Main Article Content
Aims: To determine bacterial diversity in milkfish culture ponds that contain different life-cycle stages of the milkfish (pond A: fry, pond B: juveniles and pond C: adults) by DNA sequence analysis of organisms and compare that microbial diversity to organisms found in soil adjacent to the ponds.
Study Design: Comparative metagenomic study of aquatic and terrestrial biodiversity based on DNA sequence analysis of water and soil DNA.
Place and Duration of Study: SEADEC milkfish ponds in Tingnauan, Iloilo. Philippines. All water and soil samples were collected over a three-day period.
Methodology: DNA sequence analysis of nucleic acids extracted from water samples collected from the three types of milkfish ponds along with soil adjacent to the ponds. DNA was extracted and PCR was performed using the 11F-1492R primer pair to amplify 16S rRNA gene. Purified 16S rDNA amplicons were cloned in using the TOPO-TA cloning kit for DNA sequencing. 16s rRNA gene sequences were analyzed with the use of software tools at the National Center for Biotechnology Information website and imported into the ARB phylogenetic analysis software. Distance matrices were exported using the neighbor-joining algorithm in ARB, in the form of PHYLIP-formatted lower triangular matrices. The distance matrices were then used to calculate Shannon-Weaver and Simpson diversity indices to evaluate the richness and evenness of the sampled populations. Rarefaction curves were determined to evaluate sampling efficiency.
Results: Rarefaction curves indicated that the sampling effort was sufficient to reveal the majority of phyla present in the sample. Shannon-Weaver and Simpson indices suggested that the diversities of all the groups were statistically different from each other. It was observed that pond A was least diverse, followed by pond C and pond B. The soil was most diverse. DNA sequence analysis identified the various species of bacteria in soil and water.
Conclusion: All three pond communities were significantly different in diversity. This study did not identify any significant human pathogens such as Vibrios, Salmonella or Shigella. Bacterial diversity of sites decreased in the following order: soil > fry pond > fingerling pond > adult pond.
Luckstadt C, Focken U, Coloso R, Becker K. Survey on the use of natural food and supplemental feed in commercial milkfish farms on Panay, Philippines. Poster presented at Deutscher Tropentag 2000. International Agricultural Research – A Contribution to Crisis Prevention, University of Hohenheim; 2000.
Luckstadt C. Feed intake and feed utilization of juvenile milkfish (Chanos, Forsskål, 1775) in commercially managed ponds in the Philippines. Diss. Univ. Hohenheim 2004, Aachen: Shaker Verlag. 2004;192.
Gordon SM, Hong LQ. Biology. In: Lee CS, Gordon MS, Watanabe WO. Editors. Aquaculture of milkfish (Chanos chanos): State of the art. Hawaii: The Oceanic Institute.1980;284.
Bagarinao TU. Biology of milkfish (Chanos Forsskal). Aquaculture Department Southeast Asian Fisheries Development Center, 991, Tigbauan, Iloilo, Philippines. 1991;94.
Villaluz AC, Villaver WR, Salde RJ. Milkfish fry and fingerling industry of the Philippines: Methods and practices. Aquaculture Department, SEAFDEC. International Development Research Centre. Technical Report No. 9. 2nd Edition. 1983;81.
Lee CS. Aquaculture of milkfish (Chanos chanos). Tungkang Marine Laboratory, Taiwan Fisheries Research Institute. Tungkang Marine Laboratory, TFRI, Taiwan & the Oceanic Institute, Hawaii, U.S.A. TML Aquaculture Series No. 1; 1990.
Mwakalap EB, Simukoko CK, Mmochi AJ, Mdegegela RH, Berg V, Bjorge GE, Muller MH, Lyche JL, Polder A. Heavy metals in farmed and wild milkfish along the coasts of Tanzania and associated health risk for humans and fish. Chemosphere. 2019; 224:176-186.
Ragasa LRP, Dinglasan JLN, Filipe IRE, Basiaso ZU, Verlande MC. Exposure to Aeromonas hydrophilia induced inflammation and increases expression of the gene encoding for a putative dual CTLD-containing lectin in milkfish liver. Comp Biochem Physiol B Biochem Mol Biol. 2019;230:37-47.
Pleto JVR, Arboleda MDM, Simbahan JF, Migo VP. Assessment of the effect of remediation strategies on the environmental quality of aquaculture ponds in Marilao and Meycauayan, Bulacan, Philippines. J Health Pollut. 2018;8(20): 181205.
Tey YH, Jong KJ, Fen SY, Wong HC. Genetic variation in Vibrio parahaemolyticus isolated from aquaculture environments. Lett Appl Microbiol. 2015;60(4):321-7.
Elhadi N. Prevalence and antimicrobial resistance of Salmonella spp. In raw retail frozen imported freshwater fish to Eastern Province of Saudi Arabia. Asian Pac J Trop Biomed. 2014;4(3): 234-8.
Reichardt WT, Reyes JM, Pueblos MJ, Lluisma AQ. Impact of milkfish farming in the tropics and potentially pathogenic vibrios. Mar Pollut Bull. 2013;77(1-2):325-32.
Rasheeda MK, Rangamaran VR, Srinivasan S, Ramaiah SK, Gunasekaran R, Jaypal S, Gopal D, Ramalingam K. Comparative profiling of microbial community of three economically important fishes reared in sea cages under tropical offshore environment. Marine Genomics. 2017;34:57-65.
Zeyaullah M, Kamli MR, Islam B, Atif M, Benkhayal FA, Nehal M, Rizvi MA, Ali A. Metagenomics - An advanced approach for non-cultiviable micro-organisms. Biotechnology and Molecular Biology Reviews. 2009;4(3): 49-54.
Frias-Lopez J, Shi Y, Tyson GW, Coleman ML, Schuster SC, Chisholm SW, Delong EF. Microbial community gene expression in ocean surface waters. Proc Natl Acad Sci USA. 2008;105(10): 3805-10.
Lozupone C, Hamady M, Knight R. UniFrac - An online tool for comparing microbial community diversity in a phylogenetic context. BMC Bioinformatics. 2006;7:371.
US National Center for Biotechnology Information.
The ARB Project.
Fudou R, Jojima Y, Iizuka T, Yamanaka S. Haliangium ochraceum gen. nov., sp. nov. and Haliangium tepidum sp. nov.: Novel moderately halophilic myxobacteria isolated from coastal saline environments. J Gen Appl Microbiol. 2002;48(2):109-16.
Monciardini P, Cavaletti L, Schumann P, Rohde M, Donadio S. Conexibacter woesei gen. nov., sp. nov., a novel representative of a deep evolutionary line of descent within the class Actinobacteria. Int J Syst Evol Microbiol. 2003;53(Pt 2): 569-76.
Kurahashi M, Fukunaga Y, Sakiyama Y, Harayama S, Yokota A. Iamia majanohamensis gen. nov., sp. nov., an actinobacterium isolated from sea cucumber Holothuria edulis and proposal of Iamiaceae fam. nov. Int J Syst Evol Microbiol. 2009;59(Pt 4):869-73.
Lechner U, Brodkorb D, Geyer R, Hause G, Härtig C, Auling G, Fayolle-Guichard F, Piveteau P, Müller RH, Rohwerder T. Aquincola tertiaricarbonis gen. nov., sp. nov., a tertiary butyl moiety-degrading bacterium. Int J Syst Evol Microbiol. 2007; 57(Pt 6):1295-303.
Hristova KR, Lutenegger CM, Scow KM. Detection and quantification of methyl tert-butyl ether-degrading strain PM1 by real-time TaqMan PCR. Appl Environ Microbiol. 2001;67(11):5154-60.
Liu H, Wang SJ, Zhou NY. A new isolate of Pseudomonas stutzerithat degrades 2-chloronitrobenzene. Biotechnol Lett. 2005; 27(4):275-8.