Genome editing for phage design and uses for therapeutic applications.

The over usage of antibiotics leads to antibiotic abuse which in turn eventually raises resistance mechanisms among wide range of pathogens. Due to lack of experimental data of efficacy of phages as potential antimicrobial and therapeutic agent and also more specific and cumbersome isolation process against specific pathogens makes it not so feasible technology to be looked as an alternative therapy. But, recent developments in genome editing techniques enables programmed nuclease enzymes that has effectively improvised our methodology to make accurate changes in the genomes of prokaryote as well as eukaryote cells. It is already strengthening our ability to improvise genetic engineering to disease identification by facilitating the creation of more precise models to identify the root cause. The present chapter discusses on improvisation of phage therapy using recent genome editing tools and also shares data on the methods of usage of phages and their derivatives like proteins and enzymes such as lysins and depolymerases, as a potential therapeutic or prophylaxis agent. Methods involved in recombinant based techniques were also discussed in this chapter. Combination of traditional approach with modern tools has led to a potential development of phage-based therapeutics in near future.

Vibrio harveyi biofilm as immunostimulant candidate for high-health pacific white shrimp, Penaeus vannamei farming.

The study was to develop Vibrio harveyi biofilm-based novel microbial product and its oral delivery for high health Penaeus vannamei farming. Yield of bacterial biofilm was optimized on chitin substrate (size: <360, 360-850 and 850-1250 µm; concentration: 0.3, 0.6 and 0.9%) in tryptone soy broth (0.15%). The biofilm was characterized by crystal violet assay, SEM and LSCM imaging; protein profiling by SDS-PAGE and LC-ESI-MS/MS. The immune stimulatory effect of the biofilm in yard experiments was evaluated by relative quantification of immune genes using real-time PCR effect on overall improvement on health status under field trials. The highest biofilm yield (6.13 ±â€¯0.2 × 107 cfu/ml) was obtained at 0.6% of <360 µm chitin substrate. The biofilm formation was stabilized by 96 h of incubation at 30 °C. Protein profiling confirmed expression of six additional proteins (SDS-PAGE) and 11 proteins were differentially expressed (LC-ESI-MS/MS) in biofilm cells over free cells of V. harveyi. Oral administration of the biofilm for 48 h confirmed to enhance expression of antimicrobial peptides, penaeidin, crustin and lysozyme in P. vannamei. Further Oral administration of biofilm for two weeks to P. vannamei (1.8 ±â€¯0.13 g) improved the growth (2.66 ±â€¯0.06 g) and survival (84.44 ±â€¯1.82%) compared to control (2.15 ±â€¯0.03 g; 70.94 ±â€¯0.66%) Nursery trials showed a significant reduction in occurrence of anatomical deformities like antenna cut (12.67 ±â€¯0.66%), rostrum cut (4.66 ±â€¯0.87%), and tail rot (3.33 ±â€¯0.88%), compared to animals fed with normal diet which was 24.33 ±â€¯2.72; 14 ±â€¯1.52 and 10.66 ±â€¯1.45% respectively. In vitro and in vivo studies suggest inactivated biofilm cells of V. harveyi on chitin substrate express additional antigenic proteins and when administered orally through feed at regular intervals stimulates immune response and improve growth, survival and health status of shrimp.