HPLC based method for the measurement of the reduction of aflatoxin B<Subscript>1</Subscript> by bacterial cultures isolated from different African foods

The consumption of fermented foods contaminated with aflatoxin B₁ is linked to aflatoxicosis. Aflatoxicosis is a serious problem in developing countries with environmental conditions appropriate for the biosynthesis of AFB₁ byAspergillus flavus andAspergillus parasiticus. In Africa, especially in Ghana and Nigeria, there is a very high risk of liver cancer which is caused by the consumption of AFB₁-intoxicated, traditionally fermented maize and sorghum products. It is suggested that one way to diminish this health risk might be the reduction of the AFB₁ concentration in foods by bacteria. Especially bacteria used for food fermentation processes are of great importance, with a special emphasis on lactic acid bacteria which are involved in traditionally fermented African foods based on maize and sorghum.Most publications dealing with aflatoxin degradation by microorganisms describe a phosphate buffer test system for the performance of degradation experiments. In contrast to that, a test system based on physiological active bacterial and yeast cells has been developed, to assess food fermentation organisms for their ability to reduce the AFB₁ concentration in vitro. The aflatoxin B₁ concentration in test samples was quatitatively determined by HPLC.The assessment of lactic acid bacteria originating from different German and other European culture collections only showed a very slight reduction of the AFB₁ concentration from 3% to 12%. Screening experiments in which other bacterial genera and lactic acid bacteria, isolated from different African foods have been assessed, in most cases showed the same results. However, some bacterial strains, e.g. strains of the genusBacillus derived from European culture collections and strains of the genusLactobacillus isolated from African foods, caused a release of AFB₁ which was chemically bound before to components of the test medium and which therefore could not be extracted with chloroform.A process quite similar to that may happen during food fermentations. Different experiments showed that e.g. cellulose can bind AFB₁ very effectively. Cellulose and different other food components are well known to absorb AFB₁. During fermentation the cellulose and other AFB₁-absorbing components may be degraded and the AFB₁ will be released again.The only bacterial strain known as yet which is able to reduce the AFB₁ concentration in vitro and in different food comodities isNocardia corynebacteroides (formerFlavobacterium aurantiacum). Nevertheless the mechanism of this AFB₁ reduction is actually not well understood, it still has to be investigated. In the meantime several other bacterial strains, presumably from the taxonomic group of theActinomycetes could be proved to be effective reducers of the AFB₁ concentration in our in vitro test system. Because as yet no food relevant microorganism could be found, which is able to degrade AFB₁, these new strains in general offer the possibility for a genetic modification of food relevant microorganisms. This seems to be the way to come to starter cultures which are able to degrade AFB₁ during food fermentations.     

Effects of sex and dietary phosphorus level on the apparent metabolizable energy and nutrient digestibility in broiler chickens

The influence of sex of broilers and dietary phosphorus (P) level on energy utilization and apparent ileal digestibility of N, P and phytate-P were investigated. The AME_N was determined using 3- and 6-week old broilers, while the apparent ileal digestibility were determined only with 6-week old birds. Sex of broilers had no effect on the AME_N values determined during week 3. During week 6, the AME_N values for male broilers were higher (P?0.01) than those for the females. An interaction (P?0.05) between sex and dietary P level was also observed. AME_N values determined with male broilers were lower in the adequate-P diet compared to those in the low-P diet, whereas no effect of P level was observed in females. Female broilers tended (P?0.10) to have a higher ileal N digestibility than the males, but a significant (P?0.01) sex ×?P level interaction was observed. Males receiving the adequate-P diet had a lower N digestibility than those receiving the low-P diet, whereas the opposite was true in the females. Ileal phytate P degradation in males was higher than in females (0.282 vs. 0.234), but the differences were not significant. Increasing dietary P level increased ileal P digestibility in both the males and females, but the improvements were greater in the females, as indicated by a significant sex × P level interaction.     

Duodenal mucosal risk markers in patients with familial adenomatous polyposis: effects of celecoxib/ursodeoxycholic acid co-treatment and comparison with patient controls

Background

Familial adenomatous polyposis (FAP) is a disease characterized by the development of hundreds to thousands of adenomatous polyps in the colorectum early in life. Virtually all patients with FAP will develop colorectal cancer before the age of 40 to 50 years, unless prophylactic colectomy is performed, which significantly improves their prognosis. The mortality pattern has changed and duodenal cancer now is one of the main cancer-related causes of death in these patients. Practically all patients with FAP develop premalignant duodenal adenomas, which may develop to duodenal cancer in approximately 3-7% of patients. Duodenal cancer in patients with FAP has a poor prognosis. The clinical challenge is to identify patients at high-risk for duodenal carcinoma. Chemoprevention would be desirable to avoid duodenectomy. The main goal of this study is to identify risk markers in normal duodenal mucosa of patients with FAP, that could help identify patients at increased risk for malignant transformation.

Methods

Messenger RNA (mRNA) levels of glutathione S-transferase A1 (GSTA1), glutathione S-transferase P1 (GSTP1), KIAA1199, E-cadherin, peroxisome proliferative activated receptor δ (PPARδ), caspase-3, cyclin D1, β-catenin, and cyclooxygenase-2 (COX-2) were measured in duodenal mucosa, using the QuantiGene 2.0 Plex assay. Levels in normal appearing mucosa of patients with FAP (n?=?37) were compared with levels in non-FAP patient controls (n?=?16). In addition, levels before and after treatment with either celecoxib & ursodeoxycholic acid (UDCA, n?=?14) or celecoxib & placebo (n?=?13) were evaluated in patients with FAP.

Results

mRNA levels of glutathione S-transferase A1 (28.16% vs. 38.24%, p?=?0.008) and caspase-3 (3.30% vs. 5.31%, p?=?0.001) were significantly lower in patients with FAP vs. non-FAP patient controls, respectively. COX-2 mRNA levels in normal duodenal mucosa of patients with FAP were found to be unexpectedly low. None of the potential risk markers was influenced by celecoxib or celecoxib & UDCA.

Conclusions

Protection against toxins and carcinogens (GSTA1) and apoptosis (caspase-3) is low in patients with FAP, which could contribute to increased susceptibility for malignant transformation of duodenal mucosa.

Trial registration

http://ClinicalTrials.gov number NCT00808743

     

The rate of synonymous substitution in enterobacterial genes is inversely related to codon usage bias

Genes sequences from Escherichia coli, Salmonella typhimurium, and other members of the Enterobacteriaceae show a negative correlation between the degree of synonymous-codon usage bias and the rate of nucleotide substitution at synonymous sites. In particular, very highly expressed genes have very biased codon usage and accumulate synonymous substitutions very slowly. In contrast, there is little correlation between the degree of codon bias and the rate of protein evolution. It is concluded that both the rate of synonymous substitution and the degree of codon usage bias largely reflect the intensity of selection at the translational level. Because of the high variability among genes in rates of synonymous substitution, separate molecular clocks of synonymous substitution might be required for different genes.      

Testing constraints on rRNA bases that make nonsequence-specific contacts with the codon-anticodon complex in the ribosomal A site

During protein synthesis, interactions between the decoding center of the ribosome and the codon-anticodon complexes maintain translation accuracy. Correct aminoacyl-tRNAs induce the ribosome to shift into a "closed" conformation that both blocks tRNA dissociation and accelerates the process of tRNA acceptance. As part of the ribosomal recognition of cognate tRNAs, the rRNA nucleotides G530 and A1492 form a hydrogen-bonded pair that interacts with the middle position of the codon.anticodon complex and recognizes correct Watson-Crick base pairs. Exchanging these two nucleotides (A530 and G1492) would not disrupt these interactions, suggesting that such a double mutant ribosome might properly recognize tRNAs and support viability. We find, however, that exchange mutants retain little ribosomal activity. We suggest that even though the exchanged nucleotides might function properly during tRNA recruitment, they might disrupt one or more other functions of the nucleotides during other stages of protein synthesis.     

Morphologic,cytogenetic, and enzymatic variation in tissue culture regenerated plants of apomictic old-world bluestem grasses (Bothriochloa sp.)

Somaclonal variant plants may be of use in broadening the germplasm base of plant species and providing useful stocks for cytogenetic investigations. This study was conducted to compare morphologic, cytogenetic and enzymatic characteristics of 21 R₁ (initial regenerate) bluestem,Bothriochloa sp., plants, visibly identified in a field-grown population of 522 plants as probable variants, with their respective R₀ (explant donor) progenitor. An R₂ seedling population was grown to ascertain the transmission of the variant R₁ phenotypes. All R₁ plants differed from their respective R₀ progenitors in one or more morphological characters. Foliage colour was the most pronounced difference in most cases. Four of the plants, three of which were dwarfed, produced no inflorescences. The R₁ plants tended to be shorter than R₀ progenitors and had corresponding decreases in lengths on inflorescences and lowest racemes. All R₁ plants of accessions 8911C and 8793 had an increase in chromosome number from2n=4x=40 to2n=5x=50. Three dwarfed R₁ plants, derived from accession 8873B, were aneuploids, two having2n=48 chromosomes and the third being a probable mixoploid with 55–58 chromosomes. Other plants of accession 8873B had the R₀ chromosome number. Fertility, as estimated by pollen stainability and seed set, generally was reduced in R₁ plants relative to the R₀. This reduction was not drastic, however, with all flowering plants having 45% or higher seed set. Apomixis apparently maintained fertility in all R₁ plants, including those with a pentaploid chromosome number. All R₁ plants differed from their respective R₀ plants in peroxidase and esterase banding patterns. All R₁ plants of accessions 8911C, and 8793, respectively, had identical peroxidase and esterase bands. For both enzyme systems two banding patterns were present in R₁ plants of accession 8873B, with 12 of 13 plants exhibiting common patterns. Examination of R₂ progeny plants confirmed the genetic transmission of the variant phenotypes and, by virtue of uniformity, indicated apomictic reproduction in the R₁ plants. The results demonstrate the production of potentially useful genetic and cytogenetic variant plants via tissue culture in these apomictic species.     

Genbank accession #: AJ237597

Plant Molecular Biology -     

Retention of cryptic genes in microbial populations

Cryptic genes are silenced genes that can still be reactivated by mutation. Since they can make no positive contribution to the fitness of their carriers, it is not clear why many cryptic genes in microbial populations have not degenerated into useless DNA sequences. Hall et al. (1983) have suggested that cryptic genes have persisted because of occasional strong environmental selection for reactivated genes. The present mathematical study supports their suggestion. It shows that a cryptic gene can be retained without having any selective advantage over a useless DNA sequence, if selection for the reactivated gene occasionally occurs for a substantially long time.