Identification of bacteriocin-producing bacteria from shrimp paste using 16S rRNA gene method and assay for MIC of their bacteriocin
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Abstract. Sukmawati S, Metusalach M, Syahrul S. 2025. Identification of bacteriocin-producing bacteria from shrimp paste using 16S rRNA gene method and assay for MIC of their bacteriocin. Biodiversitas 26: 3367-3377. Bacteria play a crucial role in various ecosystems, serving as key components in biogeochemical cycles and sources of bioactive compounds. Bacteriocins, a type of secondary metabolite, are antimicrobial compounds produced by bacteria to inhibit the growth of other microorganisms, notable for their specificity, stability, and eco-friendliness. These compounds have significant potential in various applications, ranging from food preservation to medical use as an alternative to antibiotics. This study aims to identify bacteriocin-producing bacteria isolated from shrimp (Acetes) paste (terasi) and evaluate the Minimum Inhibitory Concentration (MIC) of their crude bacteriocin for its antimicrobial potential. A descriptive method was employed to characterize the bacterial species producing crude bacteriocin from shrimp paste and assess the MIC ability of potential bacteria in inhibiting indicator bacteria. The results revealed that bacteriocin-producing bacteria isolated from shrimp paste, with sample codes TRS1, TRS2, and TRS3 were identified as Bacillus tropicus strain MCCC 1A01406, Bacillus nitratireducens strain MCCC 1A00732, and Bacillus paramycoides strain MCCC 1A04098, respectively. These three bacterial isolates could produce bacteriocins. The results of the Minimum Inhibitory Concentration (MIC) test showed that at the lowest bacteriocin concentration, but with the highest minimum inhibition, sample TRS1 had an inhibition value of 2.3 mm at a concentration of 0.19%, sample TRS3 had an inhibition value of 1.0 mm at a concentration of 0.19%, and sample TRS2 had an inhibition value of 1.2 mm at a concentration of 0.78%. A total of three isolates identified as Bacillus spp. demonstrated antimicrobial activity with MIC values ranging from 0.19% to 0.78%.
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References
Akolawole JS, Okoye PAC, Omokpariola D. 2022. Effect of storage on the levels of sodium benzoate in soft drinks sold in some Nigerian market with exposure and health risk assessment. Environ Anal Health Toxicol 37 (4): e2022030. DOI: 10.5620/eaht.2022030.
Apriyanto H, Harpeni E, Setyawan A. 2014. Utilization of Rhizophora sp. fruit extract as an antibacterial agent against freshwater fish pathogenic bacteria. E-Jurnal Rekayasa dan Teknologi Budidaya Perairan 3 (1): 289-296. [Indonesian].
Aziz MI, Hasan MM, Ullah R, Bari A, Khan MA, Ul Hasnain SZ, Baloch R, Akram M, Obaid A, Ullah A, Abbas K, Amin A. 2024. Potential role of Citrus bergamia flower essential oil against oral pathogens. BMC Complement Med Ther 24 (1): 157. DOI: 10.1186/s12906-024-04457-7.
Bahrami S, Andishmand H, Pilevar Z, Hashempour-Baltork F, Torbati M, Dadgarnejad M, Rastegar H, Mohammadi SA, Azadmard-Damirchi S. 2024. Innovative perspectives on bacteriocins: Advances in classification, synthesis, mode of action, and food industry applications. J Appl Microbiol 135 (11): lxae274. DOI: 10.1093/jambio/lxae274.
Barbosa AAT, de Melo MR, da Silva CMR, Jain S, Dolabella SS. 2021. Nisin resistance in Gram-positive bacteria and approaches to circumvent resistance for successful therapeutic use. Crit Rev Microbiol 47 (3): 376-385. DOI: 10.1080/1040841X.2021.1893264.
Besson S, Almeida MG, Silveira CM. 2022. Nitrite reduction in bacteria: A comprehensive view of nitrite reductases. Coord Chem Rev 464: 214560. DOI: 10.1016/j.ccr.2022.214560.
Brzoska AJ, Hassan KA. 2014. Quantitative PCR for detection of mRNA and gDNA in environmental isolates. Methods Mol Biol 1096: 25-42. DOI: 10.1007/978-1-62703-712-9_3.
Cheruvari A, Kammara R. 2024. Bacteriocins future perspectives: Substitutes to antibiotics. Food Control 168: 110834. DOI: 10.1016/j.foodcont.2024.110834.
Darbandi A, Asadi A, Ari MM, Ohadi E, Talebi M, Zadeh MH, Emamie AD, Ghanavati R, Kakanj M. 2022. Bacteriocins: Properties and potential use as antimicrobials. J Clin Lab Anal 36 (1): e24093. DOI: 10.1002/jcla.24093.
Davidson PM, Taylor TM, Schmidt SE. 2012. Chemical preservatives and natural antimicrobial compounds. In: Doyle MP, Buchanan RL (eds). Food Microbiology: Fundamentals and Frontiers. 4th eds. ASM Press, Washington, DC.
Du H, Li S, Yao H, Wang N, Zhao R, Meng F. 2024 Bacteriocin mining in Lactiplantibacillus pentosus PCZ4 with broad spectrum antibacterial activity and its biopreservative effects on snakehead fish. Foods 13 (23): 3863. DOI: 10.3390/foods13233863.
Duhan P, Bansal P, Rani S. 2020. Isolation, identification and characterizaton of endophytic bacteria from medicinal plant Tinospora cordifolia. S Afr J Bot 134: 43-49. DOI: 10.1016/j.sajb.2020.01.047.
El Mohammady EM, El-Rahim A, Mohamed W, Gohar MR, Abd El Aleem M, Elsayed TR. 2024. Applying wheat-associated bacteria's producing Volatile Organic Compounds (VOCs) antagonistic to plant pathogenic fungi. Egypt J Chem 67 (13): 1103-1115. DOI: 10.21608/ejchem.2024.325246.10557.
Elidrissi A, Ezzaky Y, Boussif K, Achemchem F. 2023. Isolation and characterization of bioprotective lactic acid bacteria from Moroccan fish and seafood. Braz J Microbiol 54 (3): 2117-2127. DOI: 10.1007/s42770-023-01077-0.
El-Refai HA, Saleh AM, Mohamed SIA, Aboul Naser AF, Zaki RA, Gomaa SK, Hamed MA. 2024. Biosynthesis of zinc oxide nanoparticles using bacillus paramycoides for in vitro biological activities and in vivo assessment against hepatorenal injury induced by CCl4 in rats. Appl Biochem Biotechnol 196 (9): 5953-5973. DOI: 10.1007/s12010-023-04817-y.
FDA [Food and Drug Administration] U.S. 2023. Code of Federal Regulations Title 21: Substances Generally Recognized as Safe. https://www.ecfr.gov/current/title-21
Felsenstein J. 1985. Confidence limits on phylogenies: An approach using the bootstrap. Evolution 39: 783-791. DOI: 10.1111/j.1558-5646.1985.tb00420.x.
Gonzalez JM, Aranda B. 2023. Microbial growth under limiting conditions-future perspectives. Microorganisms 11 (7): 1641. DOI: 10.3390/microorganisms11071641.
Heilbronner S, Krismer B, Brötz-Oesterhelt H, Peschel A. 2021. The microbiome-shaping roles of bacteriocins. Nat Rev Microbiol 19 (11): 726-739. DOI: 10.1038/s41579-021-00569-w.
Hoover DG, Steenson LR. 2014. Bacteriocins of Lactic Acid Bacteria. Academic Press, Cambridge, Massachusetts.
Hossain KM, Khan U, Rahman SM, Khan MS. 2024. Potential antimicrobial and fruit juice clarification activity of amylase enzyme from Bacillus strains. Biotechnol Rep 44: e00861. DOI: 10.1016/j.btre.2024.e00861.
Hwanhlem N, Buradaleng S, Wattanachant S, Benjakul S, Tani A, Maneerat S. 2011. Isolation and screening of lactic acid bacteria from Thai traditional fermented fish (Plasom) and production of Plasom from selected strains. Food Control 22 (3-4): 401-407. DOI: 10.1016/j.foodcont.2010.09.010.
Janda JM, Abbott SL. 2007. 16S rRNA gene sequencing for bacterial identification in the diagnostic laboratory: Pluses, perils, and pitfalls. J Clin Microbiol 45 (9): 2761-2764. DOI: 10.1128/jcm.01228-07.
Jiang Y-H, Xin W-G, Zhang Q-L, Lin L-B, Deng X-Y. 2022. A novel bacteriocin against Shigella flexneri from Lactiplantibacillus plantarum isolated From Tilapia intestine: Purification, antibacterial properties and antibiofilm activity. Front Microbiol 12: 779315. DOI: 10.3389/fmicb.2021.779315.
Karnwal A, Malik T. 2024. Exploring the untapped potential of naturally occurring antimicrobial compounds: novel advancements in food preservation for enhanced safety and sustainability. Front Sustain Food Syst 8: 1307210. DOI: 10.3389/fsufs.2024.1307210.
Kanauchi M. 2019. Lactic Acid Bacteria. Methods in Molecular Biology. Humana New York, NY. DOI: 10.1007/978-1-4939-8907-2.
Khan H, Flint SH, Yu P-L. 2014. Bacteriocins: The natural food preservatives. In: Rai VR, Bai JA (eds). Microbial Food Safety and Preservation Techniques. CRC Press, Boca Raton.
Kim D-H, Austin B. 2008. Characterization of probiotic carnobacteria isolated from rainbow trout (Oncorhynchus mykiss) intestine. Lett Appl Microbiol 47 (3): 141-147. DOI: 10.1111/j.1472-765X.2008.02401.x.
Kowalska-Krochmal B, Dudek-Wicher R. 2021. The minimum inhibitory concentration of antibiotics: Methods, interpretation, clinical relevance. Pathogens 10 (2): 165. DOI: 10.3390/pathogens10020165.
Lahiri D, Nag M, Dutta B, Sarkar T, Pati S, Basu D, Kari ZA, Wei LS, Smaoui S, Goh KW, Ray RR. 2022. Bacteriocin: A natural approach for food safety and food security. Front Bioeng Biotechnol 10: 1005918. DOI: 10.3389/fbioe.2022.1005918.
Li W, Lu H, He Z, Sang Y, Sun J. 2021. Quality characteristics and bacterial community of a Chinese salt-fermented shrimp paste. LWT 136: 110358. DOI: 10.1016/j.lwt.2020.110358.
Li Y, Ma H, Pan R, Long Y, Zhao Y, Yu M, Peng J, Ma Y. 2025. Optimisation of cultivation conditions for Bacillus velezensis G7 from mangrove plants and exploration of potential bacteriocins. Front Pharmacol 16: 1530043. DOI: 10.3389/fphar.2025.1530043.
Lim E-S. 2022. Influence of bacteriocin-producing Bacillus strains on quality characteristics of fermented soybean product with biogenic amine-forming lactic acid bacteria. Appl Biol Chem 65: 5. DOI: 10.1186/s13765-021-00664-9.
Ma H, Ding Y, Peng J, Li Y, Pan R, Long Y, Zhao Y, Guo R, Ma Y. 2025. Identification and characterization of a novel bacteriocin PCM7-4 and its antimicrobial activity against Listeria monocytogenes. Microbiol Res 290: 127980. DOI: 10.1016/j.micres.2024.127980.
Marchesi JR, Takuichi S, Andrew JW, Tracey AM, John CF, Sarah JH, William GW. 1998. Design and evaluation of useful bacterium-specific PCR primers that amplify genes coding for bacterial 16S rRNA. Appl Environ Microbiol 64: 795-799.
Muhiddin NH, Herawati N, Ramlawati R, Hidayah N. 2024. Characterization of antibiotic-producing bacteria from cassava tape fermented by traditional ragi tape. Biodiversitas 25 (9): 3359-3368. DOI: 10.13057/biodiv/d250958.
Naghavi M, Vollset SE, Ikuta KS et al. 2024. Global burden of bacterial antimicrobial resistance 1990-2021: A systematic analysis with forecasts to 2050. Lancet 404 (10459): 1199-1226. DOI: 10.1016/50140-6736(24)01867-1.
Ooi MF, Mazian N, Foo HL, Loh TC, Mohamad R, Rahim RA, Ariff A. 2015. Effects of carbon and nitrogen sources on bacteriocin-inhibitory activity of postbiotic metabolites produced by Lactobacillus plantarum I-UL4. Malays J Microbiol 11 (2): 176-184. DOI: 10.21161/mjm.13014.
Perez RH, Zendo T, Sonomoto K. 2022. Multiple bacteriocin production in lactic acid bacteria. J Biosci Bioeng 134 (4): 277-287. DOI: 10.1016/j.jbiosc.2022.07.007.
Pramono H, Suciati P, Putra TF, Andika N, Utari S. 2023. Reduction of pathogenic bacteria during fermentation of masin by protease and bacteriocin producing lactic acid bacteria. Aquasains 7 (1): 629-636. DOI: 10.23960/aqs.v7i1.p629-636. [Indonesian]
Ren W, Li P, Wang X, Che Y, Long H, Zhang X, Cai X, Huang A, Zeng Y, Xie Z. 2022. Cross-habitat distribution pattern of Bacillus communities and their capacities of producing industrial hydrolytic enzymes in Paracel Islands: Habitat-dependent differential contributions of the environment. J Environ Manag 323: 116252. DOI: 10.1016/j.jenvman.2022.116252.
Rus AMM, Winarko R, Chaerun SK, Mufakhir FR, Astuti W, Minwal WP. 2024. Bioleaching of Rare Earth Elements (REEs) from Indonesian red mud by the bacterium Bacillus nitratireducens strain SKC/L-2. E3S Web Conf 543: 02014. DOI: 10.1051/e3sconf/202454302014.
Sambrook J, Russell DW. 2001. Molecular Cloning a Laboratory Manual. 3rd eds. Cold Spring Harbor Laboratory Pr, New York.
Sardar D, Afsana S, Habib A, Hossain T. 2025. Dietary effects of multi-strain probiotics as an alternative to antibiotics on growth performance, carcass characteristics, blood profiling and meat quality of broilers. Vet Integr Sci 23 (2): 1-17. DOI: 10.12982/VIS.2025.059.
Seo Y, Sung M, Hwang J, Yoon Y. 2023. Minimum inhibitory concentration (MIC) of propionic acid, sorbic acid, and benzoic acid against food spoilage microorganisms in animal products to use MIC as threshold for natural preservative production. Food Sci Anim Resour 43 (2): 319. DOI: 10.5851/kosfa.2022.e79.
Sharma N, Kapoor RT, Gautam N, Kumari R. 2011. Purification and characterization of bacteriocin produced by Bacillus subtilis R75 isolated from fermented chunks of mung bean (Phaseolus radiatus). Food Technol Biotechnol 49 (2): 1330-1340.
Shu H, He X, Hong Z, Dong K, Zou Y, Cao M, Wang R, Xu Y, Liao L, Zuo H, Pei X. 2024. Screening and genome analysis of potential probiotic lactic acid bacteria with broad-spectrum antibacterial activity from Sichuan sun-dried vinegar grains (Cupei). LWT 202: 116288. DOI: 10.1016/j.lwt.2024.116288.
Sugrue I, Ross RP, Hill C. 2024. Bacteriocin diversity, function, discovery and application as antimicrobials. Nat Rev Microbiol 22: 556-571. DOI: 10.1038/s41579-024-01045-x.
Sukmawati S, Sipriyadi S, Rosalina F, Ratna R, Dewi NK, Zakariah MIB. 2024. Identification of 16S rRNA genes in bacteriocin-producing bacteria. Biodiversitas 25: 3720-3727. DOI: 10.13057/biodiv/d251034.
Sukmawati S, Sipriyadi S, Yunita M, Dewi NK, Noya ED. 2022. Analysis of bacteriocins of lactic acid bacteria isolated from fermentation of rebon shrimp (Acetes sp.) in South Sorong, Indonesia as antibacterial agents. Biodiversitas 23: 3852-3859. DOI: 10.13057/biodiv/d230763.
Susilowati DN, Widawati S, Suliasih S, Syah MF, Radiastuti N, Sofiana I, Anda M. 2024. Characteristics and potency of culturable bacteria from tidal and non-tidal swamps in South Kalimantan and South Sumatra, Indonesia. AIP Conf Proc 2970: 050015. DOI: 10.1063/5.0208354.
Tamura K, Peterson D, Peterson N, Stecher G, Nei M, Kumar S. 2011. MEGA5: Molecular Evolutionary Genetics Analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods. Mol Biol Evol 28 (10): 2731-2739. DOI: 10.1093/molbev/msr121.
Taylor TM, Davidson PM, Zhong Q. 2007. Extraction of nisin from a 2.5% commercial nisin product using methanol and ethanol solutions. J Food Prot 70 (5): 1272-1276. DOI: 10.4315/0362-028X-70.5.1272.
Wess L. 2023. The Curious World of Bacteria. Greystone Books Ltd, Canada.
Wu Y, Tie Y, Tang Z, Zhu H, Li Y, Zhu W, Wang X, Yin L. 2025. Isolation and identification of functional Bacillus strain from Cupei of sichuan bran vinegar and evaluation of its fermentation characteristics. Shipin Gongye Ke-ji 46 (2): 122-131. DOI: 10.13386/j.issn1002-0306.2024010069.
Ye J, Fugaban JII, Dioso CM, Bucheli JEV, Choi G-H, Kim B, Holzapfel WH, Todorov SD. 2023. Bacillus strains isolated from Korean fermented food products with antimicrobial activity against Staphylococci, an alternative for control of antibiotic-resistant pathogens. Foodborne Pathog Dis 20 (7): 279-293. DOI: 10.1089/fpd.2023.0019.
Zhang X, Xin N, Zhu Z, Li X, Dai D, Pan C, Peng D, Sun M. 2022. Three novel leaderless bacteriocins have antimicrobial activity against gram-positive bacteria to serve as promising food biopreservative. Microb Cell Fact 21 (1): 194. DOI: 10.1186/s12934-022-01912-3.