- Darriba D, Taboada GL, Doallo R, Posada D. jModelTest 2: more models, new heuristics and parallel computing. Nat Methods. 2012;9:772. DOIPubMed
- Tamura K, Stecher G, Peterson D, Filipski A, Kumar S. MEGA6: Molecular evolutionary genetics analysis version 6.0. Mol Biol Evol. 2013;30:2725–9.DOIPubMed
- Zhou F, Sun H, Wang Y. Porcine bocavirus: achievements in the past five years. Viruses. 2014;6:4946–60. DOIPubMed
- Bodewes R, Lapp S, Hahn K, Habierski A, Förster C, König M, Novel canine bocavirus strain associated with severe enteritis in a dog litter. Vet Microbiol. 2014;174:1–8. DOIPubMed
- Chen D, Wei Y, Huang L, Wang Y, Sun J, Du W, Synergistic pathogenicity in sequential coinfection with Mycoplasma hyorhinis and porcine circovirus type 2. Vet Microbiol. 2016;182:123–30. DOIPubMed
- Mori D, Ranawaka U, Yamada K, Rajindrajith S, Miya K, Perera HK, Human bocavirus in patients with encephalitis, Sri Lanka, 2009–2010. Emerg Infect Dis. 2013;19:1859–62. DOIPubMed
- Benjamin LA, Lewthwaite P, Vasanthapuram R, Zhao G, Sharp C, Simmonds P, Human parvovirus 4 as potential cause of encephalitis in children, India. Emerg Infect Dis. 2011;17:1484–7.PubMed
- Barah F, Whiteside S, Batista S, Morris J. Neurological aspects of human parvovirus B19 infection: a systematic review. Rev Med Virol.2014;24:154–68. DOIPubMed
Figure
Suggested citation for this article: Pfankuche VM, Bodewes R, Hahn K, Puff C, Beineke A, Habierski A, et al. Porcine bocavirus infection associated with encephalomyelitis in a pig. Emerg Infect Dis. 2016 Jul [date cited]. http://dx.doi.org/10.3201/eid2207.152049
1Preliminary results from this study were presented at the 3rd International One Health Congress, March 15–18, 2015, Amsterdam, the Netherlands, and at the Conference of the German Veterinary Medical Association, March 8–10, 2015, Fulda, Germany.
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Saturday, October 8, 2016
New Chimeric Porcine Coronavirus in Swine Feces Germany 2012 Volume 22 Number 7—July 2016 Emerging Infectious Disease journal CDC
New Chimeric Porcine Coronavirus in Swine Feces Germany 2012 Volume 22 Number 7—July 2016 Emerging Infectious Disease journal CDC
New Chimeric Porcine Coronavirus in Swine Feces, Germany, 2012 - Volume 22, Number 7—July 2016 - Emerging Infectious Disease journal - CDC
Volume 22, Number 7—July 2016
Letter
New Chimeric Porcine Coronavirus in Swine Feces, Germany, 2012
On This Page
- Letter
- Suggested Citation
Suggested citation for this article
To the Editor: Porcine epidemic diarrhea virus (PEDV) and transmissible gastroenteritis virus (TGEV) can cause severe enteritis in pigs accompanied by diarrhea, vomiting, and dehydration. Clinical signs are most prominent in young suckling pigs, in which high mortality rates are common. As seen in recent porcine epidemic diarrhea outbreaks in the United States and Asia, the effect on the pig industry can be tremendous.
Recently, Boniotti et al. (1) reported detection and genetic characterization of swine enteric coronaviruses (CoVs) circulating in Italy during 2007–2014. Characterization was based on sequencing and phylogenetic analyses of spike genes of TGEV and PEDV isolates. This study also reported a new recombinant CoV strain with a TGEV backbone and a PEDV spike gene (SeCoV/Italy/213306/2009; KR061459), which was identified as a swine enteric CoV (SeCoV). This chimeric virus presumably resulted from a recombination event.
Figure. Electron micrograph of a new chimeric swine enteric coronavirus (SeCoV/GER/L00930/2012), Germany, 2012. Scale bar indicates 100 nm.
Accompanying a study of recent porcine epidemic diarrhea cases in Germany caused by a new PEDV Indel strain (2), we retrospectively analyzed fecal samples from pigs that showed typical clinical symptoms of a PEDV infection. The sample set included fecal material collected from a farm in southern Germany on which an episode of diarrhea among pigs occurred in 2012. This material was shown by electron microscopy to contain CoV-like particles (Figure), but showed negative results by reverse transcription PCRs specific for the PEDV nucleocapsid gene.
Subsequent metagenomic analyses resulted in the full-genome sequence of a swine enteric CoV (SeCoV/GER/L00930/2012). We found a sequence showing high similarity (99.5% identity) with the TGEV/PEDV recombinant reported by Boniotti et al. (1). Network analysis of complete genome sequences of similar CoVs underline the chimeric nature of the genome between TGEV and PEDV genome sequences (Technical Appendix[PDF - 386 KB - 1 page] Figure, panel A). The chimeric nature of the virus strain was confirmed by RT-PCR with primers spanning possible recombination sites and analysis of overlapping reads from next-generation sequencing.
Annotation of the sequence of SeCoV/GER/L00930/2012 performed on the basis of SeCoV/Italy/213306/2009 identified a similar putative coding sequence with a TGEV backbone and a spike coding sequence similar to that for PEDV (Technical Appendix[PDF - 386 KB - 1 page] panel B). Downstream of the spike protein–coding open reading frame (ORF), an additional hypothetical ORF was identified in both SeCoV sequences. The coded amino acid sequences (27 aa in the virus from Germany and 30 aa in the virus from Italy) resembled an N- and C-terminally truncated TGEV nonstructural protein 3a. The difference of 3 aa between the 2 strains is the result of a 10-bp deletion at the 3?-end of the hypothetical ORF, which shifted the stop 3 codons to the 5?- end (Technical Appendix[PDF - 386 KB - 1 page] Figure, panel B) in SeCoV/GER/L00930/2012. This deletion is apparently located within the potential 3? recombination site (Technical Appendix[PDF - 386 KB - 1 page] Figure, panel B).
It is tempting to speculate that SeCoV/Italy/213306/2009 is a precursor of SeCoV/GER/L00930/2012, and that other members of this novel genotype are still undetected. These viruses might be targets of secondary mutation and recombination events. Therefore, more chimeric CoVs should be identified to determine the potential origin of the recombination event.
In conclusion, we detected an enteric CoV that resembled the TGEV/PEDV chimeric virus reported by Boniotti et al. (1). Although these findings support the notion that CoV genomes are subject to mutations and recombination events, problems in disease diagnosis can be foreseen. In countries where porcine epidemic diarrhea, transmissible gastroenteritis, or both of these diseases are reportable, correct diagnosis and reporting might be difficult. Thus, diagnosticians should be aware of possible recombinants of swine CoVs. Diagnostic problems can be prevented by use of a double-check strategy with techniques specific for different genome regions. Apart from diagnostic obstacles, the effect of virus recombinations in terms of virulence and organ tropism is unknown and needs further investigations.
Valerij Akimkin1, Martin Beer1 , Sandra Blome1, Dennis Hanke1, Dirk Höper1, Maria Jenckel1, and Anne Pohlmann1
, Sandra Blome1, Dennis Hanke1, Dirk Höper1, Maria Jenckel1, and Anne Pohlmann1References
- Boniotti MB, Papetti A, Lavazza A, Alborali G, Sozzi E, Chiapponi C, Porcine epidemic diarrhea virus and discovery of a recombinant swine enteric coronavirus, Italy. Emerg Infect Dis. 2016;22:83–7 . DOIPubMed
- Hanke D, Jenckel M, Petrov A, Ritzmann M, Stadler J, Akimkin V, Comparison of porcine epidemic diarrhea viruses from Germany and the United States, 2014. Emerg Infect Dis. 2015;21:493–6 . DOIPubMed
Figure
Technical Appendix
Suggested citation for this article: Akimkin V, Beer M, Blome S, Hanke D, Höper D, Jenckel M, et al. New chimeric porcine coronavirus in swine feces, Germany, 2012 [letter]. Emerg Infect Dis. 2016 Jul [date cited]. http://dx.doi.org/10.3201/eid2207.160179
1All authors contributed equally to this article.
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