Detection of non-ribosomal and polyketide biosynthetic genes in bacteria from green mud crab Scylla serrata gut microbiome and their antagonistic activities.

Multi-modular enzyme complexes known as non-ribosomal peptide synthetases (NRPSs) and polyketide synthetases (PKSs) have been widely reported in bacteria that produce secondary bioactive metabolites such as non-ribosomal peptides (NRPs) and polyketides (PKs), respectively. These NRPS/PKS pathways contribute to synthesizing several antibiotics, such as vancomycin, rifamycin, and bleomycin, which are vital in human medicine. The present study aimed to isolate gut-associated bacteria from mud crab Scylla serrata, and detect NRPS and PKS gene clusters associated with it. This study included 36 bacterial isolates from five mud crab gut samples. Biosynthetic gene clusters (NRPS and PKS), were detected by PCR using degenerative primers specific to these genes. Three isolates (FKP2-4, FKP4-1, and FKP2-16) were positive for NRPS and two for PKS (FKP2-4 and FKP4-1) genes. The isolates were subjected to 16S rRNA gene amplification and sequenced. In silico analysis of the sequences using the Basic Local Alignment Search Tool (BLAST) identified the isolates FKP2-4, FKP4-1, and FKP2-16 as Acinetobacter variabilis, Vagococcus fluvialis, and Staphylococcus arlettae, respectively, after comparing with the existing sequences available in the National Center for Biotechnology Information (NCBI) database. Compared to the control, it was observed that these isolates exhibited intriguing antagonistic activities against Escherichia coli and Staphylococcus aureus. However, these isolates failed to show significant activity against Candida albicans. Exopolysaccharide production by the isolated organisms was tested using Zobell marine agar (ZMA) with 5% sucrose, but none of the colonies were mucoid or slimy.

Association of exopolysaccharide genes in biofilm developing antibiotic-resistant Pseudomonas aeruginosa from hospital wastewater.

The study aimed to examine the relationship between antibiotic resistance, biofilm formation and genes responsible for biofilm formation. Sixty-six Pseudomonas aeruginosa isolates were obtained from hospital wastewater and analyzed for their antibiotic resistance. Biofilm production among the isolates was tested by indirect quantification method crystal violet assay. Biofilm-associated genes among these isolates psl, alg, and pel were also checked. The maximum resistance was observed for ampicillins (88.24%) followed by nalidixic (83.82%), and nitrofurantoin (64.71%), respectively. Biofilm phenotypes are distributed in the following categories: high 39.39% (n = 26); moderate 57.57% (n = 38), and weak 3.0% (n = 2). Among the total isolates, biofilm-associated genes were detected in 84.84% (n = 56) of isolates and the remaining isolates 15.15% (n = 10) did not harbor any genes. In this study, pslB was the most predominant gene observed (71.21%, n = 47) followed by pslA (57.57%, n = 38), pelA (45.45%, n = 30), algD (43.93%, n = 29), and pelD (27.27%, n = 18), respectively. The present study reveals that the majority of the isolates are multidrug resistant being moderate and high biofilm formers. The study implies that biofilm acts as a machinery for bacteria to survive in the hospital effluent which is an antibiotic stress environment.

Comparative analysis of the intestinal bacterial communities in mud crab Scylla serrata in South India.

Little is known about the functions of the crustacean gut microbiome, but environmental parameters and habitat are known to affect the composition of the intestinal microbiome, which may in turn affect the physiological status of the host. The mud crab Scylla serrata is an economically important species, and is wild-caught, and farmed across the Indo-Pacific region. In this study, we compared the composition of the gut microbiome (in terms of gut microbial species richness and abundance) of S. serrata collected from wild sites, and farms, from the east and west coast of India, and also tested the effects of the environment on the composition. The water temperature had a statistically significant effect on gut microbiome composition, with microbial biodiversity decreasing with increasing water temperature. This could have negative effects on both wild and farmed mud crabs under future climate change conditions, although further research into the effects of temperature on gut microbiomes is required. By comparison, salinity, crab mass and carapace width, geographical location as well as whether they were farmed or wild-caught crabs did not have a significant impact on gut microbiome composition. The results indicate that farming does not significantly alter the composition of the gut microbiome when compared to wild-caught crabs.

Different expression pattern of thrombospondin gene in the presence and absence of ß-glucan fed Penaeus monodon challenged with white spot syndrome virus.

Thrombospondins (TSPs) are extracellular, calcium-binding glycoproteins that play an essential role in cell homeostasis and development, wound-healing, angiogenesis, connective tissue organization, immune response etc. and it conserves from sea sponges to mammals. However, their role in shrimp immunity is poorly understood. In the present study, the differential expression profiling of TSP transcripts in Penaeus monodon tissues such as gills, lymphoid organs, hepatopancreas, and hemolymph challenged with white spot syndrome virus (WSSV), were studied by quantitative real-time PCR. Further, shrimps fed with the immunostimulant (ß-glucan) when challenged with WSSV showed significant upregulation of TSP expression in gills, hepatopancreas, and lymphoid organ at the early phase of WSSV infection. The results suggest that TSP may be an inducible acute phase response protein to WSSV infection. The possibility of differences in mRNA expression pattern seen in immunostimmulated shrimp after the viral challenge, possibility due to altered immune mechanisms getting triggered during immunostimulant administration and virus infections in the host.

Hospital wastewater treatment reduces NDM-positive bacteria being discharged into water bodies.

New Delhi metallo-ß-lactamase-1 (NDM-1) is a novel type of metallo-ß-lactamase (MBL) associated with Enterobacteriaceae constitutes an important growing public health threat. The present study aims to characterize the NDM-1 producing Gram-negative bacteria (GNB) from the effluents of two tertiary care hospitals in Mangalore, South India and to profile their antibiotic resistance pattern. A total of 134 GNB were isolated from 30 hospital wastewater samples (treated and untreated) and analyzed. High-level resistance among untreated effluent sample was found toward nalidixic acid (74.52%), followed by cefotaxime (72.64%) and ampicillin (66.03%). Among the treated effluent isolates, the high resistance was found toward ampicillin (85.71%) followed by cefotaxime (85.71%) and piperacillin-tazobactam (53.57%). From untreated effluent isolates, 9 were NDM-1 positive by PCR; no isolates from treated effluent samples harbored blaNDM-1 . Untreated hospital wastewater is found to be important reservoirs of antibiotic-resistant bacteria carrying blaNDM-1 , and the presence of such bacteria in the effluents is a matter of great concern because they can contribute the antibiotic resistance to the natural environment. However, the absence of NDM in treated effluents emphasizes the importance of effluent treatment in reducing the dissemination of antibiotic-resistant bacteria. PRACTITIONER POINTS: Hospital wastewater is the important reservoir of antibiotic-resistant bacteria especially metallo-ß-lactamase producers (NDM-1). Wastewater treatment procedures in hospitals reduce the NDM isolates in the treated effluent. Thereby reduces the risk of resistance spread in the environment.

Clinical Applications of Antimicrobial Peptides (AMPs): Where do we Stand Now?

In this era of multi-drug resistance (MDR), antimicrobial peptides (AMPs) are one of the most promising classes of potential drug candidates to combat communicable as well as noncommunicable diseases such as cancers and diabetes. AMPs show a wide spectrum of biological activities which include antiviral, antifungal, anti-mitogenic, anticancer, and anti-inflammatory properties. Apart from these prospective therapeutic potentials, the AMPs can act as food preservatives and immune modulators. Therefore, AMPs have the potential to replace conventional drugs and may gain a significant global drug market share. Although several AMPs have shown therapeutic potential in vitro or in vivo, in most cases they have failed the clinical trial owing to various issues. In this review, we discuss in brief (i) molecular mechanisms of AMPs in various diseases, (ii) importance of AMPs in pharmaceutical industries, (iii) the challenges in using AMPs as therapeutics and how to overcome, (iv) available AMP therapeutics in market, and (v) AMPs under clinical trials. Here, we specifically focus on the therapeutic AMPs in the areas of dermatology, surgery, oncology and metabolic diseases.

Protection of shrimp Penaeus monodon from WSSV infection using antisense constructs.

White spot syndrome caused by white spot syndrome virus (WSSV) is one of the most threatening diseases of shrimp culture industry. Previous studies have successfully demonstrated the use of DNA- and RNA-based vaccines to protect WSSV infection in shrimp. In the present study, we have explored the protective efficacy of antisense constructs directed against WSSV proteins, VP24, and VP28, thymidylate synthase (TS), and ribonucleotide reductase-2 (RR2) under the control of endogenous shrimp histone-3 (H3) or penaedin (Pn) promoter. Several antisense constructs were generated by inserting VP24 (pH3-VP24, pPn-VP24), VP28 (pH3-VP28, pPn-VP28), TS (pH3-TS, pPn-TS), and RR2 (pH3-RR2) in antisense orientation. These constructs were tested for their protective potential in WSSV infected cell cultures, and their effect on reduction of the viral load was assessed. A robust reduction in WSSV copy number was observed upon transfection of antisense constructs in hemocyte cultures derived from Penaeus monodon and Scylla serrata. When tested in vivo, antisense constructs offered a strong protection in WSSV challenged P. monodon. Constructs expressing antisense VP24 and VP28 provided the best protection (up to 90 % survivability) with a corresponding decrease in the viral load. Our work demonstrates that shrimp treated with antisense constructs present an efficient control strategy for combating WSSV infection in shrimp aquaculture.