Hepatopancreatic Parvovirus Research Articles

Parvoviruses of Aquatic Animals.

Family Parvoviridae consists of small, non-enveloped viruses with linear, single-stranded DNA genomes of approximately 4-6 kilobases, subdivided into three subfamilies, Parvovirinae, Densovirinae, and Hamaparvovirinae, and unassigned genus Metalloincertoparvovirus. Parvoviruses of aquatic animals infect crustaceans, mollusks, and finfish. This review describes these parvoviruses, which are highly host-specific and associated with mass morbidity and mortality in both farmed and wild aquatic animals. They include Cherax quadricarinatus densovirus (CqDV) in freshwater crayfish in Queensland, Australia; sea star-associated densovirus (SSaDV) in sunflower sea star on the Northeastern Pacific Coast; Clinch densovirus 1 in freshwater mussels in the Clinch River, Virginia, and Tennessee, USA, in subfamily Densovirinae; hepatopancreatic parvovirus (HPV) and infectious hypodermal and hematopoietic necrosis virus (IHHNV) in farmed shrimp worldwide; Syngnathid ichthamaparvovirus 1 in gulf pipefish in the Gulf of Mexico and parts of South America; tilapia parvovirus (TiPV) in farmed tilapia in China, Thailand, and India, in the subfamily Hamaparvovirinae; and Penaeus monodon metallodensovirus (PmMDV) in Vietnamese P. monodon, in unassigned genus Metalloincertoparvovirus. Also included in the family Parvoviridae are novel parvoviruses detected in both diseased and healthy animals using metagenomic sequencing, such as zander parvovirus from zander in Hungary and salmon parvovirus from sockeye salmon smolts in British Columbia, Canada.

Nucleic acid amplification‐based methods for diagnosis of shrimp viral diseases

AbstractViral diseases are one of the constraints affecting the sustainability of the shrimp farming industry worldwide. The common causes of viral infections include white spot syndrome virus, infectious hypodermal and haematopoietic necrosis virus, hepatopancreatic parvovirus, Penaeus monodon nudivirus, decapod iridescent virus 1, Taura syndrome virus, yellow head virus, infectious myonecrosis virus and Macrobrachium rosenbergii nodavirus. Accurate diagnostic methods are necessary for preventing the spread of these transboundary diseases. Many molecular diagnostic methods for shrimp viral diseases have been developed and are currently in use, including conventional polymerase chain reaction (PCR), nested PCR, loop‐mediated isothermal amplification PCR, recombinase polymerase assay and real‐time PCR. Although the World Organization for Animal Health (founded as OIE) recommends several PCR methods for shrimp disease diagnosis, there has been a lack of efforts to comparatively evaluate the performance of the available methods, resulting in a significant knowledge gap. Therefore, in this review, we compare various PCR assays and commercial PCR‐based diagnostic kits that have been developed and/or published previously, and discuss pros and cons of each of these methods. Our objective of this review is to offer enhanced clarity and a comprehensive overview of molecular diagnostic methods for shrimp viral diseases. This endeavour would serve as an important resource for researchers engaged in relevant studies and professionals within the shrimp‐farming industry, providing valuable guidance and insights.

Validation of qPCR from a crude extract for the rapid detection of white spot syndrome virus.

White spot disease (WSD) has posed a serious threat to the China and the global shrimp aquaculture. In order to diagnose white spot syndrome virus (WSSV) early and prevent the spread and outbreak of WSD, it is necessary to establish a highly sensitive WSSV diagnosis method suitable for shrimp farming sites. In this study, a pre-amplification qPCR assay from the crude extract of samples heated lysis was established, which was further compared with the universal qPCR assay to verify the shrimp samples. The limit of detection (LOD) of pre-amplification qPCR assay and universal qPCR assay was 2.80 copies and 20.57 copies per reaction at 95% CI, respectively. It had good WSSV specificity and did not show cross-detection of infectious hypodermal and hematopoietic necrosis virus (IHHNV), hepatopancreatic parvovirus (HPV), Enterocytozoon hepatopenaei (EHP), acute hepatopancreas necrosis disease (AHPND), necrotizing hepatopancreatitis bacteria (NHPB), and decapod iridescent virus 1 (DIV1). A total of 36 shrimp samples were detected as WSSV DNA positive by pre-amplification qPCR with crude extract from samples heated lysis and universal qPCR with DNA extraction. The diagnostic sensitivity and specificity were 97.22% (85.5 ~ 99.9%, 95% CI) and 100% (81.5 ~ 100%, 95% CI), respectively. The agreement Kappa value was 0.959 (0.879 ~ 1, 95% CI), and the analysis results were basically consistent. Eliminating the tedious steps of extracting DNA and using pre-amplified qPCR to detect WSSV in shrimp, it is a good choice for aquaculture farms.Graphical abstract Supplementary InformationThe online version contains supplementary material available at 10.1007/s10499-022-00920-9.

Concurrent infection of a novel genotype of hepatopancreatic parvovirus and Enterocytozoon hepatopenaei in Penaeus vannamei in Taiwan.

Hepatopancreatic parvovirus (HPV) and Enterocytozoon hepatopenaei (EHP) are emerging and reemerging pathogens in shrimps. In the present study, a novel genotype of HPV concurrently infected with EHP in Penaeus vannamei in Taiwan leading to severe atrophy and damage of hepatopancreas were confirmed by histopathology, in situ hybridization, and PCR. The novel genotype of HPV exhibited 66%-69.5% sequence identities with all known HPVs and carried unique amino acid deletions and insertions in the VP gene. According to phylogenetic analysis, the Taiwan HPV isolates were classified as the genotype IV. The present study not only provided the histopathological and molecular proof of HPV and EHP co-infection in Taiwan, but also revealed the importance of investigating the geographical expansion of novel HPV genotypes.

Multivariate genomic prediction for commercial traits of economic importance in Banana shrimp Fenneropenaeus merguiensis

Advantages of multi-trait machine and deep learning genomic prediction models for quantitative complex traits have not been documented or very limited in aquaculture species. Thus, the present study sought to understand effects of the multi-trait single-step genomic best linear unbiased prediction (ssGBLUP), Bayesian (BayesCpi), random forest (RF) and multilayer perceptron (MLP) models on genomic prediction accuracies for traits of commercial importance in banana white shrimp (Fenneropenaeus merguiensis). Our analyses were conducted in a breeding shrimp population comprising 562 individuals (offspring of 48 parental pairs) genotyped for 9472 single nucleotide polymorphisms (SNPs) and the animals had full phenotype records for five important traits (i.e., body weight, abdominal width, tail weight, raw colour of live shrimp and resistance to hepatopancreatic parvovirus). In both univariate and multi-trait analyses, machine (RF) and deep learning (MLP) models outperformed ssGBLUP for all traits studied. However, they had similar predictive performance to BayesCpi. The benefits of the multivariate relative to univariate models were trait- and method-specific. Multi-trait BayesCpi increased the prediction accuracies for growth (weight and width), carcass (tail weight) and HPV resistance by 9.3 to 17.8%. However, the multi-trait random forest models improved the predictive power for only abdominal width (14.3%) and disease resistance to hepatopancreatic parvovirus (10.0%). When the multi-trait MLP was used, the improvements in the prediction accuracies were observed for abdominal width and raw colour (4.9 and 6.0%, respectively). There were almost no differences in the predictive power between univariate and multi-trait ssGBLUP. Among the multi-trait models used, BayesCpi outperformed other methods (ssGBLUP, RF and MLP). It is concluded that either BayesCpi or machine and deep learning-based multi-trait genomic prediction models should be employed in large-scale genetic enhancement programs for banana shrimp. These approaches show enormous potential to enhance genetic progress made in this population of banana shrimp and potentially for other aquaculture species.

Environmental and Socioeconomic Impacts of Shrimp Farming in the Philippines: A Critical Analysis Using PRISMA

Shrimp aquaculture is under pressure to increase its production to meet the growing demand for food from a growing population. In the Philippines, aquaculture has experienced the shift from milkfish to prawn, with its attractive marketable price. This intensification has led to negative and positive impacts, which have raised a range of environmental and socioeconomic problems. This paper reviews the environmental and socioeconomic challenges that the shrimp aquaculture industry faces using the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) method. We examine the gaps and the changes that are required to revitalize the industry. We examine and assess the impacts of shrimp culture on the environment, e.g., shrimp farm management, marine pollution, disease outbreaks, and the social, economic, and climate change impacts. The presence of viral diseases, such as White Spot Syndrome Virus (WSSV), Monodon Baculovirus (MBV), Infectious Hypodermal and Hematopoietic Necrosis Virus (IHHNV), Hepatopancreatic Parvovirus (HPV), and Yellow Head Virus (YHV), have caused approximate losses in the industry of 40,080 mt in 1997, and 51,000 mt in 2014. Recommended strategies and policy changes are considered for the improvement of shrimp aquaculture, including disease management, the adoption of good aquaculture practices, proper environmental monitoring, sustainable practices at the farm level, and priorities for cooperation among the concerned government agencies and local governments, as well as the involvement of state universities and colleges, for better management practices.

Hepatopancreatic parvovirus disease

This datasheet on hepatopancreatic parvovirus disease covers Identity, Associated Diseases, Pests or Pathogens, Hosts/Species Affected.

Prevalence of disease outbreak from cultured whiteleg shrimp Penaeus vannamei farms located in Karnataka, India

Penaeus vannamei farming is one of the profitable businesses of the aquaculture sector in India. As a result of excessive farming, shrimps were susceptible to various exotic, transboundary, and emerging pathogens. The purpose of this study is to monitor the prevalence of shrimp pathogens in Karnataka. A total of 91 samples of P. vannamei post larvae and juveniles were collected from April 2022 to November 2022 in Karnataka. Samples were screened for pathogens as listed in Office of Internationale des Epizootics (OIE) or World Organisation for Animal Health (WOAH) which include infectious hypodermal and hematopoietic necrosis virus (IHHNV), Enterocytozoon hepatopenaei (EHP), acute hepatopancreatic necrosis disease (AHPND), hepatopancreatic parvovirus (HPV), infectious myonecrosis virus (IMNV), yellow head virus (YHV), taura syndrome virus (TSV), white spot syndrome virus (WSSV), and other diseases such as decapod iridescent virus-1(DIV-1), and monodon baculovirus (MBV). Out of 91 samples, 5 (5.5%) samples were positive for WSSV, 26 (28.6%) samples were positive for EHP, and 2 (2.2%) samples had co-infection caused by EHP and WSSV. In this study, we have reported a high prevalence of EHP than WSSV in all three coastal districts of Karnataka. Farmers were using specific pathogen-free (SPF) seeds for culture but still, their cultures are getting infected with the same pathogens which indicates poor pond preparation and bio-security. So we strongly recommend that farmers have to follow good management practices and bio-security to increase the productivity and sustainability of P. vannamei farming in India.

Monodon baculovirus (MBV) infects wild mud crab, Scylla serrata

During a survey of farmed and wild crustaceans from India for viruses, spherical baculovirosis otherwise known as Penaeus monodon-type baculovirus (MBV) was detected in field-collected juvenile/sub-adult mud crab, Scylla serrata using a nested polymerase chain reaction (PCR)-based amplification of the hepatopancreatic DNA. Eight out of 115 mud crab (7.0%) examined during the study were found to be positive in the nested PCR resulting in a 361 nt amplicon. Mud crab, S. olivacea and other crustaceans such as marine crab, Portunus sanguinolentus and farmed penaeid shrimp, Penaeus vannamei and P. monodon were tested negative for the virus. Further, degenerate primers reported to amplify polyhedrin protein gene of MBV also showed PCR amplification in one of the MBV-positive crab samples resulting in a 250 nt amplicon. Sequencing of the two target amplicons (MBV- 361 nt and MBV polyhedrin – 216 nt) revealed more than 97.5 % and 92.8% sequence identity, respectively with the Penaeus monodon nudivirus and Penaeus monodon nucleopolyhedrovirus (MBV) reported from shrimp. Further, histological analysis of mud crab revealed nuclear hypertrophy, chromatin margination and intranuclear eosinophilic/basophilic inclusions in tubule epithelium of hepatopancreas. The hepatopancreatic tissue also showed unusually large, eosinophilic/basophilic inclusion-like structures. These inclusions resembled the viral inclusions reported from S. serrata from Australia. This is the first record of monodon-type baculovirus from a crab host and the second from a non-penaeid crustacean. Interestingly, some of the crab samples also showed deeply basophilic intranuclear inclusion-like bodies resembling hepatopancreatic parvovirus group of viruses (HPV). However, none of the crab samples subjected to PCR amplification using HPV-specific primers showed any amplification. The histological observations made in the present study indicate the possibility of the presence of two hepatopancreas-infecting viruses in S. serrata from India.

Development of a PCR assay for the effective detection of Enterocytozoon hepatopenaei (EHP) and investigation of EHP prevalence in Shandong Province, China

Enterocytozoon hepatopenaei (EHP), a recently reported pathogen in the penaeid shrimp, is spreading widely and seriously threatening Penaeus (Litopenaeus) vannamei aquaculture. This study aimed to develop a new and more sensitive polymerase chain reaction (PCR) method for the effective detection of EHP. An EHP PCR assay with a pair of primers specifically amplifying a 358 bp EHP DNA fragment was developed, which was demonstrated to be capable of detecting as low as 2 × 101 copies of EHP and is specific for EHP without cross reaction with DNA samples prepared from five common shrimp pathogens, including white spot syndrome virus (WSSV), infectious hypodermal and haematopoietic virus (IHHNV), hepatopancreatic parvovirus (HPV), infectious myonecrosis virus (IMNV), and yellow head virus (YHV). This new assay is more specific and more sensitive than the previously published EHP PCR methods. With the PCR assay developed in this study, we investigated the prevalence of EHP in four areas of Shandong, China by testing a total of 639 shrimp samples collected from Yantai, Binzhou, Dongying, and Weifang. The results showed that the EHP positive rate reached 51.2%, indicating that EHP is prevalent in shrimp culture in China.

Multiple infections of Entrerocytozoon hepatopenaei and Hepatopancreatic parvovirus in pond-reared Penaeus vannamei in India

Outbreaks of infectious disease affecting Penaeus vannamei (Pacific white shrimp), the most commonly cultured shrimp species in India, has led to severe economic loss. Five randomly selected P. vannamei samples were collected from 154 grow-out ponds located at Cuddalore and Nagapattinam Districts in Tamil Nadu where shrimp showed severe growth retardation over a two-year period. In addition to growth retardation, affected shrimp displayed an atrophied hepatopancreas (HP) and a white/empty gut and stomach. Histopathology revealed the various developmental stages (plasmodia to mature spores) of massive acidophilic microsporidian spores in the cytoplasm of hepatopancreatic (HP) tubule cells. Some of the microsporidian-infected shrimp also showed distinct hypertrophied nuclei within the HP tubular epithelial cells. TEM revealed different growth stages (meront to mature spores) of intracytoplasmic microsporidian parasites as well as icosahedral viral-like particles (VLP) ranging in diameter from 22 to 24 nm in the cytoplasm and nucleus of HP tubular epithelial cells and extracellular spaces. Among the pooled (five shrimp from each farm) HP samples from 154 farms, 53 (34.4%) farms were identified to be PCR-positive for Enterocytozoon hepatopenaei (EHP), whereas 11 (7.1%) were identified to be PCR-positive for Hepatopancreatic parvovirus (HPV). Among the PCR positives, eight (5.2%) farms were found to be positive for both EHP and HPV, indicating that some P. vannamei had dual infections with EHP and HPV. This is the first such report on the simultaneous occurrence of dual infections caused by EHP and HPV.

Viral contamination of tiger shrimp Penaeus monodon broodstock in Bangladesh

Stocking specific-pathogen-free (SPF) shrimp postlarvae (PL) has been reported to have much potential to enhance production by notably reducing the infection-driven mortality. In terms of SPF PL production in Bangladesh, except white spot syndrome virus (WSSV), several potential viruses such as hypodermal and hematopoietic necrosis virus (IHHNV), monodon baculovirus (MBV), hepatopancreatic parvovirus (HPV), and yellow head virus (YHV) which may synergistically cause mortality are ignored. Therefore, this study aimed to investigate the viral contamination in hatchery-brood-shrimps in Bangladesh as the broods horizontally and/or vertically transfer the viruses to their offspring. Five randomly selected representative hatcheries were investigated over 3 months (January to March) to know the status of viral contamination. Broods in all the hatcheries were found to be contaminated with WSSV, IHHNV, MBV, HPV, and YHV. The results showed that the prevalence of WSSV, MBV, HPV, and YHV did not significantly differ between hatcheries. The highest (72%) and the lowest (30%) prevalence of MBV and WSSV, respectively, were recorded in January. In February, the highest (81%) and the lowest (28%) prevalence were found in MBV and HPV, respectively, where the highest and the lowest prevalence were 88% and 40% for IHHNV and HPV, respectively, in March. The prevalence of IHHNV and MBV were not significantly different. The relative prevalence of WSSV, HPV, and YHV were significantly low compared with that of IHHNV and MBV. Notably, increasing water temperature from January (22 ± 0.71 °C) to March (27 ± 1 °C) increased the prevalence of all the viruses and subsequently the extremity of infection. The contamination in broods forwarding to the production of multiple-virus-contaminated PL is considered as an obstacle to higher production of shrimp at farm level in Bangladesh. Hence, the present study suggests focusing on the deleterious-virus-free shrimp broods in commercial shrimp hatcheries in Bangladesh.

Genomic prediction for disease resistance to Hepatopancreatic parvovirus and growth, carcass and quality traits in Banana shrimp Fenneropenaeus merguiensis

Conventional genetic improvement of disease resistance in aquatic animal species involves challenge tests or using qPCR to quantify viral load that is costly, time-consuming and causing biosecurity concerns. Recent developments in high throughput next generation genome sequencing platforms such as genotyping by sequencing (GBS) have opened new possibilities for improving disease traits based on DNA information. The principal aim of this study was thus to examine potential application of genomic selection to improve resistance to hepatopancreatic parvovirus (HPV) in banana shrimp Fenneropenaeus merguiensis. Specifically, we used a total of 9472 single nucleotide polymorphisms (SNPs) developed de novo from GBS platforms to assess accuracy of genomic prediction for HPV resistance and growth, carcass and quality-related traits in this white shrimp species. Our multi-locus mixed model analysis showed moderate heritabilities for HPV resistance (h2 = 0.46) and other traits studied (0.10 to 0.55). Genetic correlations of HPV titre with growth and carcass traits, estimated using SNPs markers, were negative (i.e., favourable), suggesting that selection for improved growth and carcass traits may have increased HPV resistance (i.e., reduced HPV titre). More importantly, our gBLUP model demonstrated that the accuracy of gBLUP prediction was moderate for HPV disease resistance (0.46). The genomic prediction accuracy was somewhat greater for growth and carcass related traits especially for body weight (0.76) and meat or tail weight (0.77). On the other hand, the prediction accuracy was from 0.25 to 0.41 for quality traits (raw and cooked colour and flesh streaks). Collectively, it is concluded that there are prospects to apply genomic selection in the genetic improvement for increased disease resistance, carcass and quality-related traits in this population of banana shrimp F. merguiensis.

Diversity of single-stranded DNA containing viruses in shrimp.

Over the past four decades, shrimp aquaculture has turned into a major industry providing jobs for millions of people worldwide especially in countries with large coastal boundaries. While the shrimp industry continues to expand, the sustainability of shrimp aquaculture has been threatened by the emergence of diseases. Diseases caused by single-stranded DNA containing viruses, such as infectious hypodermal and hematopoietic necrosis virus (IHHNV) and hepatopancreatic parvovirus (HPV), have caused immense losses in shrimp aquaculture since the early 1980s. In fact, the disease outbreak in the blue shrimp (Penaeus stylirostris) caused by IHHNV in early 1980s ultimately led to the captive breeding program in shrimp being shifted from P. stylirostris to the white shrimp (Penaeus vannamei), and today P. vannamei is the preferred cultured shrimp species globally. To date, four single-stranded DNA viruses are known to affect shrimp; these include IHHNV, HPV, spawner-isolated mortality virus (SMV) and lymphoidal parvo-like virus (LPV). Due to the economic losses caused by IHHNV and HPV, most studies have focused on these two viruses, and only IHHNV is included in the OIE list of Crustacean Diseases. Hence this review will focus on IHHNV and HPV. IHHNV and HPV virions are icosahedral in morphology measuring 20-22nm in size and contain a single-stranded DNA (ssDNA) of 4-6kb in size. Both IHHNV and HPV are classified into the sub-order Brevidensoviruses, family Densovirinae. The genome architecture of both viruses are quite similar as they contain two completely (as in IHHNV) or partially overlapping (as in HPV) non-structural and one structural gene. Histopathology and polymerase chain reaction (PCR)-based methods are available for both viruses. Currently, there is no anti-viral therapy for any viral diseases in shrimp. Therefore, biosecurity and the use of genetically resistant lines remains as the corner stone in the management of viral diseases. In recent years, gene silencing using the RNA interference (RNAi) approach has been reported for both IHHNV and HPV via injection. However, the delivery of RNAi molecules via oral route remains a challenge, and the utility of RNAi-based therapy has yet to be materialized in shrimp aquaculture.

Investigation on the infectious nature of Running Mortality Syndrome (RMS) of farmed Pacific white leg shrimp, Penaeus vannamei in shrimp farms of India

Shrimp aquaculture in India has made significant strides especially after the introduction of Pacific white shrimp in the year 2009. However, intensification of culture practice has exacerbated several disease issues. Apart from infectious diseases, the Indian shrimp aquaculture has been witnessing certain syndromes, affecting productivity. One such syndrome associated with significant morbidity and mortality is popularly termed as running mortality syndrome (RMS) by shrimp farmers. Since 2011, RMS has been widely prevalent in the shrimp farms in Andhra Pradesh (AP) and Tamil Nadu (TN). The affected shrimp show patches of whitish musculature in the abdominal segments as a clinical sign with continuous low-level mortalities, especially after about 35–40 days of shrimp culture. We tried to investigate if this kind of mortality is infectious in nature. Investigations conducted in 34 farms comprising 25 RMS affected and 9 healthy farms were tested negative for all the major OIE listed and other pathogens such as white spot syndrome virus (WSSV), infectious hypodermal and hematopoietic necrosis virus (IHHNV), monodon baculovirus (MBV), hepatopancreatic parvo virus (HPV), infectious myonecrosis (IMNV), Taura syndrome virus (TSV), yellow head virus (YHV), and Penaeus vannamei noda virus (PvNV). Bacteriological examination of haemolymph and hepatopancreas of RMS affected shrimp showed the predominance of Vibrio spp., such as Vibrio parahaemolyticus and Vibrio azureus. Histopathological examination of the hepatopancreas was found to be largely normal, except for haemocytic infiltration in the abdominal segments. Hemolymph of affected shrimp was observed to have less concentration of major and trace minerals than healthy ones. Bioassay through feeding RMS affected shrimp tissue to healthy shrimp and co-habitation experiment of healthy shrimp with the affected animals failed to induce RMS. When maintained in water with optimal physico-chemical parameters, affected shrimps showed recovery within 6–7 days. On the other hand, environmental parameters of pond waters such as total ammonia nitrogen, nitrite nitrogen and turbidity were relatively higher than the optimal values in RMS affected farms. Multiple correspondence statistical analysis of critical factors indicated running mortality to be associated with high stocking densities, high nitrite-N, and high turbidity. Though the study could not identify any specific known aetiological agent associated with RMS affected shrimp, failure to reproduce the syndrome by bioassay, the recovery of affected shrimp under the optimal environmental conditions and the positive correlation with critical environmental parameters and the stocking densities to mortality rates clearly suggest RMS to be a pond ecosystem or pond management associated syndrome rather than infectious in nature and thus can be overcome through best management practices.

Complete Genome Sequences of Four Major Viruses Infecting Marine Shrimp in Egypt.

The genome sequences of four economically important shrimp viruses, Penaeus stylirostris densovirus 1, hepatopancreatic parvovirus, yellow head virus, and gill-associated virus, are reported here. Genome data are fundamental for epidemiological studies in determining the origins of these viruses detected for the first time in Egypt and in developing disease management strategies.

A novel real-time PCR approach for detection of infectious hypodermal and haematopoietic necrosis virus (IHHNV) in the freshwater crayfish Procambarus clarkii.

Infectious hypodermal and haematopoietic necrosis virus (IHHNV) infects many crustacean hosts, including cultured penaeid shrimp. In the present study, we aimed to develop a novel sensitive SYBR Green-based real-time PCR method to specifically amplify DNA fragments of IHHNV. Our newly developed real-time PCR method with a 195-bp amplicon specifically detected IHHNV and showed no cross reaction with white spot syndrome virus (WSSV), hepatopancreatic parvovirus (HPV), Enterocytozoon hepatopenaei (EHP), infectious myonecrosis virus (IMNV) and yellow-head virus (YHV). This method could detect as low as one single copy of IHHNV plasmid DNA, more sensitive than other SYBR Green-based real-time PCR methods and less expensive and more convenient than the TaqMan probe-based real-time PCR. Moreover, our data using the newly designed method showed that 80% of IHHNV-fed Procambarus clarkii samples were IHHNV positive. Our findings further confirmed that P. clarkii can be infected by IHHNV.

Survey of viral and bacterial pathogens in ornamental aquatic crustaceans imported into South Korea

The annual rate of ornamental aquatic crustacean importation is increasing globally. The continuous high demand for live ornamental aquatic crustaceans in many countries has also increased the frequency of imported exotic pathogens, which has caused many problems. In the present study, we examined the monthly distribution of viral and bacterial pathogen genes in ornamental aquatic crustaceans imported into South Korea during a 6-month period from February to July 2016. Among the bacterial pathogens identified, Vibrio alginolyticus was detected in cleaner shrimp in June, and Photobacterium damselae was detected in cleaner shrimp in July. Vibrio penaeicida was detected in peppermint shrimp, banded coral shrimp, hinge-beak shrimp, and orange swamp crayfish between February and June. Among the viral pathogens identified, spawner mortality virus was detected in cherry shrimp in May. Finally, hepatopancreatic parvovirus was detected in both cherry shrimp and orange swamp crayfish between February and March. Notably, the genes of two viral pathogens were also detected in cherry shrimp. Although the viruses were not isolated, they are very likely to have been introduced into South Korea and have an influence on Korean aquaculture. In South Korea, HPV is already prevalent and has been reported several times in aquaculture and in the wild; however, SMV infection has not been reported to date. These results demonstrate that many bacterial and viral pathogens are regularly introduced into South Korea via the ornamental aquatic-crustacean trade.

Detection and quantification of shrimp hemocyte iridescent virus by TaqMan probe based real-time PCR.

Shrimp hemocyte iridescent virus (SHIV) is a recently reported shrimp virus, which threats the cultured white-leg shrimp Litopenaeus vannamei and can cause huge economic loss in shrimp farming industry in China. A quantitative real time polymerase chain reactio (qPCR) assay was developed using a TaqMan probe to detect and quantify SHIV. A pair of qPCR primers, which amplify a 188 bp DNA fragment, and a TaqMan probe were selected from ATPase gene (ORF114R) of the SHIV genome. The primers and TaqMan probe used in this assay were shown to be specific for SHIV and did not react with White spot syndrome virus (WSSV), Infectious hypodermal and hematopoietic necrosis virus (IHHNV), Hepatopancreatic parvovirus (HPV), Vibrio parahaemolyticus causing acute hepatopancreatic necrosis disease (VPAHPND), and Enterocytozoon hepatopenaei (EHP), or healthy shrimp DNA. The detection limit of the qPCR method was as low as 4 copies per reaction. The diagnostic sensitivity and the diagnostic specificity of the qPCR compared with nested PCR were 95.3% and 99.2%, respectively. The resulting standard curves showed high correlation coefficient values (R2 = 0.998) in the range of 4 × 109 to 4 × 100 DNA copies/reaction. Test of farm samples showed that SHIV was detected in L. vannamei, Fenneropenaeus chinensis and Macrobrachium rosenbergii contained SHIV ranged from 1.21E+02 to 7.95E+07 copies (μg DNA)-1. Quantitative detection of different tissues in artificial infected shrimp showed that haemolymph contained the highest SHIV load and muscle contained lowest SHIV load.

Screening of Penaeus vannamei Boone, 1931 collected from east coast of India for monodon baculovirus (MBV) and hepatopancreatic parvovirus (HPV)

Aquaculture is one of the fastest growing sectors yielding more than 10% growth rate per annum over the past 10 years, with a contribution of about 16% of animal protein production globally. Aquaculture production has surpassed that of the other meat producing sectors. Diseases, especially of viral etiology, remains a stumbling block for the growth of this sector posing great threat. The major viruses affecting shrimps in India are white spot syndrome virus (WSSV) and infectious hypodermal hematopoietic necrosis virus (IHHNV). Besides this, monodon baculovirus (MBV) and hepatopancreatic parvovirus (HPV) also cause considerable loss to the shrimp farmers. MBV outbreaks can cause mass mortalities when combined with other viral diseases. Moreover, MBV and HPV infections can lead to slow growth in cultured penaeid shrimps, thereby affecting the production. Although studies pertaining to the prevalence of these two viruses in the tiger shrimp Penaeus monodon have been reported extensively, similar information on Penaeus vannamei farming is scanty. Hence a pilot study was undertaken to screen P. vannamei samples collected from various farms of Tamil Nadu, Andhra Pradesh and West Bengal. Shrimp samples (n=20) from six farms during 35-75 days of culture (DOC) were collected for screening by PCR. It was found that all the samples were negative for MBV and HPV. It could be concluded that the prevalence of both these viruses in P. vannamei are either absent or very low.