Towards the Understanding of Fermented Food Biotechnology in Congo Brazzaville

Congo Brazzaville harbor a diversity of traditional fermented foods which involve complex interactions between various microorganisms groups and different molecules in detoxification, texture and organoleptic features. This first retrospective review aims to describe the various works that have been done regarding the biochemical and microbiological features of our traditional fermented foods including Nsamba, Poto-poto, Bikedi and Ntoba-Mbodi.

Technological processes for the traditional food production vary between areas (Table 2).However, they are generally based on the same techniques (Louémbé et al., 1986(Louémbé et al., , 1996;;Louembé et al., 2003b;Malonga et al., 1995).It is done with a non-sterile equipment and non-optimized conditions, resulting in food with an unpredictable nutritional quality (Olanrewaju et al., 2009).

Poto-poto: fermented dough of maize:
In Africa, the fermentation of maize (Zea mays) is achieved through an extremely long process.This dough is called ujior akamu in Kenya (Akingbala et al., 1981), ogiorakamu inNigeria (Akinrele, 1970), koko or akasain Ghana (Andah and Muller, 1973), mahewu in South Africa (Schweigart and de Wit, 1960) and Poto-poto in Congo Brazzaville.It contains very poor nutritional value.47% of the population in rural areas and 78% in Brazzaville feed on poto-poto (PP) (Treche et al., 1992).Louembé et al. (2004) had identified the major stages of fermentation and the microbiological diversity during the fermentation process.Nutritional quality was also documented (Elenga et al., 2009).
To remove hydrogen cyanide people practice traditional fermentation including retting and browning (Dunican, 1990;Lancaster et al., 1982;Ongusua et al., 1983).In addition, this allows a significant production of organic acids and a softening of tubers essential for the subsequent operations of transformation (Malonga et al., 1996;Kobawila et al., 2005).In order to improve the quality of final product, studies have been determined the effectiveness of these traditional techniques in reducing of cyanide compounds (Louémbé et al., 1997;Kobawila et al., 2005).Many studies have been devoted to microbiology in the study of retting and softening phenomenon (Okafor et al., 1984;Oyewole and Odunfa, 1990;Cooke, 1978;Malonga et al., 1996;Conn, 1969;Ampe et al., 1995), but the understanding on detoxification is still poor and controversial.
Biotechnology of Ntoba Mbodi (NM): NS, PP and BK products are processed in acidic medium, while Ntoba Mbodi (NM) is accompanied by a significant alkalinization (pH 8.6) of fermentation medium.The fermented NM.Moreover, cassava leaves are used for human consumption due to their high nutritional value.Cassava leaves are highly rich in protein, minerals and vitamins and all essential amino acids except for methionine and phenylalanine (Eggum, 1970;Ravindran and Ravindran, 1988;Gómez and Valdivieso, 1985;Rogers and Milner, 1963; Ross and Enriquez, The transformation of cassava leaves in NM lasts only four days (Kobawila et al., 2005).Beyond 4 days, NM fermentation appears significantly altered.This had not been observed in acidifying fermentation (Malonga et al., 1993).The alkalization of the pH determines the appearance of the microbial flora.The activity of pectate-Iyases, polygalacturonase, pectinesterase, cellulases increases the softening from the first to the third day of NM fermentation (Mokemiabeka et al., 2011).As mentioned above with Clotridium butyricum, the action of pectin enzymes using polygalacturonate polymethyl substrate released by the microorganism is necessary for softening (Kéléké et al., 2007).The increase of browning is correlating with increase of the concentration of polyphenolic polymers due to the polyphenol oxidases activity.This phenomenon of browning is interesting for NM fermentation since it improves its sensory properties (Martinez and Whitaker, 1995;Whitaker, 1995).Enzymes like proteases, deaminases and decarboxylases which generate amines are responsible for the alkalinization in medium (pH 8.5).B. Subtilis and B. amylolyquefasciens isolated from NM would beinvolved for the pH increasing by producing ammonia released during the degradation of proteins and peptides (Steinkraus, 1991;Ouoba et al., 2015) (Table 5).The alkaline pH promote the development of other Bacillus species including B. pumilus, to use hydrogen cyanide for their nutrition and therefore to decrease detoxification (Knowles, 1976;Wong and Jackson, 1983).These microorganisms are commonly found in vegetable products (Etchells et al., 1975) and also some species like Micrococcus varians, B. marcerans, B. susbtilis, Staphylococcus sciuri et Staphylococcus xylosus (Pederson and Albury, 1983) (Table 5) are the more representatives.
Amplification of 16S RNA gene and Sequencing experiments of in NM isolates allowed identification some bacteria including Weissella confusa, W. cibaria, Lb. plantarum, Pediococcus pentosaceus, Enterococcus casseliflavus, E. faecium, E. faecalis, E. avium and E. hirae (Ouoba et al., 2010).The presence of this type of bacteria demonstrates the attention that should be brought to NM, regarding different stages of quality control and HACCP procedures.These bacteria are typically found at the beginning of fermentation and some of them are responsible for nosocomial infections (Khan et al., 2015).
The enzymes of biotechnological interest: fibrinolytic and pectinolytic enzymes: Bacillus sp. has been considered for their capacity to secrete more therapeutic enzymes.Fibrinolytic and pectinolytic enzymeshave been identified (Nguimbi and Wu, 2002;Nguimbi et al., 2014;Ouoba et al., 2015).Previous studies have been shown that several bacilli species are able to produce fibrinolytic enzymes including Bacillus natto, B. amyloliquefaciens DC-4, Bacillus sp.CK, B. subtilis EM 29 and B. amyloliquefaciens CH51 (Sumi et al., 1987;Peng and Zhang, 2002;Kim et al., 1996;Nguimbi and Wu, 2002;Kim et al., 2009).B. sutilis and B. amyloliquefasciens isolated from NM are able to degrade fibrin and casein (Nguimbi et al., 2014).Deep studies will be specially addressed to Lysinibacillus louembei, novel exciting bacteria (Ouoba et al., 2015).Softening of cassava leaves is the main stage which indicates the end of the fermentation.Clostridium bytirycum MGPP179 genetically close to Clostridium felcium seems to have the necessary assets for his involvement in the retting process of cassava tubers and softening (Kéléké et al., 2007).Several bacteria secrete enzymes degrading cell walls (Nambisan and Sundaresan, 1985).Alteration of pectic compounds is generally related to the action of pectinolytic enzymes that generate significant changes in cell structures.This is especially saponifying enzymes like pectin Iyase, pectate lyase, polygalacturonase and some unknown hydrolases that can hydrolyze pectin containing in cell wall (Rexová-Benková and Markovic, 1976;Thibault, 1980).These enzymes allow the destruction of plant tissues and therefore the opening of target compartments thus favoring contact between the endogenous linamarase and linamarin (Okafor et al., 1984;Nambisan and Sundaresan, 1985).Microorganisms associated in pectinolytic activities and the evolution of softening were therefore studied.Studies showed that many bacteria are able to synthesize linamarase enzyme in the presence of appropriate linamarin or lotaustralin substrate.This includes Lc. lactis, Ln. mesenteroides and Lb.plantarum (Okafor and Ejiofor, 1985).
Bacteriocins and probiotic bacteria of congo brazzaville fermented foods: Bacteriocins are antimicrobial peptides produced by bacteria and could present a bactericidal effect against closely related species (Cotter et al., 2005).LAB represents a model of choice to ensure the absence pathogenic bacteria in the final product (Konings et al., 2000).Nisin from LAB is the only bacteriocin approved for use as a food preservative (Balciunas et al., 2013).Thirty-one bacteriocins from Lactobacillus specie were identified in Congolese fermented maize product (Poto-poto).Lactobacillus sp. can inhibit pathogenic bacteria including Escherichia coli, Salmonella enterica, Enterobacter aerogenes, Bacillus cereus, Staphylococcus aureus, Listeria monocytogenes and Enterococcus faecalis.Specific studies were performed on the correlation and the involvement between LAB fermentation and the reduction of pathogenic bacteria such as Salmonella typhimurium, Staphylococcus aureus, Escherichia coli and Shigella sp.Children fed with fermented starches showed very positive effects by reducing the risk of childhood diarrhea (Svangberg, 1991a(Svangberg, , 1991b;;Svangberg and Svanberg, 1989).
Fermented foods like NM, PP and BK are a best ecological niche to isolate microorganism producing bacteriocins.The new strain Lysinibacillus louembei isolated from NN could be interesting to investigate (Ouoba et al., 2015).
Promising and sustainable future: The use of bacteriocin-producing bacteria may be interesting as well as economic and legislative level.Producing bacteria bacteriocins may be added as a starter in fermented products (Gálvez et al., 2007;Dortu and Thonart, 2009).Their use must consider the Quorum Sensing System, the product composition (available nutrients, pH and food additives) and storage conditions.Bacteria with pectynolytic and linamarasic activities isolated from Congo Brazzaville traditional fermented foods are particularly interesting as a starter in fermented products.The characterization of these enzymes, from gene isolation to the final product will be an essential biotechnological tool for use in Biocatalysis.New isolated strains such as Lysinibacillus louembei could be interesting for the discovery of new molecules of biotechnological interest.

CONCLUSION
Basing on numerous studies discussing and mentioned above, we conclude that Congo Brazzaville fermented foods contain a fascinating diversity of microorganism.Many of them have a biotechnological interest and therapeutic properties.In addition molecules delivered from bacteria could be a great use to humanity and the natural world for a sustainable future.This discovery opens a promising future for the valuation through research and development.

Table 1 :
Non-exhaustive list of various fermented foods and beverages met in Congo Brazzaville

Table 2 :
Flowchart of production of three traditional fermented foods of Congo Brazzaville

Table 3 :
Isolated microorganisms from NS

Table 4 :
Isolated microorganisms of Poto-poto, food produced by natural fermentation of the maize dough

Table 5 :
Isolated microorganisms of Bikédi and fermented rough of cassava tubers

Table 6 :
Ntoba Mbodi (NM) isolated microorganisms from the natural fermentation of cassava leaves