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ARS Home » Northeast Area » Beltsville, Maryland (BARC) » Beltsville Agricultural Research Center » Environmental Microbial & Food Safety Laboratory » Research » Publications at this Location » Publication #316966
Title: Natural antimicrobials to control biofilms formed by environmental isolates of Salmonella

Author
item KEELARA, SHIVARAMU - University Of Maryland
item Patel, Jitu

Submitted to: BARC Poster Day
Publication Type: Abstract Only
Publication Acceptance Date: 5/6/2015
Publication Date: 5/20/2015
Citation: Keelara, S., Patel, J.R. 2015. Natural antimicrobials to control biofilms formed by environmental isolates of Salmonella. BARC Poster Day. p.29.

Interpretive Summary: Fresh produce accounts for 9.5 million (12%) of the ~76 million U.S. foodborne illnesses annually. Salmonella is the leading causative agent of an estimated 35% hospitalizations and 28% deaths. Fresh produce can be contaminated by Salmonella at farm level via contaminated manure, irrigation water, wild or domestic animals and interface of farm animal waste with agricultural land. Biofilm formation by Salmonella on fresh produce and equipment surfaces is a concern because sessile bacteria within biofilms are not easily killed by treatment with antimicrobials. The objectives of this study were to determine the biofilm forming ability of swine environmental Salmonella isolates and mitigate the biofilm forming Salmonella by natural antimicrobials. Natural antimicrobials: cinnamaldehyde and sporan were evaluated for their bactericidal efficacy in killing Salmonella in biofilm using an innovative Minimum Biofilm Eradication Concentration (MBEC) assay. All Salmonella isolates formed biofilms, although the extent varied with strain. Natural antimicrobials- cinnamaldehyde and sporan at reduced Salmonella in biofilm by 6 logs at 2000ppm and below detection limits at 3000ppm. In conclusion, natural antimicrobials such as cinnamaldehyde and sporan at 3000 ppm conc. can be used to kill Salmonella in biofilms. To our knowledge, this is the first report on bactericidal efficacy of natural antimicrobials in removing biofilms as evaluated by MBEC assay.

Technical Abstract: Fresh produce account for 9.5 million (12%) of the ~76 million U.S. foodborne illnesses annually. Salmonella is the leading causative agent of an estimated 35% hospitalizations and 28% deaths. Fresh produce can be contaminated by Salmonella at farm level via contaminated manure, irrigation water, wild or domestic animals and interface of farm animal waste with agricultural land. Biofilm formation by Salmonella on fresh produce and equipment surfaces is a concern because sessile bacteria within biofilms are not easily killed by treatment with antimicrobials. The objectives of this study were to determine the biofilm forming ability of swine environmental Salmonella isolates and mitigate the biofilm forming Salmonella by natural antimicrobials. We evaluated 15 Salmonella isolates; S. Derby (2), S. Infantis (4), S. Typhimurium (9) of conventional swine farm environment (soil and lagoon) origin. Biofilm forming ability was analyzed by BATH assay, 96-well microtiter plate CV assay, Congo red dye binding curli expression assay and PCR. Natural antimicrobials: cinnamaldehyde and sporan were evaluated for their bactericidal efficacy in killing Salmonella in biofilm using innovative Minimum Biofilm Eradication Concentration (MBEC) assay. All Salmonella isolates formed biofilm; however, biofilm formation varied with strain. Curli expression was robust among strains S322 and S435 (S. Infantis), S644, S777, S931, S953 and S977 (S. Typhimurium) as observed by Congo red dye binding assay. Hydrophobicity (%) varied with strains and growth phase of the strain; the difference was not significant. In general, Salmonella strains formed significantly higher biofilms (P <0.05) in full strength growth media compared to biofilm in diluted media. Natural antimicrobials- cinnamaldehyde and sporan at 2000 ppm reduced (P< 0.05) Salmonella in biofilm by 6 log cfu from their initial populations of 7-7.5 log cfu/cm2. The bactericidal effect of these antimicrobials increased with concentrations; Salmonella were undetectable (detection limit 0.73 log cfu/cm2) when 3000 ppm of cinnamaldehyde or sporan were used. PCR was unable to detect bapA gene associated with biofilm formation in Salmonella. In conclusion, natural antimicrobials such as cinnamaldehyde and sporan at 3000 ppm conc. can be used to kill Salmonella in biofilms. To our knowledge, this is the first report on bactericidal efficacy of natural antimicrobials in removing biofilms as evaluated by MBEC assay.