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Assessment of Probiotic Application of Lactic Acid Bacteria (lab) Isolated from Different Food Items

Mahbubul Hasan Siddiqee
Microbiology Program, Department of Mathematics and Natural Sciences, BRAC University, Dhaka-1212, Bangladesh

Khandaker Mahzebin Shurovi
Department of Microbiology, Stamford University Bangladesh, Dhaka-1217, Bangladesh

Hashem Sarker
Department of Microbiology, Stamford University Bangladesh, Dhaka-1217, Bangladesh

Abstract/ Executive Summary:

Lactic acid bacteria (LAB) are regarded as effective probiotic organisms and used with a view of augmenting the safety of the food. In the present study, five food items (meat, fish, apple, milk and carrot) were selected having high nutritive and economic value, assumptive of harboring lactic acid bacteria. A total of 29 LABs were isolated from 5 different samples after 2 batches of fermentation. All of the isolates were then tested against 4 most frequently encountered pathogens, Staphylococcus aureus, Escherichia coli, Salmonella typhimurium, and Vibrio cholerae. Among all these isolates, those from meat and fish samples showed positive average inhibitory coefficient (AIC) against two target pathogens, while those from milk and carrot showed positive AIC against three pathogens each, and isolates from apple revealed positive AIC against all four pathogens used. More than 50% of the isolates were found to inhibit or mask the pathogens when allowed to grow along with the individual pathogen on the each tested food item. Out of 29 isolates, 17 were found to successfully inhibit Escherichia coli, 11 worked against S. aureus, 11 against S. typhimurium, and 13 showed significant effect against V. cholerae. Among these isolates ML4, ML8, AP5 and CR3 most notably showed the potential to inhibit or mask at least three of the target pathogenic strains.

Keywords: Lactic acid bacteria; Probiotics; Competitive inhibition

Location: Microbiology Program, Department of Mathematics and Natural Sciences, BRAC University, Dhaka

Start Date:

End Date:

Program Area: Food Safety and Security

Commodity: Fortified food

Objectives:

1. The present study was undertaken to isolate lactic acid bacteria (LAB) from some foods known to support LAB growth with a view of using the isolates in the long run to ferment those foods, and determination of their interactions with some food-borne pathogenic bacteria frequently encountered in Bangladesh.

Methodology:

Samples chosen for this study were selected on the basis of higher possibility of harboring naturally occurring LAB (30) as well as having higher promise in terms of economic and nutritional value. Samples included meat (beef), cowmilk, fish (Pangasius bocourti), apple and carrot. Sample processing. Two batches of samples were collected from Karwan Bazar and Moghbazar markets in Dhaka city. About 100g of sample for each item was collected aseptically using sterile container. Approximately 20g of each sample (except milk) were mixed with 80 ml of sterile water and then homogenized with stomacher for 5-10 minutes and then 10 fold dilution was performed once. pH was recorded for each homogenized sample before adjusting it to 5.5 to give selective advantage to the LAB. Isolation of LABs. Processed samples were kept at room temperature inside paper boxes for 3 days for natural fermentation to occur. Then, dilution up to 10-6 was performed for each fermented sample. 0.1ml from each sample was spread on Rogosa SL Agar and incubated at 37°C for 48 hours under limited oxygen concentration by sealing the agar plates with parafilm. Pathogenic microrganisms. Four clinical isolates including Staphylococcus aureus, E. coli, S. typhimurium, and V. cholerae were collected from Microbiology Laboratory of Stamford University Bangladesh. These pathogens were selected due to their involvement in food-borne gastrointestinal illnesses. Competitive inhibition assay (CIA). Four sets of test tubes (each for a target pathogen) were filled with 5 ml of sterile homogenized food sample. In each set, one test tube was assigned for each bacterium isolated from the test samples. In addition, 5 test tubes were taken as negative control. Medium used for each of the isolates was the original food type from which the LABs were isolated, i.e. the isolates from meat samples were tested for interaction with the target pathogens in autoclaved homogenized meat, milk-isolates were tested against target pathogens in milk, and so on for the others. All the test tubes (except the negative controls) were inoculated with 0.1 ml of each of the test organisms suspended in normal saline and kept for fermentation at 37⁰C. After 48 hours, 4 sets were inoculated with 4 target organisms respectively and were further incubated for 24 hours. 100-fold dilution was performed for each of the 4 sets of samples. Mannitol salt agar, MacConkey agar, Salmonella-Shigella agar, and thiosulfate-citrate-bile salt-sucrose agar were used for enumeration of S. aureus, E. coli, S. typhimurium, and V. cholerae, consecutively. Average Inhibition Coefficient (AIC) for each group of LAB isolated from a particular type of food against each of the four target pathogens was determined by using the following formula- Average Inhibition Coefficient (AIC) = [{Total count in negative control – Average count for each group of isolates} / Total count in negative control].

Source of Information: Stamford Journal of Microbiology, December 2012. Vol. 2, Issue 1; ISSN: 2074-5346

Article Link (Online Journal):

Funding Source:

Total Budget (Tk.) :

Achievement/Findings:

Average inhibition coefficient (AIC) was calculated for each food type against each of the target strains. It can be used as a gross parameter of the candidate LABs from each sample type. Isolates from apple origin showed most uniform index of AIC ranging from 0.10 to 0.33 which were found to be positive against all four target strains. Isolates from meat and fish could effectively suppress two of the target strains each; particularly, both groups were highly efficient against E. coli with AIC of 0.95 in fermented meat and 0.96 in fermented fish. Milk and carrot isolates revealed fairly positive AIC indicating the positive result.

Knowledge Management: Journal