ReviewPorcine circoviruses: current status, knowledge gaps and challenges
Introduction
In the early 70s, a German research group investigated the permanent pig kidney (PK) cell line 15 (ATCC-CCL31) by electron microscopy and recognized small, spherical virus-like particles (Tischer et al., 1974). In a follow-up study, the same research group further characterized these particles as small icosahedral viruses with a diameter of only 17 nm, containing a covalently closed circular single-stranded DNA genome and one main capsid protein (Tischer et al., 1982). The newly identified virus was designated “porcine circovirus (PCV)” as antibodies against it could only be identified in pigs but not in rabbits, mice, calves, and humans (Tischer et al., 1982). Since then, additional PCVs have been identified and there are currently four recognized types: Porcine circovirus 1 (PCV1), Porcine circovirus 2 (PCV2), Porcine circovirus 3 (PCV3) and Porcine circovirus 4 (PCV4) (Fig. 1). All four PCVs are similar in structure: they contain two main open reading frames (ORFs) oriented in opposite directions in the circular genome; the ORF1 or rep gene encodes for proteins associated with replication, Rep and Rep’ (not characterized for PCV3 and PCV4 at this point), and the ORF2 or cap gene encodes for the capsid or Cap protein (Finsterbusch et al., 2005). Basic information on genome length, protein sizes, cell lines used for propagation and information on experimental success for each PCV is summarized in Table 1.
PCV1 corresponds to the virus reported in 1974 in PK-15 cells (Tischer et al., 1974). Although PCV1 is found in both wild and farmed pigs, it is generally considered non-pathogenic as no association with disease was observed after natural or experimental infection of pigs (Allan et al., 1995; Tischer et al., 1986). A new disease in pigs described as postweaning multisystemic wasting syndrome (PMWS) was first observed in Canada in 1995 (Harding, 2004). During 1997-1998, PCV2 was discovered and associated with PMWS by multiple laboratories (Allan et al., 1998; Ellis et al., 1998; Kiupel et al., 1998; Morozov et al., 1998; Nayar et al., 1997). The first identification of PCV2 in PMWS pigs was made possible by restriction enzyme cleavage analysis, leading to identification of a novel restriction pattern different from that of PCV1 (Nayar et al., 1997). The association of PCV2 with pathogenicity was established by traditional methods of virus isolation from PMWS affected pigs, experimental infections, histopathology, in addition to immunohistochemistry (IHC) and in situ hybridization (ISH) to detect PCV2 antigen or genome (Allan et al., 1998; Ellis et al., 1998; Kiupel et al., 1998; Morozov et al., 1998). Since its discovery, PCV2 has become one of the most important pig viruses and is associated with several disease syndromes in pigs.
One of the first viral metagenomics studies performed using the shotgun sequencing technology was published in 2002, describing the viral metagenome of seawater (Breitbart et al., 2002). Since then, the developments in high throughput sequencing and associated bioinformatics computational capacities have accelerated the discovery of many previously unknown viral genomes. While traditional virology methods were used to discover PCV2, metagenomic sequencing on tissue samples from pigs with PDNS, reproductive failure, myocarditis and multi-systemic inflammation was used to discover PCV3 (Palinski et al., 2017; Phan et al., 2016). Although PCV3 has attracted a lot of attention and research, PCV3 isolation in cell culture has only recently been successful (Oh and Chae, 2020) and its role as a pathogen is still controversial. Similarly to PCV3, PCV4 was identified very recently, with the aid of sequencing, in pigs with various health conditions from two farms in Hunan province, China (Zhang et al., 2019). Analysis by PCR with primers complementary to the conserved regions of the rep gene for the genera Circovirus and Cyclovirus allowed the sequencing of PCV4 (Zhang et al., 2019). This review aims to summarize recent findings on circoviruses (CV) currently known to circulate in the global pig population.
Section snippets
Circoviruses: A growing family
Thirteen ssDNA virus families are identified by the International Committee on the Taxonomy of Viruses (ICTV, http://ictv.global/report), with the exception of Parvoviridae and Bidnaviridae, 11 families contain circular genomes (Zhao et al., 2019). Of these, six families encode a homologous Rep protein and are named circular Rep-encoding ssDNA (CRESS DNA) viruses. This Rep protein is distantly related to the Rep protein of bacterial CRESS viruses and archaeal CRESS viruses, indicating the
Circoviruses in pigs
In pigs, four CVs have been recognized so far, PCV1, PCV2, PCV3 and PCV4 (Fig. 1). PCV1, PCV2 and PCV3 are present throughout the global pig population, while the distribution of PCV4 is currently unknown. Among all PCVs, the pathogenic PCV2 is the most studied and hence the most information is available for this virus. In general, and based on available data, PCVs have likely been circulating in domestic pigs for an extensive period of time. PCV1 was first recognized in 1974 as contaminant of
Estimated origin and evolutionary rate
An overview of PCVs and CVs in other species based on 137 PCV ORF2 sequences is provided in Fig. 2. For the final data set only 41 positions were valid and results should be interpreted with care.
Based on analysis of sequences availabe in 2009, PCV1 and PCV2 were considered to have evolved from avian CVs, first infecting wild boars and then infecting domestic pigs (Firth et al., 2009). The time to most recent common ancestor (TMRCA) or mean evolutionary divergence of pig and avian CVs was
Phylogenetic diversity
There are only a few studies on the genetic diversity of PCV1, which is in general considered low (Cao et al., 2018; Cortey and Segales, 2012). In fact, a 2020 GenBank search using the search term “PCV1” resulted in 16 hits whereas a search for “PCV2” resulted in 8133 hits, a search for “PCV3” resulted in 726 hits, and a search for “PCV4” resulted in 0 hits. While sequences may have been deposited under different names this provides some insight on which CVs are prioritized by research groups.
Potential to cause cross-species infections
Two studies on PCV1 infection in species other than pigs have been published. In one of these studies conducted in Northern Ireland, PCV1 serum antibodies were not detected in cattle, sheep, chickens, turkeys, goats, mice, rabbits or humans (Allan et al., 1994). In the other study, PCV1 antibodies were detected by a German group in 30.2% of humans tested, 12-69% of mice tested and 35% of cattle tested (Tischer et al., 1995). It has been suggested that the assay utilized for the later study may
Cell culture propagation and development of infectious clones
Key cell culture results for PCV1, PCV2, PCV3 and PCV4 are summarized in Table 1. PCV1 can be readily isolated in cell culture and PK-15 cells (Tischer et al., 1982; Tischer et al., 1987). Multiplication of PCV1 is inducible by glucosamine and is cell cycle dependent i.e. PCV1 needs actively dividing cells (Tischer et al., 1987). Cell culture propagated virus has been used to produce PCV1 inoculum and to experimentally infect pigs (Allan et al., 1995; Tischer et al., 1986). PCV2 can also be
Field observations
In general, clinical signs are not associated with PCV1 infection in pigs (Table 1). While PCV1 can be found in most investigations targeting this PCV genotype, the rate of PCV1 DNA positive pigs is often low (Cao et al., 2018; Puvanendiran et al., 2011). However, PCV1 has been identified in fetuses in the 90s. Specifically, PCV1 was isolated from stillborn piglets and PCV1 antibodies were identified in 10/160 fetal serum samples indicating transplacental infections do occur occasionally (Allan
Coinfections
Coinfections of PCV2 with other pathogens are of great importance (Opriessnig and Halbur, 2012) and this became clear soon after its discovery (Allan et al., 1999; Ellis et al., 1999). PCV2 directly targets cells of the immune system and the virus is considered immunosuppressive (Darwich et al., 2004; Meng, 2013; Segales et al., 2004). Hence, infection by other pathogens is more likely in the presence of PCV2. On the other hand, coinfections with porcine parvovirus (Allan et al., 1999), PRRSV (
Tissue tropism and viral loads
In the 90s, PCV1 antigen was demonstrated using an indirect immunofluorescense assay on spleen, thymus and lung from fetal material collected from field samples and 2-day-old colostrum-deprived piglets experimentally infected with PCV1(Allan et al., 1995).
In early PCV2 studies during the 90s, the presence of PCV2 was confirmed by either ISH (Rosell et al., 1999) or IHC (Sorden et al., 1999), techniques which were developed shortly after PCV2 discovery and have been widely used since to
Economic impact and vaccines
Based on the knowledge accumulated so far, PCV2 has a high economic impact when uncontrolled (Alarcon et al., 2013a; Alarcon et al., 2013b). Currently, many commercial vaccines are available for PCV2. PCV2 vaccines have been shown to be very successful and efficacious (da Silva et al., 2014; Sidler et al., 2012), they are also widely accepted by producers and swine veterinarians (Afghah et al., 2017; Karuppannan and Opriessnig, 2017). Most PCV2 vaccines are based on inactivated PCV2 viruses or
Knowledge gaps
Circoviruses appear to be widespread in wild and domestic pigs. The non-pathogenic nature of PCV1 has been confirmed early on and while PCV1 can be found in pigs, its prevalence is considered low. However, very few research studies have confirmed the absence of pathogenicity and the significance of PCV1 overall (Allan et al., 1995; Fenaux et al., 2003; Tischer et al., 1982). There appears to be a need for further investigations into the ability of PCV1 to cause disease, including coinfection
Conclusions
CRESS viruses are one of the most rapidly expanding class of viruses by discovery, whose host range spans from archaea to mammals, including the cruciviruses (de la Higuera et al., 2019), which are thought to have evolved by recombination between an ancestral CRESS virus and an ssRNA virus (Zhao et al., 2019). Interestingly, the rep genes of eukaryotic CRESS viruses are evolutionary conserved compared to the high diversity of their cap genes (Zhao et al., 2019). PCV2 and PCV3 both have high
Funding
TO acknowledges the Biotechnology and Biological Sciences Research Council (BBSRC) for support of the Roslin Institute Strategic Programme Control of Infectious Diseases (BBS/E/D/20002173 and BBS/E/D/20002174).
Acknowledgments
The authors thank A. A. Mattei for critical review of this manuscript.
References (164)
- et al.
Ten years of PCV2 vaccines and vaccination: Is eradication a possibility?
Vet Microbiol.
(2017) - et al.
Global status of Porcine circovirus type 2 and its associated diseases in sub-Saharan Africa
Adv in Viro
(2017) - et al.
Economic efficiency analysis of different strategies to control post-weaning multi-systemic wasting syndrome and porcine circovirus type 2 subclinical infection in 3-weekly batch system farms
Prev Vet Med
(2013) - et al.
Cost of post-weaning multi-systemic wasting syndrome and porcine circovirus type-2 subclinical infection in England - an economic disease model
Prev Vet Med
(2013) - et al.
Discovery and evolving history of two genetically related but phenotypically different viruses, porcine circoviruses 1 and 2
Virus Res
(2012) - et al.
Porcine circoviruses: a review
J Vet Diagn Invest
(2000) - et al.
Experimental reproduction of severe wasting disease by co-infection of pigs with porcine circovirus and porcine parvovirus
J Comp Pathol
(1999) - et al.
Production, preliminary characterisation and applications of monoclonal antibodies to porcine circovirus
Vet Immunol Immunopathol
(1994) - et al.
Pathogenesis of porcine circovirus; experimental infections of colostrum deprived piglets and examination of pig foetal material
Vet Microbiol
(1995) - et al.
Isolation of porcine circovirus-like viruses from pigs with a wasting disease in the USA and Europe
J Vet Diagn Invest
(1998)
Absence of evidence for porcine circovirus type 2 in cattle and humans, and lack of seroconversion or lesions in experimentally infected sheep
Arch Virol
PCV3-associated disease in the United States swine herd
Emerging microbes & infections
Evaluation of the safety of four porcine circovirus type 2 tissue homogenate vaccines in a pig bioassay
Vet Rec
Retrospective study of porcine circovirus type 2 infection reveals a novel genotype PCV2f
Transbound Emerg Dis
Detection and genetic characterization of porcine circovirus 3 (PCV3) in pigs in India
Transbound Emerg Dis.
Novel circular DNA viruses in stool samples of wild-living chimpanzees
J Gen Virol
House fly vector for porcine circovirus 2b on commercial pig farms
Vet Microbiol
Identification of multiple novel viruses, including a parvovirus and a hepevirus, in feces of red foxes
J Virol
ICTV Virus Taxonomy Profile: Circoviridae
J Gen Virol
Genomic analysis of uncultured marine viral communities
Proceedings of the National Academy of Sciences of the United States of America
Identification of an emerging recombinant cluster in porcine circovirus type 2
Virus Res
Genetic variation analysis of PCV1 strains isolated from Guangxi Province of China in 2015
BMC Vet Res
Detection and genome sequencing of porcine circovirus 3 in neonatal pigs with congenital tremors in South China
Transbound Emerg Dis
Comparative analysis of the transcriptional patterns of pathogenic and nonpathogenic porcine circoviruses
Virology
Transcriptional analysis of porcine circovirus type 2
Virology
Sequence analysis of old and new strains of porcine circovirus associated with congenital tremors in pigs and their comparison with strains involved with postweaning multisystemic wasting syndrome
Can J Vet Res
Further comments on porcine circovirus type 2 (PCV2) genotype definition and nomenclature
Vet Microbiol
Low levels of diversity among genomes of Porcine circovirus type 1 (PCV1) points to differential adaptive selection between Porcine circoviruses
Virology
Mixed treatment comparison meta-analysis of porcine circovirus type 2 (PCV2) vaccines used in piglets
Prev Vet Med
Pathogenesis of postweaning multisystemic wasting syndrome caused by Porcine circovirus 2: An immune riddle
Arch Virol
Genome Sequences of Three Cruciviruses Found in the Willamette Valley (Oregon)
Microbiology resource announcements
Phylogenetic analysis of porcine circovirus type 2 in Sardinia, Italy, shows genotype 2d circulation among domestic pigs and wild boars
Infect Genet Evol
Porcine circovirus type 3 detection in a Hungarian pig farm experiencing reproductive failures
Vet Rec
Investigation of a regulatory agency enquiry into potential porcine circovirus type 1 contamination of the human rotavirus vaccine, Rotarix: approach and outcome
Human vaccines & immunotherapeutics
Isolation of circovirus from lesions of pigs with postweaning multisystemic wasting syndrome
The Canadian veterinary journal = La revue veterinaire canadienne
Reproduction of lesions of postweaning multisystemic wasting syndrome in gnotobiotic piglets
J Vet Diagn Invest
Detection and genetic characterization of Porcine circovirus type 3 in Italy
Transbound Emerg Dis
Genetic characterization of type 2 porcine circovirus (PCV-2) from pigs with postweaning multisystemic wasting syndrome in different geographic regions of North America and development of a differential PCR-restriction fragment length polymorphism assay to detect and differentiate between infections with PCV-1 and PCV-2
J Clin Microbiol
Cloned genomic DNA of type 2 porcine circovirus is infectious when injected directly into the liver and lymph nodes of pigs: characterization of clinical disease, virus distribution, and pathologic lesions
J Virol
Immunogenicity and pathogenicity of chimeric infectious DNA clones of pathogenic porcine circovirus type 2 (PCV2) and nonpathogenic PCV1 in weanling pigs
Journal of virology
Analysis of the subcellular localization of the proteins Rep, Rep’ and Cap of porcine circovirus type 1
Virology
Insights into the evolutionary history of an emerging livestock pathogen: porcine circovirus 2
J Virol
Genetic characterisation of Porcine circovirus type 2 (PCV2) strains from feral pigs in the Brazilian Pantanal: An opportunity to reconstruct the history of PCV2 evolution
Vet Microbiol
Phylodynamic analysis of porcine circovirus type 2 reveals global waves of emerging genotypes and the circulation of recombinant forms
Molecular phylogenetics and evolution
Genotyping Porcine Circovirus 3 (PCV-3) Nowadays: Does It Make Sense?
Viruses
A wild circulation: High presence of Porcine circovirus 3 in different mammalian wild hosts and ticks
Transbound Emerg Dis
A Shift in Porcine Circovirus 3 (PCV-3) History Paradigm: Phylodynamic Analyses Reveal an Ancient Origin and Prolonged Undetected Circulation in the Worldwide Swine Population
Advanced science (Weinheim, Baden-Wurttemberg, Germany)
Full-genome sequencing of porcine circovirus 3 field strains from Denmark, Italy and Spain demonstrates a high within-Europe genetic heterogeneity
Transbound Emerg Dis
Porcine circovirus 2 (PCV-2) genotype update and proposal of a new genotyping methodology
PLoS One
Porcine circovirus type 2 (PCV2) evolution before and after the vaccination introduction: A large scale epidemiological study
Sci Rep
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