Screening for White Spot Syndrome Virus (WSSV) in pink shrimp (Penaeus paulensis) from Rocha Lagoon, Uruguay.

Penaeus paulensis (pink shrimp) is an important resource for small-scale fisheries in the brackish coastal lagoons of Uruguay. No viral diseases have been detected in shrimp populations in the Uruguayan territory. The presence of viral pathogens, such as White Spot Syndrome Virus (WSSV) and Infectious Hypodermal Haematopoietic Necrosis Virus (IHHNV) in wild shrimp populations has been previously reported in Brazil and Argentina. We investigated the presence of WSSV in wild populations of penaeid shrimp from Rocha Lagoon, Uruguay. We sampled 70 specimens of juvenile P. paulensis and assessed the presence of these viral pathogens using nested PCR and histology. Gill tissue from the 70 samples was divided into 14 pools of 5 individuals for DNA extraction and PCR analysis. We also retested each pooled sample individually. The nested PCR procedure described in the WOAH aquatic animal manual was used. A subset of 20 individual specimens were also processed using standard histological techniques. The results showed that WSSV was not detected in the pooled or individually tested samples. We found no evidence of the presence of the viral genome or gill lesions in the samples analysed. This indicates that the fishery is still likely to be free of WSSV infection. The procedures and information generated can be used as a baseline study for future implementation of surveillance programmes in the country.

Use of brome mosaic virus-like particles in feed, to deliver dsRNA targeting the white spot syndrome virus vp28 gene, reduces Penaeus vannamei mortality.

Numerous strategies have been investigated to combat viral infections in shrimp, specifically targeting the white spot syndrome virus (WSSV) that has caused outbreaks worldwide since the 1990s. One effective treatment involves intramuscular application of dsRNA-mediated interference against the viral capsid protein VP28. However, this approach presents challenges in terms of individual shrimp management, limiting its application on a large scale. To address this, our study aimed to evaluate the efficacy of oral delivery of protected dsRNA using chitosan nanoparticles or virus-like particles (VLPs) synthesized in brome mosaic virus (BMV). These delivery systems were administered before, during, and after WSSV infection to assess their therapeutic potential. Our findings indicate that BMV-derived VLPs demonstrated superior efficiency as nanocontainers for dsRNA delivery. Notably, the treatment involving vp28 dsRNA mixed in the feed and administered simultaneously to shrimp already infected with WSSV exhibited the highest survival rate (48%), while the infected group had a survival rate of zero, suggesting the potential efficacy of this prophylactic approach in commercial shrimp farms.

Eco-efficiency assessment of disease-infected shrimp farming in Mexico using environmental impact assessment tools.

Mexico ranks second in shrimp (Litopenaeus vannamei) production of in Latin America with significant annual growth, however, during 2011 shrimp production fell by almost 50 % due to the presence of the white spot syndrome virus (WSSV). In this context, a life cycle analysis (LCA) and data envelopment analysis (DEA) were performed on 76 commercial farms severely affected by the presence of WSSV in northwestern Mexico. The application of this combined methodology allowed a detailed quantification of different environmental impact categories. During the presence of WSSV, there was a negative effect on the feed conversion ratio (FCR) (>40 %), higher consumption of seawater (38 %), and energy (38 %). Consequently, operational outputs related to the discharge of nitrogen and phosphorus increased by 60 and 57 %, respectively. Similarly, CO2 emissions, increased by 38 % relative to a typical year of production. Overall, the main critical points in the impact categories analyzed are related to food (98 %), use of diesel (23 %), and rearing (24 %), dominating pollutants emissions in all categories. Consequently, an improvement scenario was evaluated related to innovation in the formulation of foods supplied with immunostimulants, which confer protection against pathogenic microorganisms. This scenario lead to a reduction environmental impact of about 82 %. The results of this analysis will be a useful resource in the design of mitigation strategies with innovation processes that allow maintaining yields for shrimp producers in this region and at the same time reduce the environmental impacts generated.

Virulence and genetic differences among white spot syndrome virus isolates inoculated in Penaeus vannamei.

White spot syndrome virus (WSSV) infects several economically important aquaculture species, and has caused significant losses to the industry. This virus belongs to the Nimaviridae family and has a dsDNA genome ranging between 257 and 309 kb (more than 20 isolate genomes have been fully sequenced and published to date). Multiple routes of infection could be the cause of the high virulence and mortality rates detected in shrimp species. Particularly in Penaeus vannamei, differences in isolate virulence have been observed, along with controversy over whether deletions or insertions are associated with virulence gain or loss. The pathogenicity of 3 isolates from 3 localities in Mexico (2 from Sinaloa: 'CIAD' and 'Angostura'; and one from Sonora: 'Sonora') was evaluated in vivo in whiteleg shrimp P. vannamei infection assays. Differences were observed in shrimp mortality rates among the 3 isolates, of which Sonora was the most virulent. Subsequently, the complete genomes of the Sonora and Angostura isolates were sequenced in depth from infected shrimp tissues and assembled in reference to the genome of isolate strain CN01 (KT995472), comprising 289350 and 288995 bp, respectively. Three deletion zones were identified compared to CN01, comprising 15 genes, including 3 envelope proteins (VP41A, VP52A and VP41B), 1 non-structural protein (ICP35) and 11 other encoding proteins whose function is currently unknown. In addition, 5 genes (wsv129, wsv178, wsv204, wsv249 and wsv497) presented differences in their repetitive motifs, which could potentially be involved in the regulation of gene expression, causing virulence variations.

Shedding the Light on Litopenaeus vannamei Differential Muscle and Hepatopancreas Immune Responses in White Spot Syndrome Virus (WSSV) Exposure.

White Spot Syndrome Virus (WSSV) is one of the main threats to farming Litopenaeus vannamei, the most important crustacean commercialized in aquaculture worldwide. Here, we performed RNA-seq analyses in hepatopancreas and muscle from WSSV-negative (healthy) and WSSV-positive (unhealthy) L. vannamei, previously exposed to the virus, to obtain new insights about the molecular basis of resistance to WSSV. We detected 71% of our reads mapped against the recently described L. vannamei genome. This is the first report mapping RNA-seq transcripts from shrimps exposed to WSSV against the species reference genome. Differentially expressed gene (DEG) analyses were performed for four independent comparisons, and 13,338 DEGs were identified. When the redundancies and isoforms were disregarded, we observed 8351 and 6514 DEGs, respectively. Interestingly, after crossing the data, we detected a common set of DEGs for hepatopancreas and healthy shrimps, as well as another one for muscle and unhealthy shrimps. Our findings indicate that genes related to apoptosis, melanization, and the Imd pathway are likely to be involved in response to WSSV, offering knowledge about WSSV defense in shrimps exposed to the virus but not infected. These data present potential to be applied in further genetic studies in penaeids and other farmed shrimp species.

The white spot syndrome virus hijacks the expression of the Penaeus vannamei Toll signaling pathway to evade host immunity and facilitate its replication.

The white spot syndrome virus (WSSV), the most lethal pathogen of shrimp, is a dsDNA virus with approximately a 300,000 base pairs and contains approximately 180-500 predicted open reading frames (ORFs), of which only 6% show homology to any known protein from other viruses or organisms. Although most of its ORFs encode enzymes for nucleotide metabolism, DNA replication, and protein modification, the WSSV uses some of its encoded proteins successfully to take control of the metabolism of the host and avoid immune responses. The contribution of the shrimp innate immune response to prevent viral invasions is recognized but yet not fully understood. Thus, the role of several components of Toll pathway of the shrimp Penaeus vannamei against WSSV has been previously described, and the consequential effects occurring through the cascade remain unknown. In the current study the effects of WSSV over various components of the shrimp Toll pathway were studied. The gene expression of Spätzle, Toll, Tube, Cactus and Dorsal was altered after 6-12 h post inoculation. The expression of LvToll3, LvCactus, LvDorsal, decreased ~4.4-, ~3.7- and ~7.3-fold at 48, 24 and 48 hpi, respectively. Furthermore, a remarkable reduction (~18-fold) in the expression of the gene encoding LvCactus in WSSV infected specimens was observed at 6 hpi. This may be a sophisticated strategy exploited by WSSV to evade the Toll-mediated immune action, and to promote its replication, thereby contributing to viral fitness.

An effective white spot syndrome virus challenge test for cultured shrimp using different biomass of the infected papilla.

White spot syndrome virus (WSSV) is one of the most virulent pathogens of cultured penaeid shrimp. Several control strategies are used commonly to mitigate the economic losses caused by the pathogen, such as application of antiviral products at farm level. One of the most practical method for the screening of potential anti-WSSV products is through challenge tests. Therefore, it is essential to develop simple, reproducible and effective bioassays able to simulate specific mortality levels. The purpose of this study was to develop a simple and reproducible bioassay that simulate different mortality levels by varying the proportion of WSSV-infected and noninfected shrimp tissues administered to susceptible shrimp during a per os challenge test. This method mimics one of the natural transmission routes of WSSV infection in shrimp and could be applied to identify potential antiviral products to different cultured shrimp species susceptible to WSSV. Here we report: •A simple and economic method to evaluate therapeutic antiviral products against WSSV through a challenge test, that uses different biomass amounts of WSSV-infected papilla.•Allows to simulate a wide and reproducible range of mortalities observed in shrimp farms.•A challenge test that simulates one mode of natural WSSV infection in shrimp.

Potential immunomodulatory and protective effects of the Arthrospira-based dietary supplement on shrimp intestinal immune defenses.

Herein, we evaluated the immunomodulatory and the antiviral protective properties of a cyanobacteria-enriched diet on the immune responses of the Pacific white shrimp Litopenaeus vannamei challenged with the White spot syndrome virus (WSSV). Shrimp were fed with an Arthrospira platensis supplemented feed during 20 days, and its effects were examined by evaluating well-known standardized shrimp immune parameters (total hemocyte counts, total protein concentration, phenoloxidase activity, and serum agglutination titer). Additionally, we assessed the expression of crucial genes involved in both hemolymph- and gut-based immunities related to the shrimp capacity to circumvent viral and microbial infections. Dietary supplementation improved shrimp survival rates after challenge with a median lethal dose of WSSV. From all immune parameters tested, only the serum agglutination titer was higher in treated animals. On the other hand, the expression of some representative marker genes from different immune response pathways was only modulated in the midgut and not in the circulating hemocytes, suggesting that this feed supplementation can be used as an attractive strategy to enhance immunity in shrimp gut. Altogether, our results evidence the immunomodulatory properties of A. platensis supplemented feed in shrimp humoral and intestinal defenses and highlight the potential use of cyanobacteria-based immunostimulants in shrimp farming for protection against infectious diseases.

Proteomic profiling of the white shrimp Litopenaeus vannamei (Boone, 1931) hemocytes infected with white spot syndrome virus reveals the induction of allergy-related proteins.

To elucidate the proteomic responses of shrimp hemocytes to white spot syndrome virus (WSSV) infection at the proteome level, a quantitative shotgun proteomic analysis was performed to detect differentially synthesized proteins in infected hemocytes of white shrimp (Litopenaeus vannamei). We identified 1528 proteins associated to 203 gene ontology (GO) categories. The most representative GO categories were regulation of cellular processes, organic substance metabolic processes and nitrogen compound metabolic processes. Most of the 83 detected up-regulated proteins are involved in DNA regulation and organization and cell signaling. In contrast, most of the 40 down-regulated proteins were related to immune defense processes, protein folding, and development. Differentially induced proteins were further analyzed at the transcript level by RT-qPCR to validate the results. This work provides new insights into the alterations of L. vannamei hemocytes at the protein level at 12 h post-infection with WSSV. Interestingly, several of the up-regulated proteins are allergy-related proteins in humans. Based on our results, we suggest a deeper analysis of the effects of this interaction on the regulation of allergy related-proteins as their up-regulation during WSSV could represent a threat to human health.

Silencing of HIF-1 in WSSV-infected white shrimp: Effect on viral load and antioxidant enzymes.

Hypoxia inducible factor-1 (HIF-1) is a transcriptional factor that induces genes involved in glucose metabolism. HIF-1 is formed by a regulatory α-subunit (HIF-1α) and a constitutive ß-subunit (HIF-1ß). The white spot syndrome virus (WSSV) induces a shift in glucose metabolism and oxidative stress. HIF-1α is associated with the induction of metabolic changes in tissues of WSSV-infected shrimp. However, the contributions of HIF-1 to viral load and antioxidant responses in WSSV-infected shrimp have been not examined. In this study, the effect of HIF-1 silencing on viral load and the expression and activity of antioxidant enzymes (superoxide dismutase-SOD, glutathione S-transferase-GST, and catalase) along with oxidative damage (lipid peroxidation and protein carbonyl) in tissues of white shrimp infected with the WSSV were studied. The viral load increased in hepatopancreas and muscle after WSSV infection, and the accumulative mortality was of 100% at 72 h post-infection. The expression and activity of SOD, catalase, and GST decreased in each tissue evaluated after WSSV infection. Protein carbonyl concentrations increased in each tissue after WSSV infection, while lipid peroxidation increased in hepatopancreas, but not in muscle. Silencing of HIF-1α decreased the WSSV viral load in hepatopancreas and muscle of infected shrimp along with shrimp mortality. Silencing of HIF-1α ameliorated the antioxidant response in a tissue-specific manner, which translated to a decrease in oxidative damage. These results suggest that HIF-1 is essential for restoring the antioxidant response, which counters the oxidative injury associated with WSSV infection.

Effect of Argemone mexicana active principles on inhibiting viral multiplication and stimulating immune system in Pacific white leg shrimp Litopenaeus vannamei against white spot syndrome virus.

Argemone mexicana called as Mexican prickly poppy is a species of poppy found in Mexico and now widely naturalized in many parts of the world with broad range of bioactivities including anthelmintic, cures lepsory, skin-diseases, inflammations and bilious fevers. Plant parts of A. mexicana were serially extracted with hexane, ethyl acetate, methanol and performed antiviral and immunostimulant screening against WSSV and Vibrio harveyi respectively. The control groups succumbed to death 100% within three days, whereas the mortality was significantly (P < 0.5) reduced to 17.43 and 7.11 in the ethyl acetate extracts of stem and root treated shrimp group respectively. The same trend was reflected in the immunostimulant screening also. Different diets were prepared by the concentrations of 100 (AD-1), 200 (AD-2), 300 (AD-3) and 400 (AD-4) mg kg-1 using A. mexicana stem and root ethyl acetate extracts and fed to Pacific white leg shrimp Litopenaeus vannamei weighed about 9.0 ±â€¯0.5 g for 30 days. The control groups fed with the normal diets devoid of A. mexicana extracts. The antiviral screening results revealed that, the ethyl acetate extract of the stem and root were effectively suppressed the WSSV and it reflected in the lowest cumulative mortality of treated shrimps. After termination of feeding trials, group of shrimps from control and each experimental group were challenged with virulent WSSV by intramuscular (IM) injection and studied cumulative mortality, molecular diagnosis by quantitative real time PCR (qRT-PCR), biochemical, haematological and immunological parameters. Control group succumbed to 100% death within four days, whereas the survival was significantly (P < 0.001) increased to 30, 45, 75 and 79% in AD1, AD-2, AD-4 and AD-5 diets fed shrimp groups respectively. qRT PCR results with positive correlation analysis revealed that, the WSSV copies were gradually decreased when increasing the A. mexicana extracts in the diets. The highest concentrations (300 and 400 mg g-1) of A. mexicana extracts in the diets helped to reduce the protein level significantly (P < 0.05) after WSSV challenge. The diets AD-3 and AD-4 also helped to decrease the coagulation time of maximum 64-67% from control groups and maintained the normal level of total haemocyte, oxyhaemocyanin level after WSSV challenge. The proPO level was significantly increased (Column: F = 35.93; P ≤ 0.001 and Row: F = 37.14; P ≤ 0.001) in the AD1-AD-4 diet fed groups from the control diet fed groups. The lowest intra-agar lysozyme activity of 1.63 mm found in control diet fed group and the activity were significantly (P < 0.05) increased to 4.86, 7.89, 9.12 and 10.45 mm of zone of inhibition respectively in AD1 to AD4 diet fed groups.

Direct sequencing of the white spot syndrome virus from Brazil: Genome assembly and new insights on phylogeny.

White spot syndrome virus (WSSV) has been the cause of great economic losses in world shrimp farming. In this work the genome of a Brazilian WSSV isolate was determined from direct sequencing of total DNA extracted from an infected whiteleg shrimp, and assembled based on a chimera template approach. Comparisons between WSSV-BR and other isolates revealed that the Brazilian virus has a relatively small genome, and is very similar to isolates from Thailand and Mexico. A phylogenetic relationship using different approaches has demonstrated that these isolates share a common evolutionary history. An analysis of conflicting phylogenetic signals also considering genomes of other isolates revealed that the evolutionary history of WSSV may be related to recombination events. We observed that these events can also be traced at some level by analyzing the homologous regions in the WSSV genome. The existence of recombination events introduces a new point of view that must be considered in the evolutionary history of WSSV.

Quantitative risk assessment of WSSV transmission through partial harvesting and transport practices for shrimp aquaculture in Mexico.

This quantitative risk assessment provided an analytical framework to estimate white spot syndrome virus (WSSV) transmission risks in the following different scenarios: (1) partial harvest from rearing ponds and (2) post-harvest transportation, assuming that the introduction of contaminated water with viral particles into shrimp culture ponds is the main source of viral transmission risk. Probabilities of infecting shrimp with waterborne WSSV were obtained by approaching the functional form that best fits (likelihood ratio test) published data on the dose-response relationship for WSSV orally inoculated through water into shrimp. Expert opinion defined the ranges for the following uncertain factors: (1) the concentrations of WSSV in the water spilled from the vehicles transporting the infected shrimp, (2) the total volume of these spills, and (3) the dilution into culture ponds. Multiple scenarios were analysed, starting with a viral load (VL) of 1×102mL-1 in the contaminated water spilled that reached the culture pond, whose probability of infection of an individual shrimp (Pi) was negligible (1.7×10-7). Increasing the VL to 1×104.5mL-1 and 1×107mL-1 yielded results into very low (Pi=5.3×10-5) and high risk (Pi=1.6×10-2) categories, respectively. Furthermore, different pond stocking density (SD) scenarios (20 and 30 post-larvae [PL]/m2) were evaluated, and the probability of infection of at least one out of the total number of shrimp exposed (PN) was derived; for the scenarios with a low VL (1×102mL-1), the PN remained at a negligible risk level (PN, 2.4×10-7 to 1.8×10-6). For most of the scenarios with the moderate VL (1×104.5mL-1), the PN scaled up to a low risk category (PN, 1.1×10-4 to 5.6×10-4), whereas for the scenarios with a high VL (1×107mL-1), the risk levels were high (PN, 2.3×10-2 to 3.5×10-2) or very high (PN, 1.1×10-1 to 1.6×10-1) depending on the volume of contaminated water spilled in the culture pond (VCWSCP, 4 or 20L). In the sensitivity analysis, for a SD of 30 PL/m2, it was shown that starting with a VL of 1×105mL-1 and a VCWSCP of 12L, the PN was moderate (1.05×10-3). This was the threshold for greater risks, given the increase in either the VCWSCP or VL. These findings supported recommendations to prevent WSSV spread through more controlled transportation and partial harvesting practices.

In vitro digestibility of specific dsRNA by enzymes of digestive tract of shrimp Litopenaeus vannamei / Digestibilidad in vitro del dsRNA específico por enzimas del tracto digestivo del camarón Litopenaeus vannamei

ABSTRACT Objective. The digestibility of specific dsRNA by action of the enzymes of digestive tract of the whiteleg shrimp Litopenaeus vannamei was determined in vitro. Materials and methods. Digestive enzyme activity (amylase, lipase, protease, DNase and RNase) was measured in the stomach, digestive gland, and anterior, middle, and posterior intestine of juvenile shrimp, and the digestibility of DNA, RNA and the dsRNA-ORF89, specific to WSSV, was determined by in vitro assays, as well as electrophoretic and densitometric analyses. Results. The highest enzymatic activity was found in the digestive gland: amylase (81.41%), lipase (92.60%), protease (78.20%), DNase (90.85%), and RNase (93.14%). The highest digestive capacity against DNA, RNA, and dsRNA was found in the digestive gland (5.11 ng of DNA ​​per minute, 8.55 ng of RNA per minute, and 1.48 ng dsRNA per minute). Conclusions. The highest digestibility of dsRNA-ORF89, specific to WSSV, was found in the digestive gland, whereas the lowest digestibility was observed in the posterior intestine. This is the first report regarding the digestibility of dsRNA-ORF89 by whiteleg shrimp digestive tract enzymes, with potential therapeutic importance in shrimp culture to prevent WSSV disease through balanced feed.

RESUMEN Objetivo. La digestibilidad del dsRNA específico para el virus de la mancha blanca (WSSV) por acción de las enzimas del tracto digestivo del camarón Litopenaeus vannamei fue analizada in vitro. Materiales y métodos. Se midió la actividad de enzimas digestivas (proteasa, amilasa, lipasa, ADNasa y ARNasa) en el estómago, la glándula digestiva, el intestino anterior, medio y posterior en juveniles de camarón patiblanco y se evaluó la digestibilidad de ácidos nucleicos ADN, ARN y dsRNA-ORF89 especifico contra el virus WSSV, por análisis electroforéticos y densitometría. Resultados. La actividad enzimática más alta se encontró en la glándula digestiva del camarón: amilasa (81.41%), lipasa (92.60%), proteasa (78.20%), ADNasa (90.85%) y ARNasa (93.14%). Se evidenció la capacidad digestiva del camarón patiblanco contra el ADN, ARN y dsRNA-ORF89 encontrando en la glándula digestiva la mayor digestión (5.11 ng de ADN por minuto, 8.55 ng de ARN por minuto y 1.48 ng de dsRNA por minuto). Conclusiones. La mayor digestibilidad del dsRNA-ORF89, específico contra el virus WSSV, se encontró en la glándula digestiva y la menor en el intestino posterior. Este es el primer informe relacionado con la digestibilidad del dsRNA-ORF89 por las enzimas del camarón patiblanco con potencial importancia terapéutica en el cultivo de camarón para prevenir la enfermedad del WSSV a través del alimento balanceado.

Comparative transcriptome and potential antiviral signaling pathways analysis of the gills in the red swamp crayfish, Procambarus clarkii infected with White Spot Syndrome Virus (WSSV)

Abstract Red swamp crayfish is an important model organism for research of the invertebrate innate immunity mechanism. Its excellent disease resistance against bacteria, fungi, and viruses is well-known. However, the antiviral mechanisms of crayfish remain unclear. In this study, we obtained high-quality sequence reads from normal and white spot syndrome virus (WSSV)-challenged crayfish gills. For group normal (GN), 39,390,280 high-quality clean reads were randomly assembled to produce 172,591 contigs; whereas, 34,011,488 high-quality clean reads were randomly assembled to produce 182,176 contigs for group WSSV-challenged (GW). After GO annotations analysis, a total of 35,539 (90.01%), 14,931 (37.82%), 28,221 (71.48%), 25,290 (64.05%), 15,595 (39.50%), and 13,848 (35.07%) unigenes had significant matches with sequences in the Nr, Nt, Swiss-Prot, KEGG, COG and GO databases, respectively. Through the comparative analysis between GN and GW, 12,868 genes were identified as differentially up-regulated DEGs, and 9,194 genes were identified as differentially down-regulated DEGs. Ultimately, these DEGs were mapped into different signaling pathways, including three important signaling pathways related to innate immunity responses. These results could provide new insights into crayfish antiviral immunity mechanism.

Virulence and genotypes of white spot syndrome virus infecting Pacific white shrimp Litopenaeus vannamei in north-western Mexico.

White spot syndrome virus (WSSV) has caused substantial global economic impact on aquaculture, and it has been determined that strains can vary in virulence. In this study, the effect of viral load was evaluated by infecting Litopenaeus vannamei with 10-fold serial dilution of tissue infected with strain WSSV Mx-H, and the virulence of four WSSV strains from north-western Mexico was assessed along with their variable number of tandem repeat (VNTR) genotypes in ORF75, ORF94 and ORF125. The LD50 of the Mx-H strain was a dilution dose of 10-7.5 ; the mortality titre was 109.2 LD50 per gram. In shrimp injected with 102.5 to 106.5 LD50 , no significant virulence differences were evident. Using mortality data, the four WSSV strains grouped into three virulence levels. The Mx-F strain (intermediate virulence) and the Mx-C strain (high virulence) showed more genetic differences than those observed between the Mx-G (low-virulence) and Mx-H (high-virulence) strains, in ORF94 and ORF125. The application of high-viral-load inocula proved useful in determining the different virulence phenotypes of the WSSV strains from the Eastern Pacific.

Hypoxia increases susceptibility of Pacific white shrimp to whitespot syndrome virus (WSSV) / Condição de hipóxia aumenta a susceptibilidade do camarão branco do Pacíficoao vírus da mancha branca (WSSV)

The present study aimed to evaluate the mortality, reactive oxygen species production (ROS) and total hemocyte counts (THC) of the marine shrimp Litopenaeus vannamei infected with the white spot syndrome virus (WSSV) at three levels of oxygen saturation. For this, 360 shrimp (20±2g) were distributed in 24 tanks (60L), divided in two groups (infected and non-infected), which were subjected to 30, 60 and 100% of dissolved oxygen saturation (in quadruplicate). During 96 hours after infection, daily hemolymph samples were collected for hemato-immunological parameter evaluation (THC and ROS) and dead animals were removed and computed to assess cumulative mortality rates. In the infected group, animals subjected to 100% saturation showed higher ROS production (P<0.05) after 48 hours, while THC was significantly reduced (P<0.05), regardless of oxygen saturation. The hypoxia resulted in high mortality when compared to 100% saturation condition. In the uninfected group, no significant differences were observed in all evaluated parameters. Thus, the hypoxia condition increased the susceptibility of shrimp to the infection of WSSV, which may be partly related to the low ROS production showed by the animals subjected to 30% oxygen saturation.(AU)

O presente estudo teve por finalidade avaliar a mortalidade e a contagem total de hemócitos (CTH) e espécies reativas de oxigênio (EROs) de camarão Litopenaeus vannamei infectados com o vírus da mancha branca (WSSV) e submetidos a três níveis de saturação de oxigênio. Para tanto, 360 camarões (20±2g) foram distribuídos em 24 tanques (60L), divididos em dois grupos, infectados e não infectados e submetidos a 30, 60 e 100% de saturação de oxigênio (em quadruplicata). Após a infecção, diariamente foram coletadas amostras de hemolinfa dos animais para avaliação dos parâmetros hematoimunológicos (CTH e EROs) e foi estimada a mortalidade, por 96 horas. No grupo com infecção, os animais submetidos à saturação de 100% apresentaram um aumento na produção de EROs (P<0,05) após 48 horas, ao mesmo tempo em que a CTH demonstrou uma redução (P<0,05) independentemente da saturação do oxigênio, e a condição de hipóxia acarretou maiores mortalidades quando comparada à do grupo com 100% de saturação. No grupo sem infecção, não foram observadas diferenças significativas nos parâmetros avaliados nem mortalidade. Dessa forma, pode-se concluir que a hipóxia aumentou a susceptibilidade do camarão à infecção com o vírus da mancha branca, que pode estar, em parte, relacionada com a baixa contagem de hemócitos e produção de EROs observadas nos animais submetidos a essa condição.(AU)

Hypoxia increases susceptibility of Pacific white shrimp to whitespot syndrome virus (WSSV) / Condição de hipóxia aumenta a susceptibilidade do camarão branco do Pacíficoao vírus da mancha branca (WSSV)

The present study aimed to evaluate the mortality, reactive oxygen species production (ROS) and total hemocyte counts (THC) of the marine shrimp Litopenaeus vannamei infected with the white spot syndrome virus (WSSV) at three levels of oxygen saturation. For this, 360 shrimp (20±2g) were distributed in 24 tanks (60L), divided in two groups (infected and non-infected), which were subjected to 30, 60 and 100% of dissolved oxygen saturation (in quadruplicate). During 96 hours after infection, daily hemolymph samples were collected for hemato-immunological parameter evaluation (THC and ROS) and dead animals were removed and computed to assess cumulative mortality rates. In the infected group, animals subjected to 100% saturation showed higher ROS production (P<0.05) after 48 hours, while THC was significantly reduced (P<0.05), regardless of oxygen saturation. The hypoxia resulted in high mortality when compared to 100% saturation condition. In the uninfected group, no significant differences were observed in all evaluated parameters. Thus, the hypoxia condition increased the susceptibility of shrimp to the infection of WSSV, which may be partly related to the low ROS production showed by the animals subjected to 30% oxygen saturation.

O presente estudo teve por finalidade avaliar a mortalidade e a contagem total de hemócitos (CTH) e espécies reativas de oxigênio (EROs) de camarão Litopenaeus vannamei infectados com o vírus da mancha branca (WSSV) e submetidos a três níveis de saturação de oxigênio. Para tanto, 360 camarões (20±2g) foram distribuídos em 24 tanques (60L), divididos em dois grupos, infectados e não infectados e submetidos a 30, 60 e 100% de saturação de oxigênio (em quadruplicata). Após a infecção, diariamente foram coletadas amostras de hemolinfa dos animais para avaliação dos parâmetros hematoimunológicos (CTH e EROs) e foi estimada a mortalidade, por 96 horas. No grupo com infecção, os animais submetidos à saturação de 100% apresentaram um aumento na produção de EROs (P<0,05) após 48 horas, ao mesmo tempo em que a CTH demonstrou uma redução (P<0,05) independentemente da saturação do oxigênio, e a condição de hipóxia acarretou maiores mortalidades quando comparada à do grupo com 100% de saturação. No grupo sem infecção, não foram observadas diferenças significativas nos parâmetros avaliados nem mortalidade. Dessa forma, pode-se concluir que a hipóxia aumentou a susceptibilidade do camarão à infecção com o vírus da mancha branca, que pode estar, em parte, relacionada com a baixa contagem de hemócitos e produção de EROs observadas nos animais submetidos a essa condição.

Delayed mortality of juvenile shrimp Penaeus vannamei challenged to White spot syndrome virus (WSSV) previously exposed to Infectious hypodermal and haematopoietic necrosis virus (IHHNV) or inactivated WSSV

In Ecuador, the development and sustainability of the cultured white shrimp, Penaeus vannamei, has been threatened by the occurrence of several viral pathogens, Infectious hypodermal and haematopoietic necrosis virus (IHHNV) and White spot syndrome virus (WSSV) mainly. The aim of the present study was to evaluate the exposition of P. vannamei juveniles to IHHNV and formalin-inactivated viruses (inactWSSV or inactIHHNV) to induce a protective response in P. vannamei juveniles against WSSV infection. P. vannamei were challenged to WSSV by intramuscular injection. Shrimp mortalities appeared at day 1 post-injection (p.i.) in positive control and inactIHHNV treatment, while IHHNV and inactWSSV treatments presented onset of mortalities at day 2 p.i. Positive control and inactIHHNV treatment presented 100% mortality at day 4 p.i., while IHHNV and inactWSSV treatments reached similar mortality at day 6 p.i. Statistical analysis revealed that WSSV-induced mortalities in juvenile P. vannamei of IHHNV and inactWSSV treatments had a significant delay (P 0.05) compared to both the inactIHHNV-treatment and positive control. Our results showed thatpreliminary exposure to IHHNV or to formalin-inactivated WSSV can induce delayed mortality in Penaeus vannameifollowing challenge with WSSV via intramuscular injection. In case of IHHNV infection, viral interference could be thebiological phenomenon involved, mediated by competition between IHHNV and WSSV. Regarding to WSSV inactivatedby formalin, a “vaccination” response would be responsible for the delay, evidencing a possible specific antiviral immuneresponse from the host.(AU)