Phylogenetic identification of bacterial MazF toxin protein motifs among probiotic strains and foodborne pathogens and potential implications of engineered probiotic intervention in food

BACKGROUND: Toxin-antitoxin (TA) systems are commonly found in bacteria and Archaea, and it is the most common mechanism involved in bacterial programmed cell death or apoptosis. Recently, MazF, the toxin component of the toxin-antitoxin module, has been categorized as an endoribonuclease, or it may have a function similar to that of a RNA interference enzyme. RESULTS: In this paper, with comparative data and phylogenetic analyses, we are able to identify several potential MazF-conserved motifs in limited subsets of foodborne pathogens and probiotic strains and further provide a molecular basis for the development of engineered/synthetic probiotic strains for the mitigation of foodborne illnesses. Our findings also show that some probiotic strains, as fit as many bacterial foodborne pathogens, can be genetically categorized into three major groups based on phylogenetic analysis of MazF. In each group, potential functional motifs are conserved in phylogenetically distant species, including foodborne pathogens and probiotic strains. CONCLUSION: These data provide important knowledge for the identification and computational prediction of functional motifs related to programmed cell death. Potential implications of these findings include the use of engineered probiotic interventions in food or use of a natural probiotic cocktail with specificity for controlling targeted foodborne pathogens

Development of a Reporter System for In Vivo Monitoring of gamma-Secretase Activity in Drosophila

The gamma-secretase complex represents an evolutionarily conserved family of transmembrane aspartyl proteases that cleave numerous type-I membrane proteins, including the beta-amyloid precursor protein (APP) and the receptor Notch. All known rare mutations in APP and the gamma-secretase catalytic component, presenilin, which lead to increased amyloid betapeptide production, are responsible for early-onset familial Alzheimer\u27s disease. beta-amyloid protein precursor-like (APPL) is the Drosophila ortholog of human APP. Here, we created Notch- and APPL-based Drosophila reporter systems for in vivo monitoring of gamma-secretase activity. Ectopic expression of the Notch- and APPL-based chimeric reporters in wings results in vein truncation phenotypes. Reporter-mediated vein truncation phenotypes are enhanced by the Notch gain-of-function allele and suppressed by RNAi-mediated knockdown of presenilin. Furthermore, we find that apoptosis partly contributes to the vein truncation phenotypes of the APPL-based reporter, but not to the vein truncation phenotypes of the Notch-based reporter. Taken together, these results suggest that both in vivo reporter systems provide a powerful genetic tool to identify genes that modulate gamma-secretase activity and/or APPL metabolism

Isolation of Escherichia coli O157:H7 from Intact Colon Fecal Samples of Swine1

Escherichia coli O157:H7 was recovered from colon fecal samples of pigs. Polymerase chain reaction confirmed two genotypes: isolates harboring the eaeA, stx1, and stx2 genes and isolates harboring the eaeA, stx1, and hly933 genes. We demonstrate that swine in the United States can harbor potentially pathogenic E. coli O157:H7

Distribution and Genetic Profiles of Campylobacter in Commercial Broiler Production from Breeder to Slaughter in Thailand

Poultry and poultry products are commonly considered as the major vehicle of Campylobacter infection in humans worldwide. To reduce the number of human cases, the epidemiology of Campylobacter in poultry must be better understood. Therefore, the objective of the present study was to determine the distribution and genetic relatedness of Campylobacter in the Thai chicken production industry. During June to October 2012, entire broiler production processes (i.e., breeder flock, hatchery, broiler farm and slaughterhouse) of five broiler production chains were investigated chronologically. Representative isolates of C. jejuni from each production stage were characterized by flaA SVR sequencing and multilocus sequence typing (MLST). Amongst 311 selected isolates, 29 flaA SVR alleles and 17 sequence types (STs) were identified. The common clonal complexes (CCs) found in this study were CC-45, CC-353, CC-354 and CC-574. C. jejuni isolated from breeders were distantly related to those isolated from broilers and chicken carcasses, while C. jejuni isolates from the slaughterhouse environment and meat products were similar to those isolated from broiler flocks. Genotypic identification of C. jejuni in slaughterhouses indicated that broilers were the main source of Campylobacter contamination of chicken meat during processing. To effectively reduce Campylobacter in poultry meat products, control and prevention strategies should be aimed at both farm and slaughterhouse levels

Emerging Approaches for Typing, Detection, Characterization, and Traceback of Escherichia coli

Pathogenic Escherichia coli strains cause a large number of diseases in humans, including diarrhea, hemorrhagic colitis, hemolytic uremic syndrome, urinary tract infections, and neonatal meningitis, while in animals they cause diseases such as calf scours and mastitis in cattle, post-weaning diarrhea and edema disease in pigs, and peritonitis and airsacculitis in chickens. The different E. coli pathotypes are characterized by the presence of specific sets of virulence-related genes. Therefore, it is not surprising that pathogenic E. coli constitutes a genetically heterogeneous family of bacteria, and they are continuing to evolve. Rapid and accurate molecular methods are critically needed to detect and trace pathogenic E. coli in food and animals. They are also needed for epidemiological investigations to enhance food safety, as well as animal and human health and to minimize the size and geographical extent of outbreaks. The serotype of E. coli strains has traditionally been determined using antisera raised against the >180 different O- (somatic) and 53 H- (flagellar) antigens. However, there are many problems associated with serotyping, including: it is labor-intensive and time consuming; cross reactivity of the antisera with different serogroups occurs; antisera are available only in specialized laboratories; and many strains are non-typeable. Molecular serotyping targeting O-group-specific genes within the E. coli O-antigen gene clusters and genes that are involved in encoding for the different flagellar types offers an improved approach for determining the E. coli O- and H-groups. Furthermore, molecular serotyping can be coupled with determination of specific sets of virulence genes carried by the strain offering the possibility to determine O-group, pathotype, and the pathogenic potential simultaneously. Sequencing of the O-antigen gene clusters of all of the known O-groups of E. coli is now complete, and the sequences have been deposited in the GenBank database. The sequence information has revealed that some E. coli serogroups have identical sequences while others have point mutations or insertion sequences and type as different serogroups in serological reactions. There are also a number of other ambiguities in serotyping that need to be resolved. Furthermore, new E. coli O-groups are being identified. Therefore, there is an essential need to resolve these issues and to revise the E. coli serotype nomenclature based on these findings. There are emerging technologies that can potentially be applied for molecular serotyping and detection and characterization of E. coli. On a related topic, the genome sequence of thousands of E. coli strains have been deposited in GenBank, and this information is revealing unique markers such as CRISPR (clustered regularly interspaced short palindromic repeats) and virulence gene markers that could be used to identify E. coli pathotypes. Whole genome sequencing now provides the opportunity to study the role of horizontal gene transfer in the evolution and emergence of pathogenic E. coli strains. Whole genome sequencing approaches are being investigated for genotyping and outbreak investigation for regulatory and public health needs; however, there is a need for establishing bioinformatics pipelines able to handle large amounts of data as we move toward the use of genetic approaches for non-culture-based detection and characterization of E. coli and for outbreak investigations