Laboratory Detection of Clostridium difficile in Piglets in Australia

Knight, Daniel R.; Squire, Michele M.; Riley, Thomas V.

doi:10.1128/jcm.01225-14

Cited by 25 publications

(16 citation statements)

References 40 publications

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“…Despite reduced sensitivity compared to enrichment culture, C. difficile ChromID represents a viable and cost-effective option for detecting C. difficile in piglets, particularly in the Australian veterinary setting. It is relatively cheap, can give answers in 24 h, and, in our experience, performs significantly better than molecular-based methods for the detection of C. difficile in porcine feces (26).…”

Section: Discussionmentioning

confidence: 99%

Nationwide Surveillance Study of Clostridium difficile in Australian Neonatal Pigs Shows High Prevalence and Heterogeneity of PCR Ribotypes

Knight

Squire

Riley

2015

Appl Environ Microbiol

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Clostridium difficile is an important enteric pathogen of humans and the cause of diarrhea and enteritis in neonatal pigs. Outside Australia, prevalence in piglets can be up to 73%, with a single PCR ribotype (RT), 078, predominating. We investigated the prevalence and genotype of C. difficile in Australian pig herds. Rectal swabs (n ‫؍‬ 229) were collected from piglets aged <7 days from 21 farms across Australia. Selective culture for C. difficile was performed and isolates characterized by PCR for toxin genes and PCR ribotyping. C. difficile was isolated from 52% of samples by direct culture on chromogenic agar and 67% by enrichment culture (P ‫؍‬ 0.001). No association between C. difficile recovery or genotype and diarrheic status of either farm or piglets was found. The majority (87%; 130/154) of isolates were toxigenic. Typing revealed 23 different RTs, several of which are known to cause disease in humans, including RT014, which was isolated most commonly (23%; 36/154). RT078 was not detected. This study shows that colonization of Australian neonatal piglets with C. difficile is widespread in the herds sampled.

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Section: Discussionmentioning

confidence: 99%

Nationwide Surveillance Study of Clostridium difficile in Australian Neonatal Pigs Shows High Prevalence and Heterogeneity of PCR Ribotypes

Knight

Squire

Riley

2015

Appl Environ Microbiol

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“…In contrast to Taq DNA polymerase used in PCR, the Bst DNA polymerase is resistant to inhibitors present in crude biological samples (Kaneko et al, 2007). As four or six specific primers that recognize six or eight distinct sequences on the target DNA are required, LAMP has been shown to amplify target DNA with high specificity and is widely used in the clinical diagnosis of epidemic bacteria Knight et al, 2014), viruses (Poon et al, 2004;Curtis et al, 2014), parasites (Li et al, 2012;Ni et al, 2014) and fetal sex identification (Fu et al, 2011), as well as detection of the hypervirulent strain and toxin types of C. difficile (Kato et al, 2005;Norén et al, 2011;Boyanton et al, 2012). In the current study, five sets of LAMP primers were designed and the assay was optimized for detection of ermB.…”

Section: Introductionmentioning

confidence: 99%

Rapid detection of ermB gene in Clostridium difficile by loop-mediated isothermal amplification

Lin

Liu

Wang

et al. 2015

Journal of Medical Microbiology

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Macrolide-lincosamide-streptogramin B resistance in Clostridium difficile is mostly due to the ermB resistance determinant. Here, we describe a sensitive and rapid molecular method to detect ermB in C. difficile to contribute to the wider epidemiological study. Five sets of loop-mediated isothermal amplification (LAMP) primers were designed and optimized for rapid detection of ermB. The specificity and sensitivity of the primers for ermB were detected, and the ermB LAMP assay was compared to conventional PCR with 80 clinical isolates of C. difficile. Real-time monitoring of turbidity and chromogenic reaction were used to determine negative and positive results. A total of 26 pathogenic bacterial strains of different species were found to be negative for ermB, which indicated the high specificity of the primers. ermB was detected in 78.8 % (63/80) of the clinical isolates by both LAMP and conventional PCR. The detection limit of LAMP was 36.1 pg DNA ml 21 and its sensitivity was 10-fold greater than that of conventional PCR. This study is the first report regarding the development and application of the LAMP assay for detection of the ermB gene in C. difficile strains. The developed LAMP method is sensitive, specific and provides a user-friendly visual approach for the rapid detection of ermB-bearing C. difficile. Received 27 March 2015Accepted 13 June 2015 INTRODUCTIONClostridium difficile is a causative agent of antibiotic-associated diarrhoea and pseudomembranous colitis, usually as a consequence of antimicrobial therapy. During the last few decades, C. difficile infection (CDI) has become more severe, more recurrent, more resistant to antibiotics and more refractory to standard treatment, which poses a significant burden on patients, healthcare systems and society in general. Estimates suggest that the annual costs for management of CDI amount to more than $3 billion in the USA and J3 billion in Europe (Bouza, 2012;McGlone et al., 2012). Major risk factors for CDI include hospitalization, advanced age and antibiotic exposure, the latter including clindamycin and erythromycin (Aziz et al., 2001;Gerding, 1989;Kelly et al., 1994;Slimings & Riley, 2014). The majority of epidemic isolates are resistant to clindamycin, erythromycin, broad-spectrum cephalosporins and fluoroquinolones (Johnson et al., 1999; Spigaglia et al., 2011;Slimings & Riley, 2014). Thus, resistance to clindamycin and erythromycin, two antibiotics of the macrolidelincosamide-streptogramin B (MLSB) group, would further increase the risk of CDI.In C. difficile, resistance to the MLSB group of antibiotics is mainly encoded by the ermB resistance determinant. The ermB gene encodes a 23S rRNA methyltransferase that modifies the target site for the antibiotic and is located on a specific mobilizable non-conjugative element, Tn5398 (Farrow et al., 2001). However, the ermB gene is located on elements other than Tn5398 in the majority of clinical isolates. In total, 17 genetic organizations of such elements 3These authors contributed equally to this work...

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“…Targeted research using highly-discriminatory WGS is required to confirm this. One stumbling block to learning more about CDI in animals is that most diagnostic tests used for laboratory diagnosis of CDI in humans do not perform well in animals 42 . Further work is required to address this problem.…”

Section: Resultsmentioning

confidence: 99%

Community-acquired Clostridium difficile infection and Australian food animals

2015

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Clostridium difficile is an anaerobic Gram positive spore-forming bacterium, the leading cause of infectious diarrhoea (C. difficile infection; CDI) in hospitalised humans. The assumption that CDI is primarily a hospital-acquired infection is being questioned. Community-acquired CDI (CA-CDI) is increasing1 particularly in groups previously considered at low risk2,3. In Australia, CA-CDI rates doubled during 2011 and increased by 24% between 2011 and 20124. Two potentially high-risk practices in Australian food animal husbandry may present a risk for CA-CDI: slaughtering of neonatal animals for food, and effluent recycling to agriculture.

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Laboratory Detection of Clostridium difficile in Piglets in Australia

Cited by 25 publications

References 40 publications

Nationwide Surveillance Study of Clostridium difficile in Australian Neonatal Pigs Shows High Prevalence and Heterogeneity of PCR Ribotypes

Nationwide Surveillance Study of Clostridium difficile in Australian Neonatal Pigs Shows High Prevalence and Heterogeneity of PCR Ribotypes

Rapid detection of ermB gene in Clostridium difficile by loop-mediated isothermal amplification

Community-acquired Clostridium difficile infection and Australian food animals

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