Screening of antibacterial activity of Crown-of-thorns starfish (Acanthaster spp.) extracts using modified microtiter-plate resazurin assay
Main Article Content
Abstract
Abstract. Tabugo SRM, Balatero TP, Dalayap RM. 2025. Screening of antibacterial activity of Crown-of-thorns starfish (Acanthaster spp.) extracts using modified microtiter-plate resazurin assay. Biodiversitas 26: 1735-1742. Marine organisms have emerged as promising sources for the discovery of new compounds that have medical application potential. Their ability to thrive in environments teeming with pathogenic microorganisms and predators indicates the production of unique chemical compounds with significant biological activities. This study identified and morphologically characterized Acanthaster spp. (Crown-of-Thorns, COT starfish) from various sites, including microhabitats in Iligan and Panguil Bays, and extracted polar and non-polar crude extracts for antibacterial activity screening. Samples were collected from three locations: Buruun, Iligan City; Samburon, Linamon; and Maigo, Lanao del Norte. Crude extracts were prepared using hexane (non-polar) and methanol (polar) solvents for all samples from the three sites. A modified microtiter antimicrobial assay was performed on all crude extracts, which exhibited antibacterial activity against Pseudomonas aeruginosa (a Gram-negative bacterium) and Staphylococcus aureus (a Gram-positive bacterium). The activity of HMCOT (hexane extract of samples from Maigo, Lanao del Norte) was comparable to Ciprofloxacin (positive control) based on the post-hoc test. Qualitative screening of the crude extracts for secondary metabolites revealed the presence of alkaloids, flavonoids, and saponins, which are likely contributors to the observed antibacterial activity. The diversity of the chemical compounds reflects the biodiversity of the source organisms, which have evolved and adapted to their specific environments. Thus, Acanthaster spp. shows potential for medical research, though further tests are necessary to validate these findings. The results of this study served as baseline data for future research.
Article Details
Issue
Section

This work is licensed under a Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International License.
References
Abd El Hafez MSM, Abd El Aziz Okbah M, Ibrahim HAH, Abd El Rahim Hussein A, Abd El Moneim NA, Ata A. 2022. First report of steroid derivatives isolated from starfish Acanthaster planci with anti-bacterial, anti-cancer and anti-diabetic activities. Nat Prod Res 36 (21): 5545-5552. DOI: 10.1080/14786419.2021.2021200.
Achmad MJ, Isnansetyo A, Kasanah N, Ustadi. 2018. Macrophage immunomodulatory activity of unsaturated fatty acid isolated from the crown-of-thorns starfish (Acanthaster planci). Pharmacogn J 10 (5): 951-957. DOI: 10.5530/pj.2018.5.161.
Andriani Y, Lazim NHM, Asari A, Mohamad F, Muhammad TST, Ismail N, Taib M, Amir H, Ahmad A, Mohamad H. 2018. Evaluation of selected echinoderms from Peninsular Malaysia for cytotoxicity against HepG2 cells, antioxidant and antibacterial activities, and their metabolites profiling. J Appl Pharm Sci 8 (10): 32-38. DOI: 10.7324/JAPS.2018.81005.
Birkeland C. 2015. Coral Reefs in the Anthropocene. Springer, Dordrecht. DOI: 10.1007/978-94-017-7249-5.
Brain KR, Jones BE, Turner TD. 1975. Application of densitometry to the qualitative and quantitative evaluation of pharmaceutical colourants. J Chromatogr 109 (2): 383-388. DOI: 10.1016/S0021-9673(01)91811-4.
Breznan D, Das D, MacKinnon-Roy C, Simard B, Kumarathasan P, Vincent R. 2015. Non-specific interaction of carbon nanotubes with the resazurin assay reagent: Impact on in vitro assessment of nanoparticle cytotoxicity. Toxicol In Vitro 29 (1): 142-147. DOI: 10.1016/j.tiv.2014.09.009.
Csepregi R, Lemli B, Kunsági-Máté S, Szente L, K?szegi T, Németi B, Poór M. 2018. Complex formation of resorufin and resazurin with ?-cyclodextrins: Can cyclodextrins interfere with a resazurin cell viability assay?. Molecules 23 (2): 382. DOI: 10.3390/molecules23020382.
Deaker DJ, Byrne M. 2022. Crown of thorns starfish life-history traits contribute to outbreaks, a continuing concern for coral reefs. Emerg Top Life Sci 6 (1): 67-79. DOI: 10.1042/ETLS20210239.
Ereguero MG, Gelani C, Dalayap R, Cordero MA, Tabugo SR. 2023. Antibacterial and anti-inflammatory potential of three sea cucumber species from Tukuran, Zamboanga del Sur, Mindanao, Philippines. Biodiversitas 24 (5): 2527-2535. DOI: 10.13057/biodiv/d240504.
Evans WC. 2009. Trease and Evans' Pharmacognosy. Saunders Ltd., United States.
Foo SA, Millican HR, Byrne M. 2024. Crown-of-thorns seastar (Acanthaster spp.) feeding ecology across species and regions. Sci Total Environ 930: 172691. DOI: 10.1016/j.scitotenv.2024.172691.
Gross IP, Lima AL, Sousa EC, Sousa MS, Cunha-Filho M, da Silva ICR, Orsi DC, Sá-Barreto LL. 2024. Antimicrobial and acaricide sanitizer tablets produced by wet granulation of spray-dried soap and clove oil-loaded microemulsion. PLoS One 19 (11): e0313517. DOI: 10.1371/journal.pone.0313517.
Hassim N, Markom M, Anuar N, Baharum SN. 2014. Solvent selection in extract of essential oil and bioactive compounds from Polygonum minus. J Appl Sci 14: 1440-1444. DOI: 10.3923/jas.2014.1440.1444.
Haywood MDE, Thomson DP, Babcock RC, Pillans RD, Keesing JK, Miller M, Rochester WA, Donovan A, Evans RD, Shedrawi G, Field SN. 2019. Crown-of-thorns starfish impede the recovery potential of coral reefs following bleaching. Mar Biol 166: 99. DOI: 10.1007/s00227-019-3543-z.
Hillberg AK, Smith MK, Lausen BS et al. 2023. Crown-of-thorns starfish spines secrete defence proteins. PeerJ 11: e15689. DOI: 10.7717/peerj.15689.
Hue T, Chateau O, Lecellier G, Kayal M, Lanos N, Gossuin H, Adjeroud M, Dumas P. 2020. Temperature affects the reproductive outputs of coral-eating starfish Acanthaster spp. after adult exposure to near-future ocean warming and acidification. Mar Environ Res 162: 105164. DOI: 10.1016/j.marenvres.2020.105164.
Ibrahim F, Widhyastuti N, Savitri IKE, Sahlan M, Wijanarko A. 2013. Antibacterial investigation of phospholipase A2 from the spines venom of crown-of-thorns starfish Acanthaster planci. Intl J Pharm Bio Sci 4 (2): B1-B5.
Kamyab E, Kellermann MY, Kunzmann A, Schupp PJ. 2019. Chemical biodiversity and bioactivities of saponins in Echinodermata with an emphasis on sea cucumbers (Holothuroidea). In: Jungblut S, Liebich V, Bode-Dalby M (eds). YOUMARES 9-The Oceans: Our Research, Our Future. Springer, Cham. DOI: 10.1007/978-3-030-20389-4_7.
Labnao ANP, Sienes PMQ, Palomar-Abesamis N. 2024. Morphological and molecular analysis of Western Pacific Crown-of-thorns Starfish Acanthaster cf. solaris (Schreber, 1793) in Southern Negros Island, the Philippines. Philipp J Sci 153 (6A): 2203-2215.
Lakna P. 2017. Difference between Gram-positive and Gram-negative bacteria. The Biology Blog, Welcome to the World of Biology. https:// pediaa.com/what-is-the-difference-between-alchemy-and-chemistry/
Lee C-C, Hsieh HJ, Hsieh C-H, Hwang D-F. 2014. Antioxidative and anticancer activities of various ethanolic extract fractions from crown-of-thorns starfish (Acanthaster planci). Environ Toxicol Pharmacol 38 (3): 761-773. DOI: 10.1016/j.etap.2014.08.021.
Lee C-C, Hsieh HJ, Hsieh C-H, Hwang D-F. 2015. Plancitoxin I from the venom of crown-of-thorns starfish (Acanthaster planci) induces oxidative and endoplasmic reticulum stress associated cytotoxicity in A375.S2 cells. Exp Mol Pathol 99: 7-15. DOI: 10.1016/j.yexmp.2015.05.001.
Ling SD, Cowan Z-L, Boada J, Flukes EB, Pratchett MS. 2020. Homing behaviour by destructive crown-of-thorns starfish is triggered by local availability of coral prey. Proc Biol Sci 287 (1938): 20201341. DOI: 10.1098/rspb.2020.1341.
Nedbalcová K, Pokludová L. 2020. Laboratory investigations and result interpretation. In: Pokludová L. (eds). Antimicrobials in Livestock 1: Regulation, Science, Practice. Springer, Cham. DOI: 10.1007/978-3-030-46721-0_8.
Petie R, Garm A, Hall MR. 2016. Crown-of-thorns starfish have true image-forming vision. Front Zool 13 (1): 41. DOI: 10.1186/s12983-016-0174-9.
Pitout JDD. 2008. Multiresistant Enterobacteriaceae: New threat of an old problem. Expert Rev Anti Infect Ther 6 (5): 657-669. DOI: 10.1586/14787210.6.5.657.
Prakash A, Sarangdevot YS, Vyas B, Roy S. 2022. Extraction and Characterization of Ficus glomerata for Antioxidant activity. Asian J Pharm Clin Res 15 (6): 46-50. DOI: 10.22159/ajpcr.2022.v15i6.44404.
Sarker SD, Nahar L, Kumarasamy Y. 2007. Microtitre plate-based antibacterial assay incorporating resazurin as an indicator of cell growth, and its application in the in vitro antibacterial screening of phytochemicals. Methods 42 (4): 321-324. DOI: 10.1016/j.ymeth.2007.01.006.
Sumitha R, Parvathi VD, Banu N. 2017. Cytotoxicity testing of star fish Stellaster equestris extracts on Pa1 celline. Res J Pharm Technol 10 (9): 2851-2856. DOI: 10.5958/0974-360X.2017.00502.9.
Teh CH, Nazni WA, Nurulhusna AH, Norazah A, Lee HL. 2017. Determination of antibacterial activity and minimum inhibitory concentration of larval extract of fly via resazurin-based turbidometric assay. BMC Microbiol 17 (1): 36. DOI: 10.1186/s12866-017-0936-3.
Travnickova E, Mikula P, Oprsal J, Bohacova M, Kubac L, Kimmer D, Soukupova J, Bittner M. 2019. Resazurin assay for assessment of antimicrobial properties of electrospun nanofiber filtration membranes. AMB Express 9 (1): 183. DOI: 10.1186/s13568-019-0909-z.
Wijanarko A, Lischer K, Hermansyah H, Pratami DK, Sahlan M. 2018. Antiviral activity of Acanthaster planci phospholipase A2 against human immunodeficiency virus. Vet World 11 (6): 824-829. DOI: 10.14202/vetworld.2018.824-829.
Yadav S, Verma M. 2018. Phytochemical screening of peepal leaves. Indian J Health Wellbeing 9 (6): 872-873.
Zheng R, Li S, Zhang X, Zhao C. 2021. Biological activities of some new secondary metabolites isolated from endophytic fungi: A review study. Intl J Mol Sci 22 (2): 959. DOI: 10.3390/ijms22020959.