Expression of immune genes in Indian major carp, Catla catla challenged with Flavobacterium columnare.

Columnaris disease, caused by Flavobacterium columnare, is one of the important bacterial diseases responsible for large-scale mortalities in numerous freshwater fishes globally. This disease can cause up to 100% mortality within 24 h of infection and is considered to be a cause of concern for aquaculture industry. Despite being a serious disease, scarce information is available regarding host-pathogen interaction, particularly the modulation of different immune genes in response to F. columnare infection. Therefore, in the present study, an attempt has been made to study expression of important immune regulatory genes, namely IL-1ß, iNOS, INF-γ, IL-10, TGF-ß, C3, MHC-I and MHC-II in gills and kidney of Catla catla following experimental infection with F. columnare. The expression analysis of immune genes revealed that transcript levels of IL-1ß, iNOS, IL-10, TGF-ß, C3 and MHC-I were significantly up-regulated (p < 0.05) in both the organs of the infected catla. IFN-γ and MHC-II were up-regulated in gills of infected catla whereas, both the genes showed down-regulation in kidney. The results indicate that important immune genes of C. catla are modulated following infection with F. columnare. The knowledge thus generated will strengthen the understanding of molecular pathogenesis of F. columnare in Indian major carp C. catla.

[Identification of hypervariable regions within the 16S-23S rRNA intergenic spacer region of Flavobacterium columnare and its application in assigning genomovar group to an individual strain].

Flavobacterium columnare is an important bacterial pathogen of fish with wide ge- netic variability within species. This intraspecies diversity has been termed as genomovars and genomovar groups on the basis of Restriction Fragment Length Polymorphisms of 16S rDNA and 16S-23S rDNA Intergenic Spacer Region (ISR), respectively. In this study, we demonstrate the source of genetic heterogeneity in the F. columnare by sequence analysis of ISR. Length of ISR sequences of different genomovars varied from 553 to 592 nucleotides, while the similarity among sequences ranged from 76.1 to 92.6%. A common ISR structure with tRNAAa and tRNAne embedded within the sequence was identified in all the genomovars ofF. columnare. The results show that strains of F. columnare can be categorized into five genomovar groups based on the heterogeneity in the ISR sequences. Of these, strains belonging to Genomovar I and II can be sub-divided into two groups each; while strains of Genomovar III belonged to one group. Sequence similarity between genomovar groups was lower for ISR (76.1-92.6%) as compared to 16S rDNA (96.1-99.4%) indicating its ability to resolve closely related groups within the genomovars of F. columnare. The main source of variation between genomovar groups is the presence of three hyper variable regions (V1, V2 & V3) in the ISR Of the three, V3 was found to be the most heterogeneous region and was found to be useful in assigning genomovar group to an individual strain of F. columnare.

Proteomic analysis of outer membrane proteins of Edwardsiella tarda.

AIMS: The purpose of this study was to identify outer membrane proteins (OMPs) of Edwardsiella tarda. METHODS AND RESULTS: The OMPs from a virulent strain of E. tarda (ET-7) was extracted using lauroyl sarcosine method. The OMPs were analysed by two-dimensional polyacrylamide gel electrophoresis (2D-PAGE), and protein spots were identified using matrix assisted laser desorption/ionization-time-of-flight mass spectrometry. A total of 21 proteins were identified from 24 protein spots observed on the 2D-PAGE gel. These proteins were identified as GroEL, antigenic proteins, ABC transporters, elongation factors, OmpA, PTSINtr with GAF domain, catalase C, glycolytic enzymes, DnaJ, transcriptional regulator, proteins mraZ and ccdA. Subcellular localizations, beta-barrel OMPs and lipoproteins of identified proteins were predicted using PSORTb, PRED-TMBB and LipoP1.0 programme. CONCLUSIONS: Identification, localization and possible functions of OMPs of E. tarda were studied. SIGNIFICANCE AND IMPACT OF THE STUDY: These proteins could be used for development of novel drug targets, diagnostics or vaccine against edwardsiellosis.

Ontogeny and tissue-specific expression of immune-relevant genes in Catla catla (Hamilton).

India is the second largest fish producing country in the world with production of 12.6 million tonnes (mt) in 2017-18, and Indian major carps (IMCs) contribute bulk of this fish production. Catla, Catla catla is the fastest growing species among IMCs. However, the survival rate of catla during larval rearing is normally lower than the other IMCs i.e rohu Labeo rohita, and mrigal Cirrhinus mrigala. Continuous efforts are devoted for the identification of nutritional and environmental requirements of fish larvae in order to reduce hatchery mortalities. However, very little information is available regarding physiology of the immune system, especially during the late larval and juvenile stages. Hence, understanding the ontogenetic development of immune-relevant genes in the larval stages of catla will serve as the markers for the development of immune competence and thereby, will be beneficial in developing effective immune intervention strategies. In the present study, expression profiles of some of the important innate (IL-1ß, IL-10, iNOS and C3) and adaptive immune (RAG-1, Ikaros, IgM and IgZ) genes during ontogenetic developmental stages and in different tissues of catla were investigated using quantitative real-time PCR. The results revealed that immune genes IL-1ß, C3, IgM and IgZ were detected in the unfertilized eggs indicating their maternal inheritance. Immune genes, IL-1ß, IL-10 and iNOS were expressed significantly during initial larval developmental stages whereas C3, RAG-1, Ikaros, IgM and IgZ showed significant expression during advanced stages of larval development in catla i.e. from 23 days post hatch (dph). Study of tissue distribution pattern of the genes indicated that innate immune genes were ubiquitously expressed in different tissues with varying degree of expression, whereas adaptive immune genes were predominantly expressed in lymphoid organs of catla. The information thus generated will improve knowledge on the maturation of the immune system in catla and will aid in deciding the appropriate age for vaccination in this teleost species.

Primary cell culture from fish gills and kidney using fish serum.

A novel, cost effective and time saving technique for primary cell culture from gills and kidney using fish serum has been developed. Single cell suspension of gills and kidney of Clarias gariepinus was obtained by trypsinization. Minimum essential medium supplemented with 10% fish serum in place of 10% foetal calf serum and 20% fish muscle extract, yielded confluent monolayer on 6th and 8th day in gill and kidney tissue respectively at 28 degrees C. Fish serum can be successfully used as media supplement for cultivation and maintenance of primary cell culture from fishes.

Kocuria flava induced growth and chromium accumulation in Cicer arietinum L.

In the present investigation a chromate tolerant rhizobacterium Kocuria flava was isolated and inoculated to the Cicer arietinum L to evaluate its effects on growth and chromium accumulation upon exposure of different concentration of chromium (1-10 microg ml(-1)) as Cr (VI) for 24 d. K. flava inoculated plant of C. arietinum demonstrated luxuriant growth as compared to non inoculated plant at respective concentration of Cr (VI). K. flava found to ameliorate chromium induced phytotoxicity in terms of chlorophylls, carotenoid and protein contents and thus helps the plant in acquiring higher biomass with high chromium concentration. After 24 d, maximum concentration of chromium recorded in root of C. arietinum (4892.39 microg g(-1) dw) inoculated with K. flava as compared to non inoculated plant (1762.22 microg g(-1) dw) upon exposure of 5 microg ml(-1) Cr (VI). Therefore, application of C. arietinum in association with K. flava could be more efficient in decontamination of chromium polluted site. Moreover, K. flava may be used as a bioresource for developing microbes assisted phytoremediation system due to its compatibility.

Antioxidant and Antidiabetic Activity of Helicteres isora (L.) Fruits.

The present investigations evaluated the antioxidant and antidiabetic activity of Helicteres isora (L.) fruits belonging to the family Sterculiaceae. The hot water extract of Helicteres isora fruits was prepared and screened for its in vitro antioxidant activity using 1,1-diphenyl,2-picryl hydrazyl assay, ss-carotene-linoleate model and microsomal lipid peroxidation or thiobarbituric acid reactive species assays and the IC(50) values were calculated. Antidiabetic effect was studied using the in vitro glucose uptake in the isolated rat hemi-diaphragm model. The hot water extract of Helicteres isora showed maximum activity with IC(50) value 25.12+/-0.18 mug/ml for 1,1-diphenyl,2-picryl hydrazyl assay method, and low activity with IC(50) value 740.64+/-4.76 mug/ml for microsomal lipid peroxidation assay. In the ss-carotene-linoleate model, the extract showed 45.63% antioxidant activity. The extract produce a significant (P<0.05) uptake of glucose by isolated rat hemi-diaphragm but less effective to that of the reference drug, metformin. The hot water extract of fruit of Helicteres isora exhibited significant antioxidant activity and moderate antidiabetic activity and merits further investigation in animal models and isolation of its active constituents.

Performance of Au transmission photocathode on a microchannel plate detector.

X-ray framing cameras, employing microchannel plates (MCPs) for detection and signal amplification, play a key role in research in high-energy-density physics. These instruments convert radiographic x-rays into electrons produced by plasma during such experiments into electrons that are amplified in the channels and then detected by a phosphor material. The separation of detection from signal amplification offers potential improvements in sensitivity and noise properties. We have implemented a suspended Au transmission photocathode (160 A thick) on a MCP and are evaluating it using a 1.5 keV Al K alpha x-ray source. We find an approximately twofold increase in the ratio of detected events to incident photons when the photocathode-to-MCP voltage difference is sufficiently large. Our calculations indicate that this increase is probably caused by a combination of signal produced by the photocathode and an increase in the efficiency of detection of x-rays that reach the MCP surface through modification of the local electric field.

Hydro-chemistry of Udaipur lakes.

The hydro-chemical character of lakes around Udaipur city has been studied for a period of five years. But for the lake Udaisagar, the salinity of water in lakes Badi, Fatehsagar, Pichhola and Swaroopsagar is less than 750 microSiemens per centimeter and fluctuates marginally in dry and wet period. The waters are Ca-Mg-HCO3 type and have calcite saturation. All the lakes have shown dissolved oxygen more than 8.0 mg/L despite of varying level of pollution. The lake Udaisagar is most polluted and exhibits eutrophic character along with the lakes Swaroopsagar and Pichhola.