Bifidobacterium species viability in dairy-based probiotic foods: challenges and innovative approaches for accurate viability determination and monitoring of probiotic functionality.

Bifidobacterium species are essential members of a healthy human gut microbiota. Their presence in the gut is associated with numerous health outcomes such as protection against gastrointestinal tract infections, inflammation, and metabolic diseases. Regular intake of Bifidobacterium in foods is a sustainable way of maintaining the health benefits associated with its use as a probiotic. Owing to their global acceptance, fermented dairy products (particularly yogurt) are considered the ideal probiotic carrier foods. As envisioned in the definition of probiotics as "live organisms," the therapeutic functionalities of Bifidobacterium spp. depend on maintaining their viability in the foods up to the point of consumption. However, sustaining Bifidobacterium spp. viability during the manufacture and shelf-life of fermented dairy products remains challenging. Hence, this paper discusses the significance of viability as a prerequisite for Bifidobacterium spp. probiotic functionality. The paper focuses on the stress factors that influence Bifidobacterium spp. viability during the manufacture and shelf life of yogurt as an archetypical fermented dairy product that is widely accepted as a delivery vehicle for probiotics. It further expounds the Bifidobacterium spp. physiological and genetic stress response mechanisms as well as the methods for viability retention in yogurt, such as microencapsulation, use of oxygen scavenging lactic acid bacterial strains, and stress-protective agents. The report also explores the topic of viability determination as a critical factor in probiotic quality assurance, wherein, the limitations of culture-based enumeration methods, the challenges of species and strain resolution in the presence of lactic acid bacterial starter and probiotic species are discussed. Finally, new developments and potential applications of next-generation viability determination methods such as flow cytometry, propidium monoazide-quantitative polymerase chain reaction (PMA-qPCR), next-generation sequencing, and single-cell Raman spectroscopy (SCRS) methods are examined.

Assessing probiotic viability in mixed species yogurt using a novel propidium monoazide (PMAxx)-quantitative PCR method.

Viability is a prerequisite for any therapeutic benefits associated with the ingestion of probiotic bacteria. Current culture-based techniques are inadequate for the enumeration of probiotics in mixed-species food products. This study utilized a quantitative PCR (qPCR) method coupled with propidium monoazide (PMAxx), and novel species-specific tuf gene primers to selectively enumerate Lacticaseibacillus rhamnosus, Bifidobacterium spp., and yogurt starter cultures in mixed-species probiotic yogurt. The method was optimized for PMAxx concentration and specificity and evaluated for efficiency and applicability. PMAxx-qPCR showed high specificity to the target organisms in mixed-species yogurt, quantifying only viable cells. The linear dynamic ranges were established over five to seven orders of magnitude. The assay was reliable with an efficiency of 91-99%, R2 values > 0.99, and a good correlation to the plate count method (r = 0.882). The results of this study demonstrate the high selectivity, improved lead time, and reliability of PMAxx-qPCR over the culture-dependent method, making it a valuable tool for inline viability verification during processing and improving probiotic quality assurance for processors and consumers.

Exploration of Infant Food Microbial Composition from Formal and Informal Settings Using Viable Counts and 16S rRNA Gene Amplicon Sequencing in Johannesburg, South Africa.

Diarrhoea is a considerable agent of disease and loss of life in children below age five in South Africa. Soweto, South Africa is an urban township in Johannesburg, with most of its population living in informal settlements. Informal settlements in areas such as Soweto are often impoverished communities that do not get water easily, inadequate sanitation is pervasive, and poor hygiene common (risk factors for diarrhoeal diseases). Among the age groups, infants are most vulnerable to diarrhoeal infection, mainly through the ingestion of food and water. The presence of undesirable microbiota is a food safety and health challenge. This study investigated the microbiome of infant food samples collected from formal (n = 19) and informal (n = 11) households in Soweto. A non-culture-dependent technique was used to characterise the bacterial diversity and composition of the infant food samples. The results indicated that household type did not influence microbial diversity and composition in Soweto. South Africa. Firmicutes, Proteobacteria, Cyanobacteria, and Tenericutes dominated the phyla rank in food samples from formal and informal households. Potential pathogens of public health significance, including diarrhoeal disease agents such as Salmonella spp., E. coli, and Campylobacter spp., were detected within the foods. We concluded that the infant food samples showed rich bacterial diversity, and the presence of potential pathogens of public health significance suggests a disease risk that infants may face upon consuming the foods.

The Microbial Genetic Diversity and Succession Associated with Processing Waters at Different Broiler Processing Stages in an Abattoir in Australia.

The high organic content of abattoir-associated process water provides an alternative for low-cost and non-invasive sample collection. This study investigated the association of microbial diversity from an abattoir processing environment with that of chicken meat. Water samples from scalders, defeathering, evisceration, carcass-washer, chillers, and post-chill carcass rinsate were collected from a large-scale abattoir in Australia. DNA was extracted using the Wizard® Genomic DNA Purification Kit, and the 16S rRNA v3-v4 gene region was sequenced using Illumina MiSeq. The results revealed that the Firmicutes decreased from scalding to evisceration (72.55%) and increased with chilling (23.47%), with the Proteobacteria and Bacteroidota changing inversely. A diverse bacterial community with 24 phyla and 392 genera was recovered from the post-chill chicken, with Anoxybacillus (71.84%), Megamonas (4.18%), Gallibacterium (2.14%), Unclassified Lachnospiraceae (1.87%), and Lactobacillus (1.80%) being the abundant genera. The alpha diversity increased from scalding to chilling, while the beta diversity revealed a significant separation of clusters at different processing points (p = 0.01). The alpha- and beta-diversity revealed significant contamination during the defeathering, with a redistribution of the bacteria during the chilling. This study concluded that the genetic diversity during the defeathering is strongly associated with the extent of the post-chill contamination, and may be used to indicate the microbial quality of the chicken meat.

Listeria monocytogenes Pathogenesis: The Role of Stress Adaptation.

Adaptive stress tolerance responses are the driving force behind the survival ability of Listeria monocytogenes in different environmental niches, within foods, and ultimately, the ability to cause human infections. Although the bacterial stress adaptive responses are primarily a necessity for survival in foods and the environment, some aspects of the stress responses are linked to bacterial pathogenesis. Food stress-induced adaptive tolerance responses to acid and osmotic stresses can protect the pathogen against similar stresses in the gastrointestinal tract (GIT) and, thus, directly aid its virulence potential. Moreover, once in the GIT, the reprogramming of gene expression from the stress survival-related genes to virulence-related genes allows L. monocytogenes to switch from an avirulent to a virulent state. This transition is controlled by two overlapping and interlinked transcriptional networks for general stress response (regulated by Sigma factor B, (SigB)) and virulence (regulated by the positive regulatory factor A (PrfA)). This review explores the current knowledge on the molecular basis of the connection between stress tolerance responses and the pathogenesis of L. monocytogenes. The review gives a detailed background on the currently known mechanisms of pathogenesis and stress adaptation. Furthermore, the paper looks at the current literature and theories on the overlaps and connections between the regulatory networks for SigB and PrfA.

Systematic-review and meta-analysis on effect of decontamination interventions on prevalence and concentration of Campylobacter spp. during primary processing of broiler chickens.

Scientific advances in pathogen decontamination offer great potential to reduce Campylobacter spp. during primary processing. The aim of this study was to collate data from eligible studies using systematic review, meta-analysis followed by meta-regression. Random effect meta-analysis revealed heterogenous (τ2 = 0.6, I2 = 98 %) pooled reduction in Campylobacter concentration of 0.6 log10 CFU/carcass and a decrease in relative risk of Campylobacter spp. prevalence in broiler carcasses by 57.2 %. Decontamination interventions during Inside-Outside-Carcass-Wash were most effective on concentration (0.8 log10 CFU/carcass) while those during evisceration were most effective on prevalence (78.0 % decrease in relative risk). Physical decontamination was more effective on Campylobacter prevalence (68.7 % decrease in relative risk) compared chemical treatment (30.3 %). Application through immersion was superior on Campylobacter concentration (0.9 log10 CFU/carcass odds reduction) to spraying (0.5 log10 CFU/carcass odds reduction). Publication bias and small study effect were observed in trials on Campylobacter prevalence but not for concentration. The meta-regression revealed four and seven potential modifier variables for concentration and prevalence respectively. This meta-analysis provides an overview of the expected magnitude in Campylobacter spp. concentration and prevalence with application of decontamination interventions on broiler carcasses along the slaughter process and forms a basis of quantitative microbial risk assessment and derivation of intervention measures. Even though modest microbial concentration reduction is reported there was a large decrease in contamination prevalence during processing interventions.

Effects of vacuum packaging storage of minimally processed cassava roots at various temperatures on microflora, tissue structure, starch extraction by wet milling and granule quality.

BACKGROUND: Vacuum package storage is commonly applied to reduce postharvest deterioration in minimally processed cassava roots. However, the influence of vacuum packaging conditions on root end-use quality is poorly understood. Hence, the effects of vacuum packaged storage at ambient, refrigerated and freezing temperatures on microflora, cassava tissue structure and starch extraction by wet milling were studied. RESULTS: Vacuum packaged storage temperature strongly affected cassava root quality. Minimal adverse effects were obtained with frozen storage. With refrigerated storage, there was negligible microbial growth but some disruption of the parenchyma cell wall structure suggestive of chilling injury. With ambient temperature storage, there was considerable Lactobacilli dominated fermentation. This caused substantial cell degradation, probably due to the production of extracellular cellulolytic and other cell wall degrading enzymes. A benefit of this cell wall breakdown was that it substantially improved starch extraction with wet milling from the stored cassava pieces; by 18% with pieces that had been ambient vacuum packaged and wet milled using a 2000 µm opening screen. However, ambient temperature storage resulted in some starch granule pitting due to the action of extracellular amylases from the fermenting microorganisms. CONCLUSION: The best vacuum packaging storage conditions for minimally processed cassava depends on application and cost. For short-term storage, refrigeration would be best for vegetable-type products. For several months storage, freezing is best. For wet milling applications, this could be combined with subsequent short-term ambient temperature storage as it improves starch extraction efficiency and could reduce distribution energy costs. © 2021 Society of Chemical Industry.

Ultraviolet-C inactivation and hydrophobicity of Bacillus subtilis and Bacillus velezensis spores isolated from extended shelf-life milk.

Bacterial spores are important in food processing due to their ubiquity, resistance to high temperature and chemical inactivation. This work aims to study the effect of ultraviolet C (UVC) on the spores of Bacillus subtilis and Bacillus velezensis at a molecular and individual level to guide in deciding on the right parameters that must be applied during the processing of liquid foods. The spores were treated with UVC using phosphate buffer saline (PBS) as a suspension medium and their lethality rate was determined for each sample. Purified spore samples of B. velezensis and B. subtilis were treated under one pass in a UVC reactor to inactivate the spores. The resistance pattern of the spores to UVC treatment was determined using dipicolinic acid (Ca-DPA) band of spectral analysis obtained from Raman spectroscopy. Flow cytometry analysis was also done to determine the effect of the UVC treatment on the spore samples at the molecular level. Samples were processed for SEM and the percentage spore surface hydrophobicity was also determined using the Microbial Adhesion to Hydrocarbon (MATH) assay to predict the adhesion strength to a stainless-steel surface. The result shows the maximum lethality rate to be 6.5 for B. subtilis strain SRCM103689 (B47) and highest percentage hydrophobicity was 54.9% from the sample B. velezensis strain LPL-K103 (B44). The difference in surface hydrophobicity for all isolates was statistically significant (P < 0.05). Flow cytometry analysis of UVC treated spore suspensions clarifies them further into sub-populations unaccounted for by plate counting on growth media. The Raman spectroscopy identified B4002 as the isolate possessing the highest concentration of Ca-DPA. The study justifies the critical role of Ca-DPA in spore resistance and the possible sub-populations after UVC treatment that may affect product shelf-life and safety. UVC shows a promising application in the inactivation of resistant spores though there is a need to understand the effects at the molecular level to design the best parameters during processing.

Invited review: Probiotic yogurt quality criteria, regulatory framework, clinical evidence, and analytical aspects.

Yogurt is a milk-based product manufactured by lactic acid fermentation enabled by symbiotic yogurt cultures. Yogurt is largely considered to be a health product, and it is employed to deliver probiotics and prebiotics to the consumer. However, not all yogurts are probiotic, neither are they all functional products. There is increasing demand for health-promoting beverages, which is prompting the dairy industry to develop functional probiotic yogurts to meet the demand. However, there seems to be a scarcity of reviews providing critical information on regulatory frameworks in regions of the world, clinical trial outcomes, and methodological approaches for enumerating multiprobiotic strains in yogurt. This review, relating to functional probiotic yogurt, covers the newest information on the topic for the period mostly between 2014 and 2019. Conformance to regulations is paramount and hence, global regulatory frameworks for probiotic yogurt and prebiotic and nonprebiotic ingredients included in yogurt are reviewed. The paper emphasizes the need for convincing clinical trial outcomes that provide the dairy industry with an opportunity to market products with substantiated beneficial claims. The paper also discusses probiotic strains in functional yogurt, which is required to have population levels above the recommended therapeutic minimum during shelf life. The multiprobiotic species added to yogurt may present challenges relating to methodological and analytical approaches needed to determine viability of each strain contained in such yogurt. Hence, the review also presents the pros and cons of the culture-dependent and culture-independent approaches for the enumeration of probiotic cells in yogurt. The review is arguably valuable to the dairy industry, functional food developers, related scientists, and researchers, as well as policy makers.

Comparative Genome Analysis of Bacillus sporothermodurans with Its Closest Phylogenetic Neighbor, Bacillus oleronius, and Bacillus cereus and Bacillus subtilis Groups.

Bacillus sporothermodurans currently possesses one of the most highly heat-resistant spores (HRS), which can withstand ultra-high temperature (UHT) processing. Determination of multiple whole genome sequences of B. sporothermodurans provided an opportunity to perform the first comparative genome analysis between strains and with B. oleronius, B. cereus, and B. subtilis groups. In this study, five whole genome sequences of B. sporothermodurans strains, including those belonging to the HRS clone (SAD and BR12) normally isolated from UHT milk, were compared with the aforementioned Bacillus species for gene clusters responsible for heat resistance. In the phylogenomic analysis, B. sporothermodurans, with its closest phylogenetic neighbor, B. oleronius, clustered with B. thermoamylovorans and B. thermotolerans. Heat shock proteins GrpE, GroES, GroEL, and DnaK presented identical sequences for all B. sporothermodurans strains, indicating that differences in functional efficiency are not involved in the thermal resistance variations. However, comparing all species evaluated, B. sporothermodurans exhibited a different gene configuration in the chromosomal region of the heat shock protein GrpE. Furthermore, only B. sporothermodurans strains presented the stage II sporulation protein P gene located in this region. Multisequence alignment and phylogenetic analysis of the ClpB protein showed differences for HRS and non-HRS strains. The study identified ClpC, ClpE, and ClpX as the three ATPases putatively involved in protein disaggregation in B. sporothermodurans. Bacillussporothermodurans exhibits high homology with other Bacillus species in the DnaK, DnaJ, GroEL, and GroES cluster of genes involved in heat resistance. The data presented here pave the way to select and evaluate the phenotypic effects of genes putatively involved in heat resistance.

Consumers' perceptions of intrinsic and extrinsic attributes as indicators of safety and quality of chicken meat: Actionable information for public health authorities and the chicken industry.

Understanding consumers' perceptions toward chicken meat safety and quality could provide valuable information to public health educators since it is the most consumed meat. This study explores perceptions of a group of South African consumers on the safety and quality of chicken meat based on intrinsic and extrinsic attributes and identifies related safety risks. Data were collected through a web-based survey (863 participants). A substantial proportion of consumers considered supermarkets as the most trusted outlets to sell safe and good quality chicken (compared with butcheries, wholesalers, farmers' markets, street vendors, or "other retailers"). The majority of respondents (53%) most trusted refrigerated chicken to be of good quality compared with 36% trusting frozen chicken or 11% chicken at room temperature. Frozen chicken was considered most safe by 48% of consumers while 43% regarded refrigerated chicken as most safe. At point of purchase and home, smell, use-by date, sell-by date, and color were perceived as important attributes when judging chicken safety and quality. Consumers considered the absence of brine use and growth-promoting hormones in chicken feed as relatively important. The majority of consumers can be classified as highly involved during purchasing. It is essential that consumers apply safe chicken handling practices from point of purchase to consumption, irrespective of the type of retailer, perceived sensory characteristics, and date labels to reduce or eliminate microbial risks. Addressing consumer's knowledge and expectations regarding factors such as growth-promoting hormones and free range may improve safety and quality perceptions. PRACTICAL APPLICATION: This study gives insight into perceptions of a group of South African consumers toward safety and quality of chicken meat. Understanding consumers' perceptions can provide valuable information to public health educators since chicken meat is a common vehicle for Salmonella spp. and Campylobacter spp., which are human pathogens. Additionally, this information can assist the chicken industry to meet consumer expectations.

Solid-State Fermentation of Cassava Roots Using Cellulolytic-Type Alkaliphilic Bacillus spp. Cultures to Modify the Cell Walls and Assist Starch Release.

To improve cassava starch extraction by wet milling, solid-state fermentation of ground roots using cellulolytic-type alkaliphilic Bacilli spp., Bacillus akibai, B. cellulosilyticus and B. hemicellulosilyticus was investigated. Enzyme assay and scanning electron microscopy indicated that Bacillus spp. production of extracellular cellulase and polygalacturonase caused the formation of micropores through the root parenchyma cell walls and exposed the embedded cellulosic network. Gas chromatography data of the cell wall constituent sugars remaining after fermentation and Fourier transform infrared data indicated that the Bacillus treatments reduced the levels of pectin and, hemicellulose and to lesser extent cellulose. Wide-angle X-ray scattering data indicated that the Bacillus spp. cell wall degrading enzymes had partially hydrolysed the amorphous fractions of the cell wall polysaccharides. All the Bacillus spp. treatments improved starch extraction by 17-23% compared to fermentation with endogenous microflora. B. cellulosilyticus was most effective in disintegration of large root particles and as result, released marginally the most starch, probably due to it having the highest cellulase activity. Solid-state fermentation using cellulolytic-type Bacillus spp. is, therefore, promising to technology to improve the efficiency of cassava wet milling cell wall disintegration and consequent starch yield without use of commercial cell wall degrading enzymes or polluting chemicals.

Dysbiosis Signatures of Fecal Microbiota in South African Infants with Respiratory, Gastrointestinal, and Other Diseases.

OBJECTIVE: To determine the association between the fecal microbiota diversity of the infants with different disease conditions, and vitamin A supplementation, antibiotic, and deworming therapies. STUDY DESIGN: In this case-control study, the bacterial community variations and the potential pathogens were identified through 16S ribosomal RNA gene-based amplicon sequencing and quantitative insights into microbial ecology pipeline in fecal samples. The participants were South African infants (mean age, 16 ± 8 months; 17 male and 17 female) hospitalized and diagnosed with gastrointestinal, respiratory, and other diseases. RESULTS: The top phyla of the infants with respiratory disease were Proteobacteria, followed by Firmicutes, which were equally abundant in gastrointestinal disease. A significant difference in Shannon (alpha) diversity index (95% CI, 2.6-4.4; P = .008), among the microbiota of the fecal samples categorized by disease conditions, was observed. In beta diversity analysis of fecal microbiota, remarkable variations were found within the groups of deworming therapy (95% CI, 0.40-0.90; P = .033), disease conditions (95% CI, 0.44-0.86; P < .012) through unweighted and antibiotic therapy (95% CI, 0.20-0.75; P = .007), vitamin A intake (95% CI, 0.10-0.80; P < .033) and disease conditions (95% CI, 0.10-0.79; P = .006) through weighted UniFrac distances. The candidate pathogen associated with the disease groups were identified through analysis of the composition of microbiomes analysis. CONCLUSIONS: This study provides preliminary evidence for the fecal microbiome-derived dysbiosis signature and pathobiome concept that may be observed in young children during illness.

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.

Genome Sequences of Bacillus sporothermodurans Strains Isolated from Ultra-High-Temperature Milk.

Here, we report the draft genome sequences of 3 Bacillus sporothermodurans strains isolated from ultra-high-temperature milk products in South Africa and Brazil and the type strain MB 581 (DSM 10599). The genomes will provide valuable information on the molecular dynamics of heat resistance in B sporothermodurans.

Persistence of foodborne diarrheagenic Escherichia coli in the agricultural and food production environment: Implications for food safety and public health.

Diarrheagenic Escherichia coli (DEC) is a leading cause of foodborne illness associated with intestinal disease. While known over the years that contamination of food sources occurs via the oral faecal-route, the mechanisms underlying its persistence within the open environments including the food chain remains virtually unknown. Therefore, in this mini-review we will shed light on bacterial processes such as initial attachment, biofilm formation, horizontal gene transfer and response to environmental stresses. These factors may enable persistence of DEC as well as the emergence of potentially more virulent strains within the agricultural and food production environment. Mechanistic studies in clinical microbiology and immunology have elucidated infection pathways in the human and other animal bodies leading to diagnostic and treatment solutions. Therefore, understanding DEC behaviour in the agricultural and food production environment is crucial for ensuring food safety and public health by reducing the burden of foodborne illnesses.

Visualisation and quantification of fumonisins bound by lactic acid bacteria isolates from traditional African maize-based fermented cereals, ogi and mahewu.

Consumption of fumonisin-contaminated foods has a negative influence on the health of humans (carcinogen; oesophageal cancer in Eastern Cape in South Africa). Lactic acid bacteria (LAB) have emerged as a promising natural detoxification agent against mycotoxins. The aim of this study was to visualise the interaction between fumonisins (FB1 and FB2) and LAB: Lactobacillus plantarum FS2, L. delbrueckii subsp. delbrueckii CIP 57.8T and Pediococcus pentosaceus D39, isolated from traditional fermented maize-based products (ogi and mahewu) using confocal laser scanning microscopy (CLSM) and to then quantify the LAB-bound fumonisin using high performance liquid chromatography (HPLC). The objective was to obtain a physically visible and quantifiable binding interaction between fumonisins and LAB strains with the aim of utilising LAB as a possible detoxifying agent. Fumonisins were derivatised using naphthalene-2,3-dicarboxaldehyde (NDA) and then combined with non-fluorescent LAB cells (viable and non-viable). For the quantification of bound fumonisins, viable and non-viable cells were incubated in the presence of predetermined concentrations of fumonisins and the level of fumonisin in the suspension was determined. CLSM showed the derivatised green fluorescent fumonisins binding to the surface of each of the LAB cells. For viable cells, L. plantarum FS2 bound FB1 most effectively while P. pentosaceus D39 bound the least level of FB1. The highest levels of FB2 were bound by L. plantarum R 1096 and the least by L. delbrueckii CIP 57.8 T. For non-viable cells, L. plantarum FS2 was also the most effective for binding both fumonisins with P. pentosaceus D39 and L. delbrueckii CIP 57.8 T being the least effective for FB1 and FB2, respectively. To our knowledge, this is the first study to visualise the interaction between LAB and fumonisins. We demonstrate that LAB isolates from indigenous fermented maize-based beverages bind fumonisins and thus present a potential strategy for their reduction in these traditional foods.

Insights into the role of bacteria in vitamin A biosynthesis: Future research opportunities.

Significant efforts have been made to address the hidden hunger challenges due to iron, zinc, iodine, and vitamin A since the beginning of the 21st century. Prioritizing the vitamin A deficiency (VAD) disorders, many countries are looking for viable alternative strategies such as biofortification. One of the leading causes of VAD is the poor bioconversion of ß-carotene into retinoids. This review is focused on the opportunities of bacterial biosynthesis of retinoids, in particular, through the gut microbiota. The proposed hypothesis starts with the premise that an animal can able to store and timely convert carotenoids into retinoids in the liver and intestinal tissues. This theory is experimental with many scientific insights. The syntrophic metabolism, potential crosstalk of bile acids, lipocalins and lipopolysaccharides of gut microbiota are reported to contribute significantly to the retinoid biosynthesis. The gut bacteria respond to these kinds of factors by genetic restructuring driven mainly by events like horizontal gene transfer. A phylogenetic analysis of ß-carotene 15, 15'-mono (di) oxygenase enzymes among a selected group of prokaryotes and eukaryotes was carried out to validate the hypotheses. Shedding light on the probiotic strategies through non-genetically modified organism such as gut bacteria capable of synthesizing vitamin A would address the VAD disorders.

Short communication: Source tracking Bacillus cereus in an extended-shelf-life milk processing plant using partial sequencing of rpoB and multilocus sequence typing.

We used rpoB partial sequencing and multilocus sequence typing (MLST) to characterize 7 Bacillus cereus strains obtained at the following points: ESL milk during shelf life, pasteurized milk, raw milk, and filler nozzles after cleaning in place. The objective of the study was to determine relatedness among B. cereus isolates from several sampling points along an ESL processing plant with the aim of source tracking. The study revealed that isolates from filler nozzles shared 100% similarity with isolates from ESL milk and raw milk using rpoB sequencing. It also revealed that isolates from pasteurized milk shared 100% similarities with isolates from filler nozzles and ESL milk using MLST. We suggest 3 routes of B. cereus contamination in ESL milk. We showed that B. cereus contamination of ESL milk might be through raw milk and biofilms from filler nozzles. In addition, rpoB partial sequencing and MLST can be used as tools for source tracking in ESL milk processing.

Enteroaggregative Escherichia coli is the predominant diarrheagenic E. coli pathotype among irrigation water and food sources in South Africa.

Diarrheagenic E. coli (DEC) has been implicated in foodborne outbreaks worldwide and have been associated with childhood stunting in the absence of diarrhoea. Infection is extraordinarily common, but the routes of transmission have not been determined. Therefore, determining the most prevalent pathotypes in food and environmental sources may help provide better guidance to various stakeholders in ensuring food safety and public health and advancing understanding of the epidemiology of enteric disease. We characterized 205 E. coli strains previously isolated from producer distributor bulk milk (PDBM)(118), irrigation water (48), irrigated lettuce (29) and street vendor coleslaw (10) in South Africa. Enteropathogenic E. coli (EPEC), enterotoxigenic E. coli (ETEC), enteroaggregative E. coli (EAEC) and diffusely adherent E. coli (DAEC) were sought. We used PCR and partial gene sequencing for all 205 strains while 46 out of 205 that showed poor resolution were subsequently characterized using cell adherence (HeLa cells). PCR and partial gene sequencing of aatA and/or aaiC genes confirmed EAEC (2%, 5 out of 205) as the only pathotype. Phylogenetic analysis of sequenced EAEC strains with E. coli strains in GenBank showing ≥80% nucleotide sequence similarity based on possession of aaiC and aatA generated distinct clusters of strains separated predominantly based on their source of isolation (food source or human stool) suggesting a potential role of virulence genes in source tracking. EAEC 24%, 11 out of 46 strains (PDBM = 15%, irrigation water = 7%, irrigated lettuce = 2%) was similarly the predominant pathotype followed by strains showing invasiveness to HeLa cells, 4%, 2 out of 46 (PDBM = 2%, irrigated lettuce = 2%), among stains characterized using cell adherence. Therefore, EAEC may be the leading cause of DEC associated food and water-borne enteric infection in South Africa. Additionally, solely using molecular based methods targeting virulence gene determinants may underestimate prevalence, especially among heterogeneous pathogens such as EAEC.