Curt Doetkott

The Genome Sequence of Avian Pathogenic Escherichia coli Strain O1:K1:H7 Shares Strong Similarities with Human Extraintestinal Pathogenic E. coli Genomes

Escherichia coli strains that cause disease outside the intestine are known as extraintestinal pathogenic E. coli (ExPEC) and include human uropathogenic E. coli (UPEC) and avian pathogenic E. coli (APEC). Regardless of host of origin, ExPEC strains share many traits. It has been suggested that these commonalities may enable APEC to cause disease in humans. Here, we begin to test the hypothesis that certain APEC strains possess potential to cause human urinary tract infection through virulence genotyping of 1,000 APEC and UPEC strains, generation of the first complete genomic sequence of an APEC (APEC O1:K1:H7) strain, and comparison of this genome to all available human ExPEC genomic sequences. The genomes of APEC O1 and three human UPEC strains were found to be remarkably similar, with only 4.5% of APEC O1s genome not found in other sequenced ExPEC genomes. Also, use of multilocus sequence typing showed that some of the sequenced human ExPEC strains were more like APEC O1 than other human ExPEC strains. This work provides evidence that at least some human and avian ExPEC strains are highly similar to one another, and it supports the possibility that a food-borne link between some APEC and UPEC strains exists. Future studies are necessary to assess the ability of APEC to overcome the hurdles necessary for such a food-borne transmission, and epidemiological studies are required to confirm that such a phenomenon actually occurs.

Plasmid replicon typing of commensal and pathogenic Escherichia coli isolates.

ABSTRACT Despite the critical role of plasmids in horizontal gene transfer, few studies have characterized plasmid relatedness among different bacterial populations. Recently, a multiplex PCR replicon typing protocol was developed for classification of plasmids occurring in members of the Enterobacteriaceae. Here, a simplified version of this replicon typing procedure which requires only three multiplex panels to identify 18 plasmid replicons is described. This method was used to screen 1,015 Escherichia coli isolates of avian, human, and poultry meat origin for plasmid replicon types. Additionally, the isolates were assessed for their content of several colicin-associated genes. Overall, a high degree of plasmid variability was observed, with 221 different profiles occurring among the 1,015 isolates examined. IncFIB plasmids were the most common type identified, regardless of the source type of E. coli. IncFIB plasmids occurred significantly more often in avian pathogenic E. coli (APEC) and retail poultry E. coli (RPEC) than in uropathogenic E. coli (UPEC) and avian and human fecal commensal E. coli isolates (AFEC and HFEC, respectively). APEC and RPEC were also significantly more likely than UPEC, HFEC, and AFEC to possess the colicin-associated genes cvaC, cbi, and/or cma in conjunction with one or more plasmid replicons. The results suggest that E. coli isolates contaminating retail poultry are notably similar to APEC with regard to plasmid profiles, with both generally containing multiple plasmid replicon types in conjunction with colicin-related genes. In contrast, UPEC and human and avian commensal E. coli isolates generally lack the plasmid replicons and colicin-related genes seen in APEC and RPEC, suggesting limited dissemination of such plasmids among these bacterial populations.

Comparison of extraintestinal pathogenic Escherichia coli strains from human and avian sources reveals a mixed subset representing potential zoonotic pathogens

ABSTRACT Since extraintestinal pathogenic Escherichia coli (ExPEC) strains from human and avian hosts encounter similar challenges in establishing infection in extraintestinal locations, they may share similar contents of virulence genes and capacities to cause disease. In the present study, 1,074 ExPEC isolates were classified by phylogenetic group and possession of 67 other traits, including virulence-associated genes and plasmid replicon types. These ExPEC isolates included 452 avian pathogenic E. coli strains from avian colibacillosis, 91 neonatal meningitis E. coli (NMEC) strains causing human neonatal meningitis, and 531 uropathogenic E. coli strains from human urinary tract infections. Cluster analysis of the data revealed that most members of each subpathotype represent a genetically distinct group and have distinguishing characteristics. However, a genotyping cluster containing 108 ExPEC isolates was identified, heavily mixed with regard to subpathotype, in which there was substantial trait overlap. Many of the isolates within this cluster belonged to the O1, O2, or O18 serogroup. Also, 58% belonged to the ST95 multilocus sequence typing group, and over 90% of them were assigned to the B2 phylogenetic group typical of human ExPEC strains. This cluster contained strains with a high number of both chromosome- and plasmid-associated ExPEC genes. Further characterization of this ExPEC subset with zoonotic potential urges future studies exploring the potential for the transmission of certain ExPEC strains between humans and animals. Also, the widespread occurrence of plasmids among NMEC strains and members of the mixed cluster suggests that plasmid-mediated virulence in these pathotypes warrants further attention.

Identification of Minimal Predictors of Avian Pathogenic Escherichia coli Virulence for Use as a Rapid Diagnostic Tool

ABSTRACT To identify traits that predict avian pathogenic Escherichia coli (APEC) virulence, 124 avian E. coli isolates of known pathogenicity and serogroup were subjected to virulence genotyping and phylogenetic typing. The results were analyzed by multiple-correspondence analysis. From this analysis, five genes carried by plasmids were identified as being the most significantly associated with highly pathogenic APEC strains: iutA, hlyF, iss, iroN, and ompT. A multiplex PCR panel targeting these five genes was used to screen a collection of 994 avian E. coli isolates. APEC isolates were clearly distinguished from the avian fecal E. coli isolates by their possession of these genes, suggesting that this pentaplex panel has diagnostic applications and underscoring the close association between avian E. coli virulence and the possession of ColV plasmids. Also, the sharp demarcation between APEC isolates and avian fecal E. coli isolates in their plasmid-associated virulence gene content suggests that APEC isolates are well equipped for a pathogenic lifestyle, which is contrary to the widely held belief that most APEC isolates are opportunistic pathogens. Regardless, APEC isolates remain an important problem for poultry producers and a potential concern for public health professionals, as growing evidence suggests a possible role for APEC in human disease. Thus, the pentaplex panel described here may be useful in detecting APEC-like strains occurring in poultry production, along the food chain, and in human disease. This panel may be helpful toward clarifying potential roles of APEC in human disease, ascertaining the source of APEC in animal outbreaks, and identifying effective targets of avian colibacillosis control.

First Year Seminars Produce Long-Term Impact.

In an effort to provide insight into a strategy for increasing student retention, students enrolled in a first year seminar were paired with a matched comparison group, and retention of the 1,853 students in the two groups was compared over a period of four years. Results indicated significantly greater retention over a period of four years for students enrolled in the seminar. Review of the literature indicates that while such seminars are gaining in popularity, longitudinal studies of their effectiveness using matched comparison groups are lacking. In addition to expanding the research base of the first year seminars, this article also presents the background of the seminar and studies regarding the first year seminar that are relevant to retention.

Complement resistance-related traits among Escherichia coli isolates from apparently healthy birds and birds with colibacillosis.

In this study, 294 Escherichia coli isolates from birds with colibacillosis were collected from disease outbreaks throughout the United States and were compared with 75 fecal E. coli isolates of apparently healthy chickens by their possession of several purported virulence genes, resistance to rough-lipopolysaccharide-specific bacteriophages (rLPSr), and elaboration of capsule. Traits were selected for study on the basis of their association with complement resistance. The genes targeted in this study included those encoding colicin V (cvaC) and the outer membrane proteins TraT (traT), OmpA (ompA), and Iss (iss). No significant differences were found between the two groups of isolates in the occurrence of cvaC-, traT-, or ompA-homologous sequences or in rLPSr. Only a few isolates were encapsulated, and the isolates of healthy birds were significantly more likely to be encapsulated than were the isolates of sick birds. However, iss, whether detected through hybridization or amplification, was found in more of the disease-associated isolates than in those of healthy birds. This difference was highly significant. Further, iss sequences were widely distributed among isolates of different serotypes from various avian host species and sites within these hosts. Such results suggest that possession of the iss sequence by an avian E. coli isolate may be a good indicator of that isolates potential to cause disease. This association warrants further study because iss and the protein it encodes may be useful targets of future colibacillosis control efforts.

Effect of extrusion process parameters on the quality of buckwheat flour mixes

ABSTRACT A response surface analysis using a second-order central composite design was used to study the effect of extrusion process parameters on the extrudate quality of three blends containing buckwheat flour. The extrudates were prepared as three blends. Blend 1 was a 55:40:5 (w/w) mix of light buckwheat flour, wheat flour, and nonfat dry milk (NFDM). Blend 2 was a 40:55:5 mix of light buckwheat flour, corn meal, and NFDM. Blend 3 was a 30:60:10 mix of light buckwheat flour, corn meal, and NFDM. The blends were processed in a twin-screw extruder with factorial combinations of the parameters including: process temperatures of 95–150°C, dough moisture of 15–22%, and screw speeds of 260–390 rpm. The linear components alone significantly explained most of the variation of expansion index, bulk density, water absorption, and breaking strength. The greatest amount of variability was explained by process temperature for blend 1. Dough moisture accounted for the greatest amount of variation for blends 2 and 3...

Examination of the Source and Extended Virulence Genotypes of Escherichia coli Contaminating Retail Poultry Meat

Extraintestinal pathogenic Escherichia coli (ExPEC) are major players in human urinary tract infections, neonatal bacterial meningitis, and sepsis. Recently, it has been suggested that there might be a zoonotic component to these infections. To determine whether the E. coli contaminating retail poultry are possible extraintestinal pathogens, and to ascertain the source of these contaminants, they were assessed for their genetic similarities to E. coli incriminated in colibacillosis (avian pathogenic E. coli [APEC]), E. coli isolated from multiple locations of apparently healthy birds at slaughter, and human ExPEC. It was anticipated that the retail poultry isolates would most closely resemble avian fecal E. coli since only apparently healthy birds are slaughtered, and fecal contamination of carcasses is the presumed source of meat contamination. Surprisingly, this supposition proved incorrect, as the retail poultry isolates exhibited gene profiles more similar to APEC than to fecal isolates. These isolates contained a number of ExPEC-associated genes, including those associated with ColV virulence plasmids, and many belonged to the B2 phylogenetic group, known to be virulent in human hosts. Additionally, E. coli isolated from the crops and gizzards of apparently healthy birds at slaughter also contained a higher proportion of ExPEC-associated genes than did the avian fecal isolates examined. Such similarities suggest that the widely held beliefs about the sources of poultry contamination may need to be reassessed. Also, the presence of ExPEC-like clones on retail poultry meat means that we cannot yet rule out poultry as a source of ExPEC human disease.

Biofilm formation by avian Escherichia coli in relation to media, source and phylogeny.

Aims:  To assess the abilities of 105 avian pathogenic Escherichia coli (APEC) and 103 avian faecal commensal E. coli (AFEC) to form biofilms on a plastic surface and to investigate the possible association of biofilm formation with the phylotype of these isolates.

SPOILAGE IDENTIFICATION OF BEEF USING AN ELECTRONIC NOSE SYSTEM

A commercially available Cyranose-320. conducting polymer-based electronic nose system was used to analyze the volatile organic compounds emanating from fresh beef strip loins (M. Longisimmus lumborum) stored at 4°C and 10°C. Two statistical techniques, i.e., linear discriminant analysis (LDA) and quadratic discriminant analysis (QDA), were used to develop classification models from the collected sensor signals. The performances of the developed models were validated by two different methods: leave-1-out cross-validation, and bootstrapping. The developed models classified meat samples based on the microbial population into “unspoiled” (microbial counts 6.0 log10 cfu/g). Overall, quadratic discriminant-based classification models performed better than linear discriminant analysis based models. For the meat samples stored at 10°C, the highest classification accuracies obtained by the LDA method with leave-1-out and bootstrapping validations were 87.10% and 85.87%, respectively. On the other hand, classification by QDA and subsequent validation by leave-1-out and bootstrapping provided highest accuracies of 87.5% and 97.38%, respectively. For samples stored at 4°C, the LDA method provided highest classification accuracies of 79.17% and 85.64% using leave-1-out and bootstrapping validation, respectively. When the QDA method was used, the highest classification accuracies obtained for the samples stored at 4°C were 87.50% and 98.48%, respectively, with leave-1-out and bootstrapping validations.