Runt Deformity Syndrome Research Articles

Transcriptome analysis of IHHNV infection in Penaeus vannamei at different developmental stages

Infectious hypodermal and haematopoietic necrosis virus (IHHNV) is the smallest known virus in shrimp, which causes runt-deformity syndrome (RDS) and leads to huge economic loss every year in penaeid shrimp farming. Previous studies have shown that the juvenile Penaeus vannamei is more susceptible to IHHNV infection than the adults, but the mechanism is still unclear. In order to investigate the mechanism of pathogenic differences in IHHNV infection of P. vannamei at different developmental stages, the juvenile and adult P. vannamei were studied by transcriptome high-throughput sequencing to analyze their response to IHHNV infection. GO and KEGG enrichment were analyzed to search for differentially expressed genes (DEGs) related to immunity, growth and metabolism. The results showed that many immune-related genes of the juvenile and adult P. vannamei responded differently to IHHNV infection. For the adult P. vannamei, the expression of most immune-related genes was significantly up-regulated, which means that a cellular defense response was triggered after IHHNV infection. However, most immune-related genes in juvenile P. vannamei were inhibited, indicating that the immune system of juvenile the P. vannamei is imperfect and makes it to be more susceptible to IHHNV. Similarly, the growth-related genes of P. vannamei were changed during IHHNV infection. For the juvenile P. vannamei, the growth-related genes were significantly down-regulated, which resulted in a growth hormone disorder and prevented the juvenile P. vannamei from growth. In the adult P. vannamei, most molting-related genes were significantly up-regulated, indicating that IHHNV infection leads the adult P. vannamei to early molting to eliminate pathogen in the body. Metabolic process data showed that energy metabolism pathway was affected when P. vannamei infected with IHHNV. The adult P. vannamei infected with IHHNV can cause energetically costly and lead to the disturbance of the metabolism, activate complex immune systems to resist the invasion of pathogens. The results of this study clarified the response mechanism of P. vannamei at different developmental stages to IHHNV infection, which can provide new insights to IHHNV effective control and a reference for the study of sensitive period of different shrimp virus to host infection.

Complete Coding Genome Sequence of Infectious Hypodermal and Hematopoietic Necrosis Virus Isolated from Penaeus (Litopenaeus) vannamei Shrimp in Ecuador.

ABSTRACTInfectious hypodermal and hematopoietic necrosis virus (IHHNV), recognized as Penaeus stylirostris penstyldensovirus 1 (PstDV1), has been associated with the runt-deformity syndrome (RDS) in cultured Penaeus (Litopenaeus) vannamei and Penaeus monodon shrimp. Here, we report the second published genome sequence of IHHNV, which was isolated from P. vannamei shrimp cultured in Ecuador.

The rise of the syndrome – sub‐optimal growth disorders in farmed shrimp

AbstractThe phenomenon of slow growth in farmed shrimp was observed for the first time in 1989 in Penaeus vannamei as a clinical manifestation of runt deformity syndrome caused by infectious hypodermal and hematopoietic necrosis virus. Subsequently, it was recorded in Penaeus monodon during 2001–2002. Since then, the condition described as monodon slow growth syndrome (MSGS) has been reported from many countries. Though not leading to mortality, retarded growth at the pond level results in significant economic losses. Several potential pathogens have been identified from affected shrimp; however, no confirmed causal relationship has yet been established. Initially, P. vannamei farmed alongside slow growth‐affected P. monodon were unaffected by the condition. Further, experimental studies suggested that P. vannamei was not susceptible. However, in recent years, increasing incidence of severe growth retardation has been reported in farmed P. vannamei. Currently, slow growth is considered as one of the most impactful production‐limiting conditions affecting shrimp farming across Asia. Similar to MSGS, many infectious agents have been detected in slow growth‐affected P. vannamei, especially the microsporidian Enterocytozoon hepatopenaei. Further, genetic and environmental factors have been suggested to play a role. However, the exact cause of slow growth remains elusive, proposing that it may be a multi‐factor syndrome. This review tracks the trajectory of the condition over the period of significant expansion of shrimp farming and postulates that conventional single pathogen/single disease paradigms are insufficient to deal with chronic yield‐limiting syndromes. We suggest more realistic, multi‐dimensional aetiological consideration of syndromic and emerging diseases in shrimp aquaculture.

Studies on infectious myonecrosis virus (IMNV) and infectious hypodermal and hematopoietic necrosis virus (IHHNV) in cultured penaeid shrimp in Egypt.

The present study aimed to diagnose infectious myonecrosis virus (IMNV) and infectious hypodermal and hematopoietic necrosis virus (IHHNV) among cultured penaeid shrimp (Penaeus semisulcatus, n = 120) collected from private farms in 2 Egyptian provinces (Damietta and North Sinai) along the Mediterranean coast. The collected shrimp were subjected to clinical examination, histopathology, molecular characterization, and phylogenetic analysis. Most of the shrimp infected with IMNV showed a distinctive appearance resembling cooked shrimp and white necrosis on distal abdominal segments and tail fans. Simultaneously, IHHNV-infected cases displayed opaque abdominal muscles, white milky to buff mottling on the shell, and a pathognomonic runt-deformity syndrome. Histopathological examination of infected specimens revealed muscular edema, hemocyte infiltration, deformities, Zenker's necrosis, and eosinophilic intra-nuclear inclusion bodies (Cowdry type A). PCR results gave predictable amplicon sizes of 139 and 81 bp and confirmed the presence of IMNV and IHHNV with a total prevalence of 37.5 and 25%, respectively. A homology search by BLAST analysis showed that the retrieved isolates putatively belonged to IMNV and IHHNV based on 96.3 to 97% nucleotide identity to the corresponding open reading frame gene of each virus. The phylogenetic analysis clearly showed genetic similarity and cross-lineage between our isolates and other isolates from Egypt, the USA, Brazil, Indonesia, China, Korea, Taiwan, and Ecuador. In conclusion, gross inspection and histopathology may aid in the diagnosis of viral diseases; however, molecular tools are indispensable for confirming a possible infection. The current study recommends strict regulations during live shrimp transportation and implementing health control certificates over all imports and exports, especially in developing countries, including Egypt.

Classical Runt Deformity Syndrome Cases in Farmed Penaeus vannamei Along the East Coast of India

ABSTRACT Jagadeesan, V.; Ezhil Praveena, P.; Otta, S.K., and Jithendran, K.P., 2019. Classical runt deformity syndrome cases in farmed Penaeus vannamei along the east coast of India. In: Jithendran...

The Penaeus stylirostris densovirus capsid protein interacts with the Litopenaeus vannamei BCCIP protein

Viral capsid proteins play an important role in the viral infection process. To identify the cellular proteins in shrimp that interact with the Penaeus stylirostris densovirus capsid protein (PstDNV-CP), we constructed a yeast two-hybrid (Y2H) cDNA library of the muscle tissue of Litopenaeus vannamei, and hybridized the bait vector pGBKT7-CP with this library. Cloning and sequencing showed that the shrimp protein interacting with PstDNV-CP was a homolog of BRCA2 and CDKN1A(p21)-interacting protein (BCCIP). We named this protein L. vannamei BCCIP (LvBCCIP). Further analysis showed that LvBCCIP interacted with L. vannamei calmodulin (LvCaM). We validated the interactions between PstDNV-CP and LvBCCIP, and between LvBCCIP and LvCaM, with GST pulldown assays. The gene expression of LvBCCIP increased significantly after PstDNV challenge. In addition, the PstDNV titer of PstDNV-challenged shrimp was significantly reduced after LvBCCIP expression was inhibited using double-stranded RNA (dsRNA) interference. These results indicated that LvBCCIP is critical to PstDNV pathogenesis in L. vannamei. Interestingly, the growth rate of L. vannamei was significantly reduced when LvBCCIP gene expression was silenced, indicating that LvBCCIP may also be associated with growth regulation in L. vannamei. Thus, the interaction between PstDNV-CP and LvBCCIP might explain why PstDNV infection leads to runt-deformity syndrome in shrimp.

Real-time PCR tests to specifically detect IHHNV lineages and an IHHNV EVE integrated in the genome of Penaeus monodon.

Infectious hypodermal and hematopoietic necrosis virus (IHHNV) can cause mass mortalities in western blue shrimp Penaeus stylirostris, runt deformity syndrome in Pacific white shrimp P. vannamei and scalloped abdominal shell deformities in black tiger shrimp P. monodon. In P. monodon, however, PCR-based diagnosis of IHHNV can be complicated by the presence of a chromosome-integrated, non-replicating endogenous viral element (EVE). To facilitate high-throughput screening of P. monodon for IHHNV infection and/or EVE sequences, here we report real-time PCR tests designed to specifically detect IHHNV Lineage I, II and III but not EVE Type A sequences and vice versa. Using 108 dsDNA copies of plasmid (p)DNA controls containing either IHHNV or EVE-Type A sequences, both tests displayed absolute specificity. The IHHNV-q309 PCR reliably detected down to ≤10 copies of pDNA, at which levels a 309F/R PCR amplicon was just detectable, and the presence of an IHHNV-EVE sequence did not significantly impact its sensitivity. The IHHNV-qEVE PCR was similarly sensitive. Testing of batches of P. monodon clinical samples from Vietnam/Malaysia and Australia identified good diagnostic concordance between the IHHNV-q309 and 309F/R PCR tests. As expected for a sequence integrated into host chromosomal DNA, IHHNV-qEVE PCR Ct values were highly uniform among samples from shrimp in which an EVE was present. The highly specific and sensitive IHHNV-q309 and IHHNV-qEVE real-time PCR tests described here should prove useful for selecting broodstock free of IHHNV infection and in maintaining breeding populations of P. monodon specific pathogen free for IHHNV, and if desired, also free of IHHNV-EVE sequences.

High Occurrence of the Decapod Penstyldensovirus (PstDV1) Detected in Postlarvae of Penaeus vannamei Produced in Commercial Hatcheries of Mexico.

The decapod penstyldensovirus (PstDV1) is a widely spread shrimp pathogen that causes high mortalities in the shrimp Penaeus stylirostris, while in P. vannamei, it has been associated with induction of the runt deformity syndrome. Using shrimp post-larvae (PL, stages PL13-PL21) collected from 16 commercial hatcheries from Mexico, and a sensitive PCR protocol for its detection, a survey of the PstDV1 prevalence in larvae was undertaken. A high overall prevalence of PstDV1 (49.5%) in shrimp PL from the studied hatcheries was found. This study reveals that PstDV1 occurs persistently in PL populations, which may have significant implications for its dispersal.

Prediksi Resistensi Udang Vaname (Litopenaus vannamei) terhadap Infectious Hypodermal and Hematopoietic Necrosis Virus (IHHNV) dari Tambak Intensif dan Semi Intensif Jepara Menggunakan Marka RAPD

Infectious Hypodermal and Hematopoietic Necrosis Infectious Virus (IHHNV) is the most important DNA virus which can lead to Runt Deformity Syndrome (RDS) in vaname shrimp. The RAPD technique can be used to determine the resistance of a species to a disease. This research aimed to screen and identify RAPD markers which could distinguish the resistance of vaname shrimp to IHHNV reared at intensive and semi-intensive pond. The DNA template was amplified by PCR using 5 primers : OPA 06, OPA 08, OPA 19, OPD-02 and OPZ-15. The results showed that only the primer OPA-19 and OPZ-15 were able to produce 100% polymorphic bands with sizes from 400-1700 bp as well as showing the resistance IHHNV in vaname shrimp. Based on these results, vaname shrimp which reared at the intensive pond were more resistant to IHHNV compared with the semi-intensive pond. Key words: RAPD, shrimp vaname, IHHNV, intensive & semi-intensive pond

Rapid and sensitive detection of infectious hypodermal and hematopoietic necrosis virus by loop-mediated isothermal amplification combined with a lateral flow dipstick

Infectious hypodermal and hematopoietic necrosis virus (IHHNV) is an important shrimp pathogen that causes mortality in Penaeus stylirostris and stunting (called runt deformity syndrome or RDS) in Penaeus vannamei. Loop-mediated isothermal amplification (LAMP) allows rapid amplification of nucleic acids under isothermal conditions. It can be combined with a chromatographic lateral flow dipstick (LFD) for highly specific, rapid and simple visual detection of IHHNV-specific amplicons. Using this protocol, a 30-min amplification followed by 5 min hybridization with an FITC-labeled DNA probe and 5 min LFD resulted in visualization of DNA amplicons trapped at the LFD test line. Thus, 10 min for rapid DNA extraction followed by LAMP combined with LFD detection resulted in a total assay time of approximately 50 min. Detection sensitivity was comparable to other methods used commonly for nested PCR detection of IHHNV but had the additional advantages of reduced assay time, confirmation of amplicon identity by hybridization and elimination of electrophoresis with carcinogenic ethidium bromide.

Different responses to infectious hypodermal and hematopoietic necrosis virus (IHHNV) in Penaeus monodon and P. vannamei

Infectious hypodermal and hematopoietic necrosis virus (IHHNV) is widespread in cultured Penaeus monodon and P. vannamei in Thailand. It causes runt-deformity syndrome that is characterized by physical abnormalities and stunted growth in P. vannamei, but causes no apparent disease in P. monodon. In both species, the virus may produce Cowdry Type A inclusions in tissues of ectodermal and mesodermal origin, but these are common in P. vannamei and rare in P. monodon. The virus can be more easily detected in both species by IHHNV-specific PCR primers. By in situ hybridization (ISH) using specific IHHNV probes, fixed phagocytes associated with myocardial cells tended to show strong positive reactions in both shrimp species. Ovarian and neural tissue (neurons in the nerve ganglia and glial cells in the nerve cord) were ISH positive for IHHNV only in P. vannamei. By transmission electron microscopy, necrotic cells were found in the gills of IHHNV-infected P. vannamei, while paracrystalline arrays of virions and apoptotic cells rather than necrotic cells were found in the lymphoid organ of IHHNV-infected P. monodon. Thus, it is possible that apoptosis in P. monodon contributes to the absence of clinical disease from IHHNV. These findings reveal different responses to IHHNV infection by the 2 shrimp species. A curious feature of IHHNV infection in P. monodon was inconsistency in the comparative viral load amongst tissues of different specimens, as detected by both ISH and real-time PCR. This inconsistency in apparent tissue preference and the reasons for different cellular responses between the 2 shrimp species remain unexplained.

RUNT-DEFORMITY SYNDROME IN CULTURED GIANT TIGER PRAWN PENAEUS MONODON

Abstract A total of 24 morphological abnormalities associated with the Runt-Deformity Syndrome (RDS) is reported for the first time in 17- to 18-mo old F3 generation Penaeus monodon (>40 mm CL) belonging to three breeding families. Although diagnosed as positive for the infectious hypodermal and hematopoietic necrosis virus (IHHNV), the shrimp appeared normal at 9 to 10 mo when they were procured from a commercial facility in Antique, central Philippines, in September 1996. The abnormalities affected either specific shrimp organs and appendages (e.g., rostrum, antennae, uropods) or the whole body (e.g., shell color and hardness). A two-way analysis of variance was used to evaluate differences in the morphometric characters rostral length (RL), carapace length (CL), abdominal length (AL), body length (BL), total length (TL), carapace width (CW), 1st abdominal segment width (AS1W), and body weight (BW) and the morphometric ratios RL:CL, CL:AL, and CW:AS1W by sex and stock (3 families and wild controls). Fem...

Runt-Deformity Syndrome in Cultured Giant Tiger Prawn Penaeus Monodon

A total of 24 morphological abnormalities associated with the Runt-Deformity Syndrome (RDS) is reported for the first time in 17- to 18-mo old F3 generation Penaeus monodon (>40 mm CL) belonging to three breeding families. Although diagnosed as positive for the infectious hypodermal and hematopoietic necrosis virus (IHHNV), the shrimp appeared normal at 9 to 10 mo when they were procured from a commercial facility in Antique, central Philippines, in September 1996. The abnormalities affected either specific shrimp organs and appendages (e.g., rostrum, antennae, uropods) or the whole body (e.g., shell color and hardness). A two-way analysis of variance was used to evaluate differences in the morphometric characters rostral length (RL), carapace length (CL), abdominal length (AL), body length (BL), total length (TL), carapace width (CW), 1st abdominal segment width (AS1W), and body weight (BW) and the morphometric ratios RL:CL, CL:AL, and CW:AS1W by sex and stock (3 families and wild controls). Female P. monodon had significantly larger CL, AL, BL, TL, CW, and BW than males, regardless of stock. Wild females had longer BL and TL than cultured ones, whereas wild male shrimp had shorter CL, CW, and BW than cultured male P. monodon. Cultured shrimp had significantly higher CL:AL and CW:AS1W ratios and lower RL:CL ratio compared to wild P. monodon. The shorter and narrower abdomen relative to the carapace gave the shrimp a “runted” or dwarf appearance characteristic of RDS earlier described in Litopenaeus vannamei with IHHNV. Confirmed by parallel studies, IHHNV infection of the cultured P. monodon may account for their cuticular deformities, slower growth, and smaller sizes. The appearance of deformities in older (and bigger) P. monodon suggests that RDS expression is dependent on age and size. Examination for external abnormalities and evaluation of the three morphometric ratios may complement existing screening protocols for diseases and growth rates in shrimp breeding programs. Cultured giant tiger prawn also had lower sperm count, greater proportion of dead and abnormal sperm, and lower incidence of mating (absence of sperm in thelyca) compared to wild P. monodon.

The Penaeid Shrimp Viruses TSV, IHHNV, WSSV, and YHV

ABSTRACT Viral diseases have emerged during this decade as serious economic impediments to successful shrimp farming. While nearly 20 distinct viruses or groups of viruses are known to infect penaeid shrimp, only four, WSSV, YHV, IHHNV, and TSV, pose a threat to the future of penaeid shrimp culture in the Western Hemisphere. TSV and IHHNV have caused serious disease epizootics throughout the Americas and Hawaii. IHHNV was described nearly 20 years ago when it was found to be responsible for cumulative losses of cultured Penaeus stylirostris that often exceed 90%. The threat of high mortalities posed by IHHNV has historically curtailed interest in the culture of this species in favor of the more IHHNV-resistant P. vannamei. While relatively resistant, IHHNV nonetheless infects P. vannamei and causes runt deformity syndrome (RDS) in which affected shrimp display reduced growth, highly variable sizes, lower production, and sometimes reduced survival. Taura Syndrome in P. vannamei is the virtual “mirror image” of IHHN disease in P. stylirostris. Following its recognition in 1992 as a distinct disease of P. vannamei in Ecuador, Taura Syndrome and its viral agent TSV spread rapidly throughout many of the shrimp farming regions of the Americas. Cumulative mortalities due to TSV in affected juvenile P. vannamei populations have ranged from 40 to 95%. Because P. stylirostris was found to be innately TSV-resistant, genetically selected IHHNV-resistant lines of this species disease are being developed and marketed in the Americas. Some shrimp farmers are using domesticated, selected stocks of P. vannamei that show improved resistance to TSV, while others have continued to use wild stocks that are also showing increased resistance, perhaps through intense natural selection. The viruses of the Yellow Head Disease (YHV) and the White Spot Syndrome (WSSV) were first recognized in 1991-92 in Asia, and by 1996 the two diseases had spread and had caused major pandemics throughout much of the shrimp farming regions of East Asia, Southeast Asia, Indonesia, and India. In late 1995, WSSV was found in North America for the first time in P. setiferus at a shrimp farm in south Texas. WSSV has since been detected in cultured and wild shrimp, crabs, and freshwater crayfish at multiple sites in the eastern and southeastern U.S., as well as being commonly found (along with YHV) in imported frozen commodity shrimp. Because Western Hemisphere penaeids are highly susceptible to WSSV and YHV, the introduction and establishment of either of the viruses poses a significant threat to the shrimp farming industry. The available detection methods for IHHNV, TSV, WSSV, and YHV include traditional methods that employ gross signs, clinical history, histopathology, and bioassay with susceptible shrimp hosts. Molecular and serological methods have also been developed, and specific gene probes, monoclonal antibodies, or PCR methods are readily available for each of the four viruses.

IHHN VIRUS AND INTENSIVE CULTURE OF PENAEUS VANNAMEI: EFFECTS OF STOCKING DENSITY AND WATER EXCHANGE RATES

Producers and researchers have long suspected correlations between runt deformity syndrome (RDS), morphological anomalies, poor production, and viral infection for various penaeid shrimp species. Unfortunately, evidence has been circumstantial and little research has been focused on defining harvest size distributions as it relates to viral histopathology. The present study highlights these elements, in a factorial experimental design, using increased stocking density (60/m2 and 100/m2) and decreased water exchange (100%, 50%, and 10% per day) as factors. Twelve 29.2-m2 tanks were stocked with suspected IHHNV-infected Penaeus vannamei postlarvae from a Central American hatchery, and were harvested after 164 days. Low dissolved oxygen levels resulted in poor survival (43.5% and 53.6%) in tanks which were stocked at 100/m2 and limited to 10% water exchange per day. Density and water exchange regimes had little effect on water quality, survival, or growth in other experimental groups. A runt deformity syndrome and other physical anomalies are described and the former is correlated with increased incidence of IHHNV associated Cowdry type A inclusion bodies in 4 organ systems. A trend toward decreasing incidence of inclusion bodies was found as exchange rates were increased from 50% to 100% per day. Distributions of size at harvest indicate less skewness and kurtosis as well as lower incidence of deformities in low survival tanks, suggesting increased mortality of more severely infected RDS shrimp.

IHHN Virus as an Etiological Factor in Runt‐Deformity Syndrome (RDS) of Juvenile Penaeus vannamei Cultured in Hawaii

AbstractRuntdeformity syndrome (RDS) is an economically significant, frequent disease problem of cultured Penaeus vannamei. RDS is characterized by variable, often greatly reduced, growth rate of up to 30% of a cultured population and many shrimp with cuticle deformities of the rostrum, anterior appendages or other parts. The cause of RDS is undetermined.Nursery trials comparing histologically IHHN‐positive and histologically IHHN‐negative Penaeus vannamei cultured under identical conditions were conducted at The Oceanic Institute. The IHHN‐positive populations developed RDS symptoms including increased individual size variation, reduced population growth, and high prevalance of rostrum, antennae or cuticle deformity, while the IHHN‐negative groups displayed none of these symptoms. No other diseases or parasites were identified in the IHHN‐positive populations that would account for the RDS symptoms.A single commercialele nursery was stocked with histologidly IHHN‐positive P. vannamei and a high prevalence of RDS was observed. Shrimp growth was bimodal with a subpopulation growing normally and a subpopulation growing slowly. Significant relationships between shrimp harvest size and IHHN prevalance, selected individual organ IHHN severity grades and IHHN severity index (six organs evaluated) were found. Large, apparently normal shrimp were less severely infected with virus and did not display any cuticle deformities.In sum, these data provide epidemiological and histopathological evidence for the hypothesis that infection by IHHN virus is the cause of RDS in cultured P. vannamei.