Introduction

Porcine Parvovirus 1 (PPV1) is one of the prime causative agents associated with SMEDI (stillbirth, mummification, embryonic death, and infertility) syndrome, which causes marked loss to the swine industry worldwide (Mengeling et al. 2000). PPV was first isolated as a cell culture contaminant from a porcine primary cell culture which was used for propagation of classical swine fever virus in Germany during the early 1960s (Mayr and Mahnel 1964). The incidence of PPV associated with abortions in swine was first described by Carwright and Huck (1967). Parvoviruses are small, non-enveloped, single-stranded, and negative-sense DNA virus that belongs to the family Parvoviridae (Molitor et al. 1984). Apart from classical PPV1, six novel porcine parvoviruses (PPV2–PPV7) were described in the past two decades (Palinski et al. 2016). As per International Committee on Taxonomy of Viruses (ICTV) classification, PPV1 belongs to the genus Protoparvovirus, whereas novel parvoviruses PPV2-PPV3, PPV4–PPV6, and PPV7 belong to the genera Tetraparvovirus, Copiparvovirus, and Chapparvovirus respectively (** along with cluster E strains within PPV1 sequences (Fig. 4). NCBI-BLAST analysis of contig sequence from sample INDTNCHNT5 revealed 100% sequence homology with Chinese PPV1genome (MH183297), whereas sequence from sample INDTNCHNL17 revealed 99.43% sequence homology with Spain (MH558678), Chinese (MH183297), and German (JN400516) PPV1 sequences. Deduced amino acid sequence analysis of VP2 coding region of isolates obtained in this study with reference PPV1 amino acid residues revealed variations at position 215 (I to T) in both the isolates, variation at position 228 (Q to E) in T5 isolate and variations at position 59 (L to M) and 314 (K to E) in L17 isolate (Fig. 5).

Fig. 3
figure 3

Amplification of partial VP2 gene of PPV1 using designed primer in this study. Lane 1, 2, 3, 4, 6, and 7- field samples, Lane 5 is 100 bp ladder, Lane 8—known positive DNA, Lane 9- Non-template control,. The amplified product is 897 bp and labeled separately

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Table 2 List of PPV genome genomes used in Phylogentic analysis
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Fig. 4
figure 4

Phylogenetic analysis PPV1 genome by Maximum Likelihood method based on p-distance model with 1000 bootstraps. A Traditional rectangular dendrogram. B Circular dendrogram. The phylogenetic tree was inferred based on alignment of VP2 protein and were midpoint rooted. The trees were drawn with two PPV1 sequences from this study (labeled by red colored circle) along with 42 established PPV1 sequences from the GenBank (including seven PPV sequences from India labeled in blue diamond) one PPV2 sequence as an out-group. The analyses were conducted in MEGA X with Bootstrap replicates of 1000

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Fig. 5
figure 5

Deduced amino acid sequences of partial VP2 gene of PPV1. This alignment included deduced amino acid partial sequences of VP2 protein from two PPV1 sequences from this study (T5 and L17) and seven PPV1 sequences from different countries including one reference PPV1 sequences retrieved from GenBank. All the sequences were aligned in ClustalW and viewed in SanpGene alignment tool. The PPV1 sequences were demarcated by the red colored arrows and aminoacid variations sites were marked by rectangle shape balck box

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Discussion

PPV1 is the well-documented viral pathogen associated with reproductive failure in most swine-producing countries. The detection of PPV by PCR-based molecular technique is highly specific and sensitive in comparison to hemagglutination or immunofluorescence assays (Soares et al. 1999). NS1 gene–specific PCR assay is the best suitable approach for molecular screening of PPV infections (Xu et al. 2012). NS1 gene–based PPV1 surveillance in this study revealed 14.3% positivity which is lower when compared to global prevalence status. The global prevalence of PPV1 varies from 25.8 to 71.88% (Opriessnig et al. 2015). Each PPV1 clusters are subdivided into subgroups that include strains from both domestic and wild swine populations with varying virulence properties related to the amino acid composition of VP2 proteins (Cadar et al. 2012).

Varying amino acid substitutions have been observed in strains from several countries. Hot spots were found to be located on the capsid surface, and a surface profile distinct from the vaccine strains was observed. For PPV1, 12 linear epitopes have been proposed between amino acid positions 5–51, 85–101, 130–140, 154–167, 190–240, 260–314, 272–320, 378–458, 467–478, 502–514, 535–542, and 547–576 in the VP2 proteins numbered serially from 1 to 12, containing a huge potential role, as determined by B cell epitope prediction software. The virulent PPV1 field strains have 5 common amino acid substitutions at I-215-T, D-378-G, H-383-Q, S-436-P, and R-565-K when compared to non-virulent strains. Substitutions at 378, 383, and 436 amino acid positions of 8th B-cell epitope region determine the tissue tropism of PPV1 (Chung, et al. 2020).

Both the PPV1 (T5 and T7) sequences in this study were obtained from tissues of stillborn and mummified fetuses of swine and had amino acid substitutions at 215 positions similar to that of virulent PPV1 isolates. Additionally, one of the Tamil Nadu PPV1 sequences (T5) had host immune evasion mutations at VP2 amino acid position 228-E as like that of German 27a (AY684871) virulent field isolate (Zeeuw et al. 2007). Based on the above said evidence, the two PPV1 sequences (T5 and L17) in this study are characterized as pathogenic strains. It is hypothesized that variations in the amino acid composition of capsid could be due to viral adaptation to host and or vaccinal immune response refereed as escape mutants (Cadar et al. 2012). Commercial PPV1 (whole virus inactivated) vaccines are derived from attenuated NADL2 strains used in many countries prevent only reproductive loss and do not eliminate virus infection and dissemination (Mengeling et al. 1980). The emergence of new clusters of PPV1 in the field with vast divergence from vaccine strain necessitates the development of an alternative vaccine development approach involving suitable candidate vaccine strain circulating in the field (Streck et al. 2015). Therefore, the emergence and spread of viruses with varying amino acid profiles require close surveillance. Molecular characterization and phylogenetic analysis of PPV in India are scarce. The molecular detection of the PPV genome in the swine population of Tamil Nadu has not been reported so far. This study documents incidence of PPV1 cluster E strains for the first time in Tamil Nadu. The PPV1 isolates in this study showed homology to China and European countries isolates and these findings are supported by Cadar et al. (2012), who found that PPV1 cluster E includes the highly virulent Kresse strain along with the challenge UK and Brazilian strains of PPV. Although it could be hypothesized that these phylogenetic clustering might be due to live pig import from China and European countries to India. Additionally, investigations in contaminated commercial biological products, as porcine cell lines associated vaccines, may elucidate the origin and route of transmission of PPV across the countries. This is the foremost molecular characterization report of PPV documented from Tamil Nadu. To determine the prevalence, transmission, molecular epidemiology, and impact of PPV in commercial swine husbandry, it is necessary to extend this study to larger populations. Currently, the PPV is a very less-explored pathogen with no complete scientific data available in Indian context; hence, there is no indigenous vaccine available to control this infection; in general, the current control measures are at primitive level by adopting general hygiene.