Taura Syndrome Virus Infection Research Articles

DNA Profile of Pacific White Shrimp, L. vannamei Infected by Taura Syndrome Virus Using Single Strand Conformation Polymorphism (SSCP) Analysis

The shrimp infected by Taura syndrome virus (TSV) can survive until harvesting time. At this phase, the growth of this shrimp is relatively normal but it is referred as TSV carrier. This research is expected to provide a recommendation in effort to prevent the occurrence of TSV infection. Therefore, this study was aimed to evaluate the DNA polymorhism of Pacific white shrimp L. vannamei infected by TSV. A total of 60 samples of L. vannamei were collected from SPF (specific pathogen free)-Florida, F1-healthy, F1-tolerance (it is can life normally after infected by TSV), and F1-moribund (diying after infected by TSV). The samples were analysis using PCR with IQ-2000 KIT to determine the positive infection of samples. DNA profile of white shrimp, L. vannamei was significantly different from SPF Florida, F1-moribund, and F1-healthy shrimp. This expression may control the defense mechanism. This can be ascribed to the reason that shrimp survived from TSV infection had a higher amount of locus (3 loci) that were detected from DNA polymorphism of gene in white shrimp contributed to a gene control that was determined from allele variation. Two important alleles (200 bp and 220 bp) unique may be associated with tolerant trait. Those loci only found in F1-tolerance shrimp. However to prove that further necessary to sequences a nucleotide of three loci as responsible for resistance to TSV infection.

Molecular cloning, characterization, and immune response against white spot syndrome virus and Taura syndrome virus infections of peroxiredoxin in Litopenaeus vannamei and its antioxidant activity

Peroxiredoxin (Prx) is an important peroxidase that can protect organisms against various oxidative stresses. In this study, a member of Prx family, designated LvPrx, was cloned fromLitopenaeus vannamei. Sequence and phylogenetic analyses indicated that LvPrx belongs to the 2-Cys Prx (Prx IV) isoform. The recombinant LvPrx protein was constructed and expressed inEscherichia coli, and the purified LvPrx proteins were shown to reduce H2O2in vitro in the presence of dithiothreitol, indicating that LvPrx is a functional peroxiredoxin. Using qRT-PCR, the mRNA expression levels of LvPrx were determined in the haemocytes ofL. vannameiat different stages after being challenged with WSSV and TSV at different doses. The results showed that the expression levels of LvPrx were significantly up-regulated () during 4-24 h after both WSSV and TSV challenge, suggesting that LvPrx may participate in the shrimp’s immune response to viral infection.

Molecular Detection of Taura Syndrome Virus Infections in White Shrimp (Litopenaeus vannamei) and Giant Prawn (Macrobrachium rosenbergii)

Abstract Giant prawn (Macrobrachium rosenbergii) and vaname shrimp (Litopenaeus vannamei) are types of shrimp that became excellent commodities in the fisheries sector. However, one of the obstacles in the vaname shrimp aquaculture is a disease caused by the infection of Taura Syndrome Virus (TSV). One of the consequence of raising the vaname shrimp in Indonesia is the possibility of spreading TSV infection in another shrimp species. TSV infection in giant prawns in Indonesia has not been reported. The aims of this study were: 1. To determine the resistance of giant prawns toward TSV infection and 2. To detect molecularly using RT-PCR technique the presence of viruses TSV on vaname shrimp or giant prawns infected with 3 different doses (0.05 ml; 0.10 ml and 0.15 ml) of TSV inoculum using a pair of specific primers for TSV 9992F (5'-AAG CTT GCG TAG ACA GCC-3') and 9195R (5'-TCA AGA ATG GCT TCC TGG-3'). The research results showed that vaname shrimp mortality infected by 0.05 ml; 0.10 ml and 0.15 ml TSV inoculums were 14.28%, 42.86%, and 57.14%, respectively. Whereas the giant prawns mortality that were infected using the same dose of TSV inoculums were 0%, 8.33%, and 8.33%, respectively. Positive result was detected molecularly only from haemolymph of vaname shrimp infected using 0.15 ml of TSV inoculum. On the other hand, positive results were detected in pleopod and the gill of vaname shrimp infected using 0.05 ml; 0.10 ml or 0.15 ml of TSV inoculums. In giant prawns, infection using 3 different doses of TSV inoculums causes negative result molecularly. Based on all of the facts, it can be concluded that, giant prawns has the higher resistance to TSV infection than that of vaname shrimp.

Transcriptome Analysis of Pacific White Shrimp (Litopenaeus vannamei) Hepatopancreas in Response to Taura Syndrome Virus (TSV) Experimental Infection

BackgroundThe Pacific white shrimp, Litopenaeus vannamei, is a worldwide cultured crustacean species with important commercial value. Over the last two decades, Taura syndrome virus (TSV) has seriously threatened the shrimp aquaculture industry in the Western Hemisphere. To better understand the interaction between shrimp immune and TSV, we performed a transcriptome analysis in the hepatopancreas of L. vannamei challenged with TSV, using the 454 pyrosequencing (Roche) technology.Methodology/Principal FindingsWe obtained 126919 and 102181 high-quality reads from TSV-infected and non-infected (control) L. vannamei cDNA libraries, respectively. The overall de novo assembly of cDNA sequence data generated 15004 unigenes, with an average length of 507 bp. Based on BLASTX search (E-value <10−5) against NR, Swissprot, GO, COG and KEGG databases, 10425 unigenes (69.50% of all unigenes) were annotated with gene descriptions, gene ontology terms, or metabolic pathways. In addition, we identified 770 microsatellites and designed 497 sets of primers. Comparative genomic analysis revealed that 1311 genes differentially expressed in the infected shrimp compared to the controls, including 559 up- and 752 down- regulated genes. Among the differentially expressed genes, several are involved in various animal immune functions, such as antiviral, antimicrobial, proteases, protease inhibitors, signal transduction, transcriptional control, cell death and cell adhesion.Conclusions/SignificanceThis study provides valuable information on shrimp gene activities against TSV infection. Results can contribute to the in-depth study of candidate genes in shrimp immunity, and improves our current understanding of this host-virus interaction. In addition, the large amount of transcripts reported in this study provide a rich source for identification of novel genes in shrimp.

Effects of endosulfan exposure and Taura Syndrome Virus infection on the survival and molting of the marine penaeid shrimp, Litopenaeus vannamei

Molting in crustaceans is an important endocrine-controlled biological process that plays a critical role in growth and reproduction. Many factors can affect this physiological cycle in crustaceans including environmental stressors and disease agents. For example the pathology of Taura Syndrome Virus (TSV) of shrimp is closely related to molting cycle. Similarly, endosulfan, a commonly used pesticide is a potential endocrine disruptor. This study explores interrelationships between pesticide exposure, virus infection and their interactions with physiology and susceptibility of the shrimp. Litopenaeus vannamei (Pacific white shrimp) were challenged with increasing doses of endosulfan and TSV (TSV-C, a Belize reference strain) to determine the respective median lethal concentrations (LC50s). The 96-h endosulfan LC50 was 5.32μgL−1, while the 7-d TSV LC50 was 54.74mgL−1. Subsequently, based on their respective LC50 values, a 20-d interaction experiment with sublethal concentrations of endosulfan (2μgL−1) and TSV (30mgL−1) confirmed a significant interaction (p<0.05, χ2=5.29), and thereby the susceptibility of the shrimp. Concurrently, molt-stage of animals, both at the time of exposure and death, was compared with mortality. For animals challenged with TSV, no strong correlation between molt-stage and mortality was observed (p>0.05). For animals exposed to endosulfan, animals in the postmolt stage were shown to be more susceptible to acute toxicity (p<0.05). For animals exposed to both TSV and endosulfan, interference of endosulfan-associated stress lead to increasingly higher susceptibility at postmolt (p<0.05) during the acute phase of the TSV disease cycle.

Epidemiology and phylogenetic analysis of Taura syndrome virus in cultured Pacific white shrimp Litopenaeus vannamei B. in Taiwan

Taura syndrome virus (TSV) has spread worldwide, causing significant economic losses since Taura syndrome was first described in Ecuador in 1992. To determine the prevalence and impact of TSV infection on the shrimp farming industry in Taiwan, Pacific white shrimp Litopenaeus vannamei B. were collected from 220 farms between 2004 and 2006 for viral detection by reverse transcription polymerase chain reaction. Data showed that the overall TSV prevalence rate was 20% (43/220 farms). Comparing shrimp growth stages, TSV prevalence rates were 4% for postlarvae, 24% for juveniles, 24% for subadults, 32% for adults, and 5% for brooders. Among TSV-positive farms, average infection incidence was 35% in postlarvae farms, 55% in juvenile farms, 39% in subadult farms, 31% in adult farms, and 20% in brooder farms. Notably, TSV was also detected in Exopalaemon orientis H. from 1 of 10 farms. Tail fans and appendages had red pigmentation, which is characteristic of TSV infection. Of shrimp with pathological lesions, 100% had lesions on tail fans, 88% on appendages, and 80% in gills. Sequence comparison using the TSV VP1 (structural protein) gene showed that 9 isolates from the farms had 92.3 to 99.5% nucleotide sequence identity with strains in the GenBank database from Taiwan (AF406789 and AY355310) and Venezuela (DQ212790). This is the first broad epidemiological study of TSV infection in L. vannamei in Taiwan.

Improved sensitivity of Taura syndrome virus immunodetection with a monoclonal antibody against the recombinant VP2 capsid protein

Taura syndrome virus (TSV) is one of the major pathogens causing mortality in the whiteleg shrimp, Litopenaeus vannamei. In this study, the gene sequence encoding the VP2 capsid protein (40 kDa) of TSV was cloned into pMAL-C2 expression vector. Five monoclonal antibodies (MAbs) were produced against the VP2 capsid protein, which was expressed heterologously in the form of a fusion protein with maltose binding protein and called MBP-VP2. All MAbs belonged to the IgG1 subclass and could bind MBP-VP2 at 400–800 pg/spot in immuno-dot blot assays. The MAbs could detect VP2 both in extracts from shrimp infected naturally in western blotting and dot blotting and in shrimp tissues in immunohistochemistry. Additionally, these MAbs did not exhibit cross-reactivity to extracts from uninfected shrimp or shrimp infected with several other common viruses. However, the dot blot assay sensitivity for TSV was approximately 10,000 times lower than that of one step RT-PCR. The MAb TSV2-88 specific to VP2 obtained in this study demonstrated an approximately twofold higher sensitivity than that of the MAb specific to VP3 from a previous study. In immunohistochemistry, the MAb TSV2-88 specific to VP2 demonstrated stronger immunoreactivity than the MAb TSV3-601 specific to VP3. A combination of the VP2 and VP3 MAbs could be used to more easily detect TSV infections in field samples of L. vannamei with better sensitivity and fidelity than using a single MAb.

C-Terminal Hemocyanin from Hemocytes of Penaeus vannamei Interacts with ERK1/2 and Undergoes Serine Phosphorylation

To understand molecular immune response of Penaeus vannamei during Taura syndrome virus (TSV) infection, expression and functional proteomics studies were performed on hemocyanin, which is a major abundant protein in shrimp hemocytes. Two-dimensional electrophoresis (2-DE) revealed up-regulation of several C-terminal fragments of hemocyanin, whereas the N-terminal fragments were down-regulated during TSV infection. 2-D Western blot analysis showed that the C-terminal hemocyanin fragments had more acidic isoelectric points (pI), whereas the N-terminal fragments had less acidic pI. Further analysis by NetPhos showed a greater number of serine phosphorylation sites in the C-terminal hemocyanin. Additionally, motif scan using Scansite revealed ERK D-domain, which is required for activation of ERK1/2 effector kinase, as a kinase-binding site at the 527th valine in the C-terminal hemocyanin, whereas neither motif nor functional domain was found in the N-terminus. Co-immunoprecipitation confirmed the interaction between the C-terminal hemocyanin and ERK1/2. 1-D Western blot analysis showed that ERK1/2 was also up-regulated during TSV infection. Our findings demonstrate for the first time that ERK1/2 signaling pathway may play an important role in molecular immune response of P. vannamei upon TSV infection through its interaction with the C-terminal hemocyanin.

Comparison of Taura syndrome virus (TSV) detection methods during chronic-phase infection in Penaeus vannamei

Methods to detect Taura syndrome virus (TSV) were assessed for their ability to detect the virus during chronic phase infection in the Pacific white shrimp Penaeus vannamei. In situ hybridization (ISH), immunohistochemistry (IHC) using monoclonal antibody 1A1, conventional RT-PCR and real-time quantitative (q)RT-PCR were compared using shrimp sampled over 60 wk following experimental TSV infection. Between Weeks 7 and 60, hematoxylin-eosin histology confirmed the presence of lymphoid organ spheroids (LOS) and an absence of lesions in the cuticular epithelium. ISH detected TSV in LOS over the duration of the study. IHC was generally less sensitive than ISH, and after Week 24, was often unable to confirm TSV infection. Detection of TSV by RT-PCR was highly dependent on sample source after Week 43, where viral RNA was detected in 12 of 14 hemolymph samples but only 5 of 16 pleopod samples. qRT-PCR detected TSV over the 60 wk in both hemolymph and pleopods, although RNA copy numbers in pleopods were consistently lower throughout the study. This study demonstrates that ISH and qRT-PCR are the most reliable methods for detecting TSV during late chronic phase infection. RT-PCR was also reliable if hemolymph was used as the sample source.

Experimental infections reveal that common Thai crustaceans are potential carriers for spread of exotic Taura syndrome virus

Taura syndrome virus (TSV) was first reported as a serious cause of shrimp mortality limited to reared Penaeus (Litopenaeus) vannamei in the Americas, where it spread principally through regional and international transfer of live post larvae (PL) and broodstock. Subsequently, through importation of infected broodstock, TSV outbreaks spread to Asia, first to Taiwan and China and then to Thailand, Indonesia and Korea. Since its introduction to Thailand, outbreaks have occasionally been reported from rearing ponds stocked with batches of specific pathogen free (SPF) P. vannamei PL that tested negative for TSV by nested RT-PCR assay. Since it was possible that the outbreaks may have occurred via horizontal transfer of TSV from wild carrier species, we tested 5 common native crustaceans that live in and around shrimp ponds (2 palaemonid shrimp species, Palaemon styliferus and Macrobrachium lanchesteri, and 3 species of crabs, Sesarma mederi, Scylla serrata and Uca vocans) for susceptibility to TSV in experimental challenges. We found that U. vocans, S. serrata and S. mederi did not die but, respectively, gave strong RT-PCR reactions indicating heavy viral load at 5, 10 and 15 d post-injection of TSV and 10, 15 and up to 50 d after feeding with TSV-infected P. vannamei carcasses. Also after feeding, P. styliferus did not die, but a high proportion gave strong RT-PCR reactions at 5 d post-challenge and no reactions at 15 d. Similarly after feeding, M. lanchesteri showed no mortality and gave only light RT-PCR reactions at 2 d, moderate reactions at 5 d and no reaction at 15 d. By contrast, transmission experiments from the TSV-infected crabs and palaemonid shrimp via water or feeding resulted in death of all the exposed P. vannamei from 8 to 12 d post-challenge and all were positive for heavy viral load by RT-PCR assay. Despite the results of these laboratory challenge tests, natural TSV infections were not detected by nested RT-PCR in samples of these species taken from the wild. These results indicated that transmission of TSV from infected crabs and palaemonid shrimp via water or feeding might pose a potential risk to shrimp aquaculture.

Specific monoclonal antibodies raised against Taura syndrome virus (TSV) capsid protein VP3 detect TSV in single and dual infections with white spot syndrome virus (WSSV)

The gene sequence encoding VP3 capsid protein of Taura syndrome virus (TSV) was cloned into pGEX-6P-1 expression vector and transformed into Escherichia coli BL21. After induction, recombinant GST-VP3 (rVP3) fusion protein was obtained and further purified by electro-elution before use in immunizing Swiss mice for production of monoclonal antibodies (MAb). One MAb specific to glutathione-S-transferase (GST) and 6 MAb specific to VP3 were selected using dot blotting and Western blotting. MAb specific to VP3 could be used to detect natural TSV infections in farmed whiteleg shrimp Penaeus vannamei by dot blotting and Western blotting, without cross reaction to shrimp tissues or other shrimp viruses, such as white spot syndrome virus (WSSV), yellow head virus (YHV), monodon baculovirus (MBV) and hepatopancreatic parvovirus (HPV). These MAb were also used together with those specific for WSSV to successfully detect TSV and WSSV in dual infections in farmed P. vannamei.

Proteomic analysis of differentially expressed proteins in Penaeus vannamei hemocytes upon Taura syndrome virus infection

To understand molecular responses of crustacean hemocytes to virus infection, we applied 2-DE proteomics approach to investigate altered proteins in hemocytes of Penaeus vannamei during Taura syndrome virus (TSV) infection. At 24 h postinfection, quantitative intensity analysis and nano-LC-ESI-MS/MS revealed 11 forms of 8 proteins that were significantly up-regulated, whereas 9 forms of 5 proteins were significantly down-regulated in the infected shrimps. These altered proteins play important roles in host defense (hemocyanin, catalase, carboxylesterase, transglutaminase, and glutathione transferase), signal transduction (14-3-3 zeta), carbohydrate metabolism (acetylglucosamine pyrophosphorylase), cellular structure and integrity (beta-tubulin, beta-actin, tropomyosin, and myosin), and ER-stress response (protein disulfide isomerase). Semiquantitative RT-PCR and Western blot analysis confirmed the upregulation of 14-3-3 at both mRNA and protein levels. Interestingly, several altered protein spots were identified as fragments of hemocyanin. Mass spectrometric analysis showed that the hemocyanin spots at acidic and basic regions represented the C- and N-terminal hemocyanin fragments, respectively. As three-quarters of C-terminal fragments were up-regulated, whereas two-thirds of N-terminal hemocyanin fragments were down-regulated, we therefore hypothesize that C- and N-terminal hemocyanin fragments may have differential roles in hemocytes. Further investigation of these data may lead to better understanding of the molecular responses of crustacean hemocytes to TSV infection.

Molecular epidemiological investigation of infectious hypodermal and hematopoietic necrosis virus and Taura syndrome virus in Penaeus Vannamei cultured in China

The Infectious hypodermal and hematopoietic necrosis virus (IHHNV) and Taura syndrome virus (TSV) are two important shrimp viruses in cultured shrimp in America. These two viruses were transmitted to China at the beginning of the 21st century. In this study, 214 shrimp samples of Penaeus vannamei were collected from seven different areas of China and tested by PCR for IHHNV and TSV infection. The results showed that there were a high prevalence of IHHNV (65.42%) and low prevalence of TSV (3.27%) in the tested samples. Several samples were found to be co-infected with these two viruses. A 3 kb fragment of 7 positive IHHNV samples and a structure protein region (ORF2) of three TSV positive samples were amplified and sequenced. The sequence comparison indicated that both IHHNV and TSV sequenced in China have a low genetic variations compared with the prototype IHHNV and TSV from Hawaii. Phylogenetic analysis showed that TSV isolates were clustered into two groups, Asia and America group, which was genetically correlated to geographic distribution.

Ultrastructure of the replication site in Taura syndrome virus (TSV)-infected cells

Taura syndrome virus (TSV) is a member of the family Dicistroviridae that infects Pacific white shrimp Litopenaeus vannamei (also called Penaeus vannamei), and its replication strategy is largely unknown. To identify the viral replication site within infected shrimp cells, the viral RNA was located in correlation with virus-induced membrane rearrangement. Ultrastructural changes in the infected cells, analyzed by transmission electron microscopy (TEM), included the induction and proliferation of intracellular vesicle-like membranes, while the intracytoplasmic inclusion bodies and pyknotic nuclei indicative of TSV infection were frequently seen. TSV plus-strand RNA, localized by electron microscopic in situ hybridization (EM-ISH) using TSV-specific cDNA probes, was found to be associated with the membranous structures. Moreover, TSV particles were observed in infected cells by TEM, and following EM-ISH, they were also seen in close association with the proliferating membranes. Taken together, our results suggest that the membranous vesicle-like structures carry the TSV RNA replication complex and that they are the site of nascent viral RNA synthesis. Further investigations on cellular origins and biochemical compositions of these membranous structures will elucidate the morphogenesis and propagation strategy of TSV.

Comparison of four Taura syndrome virus (TSV) isolates in oral challenge studies with Litopenaeus vannamei unselected or selected for resistance to TSV

Taura syndrome virus (TSV) infection in TSV-resistant (TSR) and TSV-susceptible (Kona) Litopenaeus vannamei (also called Penaeus vannamei) was investigated using histology, in situ hybridization (ISH), conventional reverse transcription polymerase chain reaction (RT-PCR) assays, and SYBR-Green real-time RT-PCR analysis. The shrimp were challenged by feeding with minced tissues of L. vannamei infected with 4 genotypic variants of TSV (Bz01, Th04, UsHi94, and Ve05). Survival probabilities of TSR shrimp were higher than those for Kona shrimp with all 4 variants. Th04, UsHi94, and Ve05 gave no Taura syndrome lesions with TSR shrimp. In contrast, TSR shrimp challenged with Bz01 and Kona shrimp with all 4 TSV variants exhibited severe necrosis of cuticular epithelial cells and lymphoid organ spheroids, indicative of acute and chronic phases of TSV infection, respectively. TSV was not detected by RT-PCR in TSR shrimp infected with Th04, UsHi94, and Ve05, or in Kona shrimp infected with Ve05 but was detected in TSR shrimp infected with Bz01 and in Kona shrimp infected with Bz01, Th04, and UsHi94. Real-time RT-PCR revealed that mean TSV copy numbers in TSR shrimp infected with Bz01, Th04, and UsHi94 were significantly (p < 0.0005) lower than those in Kona shrimp. In contrast, mean TSV copy numbers in TSR and Kona shrimp infected with Ve05 were not significantly different (p > 0.4). The results show that TSR L. vannamei are susceptible to infection but give high survival rates following challenge by all 4 variants of TSV.

The molecular detection of Taura syndrome virus emerging with White spot syndrome virus in penaeid shrimps of Thailand

This study is to investigate the outbreak of the Taura syndrome (TS) in penaeid shrimps of Thailand. Following the outbreak in 2003, the Taura syndrome virus (TSV) was detected in two closely related penaeids; the Pacific white shrimp, Penaeus vannamei, and the black tiger prawn, Penaeus monodon. Multiple viral infections were detected, we found that the TSV-infected P. vannamei also presented evidence of the White spot syndrome virus (WSSV) as evidenced by histological examination and molecular diagnostic methods. Infections of TSV and WSSV were investigated over a period of ten months between October 2003 and July 2004. TSV infection in shrimps was diagnosed by RT-PCR using the OIE recommended 9195/9992 primers. Positive shrimp gave the expected amplicon of 231 bp. We also amplified and sequenced a fragment of the vp1 gene (354 bp). The pairwise comparisons of vp1 revealed that Thai TSV was phylogenetically related to the Taiwanese strains (98.3% identity).

Viral resistance in shrimp that express an antisense Taura syndrome virus coat protein gene

Taura syndrome virus (TSV) is a major cause of mortality and morbidity in shrimp, and has a profound economic impact on commercial U.S. shrimp farming. This paper describes the stable expression of an antisense Taura syndrome virus-coat protein (TSV-CP) gene construct in shrimp zygotes, via transfection using jetPEI reagent, over a period of at least 236 days. The transgenic shrimp showed no statistically significant difference from normal control shrimp in terms of weight gain or their appearance, morphology, swimming and eating activities. When challenged with live TSV, the transgenic shrimp exhibited increased resistance to the TSV infection (83% survival rate) as compared to control animals (44% survival rate). This work demonstrates that transgenic shrimp, which stably express an antisense transcript from the TSV-CP gene, are partially resistant to TSV infection. These data may have an important implication for commercial shrimp farming.

Experimental infection of Penaeus monodon with Taura syndrome virus (TSV)

Clinical signs and lesions of Taura syndrome virus (TSV) infection in Penaeus monodon have not been documented although the virus has been detected in this shrimp species by reverse transcription polymerase chain reaction (RT-PCR). This study provides the first evidence of TSV infection in P. monodon by histological and in situ hybridization (ISH) analyses. We performed experimental bioassays with groups of P. monodon using inocula of P. monodon and Litopenaeus vannamei (Th04PmTSV and Th04LvTSV, respectively), which were collected from Thailand in 2004 and found to be positive for TSV by RT-PCR. Samples of shrimp for histological and ISH analyses were collected on Days 2, 14, and 28 post-inoculation. Mortality among TSV-inoculated P. monodon appeared on Day 3, with 2 out of 10 shrimp dying. Severe necrosis of cuticular epithelial cells and lymphoid organ spheroids, indicative of acute and chronic phase lesions of TSV infection, respectively, were detected in the samples. Sequence analyses of the capsid protein 2 (CP2) gene showed that Th04PmTSV and Th04LvTSV isolates were different; however, both belonged to a phylogenetic family of Asian TSV isolates. The results of this study demonstrated that both mortality and histological lesions are associated with TSV infection in P. monodon.

Effect of chronic Taura syndrome virus infection on salinity tolerance of Litopenaeus vannamei

Taura syndrome virus (TSV) is one of the most important shrimp viruses affecting farmed shrimp worldwide. After an acute phase during which the likelihood of mortality is elevated, infected shrimp enter a chronic phase during which shrimp appear to resume normal behavior and display no gross signs of infection. This study was designed to determine if chronically TSV-infected shrimp Litopenaeus vannamei are compromised by the infection. Specifically we investigated whether chronically infected shrimp could tolerate a drop in salinity as strongly as uninfected shrimp. The study consisted of 3 trials that compared survival of uninfected and chronically TSV-infected L. vannamei after drops in salinity from 24 ppt to salinities varying from 18 to 0 ppt. Logistic regression detected a significant effect of TSV infection on survival of chronically infected shrimp (p < 0.05). Salinity drops from 24 ppt to 3 and 6 ppt resulted in statistically different survivals (p < 0.05). Survival rates were similar among groups for salinity drops to greater than 6 ppt or less than 3 ppt. Salinities at which 50% of the shrimp died (LC50) were 3.06 ppt for the uninfected and 6.65 ppt for the chronically infected groups. Moreover, histopathological analysis of chronically infected shrimp that were moribund or recently dead showed no signs of having reverted to the acute stage of the disease. These results suggest that chronically infected shrimp are not able to tolerate a salinity drop as strongly as uninfected shrimp.

The geographic distribution of Taura Syndrome Virus (TSV) in the Americas: determination by histopathology and in situ hybridization using TSV-specific cDNA probes

Representative archived Litopenaeus vannamei samples (117 total), originating from 13 different countries and collected between 1992 to 1996, were analyzed by in situ hybridization to verify the presence of Taura Syndrome Virus (TSV) within pathodiagnostic acute phase TS histological lesions. The in situ assay results showed that TSV was present in one or more representative samples analyzed from each country (76 of 117 samples or 65%), thus, confirming the original histological diagnosis of TSV infection. The false negative in situ hybridization results, obtained for 35% of the samples assayed (41 in total), were attributed to over-fixation with Davidson's AFA (acetic acid, formaldehyde, alcohol) solution and consequent acid hydrolysis of TSV genomic RNA within pathodiagnostic TSV lesions. The collective findings of this disease survey assisted in documenting the spread and current distribution of TSV over a 5-year period and definitively established the presence of TSV within TS diseased shrimp originating from Ecuador when and where the disease was first recognized in 1992. These findings further strengthen the existing evidence that TS has a viral, not a toxic, etiology and indicate that either a single TSV strain, or very similar strains of the same virus, are responsible for the TSV panzootic that has been expanding in the Americas since 1992.