Atypical Helicobacter canadensis strains associated with swine.
ABSTRACT: Forty-two Helicobacter isolates were isolated from swine feces in The Netherlands and Denmark. All 12 isolates sequenced (16S rRNA gene) formed a robust clade with Helicobacter canadensis ( approximately 99% similarity). Species-specific PCR indicated that all of the isolates were H. canadensis isolates. Although the appearance of the porcine isolates was similar to the appearance of H. canadensis, only one of these isolates was able to hydrolyze indoxyl acetate, a cardinal characteristic of this taxon. Examination of the 23S rRNA and hsp60 genes revealed high levels of similarity between the porcine isolates and H. canadensis. However, amplified fragment length polymorphism genomic typing showed that isolates recovered from swine feces were genetically distinct from H. canadensis strains obtained from humans and geese.
Project description:We recently analyzed 11 helicobacter isolates cultured from diarrhea patients in Canada. These isolates had been characterized biochemically by restriction fragment length polymorphism (RFLP; AluI, HhaI) analysis and by fatty-acid analysis as Helicobacter pullorum. However, four of the isolates differed biochemically from H. pullorum by their inability to hydrolyze indoxyl acetate and their resistance to nalidixic acid. Using complete 16S rRNA analysis, we determined that these four strains clustered near H. pullorum but had a sequence difference of 2% and therefore represent a novel helicobacter, Helicobacter canadensis. This novel helicobacter could also be distinguished from H. pullorum by RFLP analysis using ApaLI. The number of novel Helicobacter spp. associated with gastrointestinal disease in humans and animals is rapidly increasing. There are now six Helicobacter spp. isolated from diarrheic humans, the other five being H. pullorum, H. canis, "H. rappini," H. fennelliae, and H. cinaedi. This finding highlights the importance of careful molecular analysis in addition to standard biochemical tests in identifying the increasing number of Helicobacter spp. isolated from humans and animals.
Project description:Numbers of nonmigratory Canada geese have increased substantially in the past decade, and they have become a nuisance in some urban areas. Because of their close contact with humans in parks and areas adjacent to surface waterways, contact with their feces poses a zoonotic risk. A total of 97 geese from 10 separate geographic locales in the greater Boston area had their feces sampled for detection of Helicobacter spp. Identification of Helicobacter spp. based on 16S rRNA genus-specific helicobacter primers was noted in 39 of 97 (40.2%) DNA fecal extracts. Twenty-seven (27.8%) of these geese had helicobacters isolated from their feces. A urease-positive novel species, Helicobacter anseris, based on phenotypic, biochemical, and 16S rRNA analyses, was isolated from 20 geese from seven different flocks. A second, novel, urease-negative Helicobacter sp., H. brantae, was identified in seven geese. Four geese had both novel Helicobacter spp. cultured from their feces. Whether these two novel helicobacters pose a zoonotic risk, similar to other enteric helicobacters (e.g., H. canadensis, previously isolated from diarrheic and bacteremic humans and from geese in Europe), will require further studies.
Project description:We determined the genome sequence of the type strain of Helicobacter canadensis, an emerging human pathogen with diverse animal reservoirs. Potential virulence determinants carried by the genome include systems for N-linked glycosylation and capsular export. A protein-based phylogenetic analysis places H. canadensis close to Wolinella succinogenes.
Project description:UNLABELLED:Since May 2013, outbreaks of porcine epidemic diarrhea have devastated the U.S. swine industry, causing immense economic losses. Two different swine enteric coronaviruses (porcine epidemic diarrhea virus and Delta coronavirus) have been isolated from the affected swine population. The disease has been reported from at least 32 states of the United States and other countries, including Mexico, Peru, Dominican Republic, Canada, Columbia, Ecuador, and Ukraine, with repeated outbreaks in previously infected herds. Here we report the isolation and characterization of a novel mammalian orthoreovirus 3 (MRV3) from diarrheic feces of piglets from these outbreaks in three states and ring-dried swine blood meal from multiple sources. MRV3 could not be isolated from healthy or pigs that had recovered from epidemic diarrhea from four states. Several MRV3 isolates were obtained from chloroform-extracted pig feces or blood meal in cell cultures or developing chicken embryos. Biological characterization of two representative isolates revealed trypsin resistance and thermostability at 90°C. NextGen sequencing of ultrapurified viruses indicated a strong homology of the S1 segment to mammalian and bat MRV3. Neonatal piglets experimentally infected with these viruses or a chloroform extract of swine blood meal developed severe diarrhea and acute gastroenteritis with 100% mortality within 3 days postinfection. Therefore, the novel porcine MRV3 may contribute to enteric disease along with other swine enteric viruses. The role of MRV3 in the current outbreaks of porcine epidemic diarrhea in the United States remains to be determined, but the pathogenic nature of the virus warrants further investigations on its epidemiology and prevalence. IMPORTANCE:Porcine orthoreoviruses causing diarrhea have been reported in China and Korea but not in the United States. We have isolated and characterized two pathogenic reassortant MRV3 isolates from swine fecal samples from porcine epidemic diarrhea outbreaks and ring-dried swine blood meal in the United States. These fecal and blood meal isolates or a chloroform extract of blood meal induced severe diarrhea and mortality in experimentally infected neonatal pigs. Genetic and phylogenetic analyses of two MRV3 isolates revealed that they are identical but differed significantly from nonpathogenic mammalian orthoreoviruses circulating in the United States. The present study provides a platform for immediate development of suitable vaccines and diagnostics to prevent and control porcine orthoreovirus diarrhea.
Project description:During a 6-year period, 64 of 227 commercially reared cats had microaerobic bacteria isolated from their feces. All the isolates were initially identified as Campylobacter-like organisms based on biochemical and phenotypic characteristics. DNA extractions from 51 of these isolates were subjected to PCR using primers specific for Helicobacter spp. and Campylobacter spp. Of the isolates, 92% (47 of 51 isolates) were positive for Campylobacter spp., 41% (21 of 51 isolates) were positive for Helicobacter spp., 33% (17 of 51 isolates) were positive for both genera, 59% (30 of 51 isolates) were positive only for Campylobacter spp., and 8% (4 of 51) were positive only for Helicobacter spp. Sixteen of the 47 Campylobacter-positive cultures were positive for more than one Campylobacter spp. Based on a species-specific PCR assay, 83% of the isolates were identified as Campylobacter helveticus, 47% of the isolates were identified as Campylobacter upsaliensis, and 6% of the isolates were classified as Campylobacter jejuni. The 1.2-kb PCR products of the 16S rRNA genes of 19 Helicobacter species isolates were subjected to restriction fragment length polymorphism (RFLP) analysis. Of the five different RFLP patterns obtained, two clustered with Helicobacter ("Flexispira") taxon 8, one clustered with Helicobacter bilis, one clustered with Helicobacter canis, and the remaining pattern was closely related to a novel Helicobacter sp. strain isolated from a woodchuck. The sequence data for the 16S rRNA genes of 10 Helicobacter spp. validated the RFLP-based identification of these isolates. This study demonstrated that biochemical and phenotypic characteristics of microaerobic organisms in cat feces were insufficient to characterize mixed Helicobacter and Campylobacter infections. Molecular structure-based diagnostics using genus- and species-specific PCR, RFLP analysis, and 16S rRNA sequence analysis enabled the identification of multiple microaerobic species in individual animals. The clinical relevance of enteric Helicobacter and Campylobacter coinfection in cats will require further studies.
Project description:From 1997 to 1999 seven isolates of Campylobacter-like organisms from five patients that were exhibiting symptoms of gastroenteritis, including fever, stomach malaise, and diarrhea, were investigated. The organisms were isolated from stool samples and found to exhibit a diverse colony morphology; hence multiple isolates were submitted from one of the patients. All isolates were found to be identical. The organisms were catalase, urease, alkaline phosphatase, and nitrate negative but oxidase and indoxyl acetate positive. They grew at 37 degrees C but not at 42 degrees C, and three of the isolates from two different patients were sensitive to nalidixic acid and cephalothin. Full 16S rRNA sequence analysis not only grouped these organisms within the Helicobacter genus but also differentiated them from previously identified Helicobacter species. The closest relative by phylogenetic analysis was Helicobacter sp. flexispira taxon 1. Electron microscopy showed that these isolates had one or two bipolar flagella; however, the periplasmic fibers, a characteristic of the known Helicobacter sp. flexispira taxa, were not observed. The present isolates also lacked a flagellar sheath, a trait shared with four other Helicobacter spp., H. canadensis, H. mesocricetorum, H. pullorum, and H. rodentium. On the basis of the unique phenotypic properties of these isolates and 16S rRNA sequence analysis, we propose the classification of a new Helicobacter species, Helicobacter winghamensis sp. nov.
Project description:Swine are regarded as promising biomedical models, but the dynamics of their gastrointestinal microbiome have been much less investigated than that of humans or mice. The aim of this study was to establish an integrated multi-omics protocol to investigate the fecal microbiome of healthy swine. To this end, a preparation and analysis protocol including integrated sample preparation for meta-omics analyses of deep-frozen feces was developed. Subsequent data integration linked microbiome composition with function, and metabolic activity with protein inventories, i.e., 16S rRNA data and expressed proteins, and identified proteins with corresponding metabolites. 16S rRNA gene amplicon and metaproteomics analyses revealed a fecal microbiome dominated by <i>Prevotellaceae, Lactobacillaceae</i>, <i>Lachnospiraceae</i>, <i>Ruminococcaceae</i> and <i>Clostridiaceae.</i> Similar microbiome compositions in feces and colon, but not ileum samples, were observed, showing that feces can serve as minimal-invasive proxy for porcine colon microbiomes. Longitudinal dynamics in composition, e.g., temporal decreased abundance of <i>Lactobacillaceae</i> and <i>Streptococcaceae</i> during the experiment, were not reflected in microbiome function. Instead, metaproteomics and metabolomics showed a rather stable functional state, as evident from short-chain fatty acids (SCFA) profiles and associated metaproteome functions, pointing towards functional redundancy among microbiome constituents. In conclusion, our pipeline generates congruent data from different omics approaches on the taxonomy and functionality of the intestinal microbiome of swine.
Project description:Draft genomes of two strains of Escherichia coli, FP2 and FP3, isolated from the feces of the Canada goose (Branta canadensis), were sequenced. Genome sizes were 5.26?Mb with a predicted G+C content of 50.54% (FP2) and 5.07?Mb with a predicted G+C content of 50.41% (FP3).
Project description:<i>Mentha canadensis</i> is a well-known resource of traditional Chinese herbal medicine, belonging to the <i>Mentha</i> of the Labiatae family. In this study, the whole chloroplast genome of the <i>M. canadensis</i> chloroplast genome was sequenced, assembled and annotated, which contains 134 unique genes, including 89 protein-coding genes, 37 tRNA genes and 8 rRNA genes. A maximum likelihood phylogenetic tree based on 11 complete chloroplast genomes revealed that <i>M. canadensis</i> is closely related to <i>M. longifolia</i> and <i>M. spicata</i>. The chloroplast genome could be used for variety identification, genetic engineering and effective protection of germplasm resources.
Project description:The popular herbal remedy goldenseal (Hydrastis canadensis L.) is traditionally used to treat skin infections. With this study, we show activity of H. canadensis extracts in vitro against methicillin-resistant Staphylococcus aureus (MRSA). An extract from H. canadensis leaves demonstrated more potent antimicrobial activity than the alkaloid berberine alone (MICs of 75?µg/mL and 150?µg/mL, respectively). LC-MS detected alkaloids and efflux-pump inhibitory flavonoids in the extract, and the latter may explain the enhanced efficacy of the extract compared to berberine alone. We also show evidence of anti-virulence activity as a second mechanism by which H. canadensis acts against S. aureus. The H. canadensis leaf extract (but not the isolated alkaloids berberine, hydrastine, and canadine) demonstrated quorum quenching activity against several clinically relevant MRSA isolates (USA300 strains). Our data suggest that this occurs by attenuation of signal transduction through the AgrCA two-component system. Consistent with this observation, the extract inhibited toxin production by MRSA and prevented damage by MRSA to keratinocyte cells in vitro. Collectively, our results show that H. canadensis leaf extracts possess a mixture of constituents that act against MRSA via several different mechanisms. These findings lend support for the traditional application of crude H. canadensis extracts in the prevention of infection.