Comparative Transcriptomic Analysis of WSSV-Challenged Penaeus vannamei with Variable Resistance Levels.

The Pacific white shrimp, Penaeus vannamei, is highly susceptible to white spot syndrome virus (WSSV). Our study explored the transcriptomic responses of P. vannamei from resistant and susceptible families, uncovering distinct expression patterns after WSSV infection. The analysis revealed a higher number of differentially expressed genes (DEGs) in the susceptible family following WSSV infection compared to the resistant family, when both were evaluated against their respective control groups, indicating that the host resistance of the family line influences the transcriptome. The results also showed that subsequent to an identical duration following WSSV infection, there were more DEGs in P. vannamei with a high viral load than in those with a low viral load. To identify common transcriptomic responses, we profiled DEGs across families at 96 and 228 h post-infection (hpi). The analysis yielded 64 up-regulated and 37 down-regulated DEGs at 96 hpi, with 33 up-regulated and 34 down-regulated DEGs at 228 hpi, showcasing the dynamics of the transcriptomic response over time. Real-time RT-PCR assays confirmed significant DEG expression changes post-infection. Our results offer new insights into shrimp's molecular defense mechanisms against WSSV.

Feed competition reduces heritable variation for body weight in Litopenaeus vannamei.

BACKGROUND: Competition is a common social interaction among shrimp and depending on its intensity, it can affect heritable variation and response to selection. Little is known about the variance of indirect genetic effects (IGE) under competitive and non-competitive conditions in shrimp. In this study, we used extended mixed linear models to estimate genetic parameters for the direct genetic effect (DGE) and IGE on body weight in Litopenaeus vannamei raised under ad libitum (AF, non-competitive environment) and restricted (RF, competitive environment) feeding regimes. RESULTS: Estimates of heritabilities for body weight obtained with a traditional animal model (i.e. without accounting for IGE) were 0.11 ± 0.09 under AF and 0.25 ± 0.11 under RF. With extended animal models that accounted for IGE, the corresponding estimates for body weight were 0.07 ± 0.08 and 0.34 ± 0.11. Thus, heritabilities were higher under the RF regime than under the AF regime, regardless of whether IGE was accounted for or not. The log-likelihood ratio test revealed significant IGE under the RF regime. Although estimates of indirect genetic variance were low (0.0023 ± 0.0013 for AF and 0.0028 ± 0.0012 for RF), they contributed substantially to the total heritable variance: 66.8% for AF and 692.2% for RF. The total heritable variance was smaller under the RF regime (0.7 ± 1.3) than under the AF regime (5.8 ± 2.6) because of the high contribution of the negative covariance between DGE and IGE (- 7.03). Estimates of the correlation between DGE and IGE were 0.32 ± 0.47 under AF and - 0.93 ± 0.15 under RF, those of DGE and IGE for body weight between both regimes were 0.94 ± 0.07 and 0.67 ± 0.20, respectively, and those of IGE for body weight with DGE for survival were - 0.12 ± 0.22 under AF and - 0.58 ± 0.20 under RF. CONCLUSIONS: These results indicate that strong competitive interactions occurred under the RF regime in L. vannamei. Significant reranking and variation in IGE of individuals were observed between the two feeding regimes. Strong competitive interactions reduced the total heritable variation for body weight when food was restricted. These results indicate that the extent of competition among L. vannamei depends on the feeding regime applied and that this competition affects the genetic basis of body weight.

Comparative transcriptomic characterization of the eyestalk in Pacific white shrimp (Litopenaeus vannamei) during ovarian maturation.

In crustaceans, some of fundamental regulatory processes related to a range of physiological functions, including ovarian maturation, molting, glucose homeostasis, osmoregulation, etc., occur in the organs of the eyestalk. Additionally, reproduction is regulated by neuropeptide hormones and other proteins released from secretory sites (X-organ/sinus gland, XO/SG) within the eyestalk. As unilateral eyestalk ablation was the most common method used to artificially induce ovarian maturation for farmed Litopenaeus vannamei, to better understand the reproductive regulation mechanism in L. vannamei, we have investigated the transcriptomes of the eyestalk during five ovary developmental stages with or without eyestalk ablation by high-throughput Illumina sequencing technology. The raw reads were assembled and clustered into 127,031 unigenes. Meanwhile, the differentially expressed genes (DEGs) between ovarian development stages were identified. We examined, through DEG enrichment analysis, eyestalk gene expression patterns for Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways, comparing natural to artificially induced ovarian maturation. We also identified a variety of transcripts that appear to be differentially expressed throughout ovarian maturation. These include transcripts that encode G-protein coupled receptors (GPCRs) and neuropeptides, such as the crustacean hyperglycemic hormone (CHH), molt-inhibiting hormone (MIH), and crustacean female sex hormone (CFSH). Furthermore, numerous exoskeleton formation-related genes were found to be down-regulated during ovarian maturation, including cuticle-like proteins, eclosion hormone (EH), and gastrolith-like proteins, of which the latter are the first reported in L. vannamei. Our work is the first reproduction-related investigation of L. vannamei focusing on the eyestalk at the whole transcriptome level. These findings provide novel insight into the function of the eyestalk in reproduction regulation.

Isolation and characterization of a Raf gene from Chinese shrimp Fenneropenaeus chinensis in response to white spot syndrome virus infection.

Raf is a member in the Ras/Raf/MAPKK/MAPK signaling transduction pathway. To obtain a better understanding of Raf in the interaction between the Chinese shrimp Fenneropenaeus chinensis and white spot syndrome virus (WSSV), the sequence of cDNA of Raf from F. chinensis (FcRaf) was obtained. The FcRaf gene contained a 2421 bp open reading frame (ORF). The FcRaf shared most characteristic of Raf protein, such as the Raf-like Ras-binding domain (RBD), phorbol esters/diacylglycerol binding domain (C1 domain), and catalytic domain of the serine/threonine kinases, Raf (STKc_Raf). The sequence of functional domains of Raf protein was relatively conserved. The FcRaf mRNA was detected in the tissues of gill, muscle, and hepatopancreas from normal F. chinensis. The mRNA abundance level of FcRaf in the gill was the highest, which was 2.7-fold the level in the hepatopancreas. The expression level of FcRaf was significantly (P < 0.05) up-regulated in the tissues of gill, muscle, and hepatopancreas post WSSV-infection, which suggested that FcRaf might be involved in the interaction between F. chinensis and WSSV. Two SNP loci were identified in the ORF, one of which was a C-T mis-sense mutation, where an Ala was replaced by a Val, and induced the predicted protein secondary structure change. Considering the relatively low MAF (0.07), whether this mis-sense mutation was a detrimental mutation needs further investigation.

Identification of SNP markers associated with tolerance to ammonia toxicity by selective genotyping from de novo assembled transcriptome in Litopenaeus vannamei.

The high concentration of ammonia from deteriorated aquaculture environments and the intensive culture system could increase the susceptibility to pathogens and even cause high mortality in Litopenaeus vannamei. In addition, we have revealed that the ammonia-tolerant shrimp also have high disease resistance in L. vannamei. In the present study, in order to identify SNP markers associated with tolerance to ammonia toxicity, we developed and characterized SNPs from our previous transcriptome sequencing data of ammonia-stressed and control groups, and a marker-trait association analysis was performed for marker-assisted selection (MAS) to increase production in L. vannamei. A total of 318,919 SNPs were identified from the transcriptome sequences, and 25,772 SNPs were found from the 1826 ammonia-responsive genes with functional annotation. We selected 49 SNPs from 26 ammonia-responsive genes that had strong homologies to known genes in the shrimp and probably involved in immune function as candidate markers for genotyping, among which 39 SNPs were polymorphic for further marker-trait association analysis with the ammonia-tolerant (AT) and ammonia-sensitive (AS) groups. Finally, 12 out of the 49 SNP markers were identified to be associated with ammonia tolerance, containing 10 loci with significantly different allele frequencies and 10 loci with significantly different genotyping frequencies between the AT and AS groups. Among the associated markers, the G allele of TSP-1 (the first locus from the thrombospondin gene), the A allele of TSP-3, and the C allele of XBP1-5 (the fifth locus from X-box binding protein 1) only presented in the AT groups, but they were absent from the AS groups, which would be the preference of the MAS for the ammonia-tolerant shrimp. In addition, when the 12 associated SNP markers were used for analysis, the genetic diversity of the AT groups were significantly higher than that of the AS groups, but when the 39 loci were used there was no difference. This is the first report for the markers associated with ammonia tolerance in this species, indirectly with disease resistance, which provided important potential for genetic selection to increase survival rate and production in shrimp farming.

iTRAQ-based comparative proteome analysis for molecular mechanism of defense against acute ammonia toxicity in Pacific White shrimp Litopenaeus vannamei.

In the practical farming of Litopenaeus vannamei, the intensive culture system and environmental pollution usually results in a high concentration of ammonia, which brings large detrimental effects to shrimp, such as increasing the susceptibility to pathogens and even causing high mortality. We have revealed that the survival time under acute ammonia stress varied substantially among different families and obtained ammonia-tolerant (LV_T) and ammonia-sensitive (LV_S) families. In order to understand the molecular mechanism of defense against ammonia toxicity in shrimp, we performed iTRAQ LC-MS/MS proteomic analysis between LV_T and LV_S groups of L. vannamei under acute ammonia stress to identify the key proteins and pathways that play an effective role for against ammonia toxicity. By comparative proteome analysis, 202 significantly differentially proteins (DEPs) were identified in LV_T compared to LV_S, and most of the DEPs (60%) were up-regulated. Excepting for the proteins without function reporting, the meaningful finding is that 77.8% of the DEPs have been reported mainly involving in immune defense and stress tolerant in crustacean species, such as hemocyanin, Rab7, Rab GTPase, Rac1, alpha 2 macroglobulin, Bip, peroxiredoxin, Cu/Zn SOD, glutathione peroxidase, thioredoxin, calreticulin, and Elongation Factor 1-alpha, etc. These DEPs might potentially play important role in against ammonia toxicity, and it also reflected a relation between ammonia tolerance and pathogen resistance. In addition, a total of 10 significantly changed KEGG pathways were detected, and the network diagram of these KEGG pathways showed that more critical nodes were up-regulated, which involved in protein synthesis and transport, and against stress stimuli. This study provided important information for understanding the molecular mechanism of defense against ammonia toxicity in shrimp at whole protein level.

Transcriptome analysis of 'Huanghai No. 2' Fenneropenaeus chinensis response to WSSV using RNA-seq.

White spot syndrome (WSS) is one of the most damaging phenomena in the culturing of shrimp. To characterize the mechanisms of the molecular responses to WSSV infection in 'Huanghai No. 2'' Fenneropenaeus chinensis, we used next-generation sequencing to observe the transcriptome after oral infection. A total of 108.6 million clean reads were obtained and assembled into 64,103 final unigenes with an average length of 845 bp (N50 = 1534 bp). The assembled unigenes contained 14,263 significant unigenes after BLASTX against the Nr database (E-value cut-off of 10-5). After comparison of digital gene expression data between challenged and control shrimp, a total of 896 DEGs after WSSV infection were identified. Gene pathway analysis indicated that 92, 131 and 142 metabolic pathways were affected at early, peak and late phases respectively. Some pathways were related to the immune response, such as the phagosome, complement and coagulation cascades, the antigen processing and presentation pathway and so on. Many immune-related genes were also identified after pathway analysis. Interestingly, some growth-related genes, such as cathepsin L, myosin regulatory light chain 2 smooth muscle, and alpha-amylase were also differentially expressed after WSSV infection, and the correlation between growth trait and WSSV-resistance trait need further research. The expression patterns of eight DEGs were confirmed by quantitative real-time reverse transcription polymerase chain reaction, and there was good agreement between RNA-seq and qRT-PCR. These data will provide valuable information for characterizing the immune mechanism of the response of shrimp's to WSSV.

Mechanics of the folding of a nanotube.

Nanotubes fold due to the competition between their mechanical stability and van der Waals interactions. The caused dramatic morphology change promises exciting applications of nanotubes in responsive and reconfigurable nanodevices. To investigate the folding mechanism, a curvature-based finite-deformation theoretical model simultaneously considering both the folding of a nanotube and the possible collapsing of the cross-section is developed. The predicted critical condition and the profiles in both axial and transverse directions agree well with molecular dynamics (MD) solutions, demonstrating that the cross-sectional deformation should be taken into account when investigating the folding of a nanotube with a large diameter. Moreover, simple scaling laws of the critical conditions are proposed through a small-deformation theoretical model. With these scaling laws, one can easily and quickly determine both the collapsing state of the cross-section and the folding state of the nanotube with only geometrical parameters [Formula: see text] rather than the difficult-to-determine material properties [Formula: see text].

The identification of microRNAs involved in the response of Chinese shrimp Fenneropenaeus chinensis to white spot syndrome virus infection.

MicroRNA (miRNA) is a class of small noncoding RNA, which is involved in the post-transcriptional regulation in all metazoan eukaryotes. MiRNAs might play an important role in the host response to virus infection. However, miRNAs in the aquatic crustacean species were not extensively investigated. To obtain a better understanding of the response of Chinese shrimp Fenneropenaeus chinensis to white spot syndrome virus (WSSV) infection, the sequence and expression profile of miRNAs in the hepatopancreas of WSSV-infected F. chinensis were obtained by the high-throughput Illumina HiSeq 2500 deep sequencing technique. A total number of 129 known miRNAs and 44 putative novel miRNAs were identified from the deep sequencing data. The peak size of miRNAs was 22 nt (37.0%). 25 miRNAs were significantly (P < 0.05) differentially expressed post WSSV infection. Six of the differentially expressed miRNAs were randomly selected for further verification by the real-time RT-PCR technique. The results showed that there was a consistency between the deep sequencing and real-time RT-PCR assay. The target genes of differentially expressed miRNAs were predicted. Each miRNA had 4 target genes on average. The results suggested that some specific miRNAs might be involved in the response of F. chinensis to WSSV infection, and further provided basic information for the investigation of specific miRNAs in F. chinensis.

cDNA cloning and expression analysis of a phosphopyruvate hydratase gene from the chinese shrimp Fenneropenaeus chinensis.

In the present study a cDNA encoding a phosphopyruvate hydratase (enolase) was cloned from the muscle of the Chinese shrimp (Fenneropenaeus chinensis) and named as FcEnolase. The cDNA of FcEnolase encoded a protein of 434 amino acid residues with a molecular mass 47.22 kDa. The residues 342-355 constituted the signature motif "LLLKVNQIGSVTES". A SNP locus (C96T) in the ORF at 96 bp was identified. The results showed that the FcEnolase was a conserved gene. In the normal F. chinensis, the mRNA level in the muscle was much higher (P < 0.05) than the mRNA level in the gill and hepatopancreas. To verify the mRNA level of FcEnolase in the F. chinensis post WSSV infection, a real-time RT-PCR was performed. In the WSSV-infected F. chinensis, the FcEnolase mRNA level was significantly (P < 0.05) up-regulated in the muscle at 12 and 24 h post challenge (hpc) to approximately 2.7-fold and 2.7-fold the mRNA level in the controls, respectively. The FcEnolase mRNA level in the gill was significantly (P < 0.05) down-regulated at 6 hpc to approximately 0.3-fold the mRNA level in the control, followed by a significant (P < 0.05) up-regulation at 12 hpc to approximately 2.8-fold the mRNA level in the control. There was no obvious change of FcEnolase mRNA level in the hepatopancreas during the infection process. The expression profile coincided with the fact that WSSV primarily infects the tissues of muscle and gill, but hardly infects hepatopancreas. To verify the protein level of FcEnolase post WSSV infection, a Western blot was performed. The FcEnolase protein level in the muscle at 24 hpc significantly (P < 0.05) increased to approximately 2.1-fold the level in the control. These results showed the characterization of FcEnolase and suggested that the FcEnolase might be involved in the response of F. chinensis to WSSV infection.

Theoretical studies on lattice-oriented growth of single-walled carbon nanotubes on sapphire.

Due to their excellent mechanical and electrical properties, single-walled carbon nanotubes (SWNTs) can find broad applications in many areas, such as field-effect transistors, logic circuits, sensors and flexible electronics. High-density, horizontally aligned arrays of SWNTs are essential for high performance electronics. Many experimental studies have demonstrated that chemical vapor deposition growth of nanotubes on crystalline substrates such as sapphire offers a promising route to achieve such dense, perfectly aligned arrays. In this work, a theoretical study is performed to quantitatively understand the van der Waals interactions between SWNTs and sapphire substrates. The energetically preferred alignment directions of SWNTs on A-, R- and M-planes and the random alignment on the C-plane predicted by this study are all in good agreement with experiments. It is also shown that smaller SWNTs have better alignment than larger SWNTs due to their stronger interaction with sapphire substrate. The strong vdW interactions along preferred alignment directions can be intuitively explained by the nanoscale 'grooves' formed by atomic lattice structures on the surface of sapphire. This study provides important insights to the controlled growth of nanotubes and potentially other nanomaterials.

Genetic assessment of residual feed intake as a feed efficiency trait in the Pacific white shrimp Litopenaeus vannamei.

BACKGROUND: Residual feed intake (RFI) was investigated as a measure of feed efficiency in a breeding population of Litopenaeus vannamei. Shrimp from 34 families were housed individually and feed efficiency and growth traits were recorded during two successive growth periods. The objectives of this study were (1) to estimate the heritability of RFI and related traits, including feed efficiency ratio (FER), average daily gain (ADG) and daily feed intake (DFI), (2) to determine the relationships between RFI and other traits, and (3) to evaluate the variation of these traits across two growth periods. RESULTS: Shrimp displayed large inter-individual variation in RFI, FER, ADG and DFI during each growth period. Heritability estimates of all these traits during both periods reached high values (0.577 ± 0.232 to 0.707 ± 0.252). RFI showed weak and no genetic correlations with ADG during the two growth periods between days 1 to 21 (0.135 ± 0.204) and 22 to 42 (-0.018 ± 0.128), respectively, but high positive genetic correlations with DFI (>0.8). Weak and moderate negative genetic correlations were observed between RFI and FER during the two periods (-0.126 ± 0.208 and -0.387 ± 0.183). As evidenced by the high genetic correlations between the two periods for each trait (>0.6), trait performance of the shrimp tended to be consistent across periods. CONCLUSIONS: For the first time, accurate measurement of individual feed efficiency on a large scale was achieved in shrimp. Although the estimated heritability reported here for RFI may be overestimated, it is a heritable trait in L. vannamei that can be improved by genetic improvement. For L. vannamei, the biggest potential advantage in using RFI as a measure of feed efficiency is that it is independent of growth rate, and thus genetic selection on RFI has the potential to improve feed efficiency and reduce feed intake, without compromising growth performance.

Regulation of Insulin Resistance by Multiple MiRNAs via Targeting the GLUT4 Signalling Pathway.

BACKGROUND/AIMS: Type 2 Diabetes Mellitus (T2DM) is characterized by insulin resistance (IR), but the underlying molecular mechanisms are incompletely understood. MicroRNAs (miRNAs) have been demonstrated to participate in the signalling pathways relevant to glucose metabolism in IR. The purpose of this study was to test whether the multiple-target anti-miRNA antisense oligonucleotides (MTg-AMO) technology, an innovative miRNA knockdown strategy, can be used to interfere with multiple miRNAs that play critical roles in regulating IR. METHODS: An MTg-AMO carrying the antisense sequences targeting miR-106b, miR-27a and miR-30d was constructed (MTg-AMO106b/27a/30d). Protein levels were determined by Western blot analysis, and transcript levels were detected by real-time RT-PCR (qRT-PCR). Insulin resistance was analysed with glucose consumption and glucose uptake assays. RESULTS: We found that the protein level of glucose transporter 4 (GLUT4), Mitogen-activated protein kinase 14 (MAPK 14), Phosphatidylinositol 3-kinase regulatory subunit beta (PI3K regulatory subunit beta) and mRNA level of Slc2a4 (encode GLUT4), Mapk14 (encode MAPK 14) and Pik3r2 (encode PI3K regulatory subunit beta) were all significantly down-regulated in the skeletal muscle of diabetic rats and in insulin-resistant L6 cells. Overexpression of miR-106b, miR-27a and miR-30d in L6 cells decreased glucose consumption and glucose uptake, and reduced the expression of GLUT4, MAPK 14 and PI3K regulatory subunit beta. Conversely, silencing of endogenous miR-106b, miR-27a and miR-30d in insulin-resistant L6 cells enhanced glucose consumption and glucose uptake, and increased the expression of GLUT4, MAPK 14 and PI3K regulatory subunit beta. MTg-AMO106b/27a/30d up-regulated the protein levels of GLUT4, MAPK 14 and PI3K regulatory subunit beta, enhanced glucose consumption and glucose uptake. CONCLUSION: Our data suggested that miR-106b, miR-27a and miR-30d play crucial roles in the regulation of glucose metabolism by targeting the GLUT4 signalling pathway in L6 cells. Moreover, MTg-AMO106b/27a/30d offers more potent effects than regular singular AMOs.

Isolation and expression analysis of an MAPKK gene from Fenneropenaeus chinensis in response to white spot syndrome virus infection.

Mitogen-activated kinase kinase (MAPKK) is an important gene involved in the host-virus interaction process. To obtain a better understanding of MAPKK in the interaction process between the Chinese shrimp Fenneropenaeus chinensis and white spot syndrome virus (WSSV), we cloned the sequence of an MAPKK cDNA from F. chinensis (FcMAPKK) and investigated the effect of FcMAPKK on WSSV infection. The results showed that the FcMAPKK gene contained a 1227 bp open reading frame (ORF), which encoded a highly conserved protein with a serine/threonine protein kinase catalytic (S_TKc) domain. The deduced amino acid sequence of FcMAPKK shared identities between 11.9 and 92.6% with MAPKKs from vertebrate, invertebrate, plant and fungus species. The FcMAPKK was expressed in all the examined tissues in the normal F. chinensis. FcMAPKK expression level was highest in the hepatopancreas where it was approximately 2.6-fold the expression level in the gill, and lowest in the muscle where it was approximately 0.3-fold the expression level in the hepatopancreas. The FcMAPKK expression levels in the muscle, gill, and hepatopancreas were all changed post WSSV challenge. The FcMAPKK expression was significantly (P < 0.01) up-regulated in the muscle of F. chinensis at 48 h post WSSV infection. The WSSV began to replicate quickly in the normal F. chinensis at 48 h post infection, while the WSSV replication in the U0126-treated F. chinensis could be significantly (P < 0.05) inhibited. The results suggested that FcMAPKK might be involved in the WSSV infection process, and hijacking of FcMAPKK might be required for WSSV replication in F. chinensis.

Comparative microarray profile of the hepatopancreas in the response of "Huanghai No. 2" Fenneropenaeus chinensis to white spot syndrome virus.

White spot syndrome virus (WSSV) infects all shrimp species and is the greatest detriment to shrimp culture. To better understand the mechanism of molecular responses to WSSV infection in "Huanghai No. 2" Fenneropenaeus chinensis, a microarray technique was used. Microarray gene expression profiling of 59,137 unigenes identified Differentially Expressed Genes (DEGs) both in live and moribund shrimp at early, peak and late phases. In live shrimp, 1307, 1479 and 1539 DEGs were obtained in the early, peak and late phase, respectively. Meanwhile, 1536, 2181 and 1591 DEGs were obtained in moribund shrimp. Twenty known annotation genes are uniquely expressed in the late phase of live shrimp, including adhesion regulating molecule 1, arginine kinase, BUD31 homolog, and QM. Compared to WSSV-susceptible shrimp, 75 known annotation genes are uniquely expressed in WSSV-resistant shrimp, including arginine kinase, BUD31 homolog, clottable protein 2, caspase 2, cathepsin C, calnexin, HMGBb, Histone 3, and selenoprotein M. The gene expression patterns of the infected shrimp were altered by WSSV infection. To further confirm the expression of differentially expressed genes, real-time RT-PCR was performed to test six randomly selected genes. The data will provide valuable information to understand the immune mechanism of shrimp's response to WSSV.

An analysis of indirect genetic effects on adult body weight of the Pacific white shrimp Litopenaeus vannamei at low rearing density.

BACKGROUND: Our aim was to estimate the genetic parameters for the direct genetic effect (DGE) and indirect genetic effects (IGE) on adult body weight in the Pacific white shrimp. IGE is the heritable effect of an individual on the trait values of its group mates. METHODS: To examine IGE on body weight, 4725 shrimp from 105 tagged families were tested in multiple small test groups (MSTG). Each family was separated into three groups (15 shrimp per group) that were randomly assigned to 105 concrete tanks with shrimp from two other families. To estimate breeding values, one large test group (OLTG) in a 300 m(2) circular concrete tank was used for the communal rearing of 8398 individuals from 105 families. Body weight was measured after a growth-test period of more than 200 days. Variance components for body weight in the MSTG programs were estimated using an animal model excluding or including IGE whereas variance components in the OLTG programs were estimated using a conventional animal model that included only DGE. The correlation of DGE between MSTG and OLTG programs was estimated by a two-trait animal model that included or excluded IGE. RESULTS: Heritability estimates for body weight from the conventional animal model in MSTG and OLTG programs were 0.26 ± 0.13 and 0.40 ± 0.06, respectively. The log likelihood ratio test revealed significant IGE on body weight. Total heritable variance was the sum of direct genetic variance (43.5%), direct-indirect genetic covariance (2.1%), and indirect genetic variance (54.4%). It represented 73% of the phenotypic variance and was more than two-fold greater than that (32%) obtained by using a classical heritability model for body weight. Correlations of DGE on body weight between MSTG and OLTG programs were intermediate regardless of whether IGE were included or not in the model. CONCLUSIONS: Our results suggest that social interactions contributed to a large part of the heritable variation in body weight. Small and non-significant direct-indirect genetic correlations implied that neutral or slightly cooperative heritable interactions, rather than competition, were dominant in this population but this may be due to the low rearing density.

Molecular Characterization and Expression Analysis of the C-Type Lectin Domain Family 4 Member F in Litopenaeus vannamei against White Spot Syndrome Virus.

White spot disease (WSD) outbreaks pose a significant threat to the Pacific white shrimp (Litopenaeus vannamei) farming industry. The causative agent is the white spot syndrome virus (WSSV). There are no effective treatments for WSD so far. Therefore, understanding the resistance mechanisms of L. vannamei against the WSSV is crucial. C-type lectins (CTLs) are important pattern recognition receptors (PRRs) that promote agglutination, phagocytosis, encapsulation, bacteriostasis, and antiviral infections. This study cloned the C-type lectin domain family 4 member F (LvCLEC4F) from L. vannamei. LvCLEC4F contains a 492 bp open reading frame (ORF) encoding a protein of 163 amino acids, including a carbohydrate recognition domain (CRD). Following a challenge with the WSSV, the expression profile of LvCLEC4F was significantly altered. Using RNA interference (RNAi) technology, it was found that LvCLEC4F promotes WSSV replication and affects the expression levels of genes related to the regulation of apoptosis, signaling and cellular stress response, and immune defense. Meanwhile, the hemolymph agglutination phenomenon in vivo was weakened when LvCLEC4F was knocked down. These results indicated that LvCLEC4F may play an important role in the interaction between L. vannamei and WSSV.

Reproductive Ability Disparity in the Pacific Whiteleg Shrimp (Penaeus vannamei): Insights from Ovarian Cellular and Molecular Levels.

The Pacific whiteleg shrimp (Penaeus vannamei) is a highly significant species in shrimp aquaculture. In the production of shrimp larvae, noticeable variations in the reproductive capacity among female individuals have been observed. Some females experience slow gonadal development, resulting in the inability to spawn, while others undergo multiple maturations and contribute to the majority of larval supply. Despite numerous studies that have been conducted on the regulatory mechanisms of ovarian development in shrimp, the factors contributing to the differences in reproductive capacity among females remain unclear. To elucidate the underlying mechanisms, this study examined the differences in the ovarian characteristics between high and low reproductive bulks at different maturity stages, focusing on the cellular and molecular levels. Transmission electron microscopy analysis revealed that the abundance of the endoplasmic reticulum, ribosomes, mitochondria, and mitochondrial cristae in oocytes of high reproductive bulk was significantly higher than that of the low reproductive bulk in the early stages of ovarian maturation (stages I and II). As the ovaries progressed to late-stage maturation (stages III and IV), differences in the internal structures of oocytes between females with different reproductive capacities gradually diminished. Transcriptome analysis identified differentially expressed genes (DEGs) related to the mitochondria between two groups, suggesting that energy production processes might play a crucial role in the observed variations in ovary development. The expression levels of the ETS homology factor (EHF) and PRDI-BF1 and RIZ homology domain containing 9 (PRDM9), which were significantly different between the two groups, were compared using qRT-PCR in individuals at different stages of ovarian maturation. The results showed a significantly higher expression of the EHF gene in the ovaries of high reproductive bulk at the II and IV maturity stages compared to the low reproductive bulk, while almost no expression was detected in the eyestalk tissue of the high reproductive bulk. The PRDM9 gene was exclusively expressed in ovarian tissue, with significantly higher expression in the ovaries of the high reproductive bulk at the four maturity stages compared to the low reproductive bulk. Fluorescence in situ hybridization further compared the expression patterns of EHF and PRDM9 in the ovaries of individuals with different fertility levels, with both genes showing stronger positive signals in the high reproductive bulk at the four ovarian stages. These findings not only contribute to our understanding of the regulatory mechanisms involved in shrimp ovarian development, but also provide valuable insights for the cultivation of new varieties aimed at improving shrimp fecundity.

Analysis of Elimination Effects of Inbreeding on Genotype Frequency in Larval Stages of Chinese Shrimp.

Marine animals possess genomes of considerable complexity and heterozygosity. Their unique reproductive system, characterized by high fecundity and substantial early mortality rates, increases the risk of inbreeding, potentially leading to severe inbreeding depression during various larval developmental stages. In this study, we established a set of inbred families of Fenneropenaeus chinensis, with an inbreeding coefficient of 0.25, and investigated elimination patterns and the manifestations of inbreeding depression during major larval developmental stages. Reduced-representation genome sequencing was utilized to explore the genotype frequency characteristics across two typical elimination stages. The results revealed notable mortality in hatching and metamorphosis into mysis and post-larvae stages. Inbreeding depression was also evident during these developmental stages, with depression rates of 24.36%, 29.23%, and 45.28%. Segregation analysis of SNPs indicated an important role of gametic selection before hatching, accounting for 45.95% of deviation in the zoea stage. During the zygotic selection phase of larval development, homozygote deficiency and heterozygote excess were the main selection types. Summation of the two types explained 82.31% and 89.91% of zygotic selection in the mysis and post-larvae stage, respectively. The overall distortion ratio decreased from 22.37% to 12.86% in the late developmental stage. A total of 783 loci were identified through selective sweep analysis. We also found the types of distortion at the same locus could change after the post-larvae stage. The predominant shifts included a transition of gametic selection toward normal segregation and other forms of distortion to heterozygous excess. This may be attributed to high-intensity selection on deleterious alleles and genetic hitchhiking effects. Following larval elimination, a greater proportion of heterozygous individuals were preserved. We detected an increase in genetic diversity parameters such as expected heterozygosity, observed heterozygosity, and polymorphic information content in the post-larvae stage. These findings suggest the presence of numerous recessive deleterious alleles and their linkage and suggest a major role of the partial dominance hypothesis. The results provide valuable insights into the mechanisms of inbreeding depression in marine animals and offer guidance for formulating breeding strategies in shrimp populations.

Estimation of Genetic Parameters for Growth and WSSV Resistance Traits in Litopenaeus vannamei.

The current study aimed to provide a precise assessment of the genetic parameters associated with growth and white spot syndrome virus (WSSV) resistance traits in Pacific white shrimp (Litopenaeus vannamei). This was achieved through a controlled WSSV challenge assay and the analysis of phenotypic values of five traits: body weight (BW), overall length (OL), body length (BL), tail length (TL), and survival hour post-infection (HPI). The analysis included test data from a total of 1017 individuals belonging to 20 families, of which 293 individuals underwent whole-genome resequencing, resulting in 18,137,179 high-quality SNP loci being obtained. Three methods, including pedigree-based best linear unbiased prediction (pBLUP), genomic best linear unbiased prediction (GBLUP), and single-step genomic BLUP (ssGBLUP) were utilized. Compared to the pBLUP model, the heritability of growth-related traits obtained from GBLUP and ssGBLUP was lower, whereas the heritability of WSSV resistance was higher. Both the GBLUP and ssGBLUP models significantly enhanced prediction accuracy. Specifically, the GBLUP model improved the prediction accuracy of BW, OL, BL, TL, and HPI by 4.77%, 21.93%, 19.73%, 19.34%, and 63.44%, respectively. Similarly, the ssGBLUP model improved prediction accuracy by 10.07%, 25.44%, 25.72%, 19.34%, and 122.58%, respectively. The WSSV resistance trait demonstrated the most substantial enhancement using both genomic prediction models, followed by body size traits (e.g., OL, BL, and TL), with BW showing the least improvement. Furthermore, the choice of models minimally impacted the assessment of genetic and phenotypic correlations. Genetic correlations among growth traits ranged from 0.767 to 0.999 across models, indicating high levels of positive correlations. Genetic correlations between growth and WSSV resistance traits ranged from (-0.198) to (-0.019), indicating low levels of negative correlations. This study assured significant advantages of the GBLUP and ssGBLUP models over the pBLUP model in the genetic parameter estimation of growth and WSSV resistance in L. vannamei, providing a foundation for further breeding programs.