African Fermented Foods Research Articles

The predominant lactic acid bacteria and yeasts involved in the spontaneous fermentation of millet during the production of the traditional porridge Hausa koko in Ghana

Spontaneous fermentation of cereals like millet involves a diverse population of microbes from various sources, including raw materials, processing equipment, fermenting receptacles, and the environment. Here, we present data on the predominant microbial species and their succession at each stage of the Hausa koko production process from five regions of Ghana. The isolates were enumerated using selective media, purified, and phenotypically characterised. The LAB isolates were further characterised by 16S rRNA Sanger sequencing, typed using (GTG)5 repetitive-PCR, and whole genome sequencing, while 28S rRNA Sanger sequencing was performed for yeast identification. The pH of the millet grains ranged from mean values of 6.02—6.53 to 3.51—3.99 in the final product, depending on the processors. The mean LAB and yeast counts increased during fermentation then fell to final counts of log 2.77–3.95 CFU/g for LAB and log 2.10–2.98 CFU/g for yeast in Hausa koko samples. At the various processing stages, the counts of LAB and yeast revealed significant variations (p < 0.0001). The species of LAB identified in this study were Limosilactobacillus pontis, Pediococcus acidilactici, Limosilactobacillus fermentum, Limosilactobacillus reuteri, Pediococcus pentosaceus, Lacticaseibacillus paracasei, Lactiplantibacillus plantarum, Schleiferilactobacillus harbinensis, and Weissella confusa. The yeasts were Saccharomyces cf. cerevisiae/paradoxus, Saccharomyces cerevisiae, Pichia kudriavzevii, Clavispora lusitaniae and Candida tropicalis. The identification and sequencing of these novel isolates and how they change during the fermentation process will pave the way for future controlled fermentation, safer starter cultures, and identifying optimal stages for starter culture addition or nutritional interventions. These LAB and yeast species are linked to many indigenous African fermented foods, potentially acting as probiotics in some cases. This result serves as the basis for further studies into the technological and probiotic potential of these Hausa koko microorganisms.

Complete and Draft Genome Sequences of 52 Bacillus and Priestia Strains Isolated from West African Fermentations and 26 Reference Strains from a Public Culture Collection.

The whole genomes of 78 Bacillus and Priestia strains isolated from West African fermented foods (n = 52) or acquired from a public culture collection (n = 26) were sequenced using long-read sequencing and assembled into draft (n = 32) and complete (n = 46) genomes, allowing comparative genomics and taxonomic assignment of these strains with putative uses in fermented foods.

Antimicrobial Resistance of Lactic Acid Bacteria from Nono, a Naturally Fermented Milk Product.

Antimicrobial resistance (AMR) is one of the biggest threats to public health. The food chain has been recognised as a vehicle for transmitting AMR bacteria. However, information about resistant strains isolated from African traditional fermented foods remains limited. Nono is a traditional, naturally fermented milk product consumed by many pastoral communities across West Africa. The main aim of this study was to investigate and determine the AMR patterns of lactic acid bacteria (LAB) involved in the traditional fermentation of milk for Nono production, and the presence of transferable AMR determinants. One hundred (100) LAB isolates from Nono identified in a previous study as Limosilactobacillus fermentum, Lactobacillus delbrueckii, Streptococcus thermophilus, Streptococcus infantarius, Lentilactobacillus senioris, Leuconostoc pseudomesenteriodes, and Enterococcus thailandicus were investigated. The minimum inhibitory concentration (MIC) was determined for 18 antimicrobials using the micro-broth dilution method. In addition, LAB isolates were screened for 28 antimicrobial resistance genes using PCR. The ability of LAB isolates to transfer tetracycline and streptomycin resistance genes to Enterococcus faecalis was also investigated. The experiments revealed variable antimicrobial susceptibility according to the LAB isolate and the antimicrobial tested. The tetracycline resistance genes tet(S) and tet(M) were detected in isolates Ent. thailandicus 52 and S. infantarius 10. Additionally, aad(E) encoding resistance to streptomycin was detected in Ent. thailandicus 52. The conjugation experiments suggested that the tet(S) and aad(E) genes were transferable in vitro from isolate Ent. thailandicus 52 to Ent. faecalis JH2-2. Traditional fermented foods play a significant role in the diet of millions of people in Africa, yet their contribution to the burden of AMR is largely unknown. This study highlights that LAB involved in traditionally fermented foods could be potential reservoirs of AMR. It also underscores the relevant safety issues of Ent. thailandicus 52 and S. infantarius 10 for use as starter cultures as they carry transferable AMR genes. Starter cultures are an essential aspect of improving the safety and quality attributes of African fermented foods. However, AMR monitoring is an important safety aspect in the selection of starter cultures for improving traditional fermentation technologies.

Nutritional and Phytochemical Composition of Mahewu (a Southern African Fermented Food Product) Derived from White and Yellow Maize (Zea mays) with Different Inocula

Mahweu is an important indigenous beverage for many low-income and undernourished consumers in southern Africa. As a result, the nutritional and phytochemical profile of mahewu samples (obtained using optimized fermentation and boiling conditions from a previous study) as well as their related raw materials (white and yellow maize) were investigated. At these conditions, white and yellow maize mahewu (WM and YM) were prepared utilizing various inocula including sorghum malt, wheat, millet malt, or maize malt, and the pH, titratable acidity (TTA), total soluble solid (TSS), and proximate analysis were determined. The mineral content, amino acid composition, and phenolic compound profile were also investigated using inductive coupled plasma optical emission spectrometry (ICP-OES), high-performance liquid chromatography (HPLC), and ultra-high performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UHPLC/Q-TOF-MS), respectively. Fermentation was observed to have influenced the proximate composition of obtained mahewu samples compared to the raw flour with significant (p ≤ 0.05) improvement in protein from 8.59 to 9.7% (YM) and 8.78 to 9% (WM) as well as carbohydrate from 72.27 to 74.47% (YM) and 71.15 to 72.65% (WM). Sodium, magnesium, phosphorous, potassium, calcium, manganese, iron, copper, and zinc were the minerals detected in the mahewu samples, while potassium was the most abundant mineral, having values ranging from 3051.61 to 3283.38 mg/kg (YM) and 2882.11 to 3129.97 mg/kg (WM). Heavy metals detected in this study were all below the recommended tolerable levels by the Joint FAO/WHO Expert Committee on Food Additives (JECFA). Arginine and leucine with values ranging from 0.47 to 0.52 g/100 g (YM) and 0.48 to 0.53 g/100 g (WM) as well as 0.91 to 1.04 g/100 g (YM) and 0.95 to 1.01 g/100 g (WM), respectively, were the most abundant essential amino acids, whereas for non-essential amino acids, glutamic acid, aspartic acid, alanine, and proline were observed to be abundant. Based on the different inocula, the derived mahewu samples prepared using either white or yellow maize have varying nutritional and health beneficial components and the choice of inocula might still be determined by consumer preference.

Immune System and Epidemics: The Role of African Indigenous Bioactive Substances.

With over 6 million coronavirus pandemic deaths, the African continent reported the lowest death rate despite having a high disease burden. The African community's resilience to the pandemic has been attributed to climate and weather conditions, herd immunity, repeated exposure to infectious organisms that help stimulate the immune system, and a disproportionately large youth population. In addition, functional foods, herbal remedies, and dietary supplements contain micronutrients and bioactive compounds that can help boost the immune system. This review identified significant traditional fermented foods and herbal remedies available within the African continent with the potential to boost the immune system in epidemics and pandemics. Methodology: Databases, such as PubMed, the Web of Science, and Scopus, were searched using relevant search terms to identify traditional African fermented foods and medicinal plants with immune-boosting or antiviral capabilities. Cereal-based fermented foods, meat-, and fish-based fermented foods, and dairy-based fermented foods containing antioxidants, immunomodulatory effects, probiotics, vitamins, and peptides were identified and discussed. In addition, nine herbal remedies and spices belonging to eight plant families have antioxidant, immunomodulatory, anti-inflammatory, neuroprotective, hepatoprotective, cardioprotective, and antiviral properties. Peptides, flavonoids, alkaloids, sterols, ascorbic acid, minerals, vitamins, and saponins are some of the bioactive compounds in the remedies. Bioactive compounds in food and plants significantly support the immune system and help increase resistance against infectious diseases. The variety of food and medicinal plants found on the African continent could play an essential role in providing community resilience against infectious diseases during epidemics and pandemics. The African continent should investigate nutritional, herbal, and environmental factors that support healthy living and longevity.

Phylogenomics of a Saccharomyces cerevisiae cocoa strain reveals adaptation to a West African fermented food population

SummaryVarious yeast strains have been proposed as candidate starter cultures for cocoa fermentation, especially strains of Saccharomyces cerevisiae. In the current study, the genome of the cocoa strain S. cerevisiae IMDO 050523 was unraveled based on a combination of long- and short-read sequencing. It consisted of 16 nuclear chromosomes and a mitochondrial chromosome, which were organized in 20 contigs, with only two small gaps. A phylogenomic analysis of this genome together with another 105 S cerevisiae genomes, among which 20 from cocoa strains showed a geographical distribution of the latter, including S. cerevisiae IMDO 050523. Its genome clustered together with that of a West African fermented food population, indicating a wider adaptation to West African food niches than cocoa. Furthermore, S. cerevisiae IMDO 050523 contained genetic signatures involved in sucrose hydrolysis, pectin degradation, osmotolerance, and conserved amino acid changes in key ester-producing enzymes that could point toward specific niche adaptations.

English

Yeasts are very important in food production by affecting the quality and safety of different food products. Yeasts are commonly used in the production of beer, wine and bread; but many yeast species have been isolated from different African fermented foods. The present study aims to determine the biodiversity of yeasts isolated from selected Nigerian fermented foods. A combination of phenotypic and molecular tests were employed in the identification of yeasts. A total of 145 yeasts were isolated from six different food products. The yeasts belong to six genera namely Saccharomyces, Candida, Cyberlindnera, Meyerozyma, Trichosporon and Galactomyces. The most frequently encounter was Saccharomyces cerevisiae followed by Candida glabrata. The present study has confirmed the biodiversity of yeasts isolated from fermented food of Nigerian origin. Key words: Phenotypic characterization, molecular, indigenous yeasts, Nigeria.

Lactobacillus garii sp. nov., isolated from a fermented cassava product.

A novel Gram-positive, catalase negative, rod-shaped strain, FI11369T, was isolated from gari, a traditional West African fermented food derived from cassava. Based on 16S rRNA gene sequence similarity, the closest type strains were Lactobacillus xiangfangensis LMG 26013T (99.4 % similarity), Lactobacillus plajomi NBRC 107333T (99.1 %), Lactobacillus paraplantarum DSM 10667T (99.1 %), Lactobacillus pentosus DSM 20314T (99.0 %), Lactobacillus plantarum subsp. plantarum ATCC 14917T (99.0 %), Lactobacillus modestisalitolerans NBRC 107235T (98.9 %), Lactobacillus plantarum subsp. argentoratensis DSM 16365T (98.9 %) and Lactobacillus daowaiensis NCIMB 15183T (98.8 %). The genome of strain FI11369T was sequenced and the average nucleotide identity (ANI) was compared with its closest relatives. ANI analysis showed that the closest relative, L. xiangfangensis DSM 27103T, had only a 82.4 % similarity. The main fatty acids of FI11369T were saturated C16 : 0 (18.2 %), unsaturated C18 : 1 ω9c (43.8 %) and cyclopropane C19 : 0 cyclo (ω10c and/or ω6; 22.5 %). Based on the genotypic and phenotypic data obtained in this study, a novel Lactobacillus species, Lactobacillus garii sp. nov., with the type strain FI11369T (=NCIMB 15148=DSM 108249), is proposed.

Identification of lactic acid bacteria and determination of selected biochemical properties in emasi and emahewu

Fermented foods are produced at household level for personal consumption in the Kingdom of Eswatini (formerly Swaziland). In this study, we determined the biochemical aspects, enumeration, isolation and identification of lactic acid bacteria (LAB) in emasi and emahewu – two Swazi traditional fermented foods. Emasi had an average pH of 4.68, titratable acidity of 0.9% and LAB count of 8.25 log CFU/mL. Emahewu had a pH of 3.62, titratable acidity of 0.4% and LAB count of 8.10 log CFU/mL. The LAB counts were consistent with observations for similar African fermented foods. The LAB from emasi and emahewu were identified through Gram stain, catalase reaction, sugar assimilation tests using API 50 CH test strips, and sequencing of 16S rDNA. It was found (from nine isolates) that Lactococcus lactis subsp. lactis and Leuconostoc mesenteroides were the common strains in emasi. Lactobacillus plantarum, Lactobacillus paracasei ssp. paracasei and Lactobacillus brevis were also detected. Lb. plantarum, L. mesenteroides ssp. mesenteroides, Lactobacillus fermentum, Lb. brevis, Wessella confusa, Lactobacillus acidophilus and Lb. lactis were found in emahewu (from 16 isolates). This finding was consistent with LAB found in a South African fermented milk, in which common genera were Leuconostoc, Lactococcus and Lactobacillus. Strains found in emahewu – mainly Lactobacillus spp., Weissella and Enterococcus – are similar to those found in ting, a South African fermented non-alcoholic beverage. &#x0D; Significance: &#x0D; &#x0D; This study provides the first documentation of microbial and biochemical aspects of the Swazi traditional fermented foods, emasi and emahewu.&#x0D;

Beneficial Effects of a Fermented Maize product with Its Supernatant, Lactobacillus fermentum and Lactobacillus brevis in Rat Model of Colitis

Background: African fermented foods such as maize cereal (Ogi ) and its supernatant (Omidun) are reservoirs of beneficial lactobacilli and carboxylic acids. Based on their constituents, their beneficial effect in a rat model of 2,4,6-trinitrobenzene sulphonic (TNBS) acid-induced colitis was assessed in comparison with two lactobacilli in a rat model of colitis. Methods: Female Wistar rats were distributed into seven groups of 5 rats each; the rats were pretreated for 14 days prior to colitis induction with Ogi, Omidun, L. fermentum, L. brevis and 7 days post colitic induction. Colitis was induced by an intracolonic administration of TNBS. The response of the rats to treatment was assessed macroscopically and biochemically. Results: Treatment with the Dexamethasone, Ogi and Omidun resulted in a significant reduction in colonic damage score and weight/length ratio (p&lt;0.05). Treatment with Ogi, Omidun, L. brevis, and dexamethasone significantly prevented depletion of colonic glutathione and superoxide dismutase. The up-regulation of myeloperoxidase activity was inhibited in all treated colitic rats (p&lt;0.05). However, Ogi appears to produce a better protective effect than the other treatment groups. Conclusions: This study reports that Ogi protects Wistar rats against the deleterious effect of trinitrobenzene sulphonic acid better than pure lactobacilli strains. Keywords: Fermented food, lactic acid bacteria, inflammation, oxidative stress.

Comparison of the microbial composition of African fermented foods using amplicon sequencing

Fermented foods play a major role in the diet of people in Africa, where a wide variety of raw materials are fermented. Understanding the microbial populations of these products would help in the design of specific starter cultures to produce standardized and safer foods. In this study, the bacterial diversity of African fermented foods produced from several raw materials (cereals, milk, cassava, honey, palm sap, and locust beans) under different conditions (household, small commercial producers or laboratory) in 8 African countries was analysed by 16S rRNA gene amplicon sequencing during the Workshop “Analysis of the Microbiomes of Naturally Fermented Foods Training Course”. Results show that lactobacilli were less abundant in fermentations performed under laboratory conditions compared to artisanal or commercial fermentations. Excluding the samples produced under laboratory conditions, lactobacilli is one of the dominant groups in all the remaining samples. Genera within the order Lactobacillales dominated dairy, cereal and cassava fermentations. Genera within the order Lactobacillales, and genera Zymomonas and Bacillus were predominant in alcoholic beverages, whereas Bacillus and Lactobacillus were the dominant genera in the locust bean sample. The genus Zymomonas was reported for the first time in dairy, cereal, cassava and locust bean fermentations.

Occurrence and Importance of Yeasts in Indigenous Fermented Food and Beverages Produced in Sub-Saharan Africa.

Indigenous fermented food and beverages represent a valuable cultural heritage in sub-Saharan Africa, having one of the richest selections of fermented food products in the world. In many of these indigenous spontaneously fermented food and beverages, yeasts are of significant importance. Several factors including raw materials, processing methods, hygienic conditions as well as the interactions between yeasts and other commensal microorganisms have been shown to influence yeast species diversity and successions. Both at species and strain levels, successions take place due to the continuous change in intrinsic and extrinsic growth factors. The selection pressure from the microbial stress factors leads to niche adaptation and both yeast species and strains with traits deviating from those generally acknowledged in current taxonomic keys, have been isolated from indigenous sub-Saharan African fermented food products. Yeasts are important for flavor development, impact shelf life, and nutritional value and do, in some cases, even provide host-beneficial effects. In order to sustain and upgrade these traditional fermented products, it is quite important to obtain detailed knowledge on the microorganisms involved in the fermentations, their growth requirements and interactions. While other publications have reported on the occurrence of prokaryotes in spontaneously fermented sub-Saharan food and beverages, the present review focuses on yeasts considering their current taxonomic position, relative occurrence and successions, interactions with other commensal microorganisms as well as beneficial effects and importance in human diet. Additionally, the risk of opportunistic yeasts is discussed.

Improving food value chains for cereal doughs in West Africa: case study of mawè in Benin

A good understanding of the socio-demographic and economic factors which characterize the food value chains (FVC) of traditional African fermented foods is a major prerequisite for developing and implementing scientific evidence-based policies towards the improvement of Africa’s food and nutrition security and sustainable agriculture. This study aims at identifying the key factors which shape the FVC for mawe and its derived products. Mawe is a traditional fermented cereal dough, widely consumed in Benin. The findings of the study show that mawe and its derived products are processed, marketed, and consumed through both ‘traditional’ and ‘traditional-to-modern’ value chains. The traditional chain plays a key role in facilitating food affordability for low-income people. However, the traditional-to-modern value chain is increasingly predominant, especially in an urbanized environment, and consequently offers greater opportunities for value chain upgrading for value-added processing of traditional fermented foods. The study offers insights into policy and research challenges in value chain upgrading.

Probiotic potential of thermotolerant lactic acid bacteria isolated from “Gari “a cassava-based African fermented food

Probiotic potential of thermotolerant lactic acid bacteria isolated from “Gari “a cassava-based African fermented food

Potential probiotic Pichia kudriavzevii strains and their ability to enhance folate content of traditional cereal-based African fermented food

With the aim of selecting starter cultures with interesting probiotic potential and with the ability to produce folate in a food matrix, yeast strains isolated from fermented cereal-based African foods were investigated. A total of 93 yeast strains were screened for their tolerance to pH 2 and 0.3% of bile salts. Pichia kudriavzevii isolates gave the best results. Selected P. kudriavzevii strains were tested for survival to the simulated human digestion and for adhesion to Caco-2 cells. Moreover, presence of folate biosynthesis genes was verified and production of extra and intra-cellular folate determined during growth in culture medium. 31% of yeast strains could tolerate pH 2, while 99% bile salts. Survival rate after simulated digestion ranged between 11 and 45%, while adhesion rate between 12 and 40%. Folate production was mainly intracellular, maximum after 24 h of growth. To be closer to traditional cereal-based fermentations, a P. kudriavzevii strain with good probiotic potential was co-inoculated with Lactobacillus fermentum strains in a pearl millet gruel. This resulted in in situ folate production that peaked after 4 h. The use of strains with both probiotic and nutritional enrichment properties may have a greater impact for the consumers.

PCR of crtNM combined with analytical biochemistry: An efficient way to identify carotenoid producing lactic acid bacteria

Lactic acid bacteria (LAB) synthesize a wide variety of biochemical compounds during food fermentation. Carotenoids provide important biological functions for bacteria, and their consumption by humans has many beneficial effects. In this study, the presence of several genes involved in the production of carotenoids was determined by BLAST analysis and PCR in a collection of 156 LAB isolated from traditional amylaceous African fermented foods. Only the crtE gene and the crtNM operon were present and detected in Lactobacillaceae. Most of the strains with positive PCR detection of the operon crtNM produced carotenoid-like compounds when grown in MRS broth. The carotenoids produced differed from compounds previously identified in other LAB except for one peak, which was closely related to 4,4′-diaponeurosporene already reported in the literature in Lactobacillus plantarum species. Most producing strains belonged to Lactobacillus fermentum and L. plantarum species but a few Pediococcus acidilactici were also producers. Furthermore, the most efficient L. plantarum was able to synthesize carotenoids in a cereal fermented food. Genetic screening was shown to be efficient since, in all cases, it eliminated the need for biochemical analysis of strains in which no amplicons of the operon crtNM were obtained.

Production of autoinducer-2 by aerobic endospore-forming bacteria isolated from the West African fermented foods.

Autoinducer-2 (AI-2) is a quorum-sensing (QS) molecule which mediates interspecies signaling and affects various bacterial behaviors in food fermentation. Biosynthesis of AI-2 is controlled by S-ribosylhomocysteine lyase encoded by the luxS gene. The objective of this study was to investigate production of AI-2 by aerobic endospore-forming bacteria (AEB) isolated from the West African alkaline fermented seed products Mantchoua and Maari. The study included 13 AEB strains of Bacillus subtilis, B. cereus, B. altitudinis, B. amyloliquefaciens, B. licheniformis, B. aryabhattai, B. safensis, Lysinibacillus macroides and Paenibacillus polymyxa. All the tested strains harbored the luxS gene and all strains except for P. polymyxa B314 were able to produce AI-2 during incubation in laboratory medium. Production of AI-2 by AEB was growth phase dependent, showing maximum activity at the late exponential phase. AI-2 was depleted from the culture medium at the beginning of the stationary growth phase, indicating that the tested AEB possess a functional AI-2 receptor that internalizes AI-2. This study provides the evidences of QS system in Bacillus spp. and L. macroides and new knowledge of AI-2 production by AEB. This knowledge contributes to the development of QS-based strategies for better control of alkaline fermentation.

Characterization of the dominant microorganisms responsible for the fermentation of dehulled maize grains into nsiho in Ghana

Nsiho (white kenkey) is a type of kenkey, a sour stiff dumpling, produced from fermented maize meal in Ghana. The dominant microorganisms responsible for the fermentation of nsiho were characterized by analysing samples from four traditional production sites at Anum in the Eastern Region of Ghana. During 48 h of steeping dehulled maize grains, the pH values decreased from 6.05 to 5.93 to 3.59 to 3.55, whilst titratable acidity increased from 0.02 to 0.03 to 0.27 to 0.32%. In the subsequent 12 h dough fermentation, the pH decreased from 6.02 to 5.80 to 3.52 to 3.46, whilst titratable acidity increased from 0.25 to 0.27 to 0.35 to 0.38%. The lactic acid bacteria population increased by 2 to 5 log units to concentrations of 107 to 108 CFU/ml during steeping and by 2 to 3 log units from 105 to 106 CFU/g to 108 to 109 CFU/g during dough fermentation. Yeasts counts increased by 3 to 4 log units during steeping and by 2 to 4 log units during dough fermentation. The most frequently isolated lactic acid bacteria responsible for nsiho fermentation were identified as Lactobacillus fermentum (47.1%), Lactobacillus brevis (25%), Lactobacillus plantarum (14.42%), Pediococcus pentosaceus (8.65%) and Pediococcus acidilactici , (4.8%). The dominant yeasts species were Saccharyomyces cerevisiae (47.6%), Candida krusei (29.1%), Debaryomyces spp ., (15%) and Trichosporon spp ., (8.3%). This is the first study to report on the micororganisms involved in nsiho fermentation. Key words : Nsiho , dehulled maize, kenkey, lactic acid bacteria, indigenous African fermented foods.

Phytase-producing capacity of yeasts isolated from traditional African fermented food products and PHYPk gene expression of Pichia kudriavzevii strains

Phytate is known as a strong chelate of minerals causing their reduced uptake by the human intestine. Ninety-three yeast isolates from traditional African fermented food products, belonging to nine species (Pichia kudriavzevii, Saccharomyces cerevisiae, Clavispora lusitaniae, Kluyveromyces marxianus, Millerozyma farinosa, Candida glabrata, Wickerhamomyces anomalus, Hanseniaspora guilliermondii and Debaryomyces nepalensis) were screened for phytase production on solid and liquid media. 95% were able to grow in the presence of phytate as sole phosphate source, P. kudriavzevii being the best growing species. A phytase coding gene of P. kudriavzevii (PHYPk) was identified and its expression was studied during growth by RT-qPCR. The expression level of PHYPk was significantly higher in phytate-medium, compared to phosphate-medium. In phytate-medium expression was seen in the lag phase. Significant differences in gene expression were detected among the strains as well as between the media. A correlation was found between the PHYPk expression and phytase extracellular activity.

Characterization of Bacillus spp. strains for use as probiotic additives in pig feed

Bacillus spp. are commonly used as probiotic species in the feed industry, however, their benefits need to be confirmed. This study describes a high throughput screening combined with the detailed characterization of endospore-forming bacteria with the aim to identify new Bacillus spp. strains for use as probiotic additives in pig feed. A total of 245 bacterial isolates derived from African fermented food, feces and soil were identified by 16S rRNA gene sequencing and screened for antimicrobial activity and growth in the presence of antibiotics, bile salts and at pH4.0. Thirty-three Bacillus spp. isolates with the best characteristics were identified by gyrB and rpoB gene sequencing as B. amyloliquefaciens subsp. plantarum, B. amyloliquefaciens subsp. amyloliquefaciens, B. subtilis subsp. subtilis, B. licheniformis, B. mojavensis, B. pumilus and B. megaterium. These isolates were further investigated for their activity against the pathogenic bacteria, antibiotic susceptibility, sporulation rates, biofilm formation and production of glycosyl hydrolytic enzymes. Additionally, ten selected isolates were assessed for heat resistance of spores and the effect on porcine epithelial cells IPEC-J2. Isolates of B. amyloliquefaciens, B. subtilis and B. mojavensis, showed the best overall characteristics and, therefore, potential for usage as probiotic additives in feed. A large number of taxonomically diverse strains made it possible to reveal species and subspecies-specific trends, contributing to our understanding of the probiotic potential of Bacillus species.