Molecular characterization of channel catfish virus (CCV) isolated from cage-cultured pangasius hypophthalmus farmed in Pahang River, Malaysia

Abstract

Pangasius hypophthalmus (Sauvage, 1878), is a freshwater fish originated from Mekong River, which currently cultivated in most Southeast Asia countries including Malaysia. In Malaysia, these species contributed more than 20% of the whole aquaculture production from freshwater culture system. However, viral infection in Pangasius may cause high mortality in cage-cultured farm. This study aims to characterize the local isolates of channel catfish virus (CCV) and determine the cytopathic effect (CPE) formation of local CCV in brown bullhead (BB) and channel catfish ovary (CCO) cell lines. Samples of liver, kidney and spleen were collected every two weeks in three selected P. hypophthalmus cage-cultured along Pahang River, Temerloh, Pahang. A total of 250 samples were collected from three cage-cultured farm which are 90 samples from Kuala Krau 1, 50 samples from Kuala Krau 2 and 110 samples from Kg Pangsenam. All of the samples were analysed using polymerase chain reaction (PCR) for CCV (ORF27 primers). The PCR analysis showed that 17.6% of the samples were CCV positive, where Farm Kg. Pangsenam placed first with 27.3% followed by Farm 2 Kuala Krau with 12.0% and lastly Farm 1 Kuala Krau with only 8.9% prevalence of CCV detected. It was found that CCV prevalence in Farm Kg. Pangsenam was significantly higher (p< 0.01) in the cage-cultured from Pahang River downstream because of the low pH value. In addition, the CCV prevalence was also significantly higher (p<0.01) during dry season because CCV outbreaks occur at high water temperature (25-33oC). However, the CCV prevalence detected in this study show no significant difference (p>0.05) with the fish bodyweight. Sequence analysis of the amplified band (590-bp) using NCBI-BLAST showed that local CCV sequence had 100% similarity with M75136.2 Ictalurid herpesvirus-1 (IcHV1) strain Auburn. Next, the local CCV-positive samples were inoculated in BB and CCO cells for CPE formation observation and susceptibility test. As a result, CPE progression of local CCV in CCO cells took 10 days of post inoculation (dpi) to achieve 80% CPE. The pyknotic cells and loosed network of syncytia were observed and monolayer cell start to detach from the vessel surface. While in BB cells, the normal cells start to rounded and granulated, lead to formation gigantic cell followed by cells detach from surface in 14 dpi. This study shows CPE formation occurs more rapidly in CCO cells than in BB cells. This finding can be applied for early detection and future disease surveillance of CCV infection in P. hypophthalmus.