First report on the occurrence of white spot syndrome virus, infectious myonecrosis virus and Enterocytozoon hepatopenaei in Penaeus vannamei reared in freshwater systems.

Samples of white leg shrimp, Penaeus vannamei, were collected on a monthly basis from freshwater ponds with the salinity of 0 ppt located at Tiruvannamalai and Villupuram districts in Tamil Nadu, India for screening of viral and fungal pathogens. Totally, 130 shrimp samples were collected from 67 freshwater ponds and screened for white spot syndrome virus (WSSV), infectious myonecrosis virus (IMNV), infectious hypodermal and haematopoietic necrosis virus (IHHNV) and Enterocytozoon hepatopenaei (EHP) by PCR and RT-PCR using pathogen-specific primers. Among the samples screened, one sample was found to be positive to WSSV, two samples showed positive to IMNV and two samples positive for EHP. No sample showed positive to IHHNV. The WSSV detected in the sample was found to be a new strain of WSSV and highly virulent. The inoculum prepared from freshwater reared WSSV or IMNV-infected shrimp caused 100% mortality in experimental infection studies. The PCR and RT-PCR results revealed the presence of WSSV and IMNV in different organs of experimentally infected shrimp, respectively. No clinical signs were observed in experimentally EHP-injected shrimp, although the PCR results revealed the presence of EHP in experimentally infected shrimp.

An Indigenous, Field-Deployable, Lateral Flow Immunochromatographic Assay Rapidly Detects Infectious Myonecrosis in Shrimp, Litopenaeus vannamei.

Shrimp farming is an important socioeconomic activity worldwide. Infectious myonecrosis virus (IMNV) is an important shrimp virus responsible for significant mortality (up to 70%) in Litopenaeus vannamei. We produced recombinant capsid protein (r-IMNV31) and obtained a highly specific antibody, anti-r-IMNV31, which was used in WOAH-approved ELISA and Western blot to detect IMNV. Further, anti-r-IMNV31 was employed in an indigenously developed lateral flow immunoassay (LFA) with gold nanoparticles as a visual label. Using LFA, IMNV could be detected rapidly (20 min) from tissue homogenate with high specificity, reproducibility, and sensitivity (LOD = 103 viral particles). LFA was validated with "gold standard" qRT-PCR using 60 samples with high sensitivity (100%), specificity (86%). A Cohen's kappa coefficient of 0.86 suggested "good agreement" between LFA and qRT-PCR. With a shelf-life of ~ 1 year at ambient temperature, the use of LFA in the on-site detection of IMNV by shrimp farmers will be a reality.

Protective Role of Hypothermia Against Heat Stress in Differentiated and Undifferentiated Human Neural Precursor Cells: A Differential Approach for the Treatment of Traumatic Brain Injury.

INTRODUCTION: The present study aimed to explore protective mechanisms of hypothermia against mild cold and heat stress on highly proliferative homogeneous human Neural Precursor Cells (NPCs) derived from Subventricular Zone (SVZ) of human fetal brain. METHODS: CD133+ve enriched undifferentiated and differentiated human NPCs were exposed to heat stress at 42°C. Then, Western-blot quantification was performed using Hsp-70 (70 kilodalton heat shock proteins) recombinant protein. Finally, changes in pluripotency and Hsp-70 expression were measured using immunofluorescence staining and RT-qPCR (Quantitative reverse transcription PCR) analysis, respectively. RESULTS: Heat stress resulted in abnormal neurospheres development. The apoptosis rate was enhanced during long-term in vitro culture of neurospheres. Neurogenic differentiation reduced and showed aberrent phenotypes during heat stress. After hypothermia treatment significant improvement in neurospheres and neuronal cell morphology was observed. CONCLUSION: Mild-hypothermia treatment induces attenuated heat shock response against heat stress resulting in induced HSP-70 expression that significantly improves structure and function of both undifferentiated human NPCs and differentiated neurons.

In vitro quantitative and relative gene expression analysis of pancreatic transcription factors Pdx-1, Ngn-3, Isl-1, Pax-4, Pax-6 and Nkx-6.1 in trans-differentiated human hepatic progenitors.

AIMS/INTRODUCTION: Diabetes is a major health concern throughout the world because of its increasing prevalence in epidemic proportions. ß-Cell deterioration in the pancreas is a crucial factor for the progression of diabetes mellitus. Therefore, the restoration of ß-cell mass and its function is of vital importance for the development of effective therapeutic strategies and most accessible cell sources for the treatment of diabetes mellitus. MATERIALS AND METHODS: Human fetuses (12-20 weeks gestation age) were used to isolate human hepatic progenitor cells (hHPCs) from fetal liver using a two-step collagenase digestion method. Epithelial cell adhesion molecule-positive (EpCAM+ve)-enriched hHPCs were cultured in vitro and induced with 5-30 mmol/L concentration of glucose for 0-32 h. Pdx-1 expression and insulin secretion was analyzed using immunophenotypic and chemifluorescence assays, respectively. Relative gene expression was quantified in induced hHPCs, and compared with uninduced and pancreatic cells to identify the activated transcription factors (Pdx-1, Ngn-3, Isl-1, Pax-4, Pax-6 and Nkx-6.1) involved in ß-cell production. RESULTS: EpCAM+ve cells derived from human fetal liver showed high in vitro trans-differentiation potential towards the ß-cell phenotype with 23 mmol/L glucose induction after 24 h. The transcription factors showed eminent expression in induced cells. The expression level of transcription factors was found significantly high in 23 mmol/L-induced hHPCs as compared with the uninduced cells. CONCLUSIONS: The present study has shown an exciting new insight into ß-cell development from hHPCs trans-differentiation. Relative quantification of gene expression in trans-differentiated cells offers vast possibility for the production of a maximum number of functionally active pancreatic ß-cells for a future cure of diabetes.