Circulating Phylotypes of White Spot Syndrome Virus in Bangladesh and Their Virulence

Abstract

White Spot Syndrome Virus (WSSV) has emerged as one of the most prevalent and lethal viruses globally and infects both shrimps and crabs in the aquatic environment. This study aimed to investigate the occurrence of WSSV in different ghers of Bangladesh and the virulence of the circulating phylotypes. We collected 360 shrimp (Penaeus monodon) and 120 crab (Scylla sp.) samples from the south-east (Cox’s Bazar) and south-west (Satkhira) coastal regions of Bangladesh. The VP28 gene-specific PCR assays and sequencing revealed statistically significant (p < 0.05, Kruskal–Wallis test) differences in the prevalence of WSSV in shrimps and crabs between the study areas (Cox’s Bazar and Satkhira) and over the study periods (2017–2019). The mean Log load of WSSV varied from 8.40 (Cox’s Bazar) to 10.48 (Satkhira) per gram of tissue. The mean values for salinity, dissolved oxygen, temperature and pH were 14.71 ± 0.76 ppt, 3.7 ± 0.1 ppm, 34.11 ± 0.38 °C and 8.23 ± 0.38, respectively, in the WSSV-positive ghers. The VP28 gene-based phylogenetic analysis showed an amino-acid substitution (E→G) at the 167th position in the isolates from Cox’s Bazar (referred to as phylotype BD2) compared to the globally circulating one (BD1). Shrimp PL artificially challenged with BD1 and BD2 phylotypes with filtrates of tissue containing 0.423 × 109 copies of WSSV per mL resulted in a median LT50 value of 73 h and 75 h, respectively. The in vivo trial showed higher mean Log WSSV copies (6.47 ± 2.07 per mg tissue) in BD1-challenged shrimp PL compared to BD2 (4.75 ± 0.35 per mg tissue). Crabs infected with BD1 and BD2 showed 100% mortality within 48 h and 62 h of challenge, respectively, with mean Log WSSV copies of 12.06 ± 0.48 and 9.95 ± 0.37 per gram tissue, respectively. Moreover, shrimp antimicrobial peptides (AMPs), penaeidin and lysozyme expression were lower in the BD1-challenged group compared to BD2 challenged shrimps. These results collectively demonstrated that relative virulence properties of WSSV based on mortality rate, viral load and expression of host immune genes in artificially infected shrimp PL could be affected by single aa substitution in VP28.

Expression of lysozyme in diseased shrimp collected from two different coastal areas

“…Lysozyme release slowed down at pH 7 due to the exchanges between cationic muramidase and non-ionic API or Na-caseinate (Benelhadj et al, 2016). Hasan et al (2022) tested lysozyme in experimentally infected shrimps with different doses and with different phylogroups of the virus and observed variations in average relative expression between the infected shrimps with two groups of virus. Deris et al (2020) observed different patterns of lysozyme gene expressions in shrimp post-larvae (PL) after challenged with pathogenic microorganisms.…”

“…RNA extraction kit (New England Biolabs, Ipswich, MA, USA) was utilized for extracting RNA from diseased and fresh shrimps. PhotoScript II First Strand cDNA Synthesis Kit, (New England Biolabs, Ipswich, MA, USA) was used for the synthesis of cDNA (Hasan et al, 2022).…”

“…Relative expressions of lysozyme gene were observed using real-time PCR. Real-time PCR conditions followed the methods described by Hasan et al, (2022). House-keeping gene (internal control), β-actin, was used for the normalization purpose.…”

“…The economic cost of a particular single disease, known as White Spot Disease on the shrimp farming industry was assessed up to US$15 billion as it initially arose expanding at the rate of US$1 billion yearly (Stentiford et al, 2012). Nevertheless, shrimp farming had developed to be the utmost profitable division in aquaculture businesses worldwide (Hasan et al, 2022). Numerous causative agents are responsible for various shrimp diseases including white spot syndrome virus (WSSV), yellow head virus (YHV), Taura syndrome virus (TSV), and bacteria from the genus Vibrio, which create massive loss in the shrimp aquaculture industry (Lightner, 2011;Tassanakajon et al, 2013).…”

“…Wang et al (2018) found changes in the activity of lysozyme in the in-vivo assays with different salinity stressed conditions. Hasan et al (2022) mentioned lysozyme as a very critical AMP participating in the host-pathogen interactions which may change expression patterns when infected with virus. Therefore, the current study targeted to observe the relative expression of lysozyme in diseased penaeid shrimps collected from Satkhira and Cox's Bazar which are different geographical regions.…”