Identification and characterization of novel microRNAs in disease-resistant and disease-susceptible Penaeus monodon.

MicroRNAs (miRNAs), known as a translational regulator, are evolutionary conserved, small, and noncoding RNA. They have played a vital role in disease biology through the host-virus-miRNA-interaction. In this study, novel miRNAs of naturally occurring, virus-free disease-resistant and disease-susceptible Penaeus monodon were identified and characterized. In disease-susceptible samples, 45 homologous mature miRNAs and 28 homologous precursor miRNAs were identified. In disease-resistant samples, 52 homologous mature miRNAs and 87 homologous precursor miRNAs were identified. In disease-susceptible samples, 33 novel mature miRNAs and 33 novel precursor miRNAs were identified. In disease-resistant samples, 523 novel mature miRNAs and 141 novel precursor miRNAs were identified. Differential expression study revealed the up-regulated and down-regulated miRNAs in disease-resistant and disease-susceptible P. monodon. Gene ontology pathway of known and novel miRNAs revealed that P. monodon miRNAs might have a potential and specific role in signal transduction, cell-to-cell signaling, innate immune response and defense response to different pathogens.

Development and characterization of white spot disease linked microsatellite DNA markers in Penaeus monodon, and their application to determine the population diversity, cluster and structure.

Pathogens that are introduced suddenly to natural populations can potentially cause quick changes to the genetics and diversity of the host. In the past three decades, white spot syndrome virus (WSSV) has caused damaging epizootics in Penaeus monodon populations. In this study, we developed WSSV resistance- or susceptibility-linked microsatellite DNA markers, and their effectiveness was validated experimentally. WSSV-resistant marker linked retroelements and genes that may have an important role in WSSV-resistance phenomena were partially identified. Allelic data of 1,694 samples from nine distinct geographic locations in India were revealed that populations from Digha and Kochi were highly dispersed, and also showed higher genetic diversity, higher population diversity, and lower prevalence of disease resistance. A very high level of gene flow was observed within all populations and a very high level of genetic variation was present within populations. Two genetically admixture population clusters were estimated in nature. WSSV-resistance has a significant link with genetic diversity, population cluster and population diversity. Microsatellite marker analysis characterized genetic divergence, diversity and structure among wild populations.

Identification and characterisation of microsatellite DNA markers in order to recognise the WSSV susceptible populations of marine giant black tiger shrimp, Penaeus monodon.

White spot disease (WSD) which is caused by white spot syndrome virus (WSSV) creates severe epizootics in captured and cultured black tiger shrimp, resulting a huge loss in the economic output of the aquaculture industry worldwide. Performing selective breeding using DNA markers would prove to be a potential cost effective strategy for long term disease control in shrimps. In the present investigation, microsatellite DNA fingerprints were compared between naturally occurring WSSV resistant and susceptible populations of Penaeus monodon. After PCR with a set of shrimp specific primers three reproducible DNA fragments of varying sizes were found, among which 442 bp and 236 bp fragments were present in considerably higher frequencies in the WSSV susceptible shrimp population (p ≤ 0.0001). After WSSV challenge experiment the copy no. of WSSV was determined using real-time PCR, where it was found to be almost 4 × 10(3) fold higher in WSSV susceptible shrimps than in the resistant ones. Thus, these microsatellite DNA markers will be useful to distinguish between WSSV susceptible and resistant brood stocks of P. monodon. Sequencing studies revealed that these DNA markers were novel in P. monodon. Highest WSSV resistance using these DNA markers, was observed in the shrimp populations of Andaman Island and Chennai among the different coastal areas of India, suggesting these places as safe for specific pathogen resistant brood stock shrimp collection. This study will be a very effective platform towards understanding the molecular pathogenesis of WSD for generation of disease free shrimp aquaculture industry.

Assessment of WSSV prevalence and distribution of disease-resistant shrimp among the wild population of Penaeus monodon along the west coast of India.

Shrimp aquaculture is threatened by many diseases, among which white spot disease (WSD) caused by white spot syndrome virus (WSSV) is the leading one. Information related to the geographical distribution and seasonal prevalence of WSD is necessary to obtain a clear understanding of the disease biology in shrimp. Identification of WSD-resistant individual shrimp with DNA markers is also an important technique to develop better WSD-free shrimp health management. The present study aim is to estimate the occurrence of WSSV in Penaeus monodon qualitatively and quantitatively during three different seasons during the years 2011 to 2013 along the west coast of India. Additionally, the disease resistance prevalence using previously developed 71 bp microsatellite and 457 bp RAPD-SCAR DNA markers is also investigated. Samples were collected throughout the year from four locations along the west coast of India: Kochi, Kerala; Mangalore, Karnataka; Vasco-da-Gama, Goa; and Veraval, Gujarat. The results depicted that the average WSSV prevalence, as determined by the nested PCR method and taken cumulatively over the four locations, was the lowest (0%) during the post-monsoon season and the highest (31.6%) during the monsoon season. The WSD prevalence was observed to increase when the latitude was decreased along the west coast of India (from Veraval to Kochi). Out of the three different seasons, the average WSSV copy number was the highest (approximately 10(3) copies µg(-1) shrimp genomic DNA) during the monsoon season. The disease-resistant prevalence, as determined using the developed DNA markers, was found to be the highest in Vasco-da-Gama (59.5%) and the lowest in Kochi (40.9%). The present study suggests better options for the efficient collection of disease-free and disease-resistant brood stocks, which would be a more cost-effective and safer approach toward disease prevention over conventional trends of seed generation from unselected wild brood stock.