Genetic diversity and recombination of enterovirus G strains in Japanese pigs: High prevalence of strains carrying a papain-like cysteine protease sequence in the enterovirus G population

Abstract

To study the genetic diversity of enterovirus G (EV-G) among Japanese pigs, metagenomics sequencing was performed on fecal samples from pigs with or without diarrhea, collected between 2014 and 2016. Fifty-nine EV-G sequences, which were >5,000 nucleotides long, were obtained. By complete VP1 sequence analysis, Japanese EV-G isolates were classified into G1 (17 strains), G2 (four strains), G3 (22 strains), G4 (two strains), G6 (two strains), G9 (six strains), G10 (five strains), and a new genotype (one strain). Remarkably, 16 G1 and one G2 strain identified in diarrheic (23.5%; four strains) or normal (76.5%; 13 strains) fecal samples possessed a papain-like cysteine protease (PL-CP) sequence, which was recently found in the USA and Belgium in the EV-G genome, at the 2C–3A junction site. This paper presents the first report of the high prevalence of viruses carrying PL-CP in the EV-G population. Furthermore, possible inter- and intragenotype recombination events were found among EV-G strains, including G1-PL-CP strains. Our findings may advance the understanding of the molecular epidemiology and genetic evolution of EV-Gs.

First detection of novel enterovirus G recombining a torovirus papain‐like protease gene associated with diarrhoea in swine in South Korea

“…Furthermore, a previous study using reverse genetics revealed that the US EV‐G17‐PLCP strain functionally produces the exogenous PLCP gene in virus‐infected cells, demonstrating its own ALFQ↓GPPV and AEFQ↓GPPT sequences as the putative cleavage sites (Shang et al., ). Considering the sequence similarity of the putative cleavage sites including GPPT−ALFQ, GPPA−ALFQ, and GPPE−ALPQ among global strains (Conceição‐Neto et al., ; Knutson et al., ; Tsuchiaka et al., ), therefore, the cleavage of the inserted PLCP gene appears to be guaranteed using each corresponding cleavage sequence. Consistently, the recombinant ToV‐PLCP of the EV‐G KNU‐1811 strain is bordered by two analogous predicted 3C pro cleavage sites, ALFQ↓GPPV and AVFQ↓GPPT, at its N and C termini, respectively, indicating its proteolytic processing by 3C pro into a functional product (Figure a).…”

A novel defective recombinant porcine enterovirus G virus carrying a porcine torovirus papain-like cysteine protease gene and a putative anti-apoptosis gene in place of viral structural protein genes

“…We call them type 1 recombinant EV-G in this study. Our previous data demonstrated high prevalence of type 1 recombinant EV-G in the EV-G population (Tsuchiaka et al, 2018). Our previous data demonstrate high https://doi.org/10.…”

“…Overlapping PCR of type 2 recombinant EV-G. RNAs were re-extracted in this study from the pig feces using High Pure Viral Nucleic Acid Kit (Roche), and cDNA was synthesized with random primers using SuperScript III First-Strand Synthesis System (Invitrogen), as described previously (Tsuchiaka et al, 2018). Primers for PCR (Table 1) were designed to amplify the viral genome at approximately every 1 kb with overlap regions.…”

“…Our previous data demonstrate high https://doi.org/10. 1016/j.meegid.2019.103975 Received 18 April 2019; Received in revised form 17 July 2019; Accepted 19 July 2019 prevalence of type 1 recombinant EV-G in the EV-G population (Tsuchiaka et al, 2018). Type 1 recombinant EV-Gs have been discovered from feces of neonatal pigs showing clinical symptoms, such as diarrhea, in the US, Belgium, Germany and South Korea (Bunke et al, 2018;Conceição-Lee and Lee, 2019;Neto et al, 2017;Knutson et al, 2017;Shang et al, 2017), while type 1 recombinant EV-Gs identified in Japan are detected from normal as well as diarrhea pig feces (Tsuchiaka Fig.…”

“…Schematic diagram of the genome organization of EV-G, type 1 and type 2 recombinant EVGs. Genome order of EV-G, type 1 and type 2 recombinant EV-G came from the EVG/Porcine/JPN/Iba464-3-1/2015/G1 (Tsuchiaka et al, 2017), EVG/Porcine/JPN/MoI2-1-1/2015/G1-PL-CP (Tsuchiaka et al, 2018), and a newly identified type 2 EV-G (EVG/Porcine/JPN/MoI2-1-2/2015/type 2) in this study, respectively.…”