In-silico analysis and mRNA modulation of detoxification enzymes GST delta and kappa against various biotic and abiotic oxidative stressors

This study reports the comprehensive comparative information of two different detoxification enzymes such as glutathione S-transferases (GSTs) delta and kappa from freshwater giant prawn Macrobrachium rosenbergii (designated as MrGSTD and MrGSTK) by investigating their in-silico characters and mRNA modulation against various biotic and abiotic oxidative stressors. The physico-chemical properties of these cDNA and their polypeptide structure were analyzed using various bioinformatics program. The analysis indicated the variation in size of the polypeptides, presence or absence of domains and motifs and structure. Homology and phylogenetic analysis revealed that MrGSTD shared maximum identity (83%) with crustaceans GST delta, whereas MrGSTK fell in arthropods GST kappa. It is interesting to note that MrGSTD and MrGSTK shared only 21% identity; it indicated their structural difference. Structural analysis indicated that MrGSTD to be canonical dimer like shape and MrGSTK appeared to be butterfly dimer like shape, in spite of four β-sheets being conserved in both GSTs. Tissue specific gene expression analysis showed that both MrGSTD and MrGSTK are highly expressed in immune organs such as haemocyte and hepatopancreas, respectively. To understand the role of mRNA modulation of MrGSTD and MrGSTK, the prawns were inducted with oxidative stressors such as bacteria (Vibrio harveyi), virus [white spot syndrome virus (WSSV)] and heavy metal, cadmium (Cd). The analysis revealed an interesting fact that both MrGSTD and MrGSTK showed higher (P < 0.05) up-regulation at 48 h post-challenge, except MrGSTD stressed with bacteria, where it showed up-regulation at 24 h post-challenge. Overall, the results suggested that GSTs are diverse in their structure and possibly conferring their potential involvement in immune protection in crustaceans. However, further study is necessary to focus their functional differences at proteomic level

Molecular importance of prawn large heat shock proteins 60, 70 and 90

Considering the importance of heat shock proteins (HSPs) in the innate immune system of prawn, a comparative molecular approach was proposed to study the crustacean large HSPs 60, 70 and 90. Three different large HSPs were identified from freshwater prawn Macrobrachium rosenbergii (Mr) cDNA library during screening. The structural and functional characteristic features of HSPs were studied using various bioinformatics tools. Also, their gene expression and mRNA regulation upon various pathogenic infections was studied by relative quantification using 2-ΔΔCT method. MrHSP60 contains a long chaperonin 60 domain at 46–547 which carries a chaperonin 60 signature motif between 427 and 438, whereas MrHSP70 contains a long HSP70 domain at 21–624 and MrHSP90 carries a HSP90 domain at 188–719. The two dimensional analysis showed that MrHSP60 contains more amino acids (52%) in helices, whereas MrHSP70 (40.6%) and MrHSP90 (51.8%) carried more residues in coils. Gene expression results showed significant (P < 0.05) expression of MrHSP60, 70 and 90 in haemocyte, gill and hepatopancreas, respectively. Further, the expression level was up-regulated upon bacterial (Aeromonas hydrophilla and Vibrio harveyi) and viral [white spot syndrome virus (WSSV) and M. rosenbergii nodo virus (MrNV)] infections during various time periods. The gene expression results exhibited the potential involvement of these three HSPs in the immune system of prawn. The study indicated the potentiality of these molecules, thereby protecting cells against pathogens as well as severe cellular and environmental stresses in crustaceans

Green synthesis of anisotropic zinc oxide nanoparticles with antibacterial and cytofriendly properties

Zinc oxide nanoparticles (ZnONPs) exhibit abundant biomedical applications. Anisotropic ZnONPs with a defined shape and size were synthesized using Bacillus megaterium (NCIM 2326) cell free extract as a bio-reductant. The study investigated the multidimensional effect of ZnONPs on Helicobacter pylori strains and assessed its biosafety in normal human mesenchymal stem cells (hMSc). The highly stable ZnONPs were produced using B. megaterium and Zinc nitrate as a precursor. The phase of ZnONPs formation and structural characterization were performed by UV- visible (UV-Vis), Fourier transform infrared (FTIR) spectroscopy, X-ray diffraction (XRD) and Field Emission Scanning electron microscopy (FESEM) analysis. Furthermore, the ZnONPs exhibited higher biocompatibility against human mesenchymal stem cells (hMSC) and proved to be potentially safe in mammalian cells. Corroborating the current investigation, we described the anti-H. Pylori dosage of ZnONPs was safe to hMSC and could efficiently use as nano-antibiotic

Anti-biofilm properties and immunological response of an immune molecule lectin isolated from shrimp Metapenaeus monoceros

The study is carried out to understand the antimicrobial and immunological response of a potential immune molecule lectin, MmLec isolated from haemolymph of Speckled shrimp, Metapenaeus monoceros. MmLec was purified using mannose coupled Sepharose CL-4B affinity chromatography, which was further subjected on SDS-PAGE to ascertain the distribution of their molecular weight. Sugar binding specificity assay was conducted at various pH and temperatures to investigate the binding affinity of MmLec towards the specific carbohydrate molecule. Functional analysis of immune molecule MmLec included haemagglutination assays performed using human erythrocytes and yeast agglutination activity against Saccharomyces cerevisiae which, were analyzed using light microscopy. In order to study the antimicrobial activity, two Gram-negative (Vibrio parahaemolyticus and Aeromonas hydrophila) and two Gram-positive (Staphylococcus aureus and Enterococcus faecalis) bacteria were treated with purified MmLec. Moreover, these bacterial species were also treated at different concentration of the MmLec to speculate the antibiofilm properties of MmLec which was analyzed under Light Microscopy and Confocal Laser Scanning Microscopy. In addition, other functional characterization of MmLec showed the uniqueness of MmLec in agglutination of human erythrocyte as well as the cells of yeast Saccharomyces cerevisiae. Also, the phenoloxidase activity and encapsulation assay was evaluated. MTT assay displayed that MmLec are potent in anticancer activity. The study will help to understand the immunological interference and antimicrobial nature of MmLec which would be supportive in establishing a potential therapeutic tool and to develop better and novel disease control strategies in shrimp and farmed aquaculture industries as well as in health management

In vitro identification and Antifungal susceptibility of different Candida Species isolated from patients with or without Diabetes having Chronic Periodontitis

Background: Candida albicans is the commonly associated species with oral lesions, but other Candida spp., such as C. glabrata, C. tropicalis, C. parapsilosis, C. krusei, and C. dubliniensis have also been isolated from the human saliva, even irrespective to the oral candidiasis. Candidal colonization has also been documented from subgingival plaque of adults with periodontitis.Objective: Nowadays there is a great demand of natural herbal medicines. For a Diabetic person having Periodontitis, the use antifungal drugs are must. Therefore the antifungal capacity of the commonly used drug and herbs has been evaluated.Methods: For the study, the patients were screened and categorised into two study and one control group, having 20 patients in each group, on the basis of their respective Blood Glucose level and Dental status using standard clinical parameters. Oral rinse of 60 patients including both gender were selected from the Out Patient Department of Periodontitis, Rungta College of Dental Sciences and Research, Chhattisgarh. Chrome Candida Agar Media was used to identify Candida albicans and NAC. In vitro tests of the effectiveness of selected antimycotic agents against test candidal isolates recovered from the patient were performed to check the efficacy of the antifungal agents by the Kirby-Bauer Disc Diffusion method.Result: We found that by the use of this chromogenic media most of the commonly recovered NAC can be easily distinguished as each species produced the widest range of colours and morphologies. The herbal extracts Aloe Vera and Garlic failed to inhibit the growth of the candidal species but Neem, Meetha Neem, Triphala, Tulsi showed sensitivity to varying degree. Now a day there are much popularity of using herbal products, having its extracts incorporated in the cosmetics and other products used by humans.Conclusion: We conducted the sensitivity check of the candidal isolates against the dental dentrifices and found that however all the toothpastes contained herbal extracts capable of destroying bacteria but it did not showed good results against candidal species except the Patanjali Toothpaste. This may be due the fact that Patanjali Dant Kanti Dental Cream has the combination of all the herbal extract mixed altogether. DOI: http://dx.doi.org/10.3126/ijosh.v4i1.1216