Detection of PCR products of the ipaH gene from Shigella and enteroinvasive Escherichia coli by enzyme linked immunosorbent assay

Sethabutr, Orntipa; Venkatesan, Malabi M.; Yam, Sylvia; Pang, Lorrin; Smoak, Bonnie L.; Sang, Willie; Echeverria, Peter; Taylor, David N.; Isenbarger, Daniel W.

doi:10.1016/s0732-8893(00)00122-x

Cited by 39 publications

(28 citation statements)

References 16 publications

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“…Analysis of these specimens by culture techniques after shipment would have been impossible. While clinical samples of larger size (for example, stool samples) are occasionally shipped long distances on ice for culture analysis, 36 the minute quantities of micro-organisms present in ocular surface scrapings would generally not survive long range shipment.…”

Section: Discussionmentioning

confidence: 99%

Polymerase chain reaction based detection of fungi in infected corneas

Gaudio

Gopinathan

Sangwan

et al. 2002

British Journal of Ophthalmology

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Aims: To evaluate a polymerase chain reaction (PCR) based assay to detect fungi in scrapings from infected corneas. Methods: A PCR assay was developed to amplify a portion of the fungal 18S ribosome gene. Corneal scrapings from 30 patients with presumed infectious keratitis were evaluated using this assay, as well as by standard microbiological techniques, and the results were compared. Conjunctival swabs from each patient's healthy, fellow eye were also evaluated by PCR. Results: PCR and fungal culture results matched (were both positive or both negative for fungi) in 22 (74%) of 30 scrapings from infected corneas. Three (10%) of 30 samples were PCR positive but fungal culture negative; two of these appeared clinically to represent fungal infections, and the third was clinically indeterminate. Four (13%) scrapings were positive by PCR but also by bacterial and not fungal culture. One specimen (3%) was PCR negative but fungal culture positive. Of the conjunctival swabs from each patient's healthy fellow eye, five (17%) of 30 were positive by PCR, and the opposite, infected eye of all five of these harboured a fungal infection. Conclusions: PCR is promising as a means to diagnose fungal keratitis and offers some advantages over culture methods, including rapid analysis and the ability to analyse specimens far from where they are collected.

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Section: Discussionmentioning

confidence: 99%

Polymerase chain reaction based detection of fungi in infected corneas

Gaudio

Gopinathan

Sangwan

et al. 2002

British Journal of Ophthalmology

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“…The amplified regions of rotavirus, norovirus, sapovirus, astrovirus, and adenovirus were located in conserved genomic regions (21)(22)(23)(24)(25), and these assays have been used in our diagnostic laboratory for several years. Bacterial PCRs were developed with guidance from available publications with respect to suitable target regions (26)(27)(28)(29)(30)(31)(32)(33)(34)(35), usually by adapting a traditional PCR method to real-time PCR (when this study was planned, suitable real-time PCR assays were lacking for most nonviral targets). Thus, established target regions were used, and primers and probes were designed with the aim of obtaining similar melting temperatures (ϳ58 to 60°C for primers and ϳ68 to 70°C for probes).…”

Section: Study Participants (I) Patientsmentioning

confidence: 99%

Real-Time PCR Threshold Cycle Cutoffs Help To Identify Agents Causing Acute Childhood Diarrhea in Zanzibar

et al. 2014

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f Molecular assays might improve the identification of causes of acute diarrheal disease but might lead to more frequent detection of asymptomatic infections. In the present study, real-time PCR targeting 14 pathogens was applied to rectal swabs from 330 children aged 2 to 59 months in Zanzibar, including 165 patients with acute diarrhea and 165 asymptomatic control subjects. At least one pathogen was detected for 94% of the patients and 84% of the controls, with higher rates among patients for norovirus genogroup II (20% versus 2.4%; P < 0.0001), rotavirus (10% versus 1.8%; P ‫؍‬ 0.003), and Cryptosporidium (30% versus 11%; P < 0.0001). Detection rates did not differ significantly for enterotoxigenic Escherichia coli (ETEC)-estA (33% versus 24%), ETEC-eltB (44% versus 46%), Shigella (35% versus 33%), and Campylobacter (35% versus 33%), but for these agents threshold cycle (C T ) values were lower (pathogen loads were higher) in sick children than in controls. In a multivariate analysis, C T values for norovirus genogroup II, rotavirus, Cryptosporidium, ETEC-estA, and Shigella were independently associated with diarrhea. We conclude that this real-time PCR allows convenient detection of essentially all diarrheagenic agents and provides C T values that may be critical for the interpretation of results for pathogens with similar detection rates in patients and controls. The results indicate that the assessment of pathogen loads may improve the identification of agents causing gastroenteritis in children.

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“…The amplified regions of rotavirus, norovirus and adenovirus were located to conserved genomic regions, [12][13][14][15] and these assays have been used in our diagnostic laboratory several years. The bacterial PCRs were developed by guidance from available publications as regards suitable target regions, [16][17][18][19][20][21][22][23][24][25] usually by adapting a traditional PCR method to real-time PCR. Thus, established target regions were used, and primers and probes were designed with the aim of obtaining similar melting temperatures (≈58-60°C for primers, ≈68-70°C for probes).…”

Section: Microbial Agents and Target Sequencesmentioning

confidence: 99%