1. TOPIC OF RESEARCH:
“Immuno-modulatory compound from fresh water sponge Eunapius. carteri helps fishes to fight against bacterial and fungal infections”

2. ABSTRACT:

3. OUTLINE:

? Bioactive compounds from fresh water sponges and its antimicrobial activities.

• Extraction of bioactive compound from fresh water sponge.
• Purification of bioactive compound and forming crude compound.
• Isolation of bacterial pathogens from infected fishes.
• Isolation of fungal pathogens from fungal infected fishes.
• Antibacterial activity of crude compound against fish pathogens.
• Antifungal activity of crude compound against fish pathogens.
• Minimum inhibitor concentration of crude compound against fish pathogens.

4. RESEARCH DESIGN:

A. INTRODUCTION:

B. RESEARCH QUESTION:

• Replacement of exciting antibacterial and antifungal drugs with more effective and natural ones has become main question to address.
• Is fresh water sponges help freshwater fishes to increases its immunity to fight against different bacterial and fungal infections?

C. HYPOTHESIS:

Marine sponges contains variety of bioactive compounds which having immunomodulatory activities. Fresh water sponges may also contain these type of bioactive compounds having antibacterial and antifungal activities which may help fresh water fishes to fight against infections and aid them in increasing their immunity.
Aims to find antibacterial and antifungal activities from fresh water sponges.

? SUPPLEMENTARY QUESTIONS:

• From where we can isolate bioactive compound?
From fresh water sponge Eunapius. carteri.
(Porifera: Demospongiae: Spongillidae a common variety of freshwater sponge).

• Why fresh water sponges?
Marine sponges are known to produces many metabolites processing various bioactive activities, some are also went for clinical trials. Whereas fresh sponges is less studied group having relatively limited information.

• Will fresh Sponges help more than marine sponges?
Technical problems associated with cultivation and harvesting large amounts of marine sponges. Whereas Fresh water sponges are easy to cultivate and grow and harvest artificially as we can grow them in aquariums.

• What type of bacteria infect fishes? And what disease they cause?
1) Aeromonas. salmonicida causes Furunculosis forms boil-like lesions or may also appear dark in coloration at the base of their fins.
2) Flavobacterium. Branchiophilum causes gill disease, leading to gill proliferation in fishes.
3) Pseudomonas.spp and vibrio.spp can cause frayed fins with hemorrhaging at the base of the fin, sores, lethargy, and swelling of the belly.

• What type of fungus infect fishes? And what disease they cause?
1) Exophiala. spp are ubiquitous yeast and are distinct olive to black-brown color lesions on fishes.
2) Saprolegnia.spp freshwater specie of fungi which causes cottony/woolly, white growth on the skin, or gills, or on fish eggs.
3) Aphanomyces. Invadans causes deep ulcerative orange to red lesions on skin scrapes.

D. OBJECTIVE OF RESEARCH:
Considering the importance of green health management in aquaculture and concern about the contamination, toxicological and environmental risks posed by synthetic drugs has led to an increase in the popularity of developing natural products as a source of ecofriendly compounds possessing antimicrobial activities.
In addition, the evolving resistance of microorganisms to existing antibiotics is becoming major issue, not only in human research but also in aquaculture. This threat is increasing day by day causing immense economic losses resulting in food insecurity. Hence, replacement of existing antibiotics with more operational and safer ones has become an important subject to discuss.

E. LITERATURE REVIEW:
Sponges are spineless animals belong to phylum, “the pore bearers” (Porifera), serve as most primitive multicellular animals, existing for millions of year ago. They frequently produce bioactive compounds as compared to other living microorganisms. Because sponges cannot move and lack physical defenses, they are highly susceptible to marine predators such as fish, turtles, and invertebrates. Thus, it is not surprising that sponges have developed a wide suite of defensive chemicals to deter predators, (Anjum, 2016) microbial infections, biofouling, and overgrowth by other sessile organisms (M. F. Mehbub, 2018).

While natural products have traditionally been harvested from terrestrial sources, from sponges and their associates, approximately 5,300 different natural compounds are known (Bibi, 2016). A major contributing factor to this development is the fact that modern technology has made it easier to gain access to the great biodiversity of life found in the oceans (Margey Tadessea, 2008).

Oceans are most primitive, important and unique form of life on the earth. It provides a huge diversity of living organisms inhabiting diverse microflora. The marine resources are widely studied nowadays because of numerous reasons. One of the reason is, the oceans cover more than 70% of planet surface and among 36 living phyla known yet, 34 of them are found in marine environments with more than 300000 known species of fauna and flora (Bibi, 2016).

Compounds isolated from sponges contains anticancer, anti-inflammatory, antiviral, antibacterial, anticoagulant (Roberta J. Melander, 2016), antitumour (Margey Tadessea, 2008), antifungal (Prabha Devi, 2013), cytotoxic, antidiabetic, antimalarial, antiplatelet, antiprotozoal, antileukemic, anti- tuberculosis, (G. Annie Selva Sonia, 2008) and immunomodulatory activities (Soumalya Mukherjee, 2016). Considering their scope of antibiotic activity against fish pathogenic bacteria, marine sponge extracts are prime candidates as sources of bioactive metabolites (G. Annie Selva Sonia, 2008)

The discovery of penicillin in the mid-twentieth century revolutionized the treatment of infectious disease. Since then, antimicrobial agents have saved the lives and eased the suffering of millions of people. Multi-resistant bacteria threaten to cause new epidemics (Bibi, 2016).

Evidence suggest that development of resistance to any new antimicrobial agents is inevitable (Prabha Devi, 2013). So the evolving resistance has made necessary a search for new antibiotics for human as well as aqua cultural purposes. In the aquatic environment, competition for space and nutrients leads to evolution of antimicrobial defense strategies. This, along with possibly adverse effects on the ecosystem and human health problems, has resulted in restrictions on the use of commercial antibiotics and chemicals in the aquatic environment (G. Annie Selva Sonia, 2008).

Emerging infectious diseases (EIDs) caused by fungi are increasingly recognized as presenting a worldwide threat to food security. This is not a new problem and fungi have long been known to constitute a widespread threat to plant species. However, pathogenic fungi (mycoses) have not been widely recognized as posing major threats to animal health. This perception is changing rapidly owing to the recent occurrence of several high-profile declines in wildlife caused by the emergence of previously unknown fungi (Matthew C. Fisher, 2012). For more than two decades worldwide and fungal infections are amongst the common diseases in hatchery and aquaculture systems leading to the demise of fish population resulting in great economic loss (Prabha Devi, 2013).

Many structurally diverse marine sponge secondary metabolites have been shown to exhibit antibiotic activities against several Gram-positive bacteria including Streptomyces. pyogenes, Staphylococcus.aureus and Bacillus. subtilis. However many of these natural products are in active against Gram-negative bacteria (Roberta J. Melander, 2016).
In most cases development and production of sponge derived drugs is hindered by environmental concerns and technical problems associated with harvesting large amounts of sponges. But now presence of sustainable source of sponge-derived drug candidates could be generated by establishing a symbiont culture or by transferring its biosynthetic genes into culturable bacteria (Anjum, 2016). There are a few examples of marine derived compounds which have successfully reached the market as therapeutic drugs (Margey Tadessea, 2008).

Multi drug resistant Staphylococcus. aureus (MRSA) formerly particularly problematic in places such as hospitals and nursing homes, is now found in commonly-used places. Scientists have isolated an extract from a sponge found in Antarctica, tested it on MRSA biofilm and found that it eliminate more than 98 percent of MRSA cells. The highly-resistant MRSA infection (USF, 2016). Several strains were identified for their potent antifungal activity, and for both antifungal and antibacterial activities (University, 2018).

Benthic marine invertebrates (Sponges) were found to be a promising source of novel bioactive compounds against human and fish pathogenic bacteria and fungi (Margey Tadessea, 2008). Freshwater poriferans are relatively a less studied group with limited scientific information (Soumalya Mukherjee, 2016). Eunapius carteri (Porifera: Demospongiae: Spongillidae a common variety of freshwater sponge) is distributed in seasonal ponds and lakes.

F. METHODOLOGY:

G. TIME FRAME:

5. REFERENCES

1) Anjum, K. A. (2016). Marine Sponges as a Drug Treasure . Biomolecules & Therapeutics, 24(4), 347–362. doi:http://doi.org/10.4062/biomolther.2016.067
2) Bibi, F. &. (2016). Bacteria from marine sponges: A source of new drugs. Current Drug Metabolism, 17. doi:10.2174/1389200217666161013090610.
3) G. Annie Selva Sonia, A. L. (2008). Antibacterial Activity of Marine Sponge Extracts against. The Israeli Journal of Aquaculture , 60(3), 172-176. Retrieved from https://pdfs.semanticscholar.org/c894/5e6c27890a0b5b9e7e8b10261e989940f760.pdf
4) M. F. Mehbub, J. E. (2018). A controlled aquarium system and approach to study the role of sponge-bacteria interactions using Aplysilla rosea and Vibrio natriegens. Nature. doi:10.1038/s41598-018-30295-y
5) Margey Tadessea, B. G. (2008). Screening for antibacterial and antifungal activities in marine benthic invertebrates from northern Norway. Journal of Invertebrate Pathology , 99(3), 286-93. doi:http://dx.doi.org/10.1016/j.jip.2008.06.009
6) Matthew C. Fisher, D. A. (2012). Emerging fungal threats to animal, plant and ecosystem health. Nature, 484, 186–194. doi:10.1038/nature10947
7) Prabha Devi, R. S. (2013). Antifungal Potential of Marine Sponge Extract against Plant and Fish. Oceanography, 1(3), 112. doi:http://dx.doi.org/10.4172/2332-2632.1000112
8) Roberta J. Melander, H.-b. L. (2016). Marine sponge alkaloids as a source of anti-bacterial adjuvants. Bioorg Med Chem Lett. , 26(24), 5863–5866. doi: doi:10.1016/j.bmcl.2016.11.018
9) Soumalya Mukherjee, A. S. (2016). Immunomodulatory effects of temperature and pH of water in an Indian freshwater sponge. Journal of Thermal Biology, 59, 1-12. doi:https://doi.org/10.1016/J.JTHERBIO.2016.04.005
10) University, F. A. (2018). Deep-sea marine sponges may hold key to antibiotic drug resistance. ScienceDaily. Retrieved from www.sciencedaily.com/releases/2018/06/180619123013.htm
USF, U. o. (2016, May). Scientists discover Antarctic sponge extract can help kill MRSA: New findings may provide opportunity for developing new drugs to fight dangerous bacteria currently highly resistant to treatment. ScienceDaily. Retrieved from www.sciencedaily.com/releases/2016/05/160518094706.htm