Probiotic and technological properties of lactic acid bacteria from bovine origin

Abstract

Lactic acid bacteria are generally regarded as safe microorganisms, and they have been used as food additives and therapeutic supplements in functional foods. In 2013 the world market for probiotics was USD 36 million and increased to 35.9 billion in 2018. The probiotic preference is because of extreme interest in probiotic yoghurt and development in the utilisation of functional foods. This has led to new bacterial strains being distinguished and added to pharmaceutical and nutrition products. Most of the probiotics in use are obtained from the gastrointestinal tract of healthy human individuals. However, there are other probiotics in use which are of non- human origin. The probiotics which are intended to be used in humans require proof that they work in human hosts. Identification of probiotics at the strain level, evaluation of biosafety by antibiotic susceptibility testing and absence of virulence factors as well as an assessment of probiotic characteristics must be carried out before use in humans.

Therefore, the current study focused on characterising probiotic lactic acid bacteria from bovine and dairy sources for application in yoghurt. Lactobacillus acidophilus (strain D) and Lactobacillus rhamnosus (strain V) were from the dairy origin, Lactobacillus plantarum (strain VLL1) and Lactobacillus pentosus (strain LIP), were from the bovine origin and Lactobacillus acidophilus ATCC 4536 from American Type Culture Collection. The first phase of the study aimed at identifying and characterising the lactic acid bacteria. The identification was done using phenotypic methods which included Gram staining, catalase test, oxidase test, and carbon dioxide gas production from glucose. The isolates were gram-positive, catalase-negative, and oxidase negative, which proved them to be lactic acid bacteria. They were also able to produce carbon dioxide gas from glucose. The production of carbon dioxide gas classified the bacteria as heterofermentative, and ethanol, and lactic acid were produced as by-products of glucose metabolism.

The second phase included safety assessment and determination of probiotic properties. The safety assessment included haemolytic activity, lipase production, gelatinase production, and antibiotic resistance. The probiotic properties which were determined included acid and bile resistance, bile salt hydrolase activity and antimicrobial activity. The antimicrobial activity was against enteroinvasive Escherichia coli (EIEC), enterohaemorrhagic E. coli (EHEC), enteroaggregative E. coli (EAEC), enterotoxigenic E. coli (ETEC), Candida albicans ATCC 10231 (C1), C. albicans 1051255 (C4), and C. albicans M0826 (C7). On exposure to the acidic environment, the lactic acid bacteria significantly reduced their viability at pH 2. However, they were able to survive pH 5 and pH 3. The isolates were able to survive a 0.3% bile salt concentration and released the bile salt hydrolase enzymes. The lactic acid bacteria were able to inhibit diarrheagenic E.coli and pathogenic Candida spp. On antibiotic susceptibility testing, the strains were resistant to vancomycin and kanamycin and susceptible to gentamicin, clindamycin, erythromycin, tetracycline, ampicillin, and chloramphenicol. The isolates from bovine origin were moderately sensitive to tetracycline. The isolates were gelatinase and lipase negative, but they showed α- hemolysis which is the partial hydrolysis of red blood cells.

The third phase was to determine the fate of the bovine and dairy probiotic strains when used with regular fermenting bacteria during fermentation as well as storage. L. plantarum, L. rhamnosus, and L. acidophilus were selected for yoghurt processing. The yoghurt was stored for 28 days under refrigeration at 4 °C. The probiotic cells were selectively enumerated using MRS- bile agar. The cell population levels of the probiotics were positively affected in relation to the control up to day 28 of storage at 4 °C. Overall, this study exemplifies the probiotic potential of bovine and dairy isolates, where the isolates from bovine origin have slightly increased tolerance to human gastrointestinal conditions such as acid and bile tolerance, pathogenic inhibition, α-haemolytic and negative to lipase and gelatinase enzyme activities.