Abstract

Probiotics play and important role in maintaining a healthy gut and consequently promote good health. With a considerably large number people consuming fermented foods (containing a wide variety of lactic acid bacteria), very little research has been carried out to assess their probiotic properties.

The experiment included the isolation of Lactic Acid Bacteria (LAB) from three (3) randomly selected fermented samples of Palm wine, Yoghurt and Garri, cultured on MRS Agar , sub-cultured on nutrient Agar and assessed for potential probiotic properties such as acid and bile tolerance, antimicrobial susceptibility of the isolated strains of the pure colonies.

Tolerance to low pH values (2.0 to 3.0) and high bile concentrations (0.2 to 2.0) varied through-out the six (6) treatments where antibiotics; vancomysin, ciprofloxacin, doxycycline, chloramphenicol, azithromycin, erythromycin, streptomycin, penicillin where administered to each sample treatment. Kirby-Bauer’s antimicrobial susceptibility test was carried out on the six isolates and results showed that, the isolates were 90% sensitive and 5% resistant to all antibiotics.

CHAPTER ONE

INTRODUCTION AND LITERATURE REVIEW

1.1 INTRODUCTION:

The first discovered probiotic was a certain strain of bacillus in Bulgaricus yoghurt called lactobacillus bulgaricus. The discovery was made in 1905 by Bulgarian physician and microbiologist stamen Grigorov. The modern day theory is generally attributed to Russian Nobel Laureate Elie. A growing probiotics market has led to the need for stricter requirements for scientific substantiation of putative benefits conferred by microorganisms claimed to be probiotic.

Probiotics are defined as “living micro-organisms, which upon ingestion in certain numbers, exert health benefits beyond inherent basic nutrition” (Guarner and Schaafsma,1998; Tannock, 2002) but interest in this area was initiated by Metschnikov 100 years ago (Metschnikoff,1907).Most probiotic  microorganisms belong to Lactic Acid Bacteria (LAB), such as Lactobacillus sp, Bifidobacterium sp and Enterococcus sp (Klein et al., 1998).

In recent years, multiple reports have described beneficial effects from various aspects on important diseases, like intestinal infections, inflammatory bowel disease (IBD), and allergy by addition of selected strains to food products, often together with fibre or a prebiotic substance. In many countries, there are now several probiotic products on the market but the documentation is often based upon case reports, animal studies or uncontrolled small clinical trials.

Probiotics are live microorganisms promoted with claims that they provide health benefits when consumed, generally by improving or restoring the gut flora. Probiotics are considered generally safe to consume, but may cause bacteria host interactions and unwanted side effects in rare cases

Lactobacillales are an order of gram positive, low gc, acid tolerant, generally non sporulating, non-respiring either rod shaped (bacilli) or spherical(cocci) bacteria that share common metabolic and physiological characteristics. These 3 bacteria usually found in decomposing plants and milk products, produce lactic acid as the major metabolic end product of carbohydrate fermentation giving them the common name lactic acid bacteria (LAB)

Production of lactic acid has linked lactic acid bacteria with food fermentation, as acidification inhibits the growth of spoilage agents. Proteinaceous bacteriocins are produced by several lactic acid bacteria strains and provide an additional hurdle for spoilage and pathogenic microorganisms. Furthermore, lactic acid and other metabolic products contribute to the organoleptic and textural profile of a food item.

The industrial importance of lactic acid bacteria is further evidenced by their Generally Recognized as Safe (GRAS) status, due to their ubiquitous appearance in food and their contribution to the healthy microbiota of human and animal mucosal surfaces. The genera that comprises the lactic acid bacteria are at its core: lactobacillus, leuconostoc, pediococcus, lactococccus and streptococcus.

Probiotic properties of lactic acid bacteria functional foods remain the hottest topic in the food industry and nutrition especially food products containing probiotic microorganism. Lactic acid bacteria stains of genera lactobacillus have been used for a long time for the production of wide range of fermented diary and cereal based products. Probiotics are a desirable natural tool to maintain the healthy balance of the human intestinal microflora. They are consumed either as food components or as non-food preparations.

New species and more specific stains of probiotic bacteria are constantly identified. However, prior to incorporating strains into products, their efficacy should be carefully assessed. To prove scientifically the probiotic properties of these stains they have to fulfil series of selective criteria.

First the probiotic strains have to be safe for human consumption. Research has focused on natural isolates with a long history of safe use (e.g they originate either from traditionally consumed foods with proven health effects or from the gastrointestinal system of healthy individuals).

Acid and bile stability are self-evident properties for every strain expected to have effect in the intestinal tract. The stain should as well have characteristic antimicrobial activity and antibiotic resistance.

Acid conditions in the stomach are a natural barrier preventing most microorganisms from passing into the intestines. Normally pH in the stomach of healthy individuals is lower than 3.0 the ability to survive and grow in low pH environment is characteristic for LAB. Production of organic acids by LAB decrease pH of the environment, which become3s selective against the micro-organisms sensitive to low pH.

Bile tolerance of microorganisms has been used as a selective criterion for potential probiotics. The stress effect of bile on Lactobacillus strains is complex because of the variation of bile concentration and passage time within the different parts of the gastrointestinal tract. Bile resistance of some strains is related to specific enzyme activity, bile salt hydrolase (BSH) which helps hydrolyze conjugated bile thus reducing its toxic effect. In addition to Ganzle et al bile resistance can be increased due to the protective effect of some food components.

Another important property of probiotic strains is their antagonistic activity against pathogenic bacteria either by production of antimicrobial substances include organic, either inhibitory primary metabolites such as hydrogen peroxide and special compounds like bacteriocins and antibiotics. This property plays significant role in the ability of probiotics to compete against the resident flora and beneficially modify it. Numerous clinical studies demonstrate that organisms of lactobacillus genus are both preventative and therapeutic when administered with milk containing these organisms (GOTCHEVA et al., )

1.2 LITERATURE REVIEW

1.2.1 Microbiology of lactic acid bacteria

Lactic acid bacteria (LAB) constitute a group of gram positive bacteria united by a constellation of morphological, metabolic and physiological characteristics.

The general description of the bacteria included in the group is gram positive, nonspore forming, non-respirin, cocci, coccobacilli or rod, catalase negative which produce lactic acid as a major end product during the fermentation of carbohydrates. The LAB term is intimately associated with bacteria involved in the food and feed fermentation.

Although many genera of bacteria produce lactic acid as a primary or secondary end product of fermentation, the term Lactic Acid Bacteria is conventionally reserved for genera in the order Lactobacillales, which includes Lactobacillus, Leuconostoc, Pediococcus, Lactococcus and Streptococcus, in addition to Carnobacterium, Enterococcus, Tetragenococcus, Vagococcus and Weisella.

1.3 RATIONAL

Food products containing probiotic organisms remain the hottest topic in the food industry and nutrition as probiotics play an important role in the production of fermented food. Lactic acid bacteria (LAB) strains have been used for a long time for production of wide range fermented diary and cereal based products.

The growing probiotic market and the putative benefits conferred by Lactic acid bacteria (LAB) has led to the need for stricter study of the probiotic properties of Lactic acid bacteria (LAB).

1.4 PROBLEM STATEMENT

There is variety of lactic acid bacteria but little or no study of it has been done on the topic.

1.5 OBJECTIVES

The main objective of   the study is to identify, isolate and characterize Lactic acid bacteria as a probiotic in fermented food in Buea Municipality.

This objective will be achieved through the following specific objectives;

1. To isolate lactic acid bacteria from diluted food samples in a laboratory
2. To identify and characterize lactic acid bacteria by gram and catalase
3. To screen antimicrobial activity (in vitro)

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