Usage Of Gelatin-Virus Balls And Liquid Virus Filled Gelatin Capsules To Control Coral Reef Diseases: Model For Phage Therapy

European Scientific Journal June 2016 edition vol.12, No.18 ISSN: 1857 – 7881 (Print) e - ISSN 1857- 7431 Usage Of Gelatin-Virus Balls And Liquid Virus Filled Gelatin Capsules To Control Coral Reef Diseases: Model For Phage Therapy Bhagwan N. Rekadwad, PhD Chandrahasya N. Khobragade, PhD School of Life Sciences, Swami Ramanand Teerth Marathwada University, Nanded, India doi: 10.19044/esj.2016.v12n18p320 URL:http://dx.doi.org/10.19044/esj.2016.v12n18p320 Abstract Coral reefs are very sensitive to environmental pollution. Coral reefs frequently get infected by various bacteria, fungi, marine algae and protozoa. The diseases include Bacterial Infections (BI), Fungal Infections (FI), Black Band Disease (BBD), Black Overgrowing Cyanophyta (BOC), Black Aggressive Band (BAB), Lethal Orange Disease (LOD), Skeleton Eroding Band (SED), PEYssonnelia (PEY), PNEophyllum (PNE) and White Syndromes (WS). Here in we have proposed a proposed model in which cold water soluble gelatin will be used to prepare Gelatin virus balls (GVB) and Liquid virus filled and sealed gelatin capsules (LVFSGC). GVB and LVFSGC will be prepared as per standard protocol in the form of paintballs and capsules. Above mentioned infecting agents of Coral reefs will be used as inoculum for production their production on the pilot scale. These produced infecting agents will be added with specific viruses of infecting agent (host-specific viruses). After the lysis of cell (naturally/artificially), lysate containg host-specific viruses will be used as infecting viruses to the Coral reef infecting agents. This lysate will be used for preparation of GVB and LVFSGC. These paintballs and capsules contain host-specific viruses can be made to release on a surface of sea water and dispersed on affected coral reefs zone naturally by Sea water current/waves. The dispersed viruses from GVB and LVFSGC will attach to their host. Ultimately, the diseasecausing agent may be killed and the coral reef infection will be removed from sea water without any harm to the environment. GVB and LVFSGC will be used for the release of viruses against disease-causing agents. The GVB and LVFSGC will systematically kill and save the coral reefs. Keywords: Corals; Cold water soluble gelatin; Phages; Sea Bed; Viral DNA, biological pollutants 320 European Scientific Journal June 2016 edition vol.12, No.18 ISSN: 1857 – 7881 (Print) e - ISSN 1857- 7431 Introduction Coral reefs provide coastal protection, supply food and natural products useful in pharmaceutical and cosmetic industries and provide foreign currency to the country by attracting the tourists from the rest of the world. Thus, Coral reefs play a key role in the economy of many countries having coastal region. Coral reefs have hundreds of species in a natural marine ecosystem (Kelman et al., 2006; Charpy et al., 2012. Saxena, 2015; Rekadwad and Khobragade, 2015). The demand of coral reefs increasing steadily due to their useful natural products. As the economy of any country grows, the adverse impacts slowly get imprinted on the ecosystem (Hunter et al., 1995; Haapkyla et al., 2007; Olsen et al., 2015). The marine ecosystem nowadays gets contaminated by various types of pollutants. Damage caused by both the physicochemical and biological pollutants to some extent can be controlled by adopting preventive measures (Nicolet et al., 2013). On the other hand, those host-specific biological pollutants are very harmful than any other pathogens which include bacteria (Vidaver et al., 1973; Comeau et al., 2005; Kulakov et al., 2009), fungi (Barrero-Canosa et al., 2013; Mann et al., 2014) and marine algae (James, 2012). Few species of protozoa also cause harm to coral reefs. All these pathogens also have their specific infection-causing agents called host-specific viruses. They systematically kill their host by utilizing host synthesizing machinery without any harm to useful biota and will not be a cause for the negative change in native ocean environment (Pratte, 2014). In the present study, an attempt has been made to design a virtual experiment to save coral reefs from the variety of infections. Host-specific viruses encased in Gelatin virus balls and Liquid virus filled and sealed gelatin capsules may kill specifically the host’s infecting organisms. Material and methods Isolation of organisms infecting coral reefs and cultivation of viruses in laboratory Infected Coral reef species is a good source of infection-causing organisms and can be isolated using the specific medium. Nutrient broth will be used for isolation of bacteria, potato dextrose broth, and malt medium will be used for isolation of fungi, Bold’s basic medium and Bristol’s medium will be used for cultivation of algae while protozoa will be cultivated in serum-free and serum containing media for the isolation and enrichment purposes (Atlas et al., 2010; Freshney, et al., 2005). In enrichment medium, host-specific viruses will be added. For example, bacteriophage phi6 and phiKMV-like viruses will be added for Pseudomonas, Vibrio phage will be added for Vibrio, cyanophage, algal viruses will be added for Chlorella and Ostreococcus etc. These viruses will attack their host. When cell burst, the 321 European Scientific Journal June 2016 edition vol.12, No.18 ISSN: 1857 – 7881 (Print) e - ISSN 1857- 7431 new progeny of viruses will be released in the medium. The released viruses again infect the new host cell and the cycle will be repeated. Thus, culture medium has a huge number of cultivated viruses (Bosch et al., 2015). Isolation of viruses The enriched media prepared as above will be used for isolation of viruses. After incubation, enriched broths/culture media will be cold lysed. The enriched cultures added with lytic enzymes such as trypsin and collagenase are allowed to stand for 1-2 hr in shaking incubator (at 100 rpm) at 4 oC. From these treated cultures, cell debris will be removed by centrifugation. The clear supernatant will be filter sterilized using bacterial filters. This filter sterilized supernatant directly will be used as virus suspension (Matsushita et al., 1995). Preparation of Gelatin Virus Balls and Liquid virus filled and sealed gelatin capsules Virus suspension will be used to prepare Gelatin virus balls (GVB)/ Liquid virus filled and sealed gelatin capsules (LVFSGC). Preparation of GVB GVB will be prepared using cold water soluble gelatin. Briefly preparation will be done by drying and aqueous solution containing virus suspension, gelatin and a hydrolyzed corn starch having dextrose, an acid (fumaric, citric, malic, adipic, ascorbic, tartaric, succinic, and phosphoric acids), and surface active agent containing a polysorbates, hydroxylated lecithin, acetylated monoglycerides, succinylated monoglycerides, ethoxylated mono- and diglycerides, sodium stearoyl 2-lactylate. These surface active agents render less drying period and more easily dispersible in cold water. GVB could be prepared (each has weight (...truncated)