ANTIBACTERIAL POTENTIAL OF BROWN ALGAE (SARGASSUM POLYCYSTUM) BACTERIAL SYMBIONT FROM COASTAL AREA IN BANTEN BAY, SERANG MUNICIPALITY, BANTEN OF INDONESIA

Brown algae are less popular and very few people pay attention to the plant; the algae are scattered on coastal areas and people treat them like garbage. Brown alga has as many nutrient contents and benefits as other types of algae. Objective of this research is to describe antibacterial potential of brown algae (Sargassum polycystum) bacterial symbiotic by isolating the brown algae bacterial symbiont with biossay method and conducting the challenge test using S.aureus and E.coli bacteria. The findings showed that brown algae (Sargassum polycystum) have potential for antibacterial by isolating a total of isolate 24 antibacterial compound-producing isolates, namely 7 isolates from surface of the algae and 17 isolates from inside of the algae. Qualitative challenge test found that 1 isolate (sa B1L) from the surface of the algae and 4 isolates (Sa B2D, sa B3D, sa B4D, sa B5D) from inside the algae can inhibit growth of E.coli and S.aureus.

We are blessed with various types of marine organism because 70% of the world is covered by sea. A type of marine organism that human being can use is macro algae or commonly known as seaweed (Ritonga, et.al, 2014). Seaweed is categorized into 3 classes, namely Chlorophyta (green seaweed), Phaeophyta (brown seaweed) and Rhodophyta (red seaweed). Each class contains certain pigment. Seaweed has primary and secondary metabolites. Primary metabolites consist of vitamin, mineral, fiber, alginate, carrageenan and agar. Besides the primary metabolite that has economic value, seaweed secondary metabolites such as tannins, phenol, saponins, alkaloids, flavonoids, and steroid produce various bioactive metabolite, which has several activities, namely as antibacterial, antiviral, antifungal and cytotastic (Zainunddin and Malina, 2009).
Marine algae try to develop a self-defense mechanism to fight pathogenic organisms in the sea. One general mechanism is to produce chemical compounds that are toxic to predators, including antibacterial active compounds. It is very likely that host organism, symbiont which is symbiotic with the host or both the host and the symbiont is responsible for producing the active compound (Nendissa, M, 2012).
A lot of research investigating benefits of seaweed based on microbiological perspective investigated microorganisms that are symbiotic with seaweed and can produce certain metabolites such as antimicrobials (Jafriati, 2014). Several recent studies have shown that seaweed micro-symbiotant can be one of the sources of bioactive compounds, but seaweed cells themselves can also produce these metabolites (Jafriati, 2014). Previous research on brown algae (Sargassum sp) reported that brown algae has antimicrobial potential in the form of bioactive protein from bacteria that are symbiotic with the algae (Sartika, et.al, 2014). Based on Pigment Test analyzing total carotenoid pigment extract and echinenone-type pigment of brown algae (Sargassum polycystum), 4 out 15 bacteria tested showed has antibacterial activity (Paransa, et.al, 2014). Sargassum polycystum extract from ethyl acetate solvent has the best antibacterial activity against Vibrio harveyi and micrococcus luteus bacteria (Riyanto, et.al, 2013).
Objective of this study is to describe antibacterial potential of brown algae (Sargassum polycystum) bacterial symbiont by isolating and selecting which brown algae (Sargassum polycystum) bacterial symbiont that produces antibacterial compounds and to describe characteristics of the bacterial symbiont obtained.

MATERIALS AND METHODS OF RESEARCH
Equipments used in this study were autoclave (Ikemoto scientific 10- The study was conducted between February 6 and May 6, 2017 in Microbiology Laboratory of Higher Institute of Fisheries of Jakarta. Sargassum polycystum samples were taken from coastal are of Banten Bay, Serang Municipality, Banten. Seaweed identification was conducted in Center of Oceanographic Research -Indonesian Institute of Science, DKI Jakarta.
Sargassum polycystum was obtained from coastal area of Banten Bay located in Pulau Lima, Serang Municipality. The samples were stored in clean containers filled with salt water from Sargassum polycystum habitat. The samples were then taken to the Microbiology Laboratory of Higher Institute of Fisheries of Jakarta.
Surface of algae: 15 grams of the samples were rinsed with 30 ml of sterilized salt water. The water was poured into 30 ml of Nutrien Broth and shaken using a shaker at 140 rpm at room temperature for 24 hours.
Inside of Algae: 15 grams of the samples were rinsed with 30 ml of sterilized salt water, ground using a mortar and pestle and sterilized salt water was poured into the pestle. Suspension was poured into 30 ml of NB and shaken using a shaker at 140 rpm at room temperature for 24 hours.
Having refreshed in NB, the samples were diluted gradually between 10 -1 and 10 -5 . Each dilution was grown in PCA with temperature of 37 o C for 2x24 hours. Bacteria with stable transparent zone were harvested and scratched with the quadrant method to obtain a single colony.
Took ± 1 ose of E.coli and S.aureus pure culture and inoculated the culture on 10 ml of Nutrient Broth (NB). Incubated the culture at 37⁰C for 24 hours.
Took ± 1 ose of bacterial symbiont pure culture from each code, and inoculated the culture on tilted Plate Count Agar (PCA). Incubated the culture at 37⁰C for 24 hours.
The following step was to conduct qualitative direct challenge test by scratching and spraying isolate onto Muller Hinton Agar (MHA) surface. 10 µl bacteria required 10 ml of liquid muller hinton agar (MHA). Bacteria-filled agar was vortexed or homogenized, poured into sterilized petri dish immediately and shaken to form number eight in order that the bacteria spread evenly. The media is sat to harden. The next step is conducting challenge test by scratching and spraying isolate on surface of media in which testing bacteria (E.coli and S.aureus) had been spread previously. The media were incubated for 2x24 hours under temperature of 37 0 C.

Macroscopic observation elevation and color) of bacterial
Gram staining provides hour culture). Gram staining crystal violet solution and resting solution and resting it for 1 minute, water, pouring Safranin solution drying and observing it under Brown Algae (Sargassum findings showed that brown antibacterial compound-producing 17 isolates from inside of the samples, the researchers identified the largest transparent zone side of the samples, the researchers isolates with the largest transparent B3D, sa B4D, sa B5D). Objective pure microbes. Pastra, et.al (2012) bacterial isolation result that consists bacterial colony from each bacterial and purified. Scratching was method Antibacterial Compound challenge test, 5 bacteria developed further. 1 bacterial isolate from isolates from the inside of the antibacterial activity against pathogenic isolate. Bacterial isolates on the surface of the population are fewer higher defense power to overcome bacteria from the inside of the the algae has a lot more and various Bacterial isolate obtained from the inside of the than that obtained from the surface of the sample bacterial symbiont obtained from the inside of algae is generally higher in number and is a specific microbe because it has direc produce. At the opposite, bacterial symbiont obtained from the surface of algae is fewer in better results when researchers use fresh was conducted by making preparate, fixing resting it for 1 minute, washing it with running water, minute, pouring bleach solution for 10-20 seconds, solution and resting the solution 15 seconds, wash a 100x microscope.

RESULTS AND DISCUSSION
Sargassum polycystum) Bacterial Symbiont Isolation algae (Sargassum polycystum) brown algae producing isolates, namely 7 isolates from surface algae; each is obtained from 10 -4 dilution. From identified 7 isolat and they were inoculated into on the surface of the algae (sa B1L, sa B2L researchers identified 17 isolates and they were transparent zone on the inside of the algae (sa Objective of inoculation is to grow, rejuvenate microbes (2012) stated that bacterial isolate purification consists of different types of bacterial colonies bacterial culture. Dominant colonies are those method for purification. Figure 1  Test). Based on the S.aureus from spreading code) and 4 bacterial and sa B5D) showed a clear zone in the seaweed. Bacteria from the nutrients and they require On the other hand, because the inside part of sample has better inhibitory zone rather . Abubakar, et.al (2011) stated that bacterial symbiont obtained from the inside of algae is generally higher in number and is a t interaction with bioactive compound the algae At the opposite, bacterial symbiont obtained from the surface of algae is fewer in number because it requires higher immunity to overcome pathogen and predator around the algae.
Cappucino and Sherman (1986) morphological colony of the bacteria with each code, either from the surface or inside part of the algae.  The table above showed B4D, sa B5D) were round and isolate colony of which code surface of the algae. In other colony is obtained. According occur due to differences in intracellular Based on the largest inhibition each bacterial symbiont isolate bacterial isolates with sa B3D S.aureus (5.55 mm) compared diameter of inhibitory zone indicate Furthermore, the sa B3D code algae were used as selected According to Rizka (2008), an B RJOAS, 12(84), December 2018 285 number because it requires higher immunity to overcome pathogen and predator around the Sherman (1986) as cited in Rizka (2008)  Sargassum polycystum symbiont of algae. ability of symbionous bacterial isolates to kill pathogenic bacteria. Ability to kill pathogenic bacteria can be characterized by clear zones formed around bacterial scratches; it proved that a tested pathogenic bacterium cannot grow anymore.
Macroscopic observation helps identifying shape, edge, elevation and color of bacterial colony. The researchers used this method to identify morphological characteristics of Sargassum polycystum bacterial symbiont isolates including its color. Table 1 showed result of the observation.
Gram staining is used to identify clear zones developed on the isolate. Table 3 showed result of the Gram staining.

CONCLUSION
Bacterial symbiont isolate from Sargassum polycystum (brown algae) from the water in Banten Bay consists of 24 isolates, 7 isolates from the surface of the algae and 17 isolates from the inside of the algae.
Based on challenge test, there are 5 isolates that can inhibit growth of S.aureus and E.coli, 1 isolate from the surface of the algae and 4 isolates from the inside of the algae.
Isolate obtained from Sargassum polycystum (brown algae) is categorized as gram positive bacteria.
Further researchers should identify phenotypes, genotypes and characteristics of secondary metabolites produced by Sargassum polycystum (brown algae) bacterial symbiont.