Aspergillus flavus growth inhibition by Lactobacillus strains isolated from traditional fermented Kenyan milk and maize products.

Certain strains of lactic acid bacteria have been reported to inhibit fungal growth and may so be potential as biocontrol agents. In this study, 171 LAB strains were isolated from traditional fermented Kenyan milk and maize products and tested against aflatoxin-producing A. flavus fungi. The three LAB strains showing highest antifungal activity were identified as Lactobacillus plantarum. None of the strains were able to completely inhibit fungal growth under conditions favorable for fungi and suboptimal for LAB. These conditions probably reduced the growth and metabolic activity of some LAB isolates, as several growth-related aspects like production of antifungal biomolecules and other metabolites contribute to the inhibiting activity. The results suggest that certain LAB strains could be employed in food to control the growth of aflatoxigenic fungi. Further studies to establish the efficacy of the potential LAB strains in fermented products are in progress.

Dynamics of microbial communities in untreated and autoclaved food waste anaerobic digesters.

This study describes the microbial community richness and dynamics of two semi-continuously stirred biogas reactors during a time-course study of 120 days. The reactors were fed with untreated and autoclaved (160 °C, 6.2 bar) food waste. The microbial community was analysed using a bacteria- and archaea-targeting 16S rRNA gene-based Terminal-Restriction Fragment Length Polymorphism (T-RFLP) approach. Compared with the archaeal community, the structures and functions of the bacterial community were found to be more complex and diverse. With the principal coordinates analysis it was possible to separate both microbial communities with 75 and 50% difference for bacteria and archaea, respectively, in the two reactors fed with the same waste but with different pretreatment. Despite the use of the same feeding material, anaerobic reactors showed a distinct community profile which could explain the differences in methane yield (2-17%). The community composition was highly dynamic for bacteria and archaea during the entire studied period. This study illustrates that microbial communities are dependent on feeding material and that correlations among specific bacterial and archaeal T-RFs can be established.

Comparative study of sugar fermentation and protein expression patterns of two Lactobacillus plantarum strains grown in three different media.

A comparative study of two strains of Lactobacillus plantarum (REB1 and MLBPL1) grown in commercial medium (MRS broth), cucumber juice, and liquid pig feed was performed to explore changes to the metabolic pathways of these bacteria, using a proteomics approach (two-dimensional electrophoresis and liquid chromatography-tandem mass spectrometry) combined with analyses of fermentable sugars and fermentation end products. The protein expression showed that even with an excess of glucose in all media, both strains could metabolize different carbohydrates simultaneously and that hexoses could also be used via a phosphoketolase pathway with preferential expression in liquid feed. Sugar analyses showed that the fermentation of sugars was homolactic for all media, with some heterolactic activity in liquid feed, as shown by the production of acetate. Cucumber juice (the medium with the highest glucose content) showed the lowest hexose consumption (10%), followed by liquid feed (33%) and MRS broth (50%). However, bacterial growth was significantly higher in cucumber juice and liquid feed than in MRS broth. This discrepancy was due to the growth benefit obtained from the utilization of the malate present in cucumber juice and liquid feed. Despite different growth conditions, the synthesis of essential cellular components and the stress response of the bacteria were unaffected. This study has improved our understanding of the mechanisms involved in the growth performance of an appropriate lactic acid bacterium strain to be used for food and feed fermentation, information that is of crucial importance to obtain a high-quality fermented product.

Potential of lactic acid bacteria in aflatoxin risk mitigation.

Aflatoxins (AF) are ubiquitous mycotoxins contaminating food and feed. Consumption of contaminated food and feed can cause a severe health risk to humans and animals. A novel biological method could reduce the health risks of aflatoxins through inhibiting mold growth and binding aflatoxins. Lactic acid bacteria (LAB) are commonly used in fermented food production. LAB are known to inhibit mold growth and, to some extent, to bind aflatoxins in different matrices. Reduced mold growth and aflatoxin production may be caused by competition for nutrients between bacterial cells and fungi. Most likely, binding of aflatoxins depends on environmental conditions and is strain-specific. Killed bacteria cells possess consistently better binding abilities for aflatoxin B1 (AFB1) than viable cells. Lactobacilli especially are relatively well studied and provide noticeable possibilities in binding of aflatoxin B1 and M1 in food. It seems that binding is reversible and that bound aflatoxins are released later on (Haskard et al., 2001; Peltonen et al., 2001). This literature review suggests that novel biological methods, such as lactic acid bacteria, show potential in mitigating toxic effects of aflatoxins in food and feed.

Construction of a Stable alpha-Galactosidase-Producing Baker's Yeast Strain.

Molasses is widely used as a substrate for commercial yeast production. The complete hydrolysis of raffinose, which is present in beet molasses, by Saccharomyces strains requires the secretion of alpha-galactosidase, in addition to the secretion of invertase. Raffinose is not completely utilized by commercially available yeast strains used for baking, which are Mel. In this study we integrated the yeast MEL1 gene, which codes for alpha-galactosidase, into a commercial mel baker's yeast strain. The Mel phenotype of the new strain was stable. The MEL1 gene was expressed when the new Mel baker's yeast was grown in molasses medium under conditions similar to those used for baker's yeast production at commercial factories. The alpha-galactosidase produced by this novel baker's yeast strain hydrolyzed all the melibiose that normally accumulates in the growth medium. As a consequence, additional carbohydrate was available to the yeasts for growth. The new strain also produced considerably more alpha-galactosidase than did a wild-type Mel strain and may prove useful for commercial production of alpha-galactosidase.

Molecular characterization of spoilage bacteria as a means to observe the microbiological quality of carrot.

This study characterized the bacteria causing decay of carrots during storage and marketing. Spoilage strains were identified by 16S-amplified rDNA restriction analysis and intergenic transcribed spacer-PCR-restriction fragment length polymorphism (ITS-PCR-RFLP). Genotypic fingerprinting by RFLP-pulsed-field gel electrophoresis was used to assess the genetic diversity of the isolates. A total of 252 Pseudomonas isolates from carrots were identified and classified into eight separate groups. Most strains belonged to group A (Pseudomonas fluorescens, Pseudomonas marginalis, and Pseudomonas veronii) and group B (Pseudomonas putida). The strains identified as Pectobacterium carotovorum subsp. carotovorum, Pectobacterium atrosepticum, Dickeya chrysanthemi, and Erwinia rhapontici were distinguished by ITS-PCR-RFLP. All isolates belonging to the genera Pectobacterium and Erwinia were responsible for carrot spoilage. This work has led to the development of new strategies for the identification and genotyping of vegetable-spoiling strains of Pseudomonas, Pectobacterium, and Erwinia. This is also the first report describing the occurrence of carrot-spoiling E. rhapontici. Early recognition of spoilage bacteria in vegetables is important for the implementation of effective handling strategies. Pectolytic bacteria may cause considerable financial losses because they account for a large proportion of bacterial rot of fruits and vegetables during storage, transit, and marketing.

Toxigenic potential of Aspergillus species occurring on maize kernels from two agro-ecological zones in Kenya.

Two agro-ecological zones in Kenya were selected to compare the distribution in maize of Aspergillus spp. and their toxigenicity. These were Nandi County, which is the main maize growing region in the country but where no human aflatoxicoses have been reported, and Makueni County where most of the aflatoxicosis cases have occurred. Two hundred and fifty-five households were sampled in Nandi and 258 in Makueni, and Aspergillus was isolated from maize. Aspergillus flavus and A. parasiticus isolates were tested for the presence of aflD and aflQ genes. Positive strains were induced to produce aflatoxins on yeast extract sucrose and quantified using liquid chromatography-tandem mass spectrometry (LCMSMS). Aspergillus flavus was the most common contaminant, and the incidence of occurrence in Nandi and Makueni was not significantly different (82.33% and 73.26%, respectively). Toxigenic strains were more prevalent than non-toxigenic strains. All the toxigenic strains from Makueni were of the S-type while those from Nandi belonged to the l-type. Quantitative differences in aflatoxin production in vitro between isolates and between strains were detected with S strains producing relatively larger amounts of total aflatoxins, B toxins and lower values for G toxins. This was in accord with the frequent aflatoxicosis outbreaks in Makueni. However some L strains produced considerable amounts of B toxins. Given the widespread distribution of toxigenic strains in both regions, the risk of aflatoxin poisoning is high when favorable conditions for toxin production occur.

Determination of aspartase activity in dairy Propionibacterium strains.

Propionic acid bacteria (PAB) are important as starter cultures for the dairy industry in the manufacture of Swiss-type cheeses, in which they are involved in the formation of eyes and are responsible for the typical flavour and aroma. These characteristics are mainly due to the classical propionic acid fermentation, but also the conversion of aspartate to fumarate and ammonia by the enzyme aspartase and the subsequent reduction of fumarate to succinate, which occur in dairy Propionibacterium freudenreichii ssp. shermanii and ssp. freudenreichii starter strains. Additionally, the metabolism of free amino acids may be partly responsible for secondary fermentation and the subsequent split defects in cheese matrix. Here a method for aspartase activity was established and a number of dairy propionibacteria belonging to P. freudenreichii ssp. shermanii and freudenreichii were screened for this enzyme activity. A wide range of aspartase activity could be found in PAB isolates originating from cheese. The majority, i.e. 70% of the 100 isolates tested, showed very low levels of aspartate activity.