The Role of Probiotic Bacteria on Microbiological and Acceptability of Sudanese White Soft Cheese

: The objectives of this investigation is to study the effect of three types of probiotic bacteria ( Lactobacilus rhamnosus , Lactobacillus casei , Bifidobacterium bifidum ) on the microbiological and acceptability of Sudanese white soft cheese during storage period 0, 15, 30, 45 and 60 days. Microbiological analysis revealed that, the highest count of probiotic bacteria (87x10 8 cfu/g) was obtained by sample containing L.rhamnosus and the lowest (39x10 6 cfu/g) by sample containing B.bifidum , while the sample containing L.casei ranked in intermediate position. Storage period affected the total probiotic bacteria count, the highest count at 2 weeks for L.casei and B.bfidum and after 4 weeks for L.rhamnosus , while the lowest at the end. The microbial analysis did not detect any pathogenic bacteria ( coliform bacteria , salmonella and staphylococcus aureus ) or yeast and molds. The sensory evaluation quality revealed that the cheese containing L.rhamnosus gave the best appearance, texture, flavour and overall acceptability, followed by L.casei and B.bifidum compared with the control samples. The storage period significantly (p≤0.05) affected the acceptability of the cheese, where the highest score was obtained at day 30 and the lowest at the beginning of the storage. The study recommends further studies and tests to improve the quality of the Sudanese white soft cheese treated by probiotic bacteria.


INTRODUCTION
Probiotic as a term is a relatively new word meaning for life and it is currently used to describe a group of bacteria when administered insufficient quantity, confer beneficial effects for humans and animal (Kopp-Hoolihan, 2001). Probiotic bacteria are applied to balance disturbed intestinal microflora and important in the treatment of a wide range of human disorders including lactose intolerance, diarrhea, food allergies, intestinal infection, constipation gastroenteritis, hepatic, flatulence, colitis, gastric acidity, osteoporosis, high blood cholesterol and cancer (Pitino et al., 2012).
The most organisms used as probiotic belong to bifidobacteria, Lactobacillus and some Enterococcus ssp. (Fooks et al., 1999). Lactic acid bacteria (LAB) and bifidobacteria are amongst the most important groups of microorganisms used in the food industry, used in the production of fermented products, such as yoghurts, cheese and pickled vegetables (Venema et al., 2015).
Bifidobacteria spp. beneficially affects human health by improving the balance of intestinal microflora and improving mucosal defenses against pathogens. Additional health benefits include enhanced immune response, reduction of serum cholesterol, vitamin synthesis, anti-carcinogenic activity, and anti-bacterial activity (Blanchette et al., 1996;Robinson et al., 1992). Lacticobacillus casei improvement of balance of intestinal microbiota and volatile fatty acid, antitumor action, stimulation of the immune system, and antimicrobial activity (Fujimoto et al., 2008).
Lactobacillus rhamnosus benefits are reducing the activity of fecal enzymes such as β-glucuronidase and azoreductase which contribute to the risk of colon, mammary, and prostate cancer (Tanock et al., 2000). Lactobacillus rhamnosus increases the number and the activity of natural killer cells and exerts immunostimulating effects, including on fetuses (Ibrahim et al., 2010).
Cheese is known as a complete nutritious food product and excellent source of many key nutrients, suitable for many ages. It is rich in protein and minerals such as calcium.
There are different types of cheese: soft cheese, semi hard and hard cheese, the difference in these types is mainly due to water content or water activity and the methods and technology for cheese making Total count of probiotic bacteria  Table 1. Shows the total count of probiotic bacteria of Sudanese white soft cheese during storage period. The highest (87x10 8 cfu/g) was obtained by the cheese samples containing Lactobacillus rhamnosus. The lowest (39x10 6 cfu/g) by the Bifidobacterium bifidum samples, while the sample containing L.casei ranked in an intermediate position (p≤0.05). In our study the number of probiotic bacteria L.rhamnosus increased from the beginning today 30 (87x10 8 cfu/g) then decreased to (47x10 7 cfu/g) at the end of storage period. L.casei, B.bifidium numbers increased until day 15 (67x10 8 and 52x10 8 cfu/g) then decreased to (4x10 7 and 39x10 6 cfu/g) respectively at the end of storage period. The viability of probiotic bacteria decreased during storage period. Due to the decrease in the pH of the medium and accumulation of organic acids as a result of growth and fermentation (Randazzo et al., 2013). Kebary et al., (2011) observed that L.

RESULTS AND DISCUSSION
rhamnousus counts in all Ras cheese increased during the first month of ripening period then decreased up to the end of storage period. Ozer et al., (2009) conducted that probiotic Beyaz cheese with B. bifidum decreased during ripening periods for 90 days at significant level. Fritzen-Freire (2010) reported that counts of B. Bifidum in minas frescal cheese did not decrease during ripening periods for 28 days.
Bergamini et al., (2010) stated that the viability of different probiotic cultures was assessed during Pategra´s cheese ripening. All the probiotics tested maintained counts above 107 cfu/g during the shelf-life settled for the product. Kebary et al., (2015) observed that the count of L. rhamnousus in Domiati cheese increased in the first 4 weeks reached 10 11 cfu/g, then decreased gradually during storage period to reached 10 8 cfu/g. The probiotic dairy product, contain at least 10 6 -10 7 cfu/g of viable probiotic bacteria at the time of consumption (FAO/WHO, 2002). IJMRA, Volume 4 Issue 5 May 2021 www.ijmra.in Page 498 Shah et al., (1995) mentioned that initial count of B.bifidum was 10 6 -10 7 cfu/ml at the time of consumption. Cheeses presented populations as high as 1x10 6 cfu/g during storage. Sua´rez-Solı´s et al., (2002) pointed that in manufactured fresh cheese supplemented with B. bifidum and L. casei the viable counts of B.bfidum and L.casei reached 1x10 7 cfu/g during storage. Yilmaztekin et al., (2004) observed survival of Bifidobacterium bifidum in white brined cheese, B.bifidum inoculated at levels of 2.5% and 5.0% were 7.0x10 8 cfu/g and 1.2x10 9 cfu/g respectively, these figures dropped to 4.0x10 6 cfu/g and 1.1x10 7 cfu/ g after 90 days. Mahmoud et al., (2013) noticed that viability of B. bifidum and L.rhamnosus in Karish Cheese effected of probiotic Bifidobacterium bifidum and Lactobacillus rhamnosus decreased markedly from 38x10 7 to 60x10 2 and from 40x10 7 to 51x10 2 cfu/g respectively at the end of storage period. The growth of B. bifidum and L.
rhamnosus during storage of Kareish cheese increased with a peak at 7 days of storage then decreased. Dave and Shah (1997) conducted that, the viability of bacteria from commercial starter culture during storage of dairy product, contained bifidobacteria observed on the level 10 6 -10 9 cfu/ml depending on the kind of starter culture and the medium used for enumeration. Decreased of number of these bacteria was observed but it was above the recommended limit 10 6 through the storage period. Zomorodi et al., (2011) stated that in Iranian white cheese supplemented with free and microencapsulation probiotics (L.casei, B.bifidum) the number of B.bifidum colonies did not decrease remarkably in the free and capsulated form.
Ozdemir and Yangilar (2013) enumerated that in Turkish Beyaz cheese samples produced with different probiotic cultures B. bifidum decreased at 1 log levels from the beginning to the end of ripening times. Aljewicz et al., (2014) stated that in Edam Cheese supplemented with Probiotic Lactobacillus rhamnosus viability of probiotics (approx. 1 log cfu/g) reduction of probiotic bacteria population was observed in the analyzed experimental cheese samples between the 2nd and the 4th week of ripening. The viability of probiotic bacteria remained stable in all experimental cheese samples from the 4th week. Gobbetti et al.,(1998) said that addition of bifidobacteria to the Crescenza cheese affected when added individually, B. bifidum was present at the same concentration in the milk (log10 6.0 cfu/ml) but reached final cell numbers of log10. Bifidobacterium bifidum adapted well to the cheese environment cell numbers decreased throughout ripening. Abadía-García et al., (2013) observed that incorporated L.casei, L. rhamnosus and a commercial mix of probiotics in cottage cheese. L. casei count was 10 8 cfu g−1, while the other probiotic populations remained at levels of about 10 6 cfu g−1 during 28 days of storage at 8 °C.
Bezerra et al., (2016) observed the lactic acid bacteria counts in the cheeses were higher than 6.5 log cfu/g cheese and 7 log cfu/g cheese at the 1st and 28th days of storage, respectively. The addition of selected bacterial in simple and cheeses can improve bacterial viability during storage (Buriti et al., 2005 a,b ). Probiotic bacteria maintained their viability throughout the storage period with a negligible loss or less than 1 log cycle loss, or even with increase in the viable counts (Karimi et al., 2011).

Coliforms, S.aureus, Salmonella, yeast and molds count
The Coliform, S.aureus, Salmonella, yeast and molds were not detected in all Sudanese soft white cheese samples in control samples or samples containing probiotic bacteria (L.rhamnosus, L.casei and B.bifidum) during ripening period. This may be due to high hygienic condition during making cheese and pickling period and the development of acidity in cheese during the ripening period (Kebary et al., 2015). Intestinal lactobacilli and bifidobacteria produce antimicrobial substances that are active against pathogenic bacteria (Servin et al., 2004).
rhamnosus used for manufacturing soft cheese resulted in reduction in counts of pathogens, as well as molds and yeasts. Probiotic L.rhamnosus inhibited the growth of pathogenic Salmonella enterica by producing lactic acid and other secreted antimicrobial molecules (Marianelli et al., 2010). El-Shafei et al., (2008) found that L. rhamnosus in Kareish cheese were able to totally inhibit the growth of tested yeasts during storage for 30 days. The yeast counts increased sharply in control samples and spoiled after 15 days of storage. Kebary et al., (2015) conducted that in probiotic Domiati cheese made by L.rhamnosus the coliform, aerobic spore forming bacteria, yeast and mold did not detected in all cheese treatments either when fresh or during pickling period.
Bacteriocins are also present in species of genus Lactobacillus, the L.casei produce casein80 (Klaenhammer, 1993).  Hussein and Shalaby (2014) stated that Kareish cheeses made with probiotic starter bifidobacterium had more accepted. Gabir (2016) conducted that the dairy product had a higher acceptability with the addition of B.bifidum compared with the control. Ramzan et al., (2010) found that L.rhamnosus on cheddar cheese enhanced the quality of Cheddar cheese. Abdalla (2005) observed that organoleptic evaluated of white soft cheese (Gibna beyda) made using different rates of starter culture had a high quality standard more than the control. The sensory parameters for probiotic Beyaz cheese samples during their storage period, cheese batch containing B.bifidum received the highest sensory scores (Yangilar and Ozdemir, 2014). Oliveira et al., (2012) noted that Coalho cheese with the added probiotic lactic acid strains alone and in co-culture were had better accepted in the sensory evaluation than cheeses without the probiotic strains. Escobar et al. (2012) reported that probiotic supplementation of Panela Cheese containing L.rhamnosus showed greater consumer acceptance. Kebary et al., (2015) noticed that acceptability increased in all Domiati cheese supplemented with different levels of L.rhamnosus during the early stage of storage and by extending of the pickling period. Mahmoudi et al., (2012) mentioned that white Iranian probiotic cheese with starter had the highest of sensory acceptability. Sidig et al., (2016) noticed that white soft cheese (jibna-beida) prepared by using starter culture (LAB) had a higher acceptability. Probiotic bacteria in cheese especially lactobacilli possess several peptidases, which can hydrolyze peptides to oligopeptides and amino acid and induce change in flavour, body and texture and, consequently, in sensory properties of the cheese (Santillo and Albanzia, 2008; Shihata and Shah, 2000; Soufa and Saad, 2009).

CONCLUSION
The effect of probiotic bacteria on the quality, the growth and survival of probiotics strains (L.rhamnosus, L.casei and B.bifidium) in Sudanese white soft cheese made from cow's milk was investigated. Microbial analysis and sensory evaluation were carried out at 0,15,30,45 and 60 days interval.