Shrimp Pathogens Research Articles

Genomic identification and characterization of co-occurring Harveyi clade species following a vibriosis outbreak in Pacific white shrimp, Penaeus (litopenaeus) vannamei

Shrimp farming is one of the fastest-growing food production sectors but disease related mortality remains a limitation to industry expansion. Bacterial species belonging to the Harveyi clade, such as Vibrio harveyi and Vibrio parahaemolyticus, have long been implicated as agents of disease in shrimp aquaculture but traditional methods cannot accurately differentiate these species. Following a vibriosis outbreak in Pacific white shrimp, Penaeus (litopenaeus) vannamei, we hypothesized that infection promoted the co-occurrence of multiple Harveyi clade species. The shrimp were reared in a zero-exchange raceway using biofloc technology and presumptive Vibrio species were isolated from the hepatopancreases of moribund shrimp by culture on a Vibrio-selective agar. Isolates were identified and characterized by PCR-based genotyping and whole-genome sequencing. A phylogenetic tree based on a 43-genome pangenome analysis revealed the presence of five known shrimp pathogens: Photobacterium damselae, Vibrio alginolyticus, V. harveyi, V. parahaemolyticus, Vibrio rotiferianus, and a potentially novel Vibrio species (Vibrio sp. Hep-1b-8). Multilocus sequence typing revealed that P. damselae and V. parahaemolyticus isolates exhibited novel allelic profiles, although the V. parahaemolyticus isolates were closely related to strains isolated from Southeastern Asia. Further, a novel comparative analysis of 23 type VI secretion systems revealed three phylogenetically distinct systems, which suggests that co-occurring bacteria possess diverse mechanisms for interspecies competition. These results highlight whole-genome sequencing as an invaluable tool for identifying and characterizing co-occurring, infection-associated bacteria at the species, strain and molecular level.

The aquatic animal pandemic crisis.

The growth of aquaculture over the past 50 years has been accompanied by the emergence of aquatic animal diseases, many of which have spread to become pandemic in countries or continents. An analysis of 400 emerging disease events in aquatic animals that were logged by the Centre for Environment, Fisheries and Aquaculture Science between 2002 and 2017 revealed that more than half were caused by viruses. However, in molluscs, most events were parasitic. Categorising these events indicated that the key processes underpinning emergence were the movement of live animals and host switching. Profiles of key pathogens further illustrate the importance of wild aquatic animals as the source of new infections in farmed animals. It is also clear that the spread of new diseases through the largescale movement of aquatic animals for farming, for food and for the ornamental trade has allowed many to achieve pandemic status. Many viral pathogens of fish (e.g. infectious salmon anaemia, viral haemorrhagic septicaemia) and shrimp (e.g. white spot syndrome virus) affect a large proportion of the global production of key susceptible species. Wild aquatic animal populations have also been severely affected by pandemic diseases, best exemplified by Batrachochytrium dendrobatidis, a fungal infection of amphibians, whose emergence and spread were driven by the movement of animals for the ornamental trade. Batrachochytrium dendrobatidis is now widespread in the tropics and subtropics and has caused local extinctions of susceptible amphibian hosts. Given the rising demand for seafood, aquacultural production will continue to grow and diseases will continue to emerge. Some will inevitably achieve pandemic status, having significant impacts on production and trade, unless there are considerable changes in global monitoring and the response to aquatic animal diseases.

Effect of biosurfactant derived from Vibrio natriegens MK3 against Vibrio harveyi biofilm and virulence.

Vibrio harveyi is a marine luminous pathogen, which causes biofilm-mediated infections, pressures the search for an innovative alternate approach to strive against vibriosis in aquaculture. This study anticipated to explore the effect of glycolipid biosurfactant as an antipathogenic against V. harveyi to control vibriosis. In this study, 27 bacterial strains were isolated from marine soil sediments. Out of these, 11 strains exhibited surfactant activity and the strain MK3 showed high emulsification index. The potent strain was identified as Vibrio natriegens and named as V. natriegens MK3. The extracted biosurfactant was purified using high-performance liquid chromatography and it was efficient to decrease the surface tension of the growth medium up to 21 mN/m. The functional group and composition of the biosurfactant were determined by Fourier-transform infrared spectroscopy and nuclear magnetic resonance spectroscopy spectral studies and the nature of the biosurfactant was identified as glycolipid. The surfactant was capable of reducing the biofilm formation, bioluminescence, extracellular polysaccharide synthesis, and quorum sensing in marine shrimp pathogen V. harveyi. The antagonistic effect of biosurfactant was evaluated against V. harveyi-infected brine shrimp Artemia salina. This study reveals that biosurfactant can be considered for the management of biofilm-related aquatic infections.

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.

IMPACT OF WHITE SPOT SYNDROME VIRUS DISEASE ON GILLS IN SHRIMP, PENAEUS MONODON

Protozoan parasites cause problems in Penaeid shrimp culture and major impact on the shrimp farming industry.White spot syndrome virus (WSSV) is a major shrimp pathogen that also infects Penaeus monodon species was selected. Histological changes in the gill cells were studied. Histological techniques using paraffin embeded tissues as well as frozen tissues were used for identification of WSSV infection. Histological manifestation such as gill cells could be detected. The gill cells were marked and characteristic of WSSV infections were observed. The present investigation was focused on the virus infecting shrimp. The microscopic examinations of the histological preparation are presently used to detect WSSV zoothamnium.

Detection of Enterocytozoon hepatopenaei using an invasive but non-lethal sampling method in shrimp (Penaeus vannamei)

The detection of enteric pathogens that cause diseases in shrimp involves the sacrifice of the host to obtain tissue samples for diagnosis. In this study, we describe an invasive but non-lethal sampling methodology using a syringe to collect biopsy samples from the hepatopancreas (HP) of Penaeus vannamei to detect the microsporidian pathogen, Enterocytozoon hepatopenaei (EHP), by qPCR and transmission electron microscopy. EHP was detected in all the infected shrimp by qPCR. The shrimp infected by the microsporidian showed 65% survival at 7 days post-sampling. Transmission electron microscopic examination of the biopsy samples revealed numerous spores of the pathogen. The presence of EHP was further confirmed by histology and in situ hybridization from HP tissue samples. The data shows that a hepatopancreas biopsy could be a viable means of detecting enteric pathogens in shrimp, and the method could be valuable in sampling broodstock and natural populations without the need to sacrifice the animals.

A future vision for disease control in shrimp aquaculture

Aquaculture is the fastest‐growing animal production sector, and shrimp production already exceeds that of the capture fishery. Viruses and bacteria account for the majority of disease losses for shrimp farmers. Viral pandemics in the mid 1990s and, more recently, a bacterial pandemic from 2009 to 2015 have led to the conclusion that future, sustainable shrimp aquaculture will depend on the development of more efficient, biosecure production facilities that cultivate specific pathogen‐free (SPF) shrimp, genetically improved for growth and disease tolerance or resistance. Major requirements for development, maintenance, and use of SPF stocks in aquaculture are effective pathogen surveillance and disease prevention methods. When protective measures fail and diseases occur in production ponds, there are currently only a few approved and practical therapeutic methods available for use with bacterial pathogens and none so far for viral pathogens. To improve existing methods of prevention and therapy and to develop new ones, research is being carried out on the nature of shrimp–pathogen interactions. Promising results have been obtained at the laboratory level for possible applications involving the use of immunostimulants for “immune priming” or “trained immunity” of RNA interference and of endogenous viral elements. Some of these promising new directions are discussed.

Crustacean Genome Exploration Reveals the Evolutionary Origin of White Spot Syndrome Virus.

White spot syndrome virus (WSSV) is a crustacean-infecting, double-stranded DNA virus and is the most serious viral pathogen in the global shrimp industry. WSSV is the sole recognized member of the family Nimaviridae, and the lack of genomic data on other nimaviruses has obscured the evolutionary history of WSSV. Here, we investigated the evolutionary history of WSSV by characterizing WSSV relatives hidden in host genomic data. We surveyed 14 host crustacean genomes and identified five novel nimaviral genomes. Comparative genomic analysis of Nimaviridae identified 28 "core genes" that are ubiquitously conserved in Nimaviridae; unexpected conservation of 13 uncharacterized proteins highlighted yet-unknown essential functions underlying the nimavirus replication cycle. The ancestral Nimaviridae gene set contained five baculoviral per os infectivity factor homologs and a sulfhydryl oxidase homolog, suggesting a shared phylogenetic origin of Nimaviridae and insect-associated double-stranded DNA viruses. Moreover, we show that novel gene acquisition and subsequent amplification reinforced the unique accessory gene repertoire of WSSV. Expansion of unique envelope protein and nonstructural virulence-associated genes may have been the key genomic event that made WSSV such a deadly pathogen.IMPORTANCE WSSV is the deadliest viral pathogen threatening global shrimp aquaculture. The evolutionary history of WSSV has remained a mystery, because few WSSV relatives, or nimaviruses, had been reported. Our aim was to trace the history of WSSV using the genomes of novel nimaviruses hidden in host genome data. We demonstrate that WSSV emerged from a diverse family of crustacean-infecting large DNA viruses. By comparing the genomes of WSSV and its relatives, we show that WSSV possesses an expanded set of unique host-virus interaction-related genes. This extensive gene gain may have been the key genomic event that made WSSV such a deadly pathogen. Moreover, conservation of insect-infecting virus protein homologs suggests a common phylogenetic origin of crustacean-infecting Nimaviridae and other insect-infecting DNA viruses. Our work redefines the previously poorly characterized crustacean virus family and reveals the ancient genomic events that preordained the emergence of a devastating shrimp pathogen.

A Magnetic Bead-Based DNA Extraction Protocol Suitable for High-Throughput Genotyping in Shrimp Breeding Programs

Due to their convenience, magnetic bead-based nucleic acid extraction kits are commonly used in shrimp genotyping and pathogen screening applications. However, in advanced breeding programs requiring the testing of many thousands of shrimp, their cost can be prohibitive. Various permutations of different Proteinase K digestion, tissue lysis and bead washing buffers as well as magnetic bead types were thus evaluated to devise a high-throughput shrimp DNA extraction (SDE) protocol capable of recovering high-purity DNA using a KingFisher™ Flex Magnetic Particle processor. When genotyped using a MassARRAY® platform (Agena Bioscience) requiring 60-61 genome regions to be co-amplified in a single multiplexed PCR, DNA extracted from shrimp muscle tissue using either the SDE protocol or a commercial kit generated comparable single-nucleotide polymorphism (SNP) call data. The SDE protocol also extracted high-purity DNA from salmon fin clips. It thus offers potential to markedly reduce the costs of large-scale genotyping in shrimp and salmon breeding programs.

Polychaetes as Potential Risks for Shrimp Pathogen Transmission

Polychaetes comprise the benthic meiofauna of the soft-bottom intertidal zone and of shrimp ponds in coastal areas. While polychaetes provide benefits to the shrimp farming industry as (i) natural food in traditional shrimp ponds, (ii) nutrient regenerators through bioturbation and removal of organic waste in the sediment through feeding, and (iii) feed supplement to enhance maturation of shrimp brooders, the conditions present in aquaculture ponds may increase the opportunity for polychaetes to transfer pathogens to shrimp through the food chainThere is growing concern that internationally traded polychaetes, which are fed to shrimp brooders, are potential vectors for the transmission of other shrimp pathogens. The detection of the aetiological agents of two newly emerging diseases of shrimp in polychaetes, Enterocytozoon hepatopenaei (EHP) and Vibrio parahaemolyticusAHPND causing hepatopancreatic microsporidiosis (HPM) and acute hepatopancreatic necrosis disease (AHPND), respectively, suggests that these worms can be a host or/and passive carrier of these pathogens. This review discusses the benefits of polychaetes to shrimp farming, the risk of shrimp pathogen transmission by polychaetes at the pond, hatchery and global level, and calls for closer observation on shrimp pathogens in polychaetes used as shrimp feed.

Multiplex SYBR Green and duplex TaqMan real-time PCR assays for the detection of Photorhabdus Insect-Related (Pir) toxin genes pirA and pirB

Acute hepatopancreatic necrosis disease (AHPND), also known as Early mortality syndrome (EMS), is a recently emerged lethal disease that has caused major economic losses in shrimp aquaculture. The etiologic agents are Vibrio spp. that carry Photorhabdus Insect-Related (Pir) toxin genes pirA and pirB. A multiplex SYBR Green real-time PCR was developed that detects pirA, pirB, and two internal control genes, the shrimp 18S rRNA and the bacterial 16S rRNA genes in a single reaction. The pirB primers amplify the 3'-end of the pirB gene allowing the detection of Vibrio spp. mutants that contain a complete deletion of pirA and the partial deletion of pirB. The assay also detects mutants that contain the entire pirA gene and the deletion of the pirB gene. Since both toxin genes are needed for disease development, this assays can distinguish between pathogenic strains of Vibrio spp. that cause AHPND in shrimp and mutants that do not cause disease. The amplicons for pirA, pirB, 18S rRNA and 16S rRNA showed easily distinguishable melting temperatures of 78.21 ± 0.18, 75.20 ± 0.20, 82.28 ± 0.34 and 85.41 ± 0.21 °C respectively. Additionally, a duplex real-time PCR assay was carried out by designing TaqMan probes for the pirA and pirB primers. The diagnostic sensitivity and specificity was compared between the SYBR Green and TaqMan assays. Both assays showed similar sensitivity with a limit of detection being 10 copies for pirA and pirB, and neither assays showed any cross reaction with other known bacterial and viral pathogens in shrimp. The high sensitivity of both assays make them suitable for the detection of low copies of the pirA and pirB genes in AHPND causing Vibrio spp. as well as for detecting non-pathogenic mutants.

The Penaeus stylirostris densovirus capsid interacts with Litopenaeus vannamei troponin I

The Penaeus stylirostris densovirus (PstDNV) (also known as infectious hypodermal and hematopoietic necrosis virus, IHHNV), a very small DNA virus, is a major shrimp pathogen. The PstDNV genome encodes only two nonstructural proteins and one capsid protein. This virus is thus an ideal, simple model for the investigation of virus-host interactions. To explore the role of the PstDNV capsid in viral infections, a yeast two-hybrid (Y2H) cDNA library was constructed based on Pacific white shrimp, Litopenaeus vannamei mRNA. The Y2H library was then screened, using the PstDNV capsid protein as bait. We identified a host protein that interacted strongly with the PstDNV capsid as L. vannamei troponin I (LvTnI). An in vitro co-immunoprecipitation experiment further supported this interaction. In addition, an in vivo neutralization experiment showed that the vaccination with anti-LvTnI significantly reduced PstDNV copies in PstDNV-challenged shrimp, indicating that the interaction between the PstDNV capsid and cellular LvTnI is essential for PstDNV infection. This result has important implications for our understanding of the mechanisms by which PstDNV infects shrimp.

Facts, truths and myths about SPF shrimp in Aquaculture

AbstractShrimp domestication and genetic improvement programmes began in late 1980s, in the United States of America, under the United States Marine Shrimp Farming Program (USMSFP), using the Pacific whiteleg shrimp Penaeus vannamei. The USMSFP was based on proven concepts from the livestock and poultry industries and began with establishing a specific pathogen‐free (SPF) shrimp stock. The original shrimp stock was obtained using rigorous screening of captured wild shrimp for selection of individuals naturally free of major shrimp pathogens. Although the concept of SPF animals was well defined for terrestrial animals, it was relatively new for aquaculture, and it took some time to be adopted by the aquaculture community. In the early 1990s, parallel to USMSFP, several other programmes on genetic improvement of shrimp were also initiated in Latin America. Subsequently, several new terminologies and products, such as specific pathogen resistant (SPR) shrimp, specific pathogen tolerant (SPT) shrimp and even ‘all pathogen exposed’ (APE) shrimp, entered the shrimp industry vocabulary and became commercial. This led to confusion in the shrimp industry about the meaning, relationship and significance of these new terms with respect to SPF. This position paper attempts to clarify these concepts, provide science‐based definitions, reconfirms the importance of developing, maintaining and using domesticated, specific pathogen‐free (SPF) shrimp stocks (which may also achieve SPR and/or SPT status) to reduce the risk of disease outbreaks and increase production and profit. The same principles would apply to development of domesticated SPF stocks for other species used in aquaculture. The paper also discusses the difficulties of confirming and certifying SPF status due to the presence of endogenous viral elements (EVEs) and calls for internationally agreed science and evidence‐based technical guidelines for producing healthy shrimp.

Screening of intestinal probiotics and the effects of feeding probiotics on the growth, immune, digestive enzyme activity and intestinal flora of Litopenaeus vannamei

In this experiment, 426 strains were isolated from the intestinal tract of Litopenaeus vannamei, and 11 strains showed strong digestive enzyme production activity and antagonistic effect against common bacterial pathogens of shrimp. After hemolysis activity test and drug sensitivity test, 2 candidate probiotics with good bacteriostatic activity, strong enzyme production ability and relatively sensitive to antibiotics were screened out, and were identified by 16s rDNA molecular identification and Biolog-System as Enterobacter hominis (E3) and lactobacillus (L3). First, the biological characteristics of 2 candidate probiotics were studied. The optimum growth conditions of E3: temperature, 30 °C; pH, 8.0; NaCl, 2.5%; bovine bile salt, 0.15%; and the optimum growth conditions of L3: temperature, 40 °C; pH, 6.0; NaCl, 0.5%; bovine bile salt, 0.0015%. Secondly, a 28-day feeding experiment was conducted using probiotic concentration of 107 CFU g−1 to determine the changes of the activities of blood related immune enzymes (SOD, PPO, ACP, POD, CAT, LZM) and intestinal digestive enzymes (NP, AL, LPS) during the feeding process of shrimp, the results showed that during the course of feeding, the activities of immune enzyme and digestive enzyme of shrimp fed with probiotics showed an increasing trend, and the growth rate of body weight of shrimp was higher than that of control group. After feeding, the cumulative mortality of probiotics groups were significantly lower than that of the control group after WSSV infection. And the mid-gut of L. vannamei was observed by electron microscope, the results showed that the intestinal mucosa was tight and the epithelium cells showed an active secretory state in probiotics group. Finally, the intestinal microbial communities of shrimp were compared and analyzed by using Biolog-ECO method in the later period of feeding, the results showed: compared with the control group, the average color change rate of the experimental group fed with probiotics increased significantly, indicating that probiotics enhanced the intestinal microorganism activity; The ability of intestinal microorganism to utilize carbon source was significantly enhanced in the experimental group, which indicated that the digestive enzyme secreted by probiotics could improve the digestion and absorption rate of prawn feed, thus promoting the rapid growth of shrimp; The Shannon index, Simpson index and McIntosh index of probiotics groups showed significant difference in 1st and 5th days, but tended to be the same in the 10th day, the results showed that probiotics could maintain in L. vannamei intestines at least 5 days.

Comparison of bacterial community structures in digestive tract between healthy and disease whiteleg shrimp (Litopenaeus vannamei) in Soc Trang, Vietnam

Gut bacteria comprise a complex bacterial community related to many functions in a host. The stability of gut bacteria plays important models in the health and immunology of a host. Many studies on intestine bacteria constructed via cultivation and Denaturation Gradient Gel Electrophoresis (DDGE) methods have proved a limited efficiency. In order to tackle these drawbacks, the next generation sequencing method was developed on 16S-rRNA-based sequences (Metabarcoding). The composition of bacterial communities was revealed based on the analysis of 16S rRNA sequences of intestine bacteria in Litopenaeus vannamei ponds in comparison with microbial communities in a Penaeus monodon pond and a muscle of shrimp. These results showed that the dominant phyla of intestine bacteria in Litopenaeus vannamei were Proteobacteria (49.3–57.4%), Firmicutes (15.6–34.4%) and Bacteroidetes (0.1–16.9%). Rhizobium(0.4%-26.1%), Vibrio(0–23.9%) and Spongiimonas(0–16,7%) were dominant genera in Litopenaeus vannamei gut. A higher proportion of Fusobacterium (10%), a shrimp pathogen group, was found in a disease shrimp pond (ST4) in comparison with a low growth shrimp pond (ST3) (0%) and a healthy shrimp pond (ST1) (0.6%). Vibrio was marked as shrimp pathogen genus accounted for 22.3% of total genera in ST4 in comparison with 2.4% in ST3 and 3.5% in ST1. Interestingly, a higher percentage of Vibrio rotiferianus (7.98%) was found in ST4 compared to ST3 (1%) and ST1 (0%). Fusobacterium and Vibrio will be the objects for the next experiments to discover shrimp pathogens specifically.

Exploring the Impact of the Biofloc Rearing System and an Oral WSSV Challenge on the Intestinal Bacteriome of Litopenaeus vannamei

We provide a global overview of the intestinal bacteriome of Litopenaeus vannamei in two rearing systems and after an oral challenge by the White spot syndrome virus (WSSV). By using a high-throughput 16S rRNA gene sequencing technology, we identified and compared the composition and abundance of bacterial communities from the midgut of shrimp reared in the super-intensive biofloc technology (BFT) and clear seawater system (CWS). The predominant bacterial group belonged to the phylum Proteobacteria, followed by the phyla Bacteroidetes, Actinobacteria, and Firmicutes. Within Proteobacteria, the family Vibrionaceae, which includes opportunistic shrimp pathogens, was more abundant in CWS than in BFT-reared shrimp. Whereas the families Rhodobacteraceae and Enterobacteriaceae accounted for almost 20% of the bacterial communities of shrimp cultured in BFT, they corresponded to less than 3% in CWS-reared animals. Interestingly, the WSSV challenge dramatically changed the bacterial communities in terms of composition and abundance in comparison to its related unchallenged group. Proteobacteria remained the dominant phylum. Vibrionaceae was the most affected in BFT-reared shrimp (from 11.35 to 20.80%). By contrast, in CWS-reared animals the abundance of this family decreased from 68.23 to 23.38%. Our results provide new evidence on the influence of both abiotic and biotic factors on the gut bacteriome of aquatic species of commercial interest.

A fast and visual method for duplex shrimp pathogens detection with high specificity using rapid PCR and molecular beacon

Molecular diagnosis of genome is one of the major methods for pathogens detection. The commonly used PCR method can realize an exponential amplification of the target gene but is time-consuming. In this work, we proposed a duplex and visual method using rapid PCR combined with molecular beacons to specifically detect two kinds of shrimp pathogens in one reaction tube. We only need to observe the fluorescence change of the reaction tube with naked eye to determine the result. A home-made automatic transfer equipment allows reaction tubes shuttling quickly between two water baths to achieve rapid PCR amplification. A simple device was also designed to present the detection results easily determined with naked eye. This duplex and visual detection method is fast, low-cost and of high specificity. From DNA extraction to results judgment, only 15 min was enough. Infectious Hypodermal and Hematopoietic Necrosis Virus (IHHNV) and Vibrio parahaemolyticus (VP) are two common shrimp pathogens which were chosen as our detection objects. This method may give a possibility to conduct end-point visual duplex detection, which may make a positive influence on the pathogen prevention.

Promotion of Lactobacillus plantarum on growth and resistance against acute hepatopancreatic necrosis disease pathogens in white-leg shrimp (Litopenaeus vannamei)

Promotion of Lactobacillus plantarum on growth and resistance against acute hepatopancreatic necrosis disease pathogens in white-leg shrimp (Litopenaeus vannamei)

First Report of a Complete Genome Sequence of White spot syndrome virus from India.

ABSTRACTWhite spot syndrome virus is a major pathogen of shrimp, causing economic loss to the aquaculture industry. For the first time, a complete de novo genome of an Indian isolate of this virus has been deciphered using Illumina and Nanopore sequencing technologies. The genome has 280,591 bp with 442 predicted coding genes.

Specific Molecular Signatures for Type II Crustins in Penaeid Shrimp Uncovered by the Identification of Crustin-Like Antimicrobial Peptides in Litopenaeus vannamei.

Crustins form a large family of antimicrobial peptides (AMPs) in crustaceans composed of four sub-groups (Types I-IV). Type II crustins (Type IIa or “Crustins” and Type IIb or “Crustin-like”) possess a typical hydrophobic N-terminal region and are by far the most representative sub-group found in penaeid shrimp. To gain insight into the molecular diversity of Type II crustins in penaeids, we identified and characterized a Type IIb crustin in Litopenaeus vannamei (Crustin-like Lv) and compared Type II crustins at both molecular and transcriptional levels. Although L. vannamei Type II crustins (Crustin Lv and Crustin-like Lv) are encoded by separate genes, they showed a similar tissue distribution (hemocytes and gills) and transcriptional response to the shrimp pathogens Vibrio harveyi and White spot syndrome virus (WSSV). As Crustin Lv, Crustin-like Lv transcripts were found to be present early in development, suggesting a maternal contribution to shrimp progeny. Altogether, our in silico and transcriptional data allowed to conclude that (1) each sub-type displays a specific amino acid signature at the C-terminal end holding both the cysteine-rich region and the whey acidic protein (WAP) domain, and that (2) shrimp Type II crustins evolved from a common ancestral gene that conserved a similar pattern of transcriptional regulation.