Bactericidal activity of Piper betle L. extract against antibiotic resistant Salmonella spp. isolated from pig farms in Southern Thailand

Main Article Content

Ratchadaporn Boripun
Panida Paopradit
Jindarha Prampramote
Ruethai Narinthorn
Sutsiree Intongead
Suthinee Sangkanu
Tanapat Narongrit
Watcharapong Mitsuwan


Piper betle L. leaves have traditionally been used to treat various infectious diseases and to possess a wide spectrum of pharmacological effects. This study aimed to determine the antibacterial activity of the Piper betle leaf extract against antibiotic resistant Salmonella spp. isolated from pig farms located in Southern Thailand. Of this, 12 Salmonella spp. isolates were isolated from 24 pig fecal samples from 24 pig farms. The isolates were resistant to ampicillin (91.67%), penicillin (91.67%), tetracycline (81.81%), and doxycycline (81.81%). Antibacterial activity of the Piper betle ethanolic leaf extract against Salmonella spp. was carried out by disc diffusion assays, followed by Minimal inhibitory concentration (MIC) and Minimal bactericidal concentration (MBC) determination, as well as Time kill study. Piper betle extract exhibited antibacterial activity against all the isolates and S. Typhimurium with the inhibition zone ranged from 15.11 ± 0.34 to 20.30 ± 0.50 mm as observed by disc diffusion assay. The extract showed bactericidal activity against the isolates with the MIC and MBC values ranging from 0.5-1.0 mg/mL. Furthermore, the extract at 4 × MIC showed the killing activity with the reduction of the pathogen at least 3 logs within 8 h. The information suggests potential medicinal benefits of the Piper betle leaf extract to inhibit the growth of antibiotic resistant Salmonella spp. isolated from pig farms. 

Article Details

How to Cite
Boripun, R., Paopradit, P. ., Prampramote, J. ., Narinthorn, R. ., Intongead, S. ., Sangkanu, S. ., Narongrit, T. ., & Mitsuwan, W. . (2022). Bactericidal activity of Piper betle L. extract against antibiotic resistant Salmonella spp. isolated from pig farms in Southern Thailand: Veterinary Integrative Sciences, 20(3), 557–569. Retrieved from
Research Articles


Choudhary, D., Kale, R.K., 2002. Antioxidant and non-toxic properties of Piper betle leaf extract: in vitro and in vivo studies. Phytother Res. 16, 461-466.

Clinical and Laboratory Standards Institute, 2019. Performance Standards for Antimicrobial Susceptibility Testing CLSI supplement M100, 29th edition. CLSI, Pennsylvania.

Cowan, M.M., 1999. Plant products as antimicrobial agents. Clin. Microbiol. Rev. 12(4), 564-582.

Cushnie, T.P., Lamb, A.J., 2005. Antimicrobial activity of flavonoids. Int. J. Antimicrob. Agents. 26(5), 343-356.

Ermawati, F.U., Sari, R., Putri, N.P., Rohmawati, L., Kusumawati, D.H., Munasir, Supardi, Z.A.I., 2021. Antimicrobial activity analysis of Piper betle linn leaves extract from nganjuk, sidoarjo and batu against Escherichia coli, Salmonella sp., Staphylococcus aureus and Pseudomonas aeruginosa. J. Physics Conference Series. 1951(1), 012004.

Erwiyani, A. R., Luhurningtyas, F. P., Sunnah, I., 2017. Optimasi formula sediaan krim ekstrak etanol daun alpukat (Persea americana Mill) dan daun sirih hijau (Piper betle Linn). Cendekia. J. Pharm. 1(1), 77-86.

Fakruddin, M., Sultana, R., Hossain, M. N., Rahaman, M. M., Islam, M. K., Ahmed, M. M., 2017. Occurrence of ingression of Salmonella spp. in Betel leaf (Piper betle L.). Int. J. Food Contam. 4(1), 1-10.

Gal-Mor, O., Boyle, E. C., Grassl, G. A., 2014. Same species, different diseases: how and why typhoidal and non-typhoidal Salmonella enterica serovars differ. Front. Microbiol. 5, 391.

Gavriil, A., Zilelidou, E., Papadopoulos, A.E., Siderakou, D., Kasiotis, K.M., Haroutounian, S.A., Gardeli, C., Giannenas, I., Skandamis, P.N., 2021. Evaluation of antimicrobial activities of plant aqueous extracts against Salmonella Typhimurium and their application to improve safety of pork meat. Sci. Rep. 11(1), 21971.

Gong, B., Li, H., Feng, Y., Zeng, S., Zhuo, Z., Luo, J., Chen, X., Li, X., 2022. Prevalence, serotype distribution and antimicrobial resistance of non-Typhoidal Salmonella in hospitalized patients in conghua district of Guangzhou, China. Front. Cell Infect. Microbiol. 12, 805384.

Holohan, N., Wallat, M., Hai Yen Luu, T., Clark, E., Truong, D.T.Q.,, Xuan, S.D., Nguyen-Viet, H., 2022. Analysis of antimicrobial resistance in Non-typhoidal Salmonella from retail stores in Vietnam. Front. Vet. Sci. 214.

Jaber, H., Oubihi, A., Ouryemchi, I., Boulamtat, R., Oubayoucef, A., Bourkhiss, B., Ouhssine, M., 2021. Chemical composition and antibacterial activities of eight plant essential oils from Morocco against Escherichia coli strains isolated from different Turkey organs. Biochem. Res. Int. 6685800.

Kulnanan, P., Chuprom, J., Thomrongsuwannakij, T., Romyasamit, C., Sangkanu, S., Manin, N., Kitpipit, W., Mitsuwan, M., 2022. Antibacterial, antibiofilm, and anti-adhesion activities of Piper betle leaf extract against avian pathogenic Escherichia coli. Arch. Microbiol. 204(1), 1-14.

Lauteri, C., Maggio, F., Serio, A., Festino, A. R., Paparella, A., Vergara, A., 2021. Overcoming multidrug resistance in Salmonella spp. isolates obtained from the swine food chain by using essential oils: an in vitro Study. Front. Microbiol. 12, 808286.

Lubis, R.R., Marlisa, Wahyuni, D.D., 2020. Antibacterial activity of betle leaf (Piper betle L.) extract on inhibiting Staphylococcus aureus in conjunctivitis patient. Am. J. Clin. Exp. Immunol. 9, 1-5.

Meurens, F., Berri, M., Auray, G., Melo, S., Levast, B., Virlogeux-Payant, I., Salmon, H., 2009. Early immune response following Salmonella enterica subspecies enterica serovar Typhimurium infection in porcine jejunal gut loops. Vet. Res. 40(1), 5.

Mitsuwan, W., Sangkanu, S., Romyasamit, C., Kaewjai, C., Jimoh, T.O., de Lourdes Pereira, M., Siyadatpanah, A., Kayesth, S., Nawaz, M., Rahmatullah, M., Butler, M.S., Wilairatana, P., Wiart, C., Nissapatorn, V., 2020. Curcuma longa rhizome extract and curcumin reduce the adhesion of Acanthamoeba triangularis trophozoites and cysts in polystyrene plastic surface and contact lens. Int. J. Parasitol. Drugs Drug Resist. 14, 218-229.

Mitsuwan, W., Wintachai, P., Voravuthikunchai, S.P., 2020. Rhodomyrtus tomentosa leaf extract and rhodomyrtone combat Streptococcus pneumoniae biofilm and inhibit invasiveness to human lung epithelial and enhance pneumococcal phagocytosis by macrophage. Cur. Microbiol. 77, 3546-3554.

Mukherjee, A., Cao, C., Lutkenhaus, J., 1998. Inhibition of ftsz polymerization by sula, an inhibitor of septation in Escherichia coli. Proc. Natl. Acad. Sci. U. S. A. 95(6), 2885-2890.

Mukherjee, P.K., Maity, N., Nema, N.K., Sarkar, B.K., 2011. Bioactive compounds from natural resources against skin aging. Phytomedicine. 19, 64-73.

Nayaka, N., Sasadara, M.M.V., Sanjaya, D.A., Yuda, P., Dewi, N., Cahyaningsih, E., Hartati, R., 2021. Piper betle (L): recent review of antibacterial and antifungal properties, safety profiles, and commercial applications. Molecules. 26(8), 2321.

Naz, S., Alam, S., Ahmed, W., Khan, S. M., Qayyum, A., Sabir, M., Nisa, S., 2022. Therapeutic potential of selected medicinal plant extracts against multi-drug resistant Salmonella enterica serovar Typhi. Saudi J. Biol. Sci. 29(2), 941-954.

Pecková, R., Doležal, K., Sak, B., Květoňová, D., Kváč, M., Nurcahyo, W., Foitová, I., 2018. Effect of Piper betle on Giardia intestinalis infection in vivo. Exp. Parasitol. 184, 39-45.

Pereira, W.A., Pereira, C.D.S., Assunção, R.G., da Silva, I.S.C., Rego, F.S., Alves, L.S.R., Santos, J.S., Nogueira, F.J.R., Zagmignan, A., Thomsen, T.T., Løbner-Olesen, A., Krogfelt, K.A., da Silva, L.C.N., Abreu, A.G., 2021. New insights into the antimicrobial action of cinnamaldehyde towards Escherichia coli and its effects on intestinal colonization of mice. Biomolecules. 11(2), 302.

Qin, X., Yang, M., Cai, H., Liu, Y., Gorris, L., Aslam, M.Z., Jia, K., Sun, T., Wang, X., Dong, Q., 2022. Antibiotic resistance of Salmonella Typhimurium monophasic variant 1,4,[5],12:I:-in china: a systematic review and meta-analysis. Antibiotics (Basel). 11(4), 532.

Singh, D., Narayanamoorthy, S., Gamre, S., Majumdar, A. G., Goswami, M., Gami, U., Subramanian, M., 2018. Hydroxychavicol, a key ingredient of Piper betle induces bacterial cell death by DNA damage and inhibition of cell division. Free Radic. Biol. Med. 120, 62-71.

International Organization for Standardization, 2007. Microbiology of food and animal feeding stuffs-horizontal method for the detection of Salmonella spp. amendment 1: Anex D: Detection of Salmonella spp. in animal faeces and in environmental samples from the primary production stage. ISO, Geneva.

Tadee, P., Patchanee, P., Pascoe, B., Sheppard, S.K., Meunsene, D., Buawiratlert, T., Tadee, P., 2021. Occurrence and sequence type of antimicrobial resistant Salmonella spp. circulating in antibiotic-free organic pig farms of northern-Thailand. Thai J. Vet. Med. 51(2), 311-319.

Taukoorah, U., Lall, N., Mahomoodally, F., 2016. Piper betle L. (betel quid) shows bacteriostatic, additive, and synergistic antimicrobial action when combined with conventional antibiotics. S. Afr. J. Bot. 105, 133-140.

Valle, D.L., Andrade, J.I., Puzon, J.J.M., Cabrera, E.C., Rivera, W.L., 2015. Antibacterial activities of ethanol extracts of Philippine medicinal plants against multidrug-resistant bacteria. Asian Pac. J. Trop. Biomed. 5(7), 532-540.

Valle, D.L., Jr., Cabrera, E.C., Puzon, J.J., Rivera, W.L., 2016. Antimicrobial activities of methanol, ethanol and supercritical co2 extracts of Philippine Piper betle L. on clinical isolates of gram positive and gram negative bacteria with transferable multiple drug resistance. PLoS One. 11(1), e0146349.

Vidayanti, I.N., Sukon, P., Khaengair, S., Pulsrikarn, C., Angkittitrakul, S., 2021. Prevalence and antimicrobial resistance of Salmonella spp. isolated from chicken meat in upper northeastern Thailand. Vet. Integr. Sci. 19(2), 121-131.

Wu, L.J., Luo, Y., Shi, G.L., Li, Z.Y., 2021. Prevalence, clinical characteristics and changes of antibiotic resistance in children with nontyphoidal Salmonella infections from 2009–2018 in Chongqing, China. Infect. Drug Resist. 14, 1403-1413.