Nikita and Hemangi (2012) reported Lactobacillus as catalase negative with gas production from glucose and 45OC as ideal temperature for growth except L. brevis that grew at 15o C. Lei et al. (2009) reported that Lactobacillus fermentum and Saccharomyces cerevisiae improved the intestinal balance of the diverse microflora species in the rectum of broiler chickens. Stern et al. (2006) reported Lactobacillus salivarius (NRRL B-30514) as a gram-positive, facultative-anaerobe, catalase-negative, nonmotile, pleomorphic rod.
The strain produced lactic acid and H2O2.
Pyar and Peh (2014) observed Lactobacillus acidophilus as non motile in hanging drop wet method, catalase negative with no bubble observed due to decomposition of H2O to produce O2 and it is known for fermenting maltose, sucrose, glucose and grow at pH ranging from 4.0 – 7.0, while viable count in bile salt concentration decrease with an increase in bile salt concentration. Qing et al. (2014) reported that Lactobacillus plantarum had great tolerance to stimulate gastrointestinal juices and had strong antimicrobial activity against four pathogenic bacteria: Escherichia coli, Staphylococcus aureus, Salmonella sp.
and Shigella sp and good adhesion sites.
Bifidobacterium are gram positive, non-spore forming, non-motile, rod shape saccharolytic anaerobic bacteria that produce lactic acid and acetic acid from carbohydrate with generation of CO2 as end products of carbohydrate fermentations (Scardovi, 1986; Felis and Dellaglio, 2007). Bifidobacteria are catalase negative with the exception of B. indicum and B. asteroids from hindgut of honey bee (Felix and Dellaglio, 2007). Bifidobacteria are saccharolytic organisms by generating acetic and lactic acids.
CO2 is not produced by bifidobacteria except with the degradation of gluconate (Biavati and Mattarelli, 2012).
Demonstration of fructose-6-phosphate phosphoketolase (F6PPK) activity in cellular extracts has been a useful means for distinguishing bifidobacteria from morphologically similar bacteria like Lactobacillus, Actinomyces, Propionibacterium and Eubacterium. Another reliable means of distinguishing bifidobacteria from other bacterial types is gas liquid chromatography of fermentation products (Scardovi, 1986). Kok (1996) reported that PCR primers supersede biochemical test for the identification of bifidobacteria to species level.
Jin et al. (1997) reported growth at 45 0C for L. acidophilus isolated from GIT of chickens is most positive while, L. fermentum, L. brevis, L. crispantus, L. salivarus and L. plantarum are positive and all were catalase negative. Mesgari and Hosseinzadeh (2016) reported 450C with no gas production and catalase negative for L.fermentum, L. reuteri and L. lactis and Khedid et al. (2006) reported 150C and a pH of 6.5 with no motility and gas production while Kram et al. (2012) reported growth at 150C for L. plantarum and 450C for L. ferementum.
Scardovi (1984) reported that fresh bifidobacterium possess V and Y shaped and survived strictly in an anaerobic environment with a temperature of 37 – 410C with no growth below 20 0C and above 46 0C and optimal pH of 6.5 – 7.3 with no growth below 4 and above 8. Colonies of bifidobacterium appeared smooth, convex and cream to white in colour. Gavini et al. (1991) reported 41- 43oC as ideal temperature for growth of Bifidobacteria in animals and the temperature of 46oC can be used to differentiate between animals and humans strains and no growth when the temperature is less than 20oC.
Biavati and Mattarelli (2015) reported ideal growth temperature of 37–41°C for bifidobacteria, except for Bifidobacterium mongoliense with 30°C for growth and a minimum growth temperature of 25–28°C but Bifidobacterium mongoliense and Bifidobacterium psychraerophilum grows at 15°C and 8°C, and Bifidobacterium thermacidophilum grow at a temperature of 49.5°C. Watabe et al. (1983) reported 370C as growth temperature for Bifidobacterium gallinarium which is also gram positive, non spore forming, non-motile in nature with a short rod shaped and slightly curved. Dong et al. (2000) stated that human and animals strains of bifidobacteria grow at an optimum temperature range of 36–38 oC and 41–43oC respectively.
Bile tolerance plays the role of reduction of bacterial pH in the presence of different percentages of bile salts. Resistance to pH and bile salts is of immense significance in survival and growth of bacteria in the intestinal tract and this serve as a requirement for probiotic properties (Havenaar et al., 1992).
Therefore, bile tolerance is as an important characteristic of the LAB which permits it to grow, survive and to put forth its action in gastrointestinal transit. Species that could grow well and metabolize in normal physical bile concentration could survive in gastrointestinal transit as reported by Sanders et al. (1996). Raja et al. (2009) reported that Lactobacillus Fermentum strain from chicken gut showed tolerance to bile salts at 0.3% and 10% and Maldonado-Valderrama (2011) reported that bile salts play critical functions in the digestion and absorption of nutrients in the GIT of chickens. They are also responsible in precise and non-precise defense mechanism of the gut since bile salts concentration decide the extent of inhibitory effect (Marteau et al., 1997).
The liver produced and stored bile salts in the gall bladder and secreted into the upper small intestine with major excretory pathway for cholesterol which is important in activating pancreatic lipase and to facilitate absorption of the fat soluble vitamins from the digestive tract (McDonald et al., 2010). Bile acids have superior inhibitory effect on lactobacilli than bifidobacteria strains (Yazid et al., 1999).
Boylston et al. (2004) reported that L. acidophilus is more resistant compared to Bifidobacterium spp. in terms of high acidity, while Liong and Shah (2005) reported pH 3 as standards for acid tolerance of probiotic culture. Hassanzadazar et al. (2012) reported significant decrease in viability of lactobacilli after incubation at pH 3.0. The ability of probiotic microorganisms to survive passage through the stomach is an importance property of acid tolerance (Prasad et al., 1998; Park et al., 2002).
Survival of bacteria in the gastric juice is hinge on their capacity to bear low pH. The pH excreted HCl in the stomach is 0.9, but ingestion of food raises the pH value to 3.0 (Erkkila and Petaja, 2000). Prasad et al. (1998) in a similar experiment reported that three strains of Lactobacillus and one strain of Bifidobacterium grow in presence of bile salts, after pre-exposure to low pH (3.0).
Low pH environment impedes the growth, metabolisms processes and reduced the viability of L. acidophilus and viability count of bacterial decline by pH of 1.5 after a period of 3 hours incubation (Madal et al., 2006). Fermandez et al. (2003) and Liong and Shah (2005) reported pH of 3.0 as standard for acids tolerance good probiotic culture. It has been reported in literatures that L. acidophilus is more resistance than Bifidobacterium (Boylston et al., 2004). They lower the pH of the gut by converting sugar to lactic acid inhibits the growth of enteropathogens (Strompfova et al., 2005).
The best pH for growth of Bifidobacteria is 6.5 – 7.0; except for Bifidobacterium thermacidophilum, which can grow at a pH 4.5 or pH 8.0 – 8.5 (Biavati and Mattarelli, 2012; Biavati and Mattarelli, 2015). Bifidobacteria are acid-tolerant microbes and their optimum pH for growth is between 6.5 and 7.0 and they donot survine above pH of 8.5 (Biavati et al. 2000)