Comparative Detection and Quantification of Arcobacter butzleri in Stools from Diarrheic and Nondiarrheic People in Southwestern Alberta, Canada

Abstract

dArcobacter butzleri has been linked to enteric disease in humans, but its pathogenicity and epidemiology remain poorly understood. The lack of suitable detection methods is a major limitation. Using comparative genome analysis, we developed PCR primers for direct detection and quantification of A. butzleri DNA in microbiologically complex matrices. These primers, along with existing molecular and culture-based methods, were used to detect A. butzleri and enteric pathogens in stools of diarrheic and nondiarrheic people (n ‫؍‬ 1,596) living in southwestern Alberta, Canada, from May to November 2008. In addition, quantitative PCR was used to compare A. butzleri densities in diarrheic and nondiarrheic stools. Arcobacter butzleri was detected more often by PCR (59.6%) than by isolation methods (0.8%). Comparison by PCR-based detection found no difference in the prevalence of A. butzleri between diarrheic (56.7%) and nondiarrheic (45.5%) individuals. Rates of detection in diarrheic stools peaked in June (71.1%) and October (68.7%), but there was no statistically significant correlation between the presence of A. butzleri and patient age, sex, or place of habitation. Densities of A. butzleri DNA in diarrheic stools (1.6 ؎ 0.59 log 10 copies mg ؊1 ) were higher (P ‫؍‬ 0.007) than in nondiarrheic stools (1.3 ؎ 0.63 log 10 copies mg ؊1 ). Of the 892 diarrheic samples that were positive for A. butzleri, 74.1% were not positive for other bacterial and/or viral pathogens. The current study supports previous work suggesting that A. butzleri pathogenicity is strain specific and/or dependent on other factors, such as the level of host resistance. N early 1.7 billion cases of diarrheal disease are reported globally each year (1), although this is an underestimation of true rates of enteritis, as many afflicted individuals do not have access to or choose not to pursue medical assistance (2). For those seeking diagnosis, the majority of cases of acute enteritis are not linked to an identified etiological agent (3, 4). Ascertaining the etiology of enteric disease is essential for the development of effective therapeutics and preventative mitigation strategies. Direct contact with animals and ingestion of untreated water and/or undercooked animal products are recognized risk factors for acute enteritis (3), which suggests that a significant number of cases of enteritis are caused by unidentified biotic pathogens of human or zoonotic origin. Critical components of the epidemiology of arcobacteriosis and the population structure of Arcobacter butzleri have yet to be resolved, in large part because effective culture and/or molecular-based detection methods for this bacterium have yet to be developed.Arcobacter butzleri is ubiquitous in the environment (e.g., river water contaminated with human and/or nonhuman animal feces) (5-7). That the bacterium is detected in such a variety of sources suggests that pathways for transmission among animals and environmental sources exist, but accurate source tracking of A. butzleri is hampered by a ...

Occurrence of putative virulence genes onArcobacter butzleriisolated from three different environmental sites throughout the dairy chain

“…; Webb et al . ). Completion of the A. butzleri RM4018 genome sequence identified several putative virulence genes, some of them presenting homology to Campylobacter jejuni virulence determinants (Miller et al .…”

Arcobacter: an emerging food-borne zoonotic pathogen, its public health concerns and advances in diagnosis and control – a comprehensive review

“…A multiplex-PCR assay with seven primers was developed by Douidah et al (2010) for the identification of the five human and mammal related species of Arcobacter (A. butzleri, A. cryaerophilus, A. skirrowii, A. thereius and A. cibarius). Recently, Webb et al (2016) developed the PCR primers for direct detection and quantification of A. butzleri DNA in microbiologically complex matrices. Densities of A. butzleri DNA in diarrheic stools were higher when compared to non-diarrheic stools in southwestern Alberta, Canada.…”

“…have gained attention of worldwide researchers as an important emerging food-borne enteropathogen. Among all Arcobacters, A. butzleri is the most common species and has been associated with human disease, such as enteritis, severe diarrhea, bacteremia and septicemia (Engberg et al 2000;Lau et al 2002;Collado et al 2014;Figueras et al 2014;Van den Abeele et al 2014;Webb et al 2016). Knowledge gap in the field of emergence and evolution of Arcobacters is very wide as several themes and questions remained unsolved.…”

“…For this purpose, further strengthening of the immunodiagnostics, molecular detection and characterization, genotyping and differentiation of Arcobacters is suggested by employing advances in PCR and its allied versions, LAMP, recombinant diagnostics, lateral flow assay, biochips, biosensors, microarrays, gene sequencing and phylogenetic analysis, genomic fingerprinting and nanotechnology-based diagnostics (Schmitt & Henderson 2005;Deb & Chakraborty 2012;Dhama et al 2012Dhama et al , 2014aNum & Useh 2013;Gilbert et al 2014;Ramees et al 2014d;Webb et al 2015Webb et al , 2016Whiteduck-L eveill ee et al 2016a). …”

Housing Gnotobiotic Mice in Conventional Animal Facilities

“…Genomic DNA from E. coli (ATCC 25922) possesses seven copies of the 16S rRNA gene. The use of an internal amplification control is recommended to guard against false negative results, e.g., due to PCR inhibition (Webb et al, 2016).…”