scholarly journals Bayesian estimation of diagnostic accuracy of fecal culture and PCR-based tests for the detection of Salmonella enterica in California cull dairy cattle

PeerJ ◽  
2020 ◽  
Vol 8 ◽  
pp. e8310
Author(s):  
John M. Adaska ◽  
Pius S. Ekong ◽  
Kristin A. Clothier ◽  
Deniece R. Williams ◽  
Paul V. Rossitto ◽  
...  
Keyword(s):  
Sensitivity And Specificity ◽  
Diagnostic Testing ◽  
Relative Sensitivity ◽  
Epidemiological Studies ◽  
Culture Methods ◽  
Time Saving ◽  
Enrichment Broth ◽  
Fecal Samples ◽  
Pcr Screening ◽  
Relative Specificity

Epidemiological studies of low prevalence disease problems are often hindered by the high cost of diagnostic testing. The objective of this study was to evaluate PCR screening of both individual and pooled fecal samples from culled dairy cows for the invA gene of Salmonella followed by culture to determine if the sensitivity and specificity were comparable to the results from traditional culture methods applied to individual samples. Cows from six different dairies were sampled in all four seasons. A total of 240 individual cow fecal samples, 24 fecal pools and 24 pools of 24-hour tetrathionate enrichment broth were tested. Diagnostic sensitivity of PCR screening followed by culture of PCR positive or indeterminate samples (i.e PCR-CUL method) was lower than that of culture (CUL) when applied to individual fecal samples (94.8%, 99.5%), however the specificity was comparable (99.6% and 97.7% respectively). For pools of five fecal samples and pools of five, 24 h tetrathionate broth samples, the specificity of both tests were comparable (∼98%); however, their sensitivity was only comparable in pooled fecal samples (∼93%) but greater for culture compared to PCR-CUL in pooled broth samples (∼99% versus ∼93%). Compared to culture results from testing of individual fecal samples, testing pooled fecal samples by culture had a relative sensitivity of 74% and relative specificity of 96%, testing pooled fecal samples by PCR-CUL resulted in relative sensitivity of 90% and relative specificity of 96%. Testing of pooled 24-hour enrichment broth by PCR-CUL increased the relative sensitivity and specificity to 100%. PCR testing followed by culture of positive or indeterminate samples is a time saving alternative to traditional methods. In addition, pooling of samples may be a useful method for decreasing cost if study aims can accommodate a moderate loss of relative sensitivity.

scholarly journals Gene panel for Mendelian strokes

2020 ◽  
Vol 5 (4) ◽  
pp. 416-421
Author(s):  
Fang Fang ◽  
Zhe Xu ◽  
Yue Suo ◽  
Hui Wang ◽  
Si Cheng ◽  
...  
Keyword(s):  
Sensitivity And Specificity ◽  
Genetic Variants ◽  
Sanger Sequencing ◽  
Large Scale ◽  
Target Genes ◽  
Diagnostic Testing ◽  
Cryptogenic Stroke ◽  
Illumina Hiseq ◽  
Time Saving ◽  
Causal Genes

BackgroundMendelian stroke causes nearly 7% of ischaemic strokes and is also an important aetiology of cryptogenic stroke. Identifying the genetic abnormalities in Mendelian strokes is important as it would facilitate therapeutic management and genetic counselling. Next-generation sequencing makes large-scale sequencing and genetic testing possible.MethodsA systematic literature search was conducted to identify causal genes of Mendelian strokes, which were used to construct a hybridization-based gene capture panel. Genetic variants for target genes were detected using Illumina HiSeq X10 and the Novaseq platform. The sensitivity and specificity were evaluated by comparing the results with Sanger sequencing.Results53 suspected patients of Mendelian strokes were analysed using the panel of 181 causal genes. According to the American College of Medical Genetics and Genomics standard, 16 likely pathogenic/variants of uncertain significance genetic variants were identified. Diagnostic testing was conducted by comparing the consistency between the results of panel and Sanger sequencing. Both the sensitivity and specificity were 100% for the panel.ConclusionThis panel provides an economical, time-saving and labour-saving method to detect causal mutations of Mendelian strokes.

scholarly journals Clinical and Analytical Evaluation of the Anyplex II HPV HR Detection Assay within the VALGENT-3 Framework

2018 ◽  
Vol 56 (11) ◽  
Author(s):  
Anja Oštrbenk ◽  
Lan Xu ◽  
Marc Arbyn ◽  
Mario Poljak
Keyword(s):  
Cancer Screening ◽  
Sensitivity And Specificity ◽  
Screening Program ◽  
Relative Sensitivity ◽  
Cervical Intraepithelial Neoplasia Grade ◽  
Clinical Sensitivity ◽  
Dna Test ◽  
Hpv Dna ◽  
Hpv Test ◽  
Relative Specificity

ABSTRACT In 2012, VALidation of human papillomavirus (HPV) GENotyping Tests (VALGENT) was initiated to provide a formalized and uniform study framework for comparison and validation of HPV assays with genotyping capability. In VALGENT-3, the clinical and analytical performance of Anyplex II HPV HR detection (Anyplex) was compared to that of the Hybrid Capture 2 HPV DNA test (hc2) and the cobas 4800 HPV test (cobas). The panel comprises 1,300 stored samples that were obtained from women 25 to 64 years old who participated in the Slovenian cancer screening program, enriched with 300 samples from women with abnormal cervical cytology. The sensitivity and specificity of Anyplex were noninferior to those of hc2, with a relative sensitivity of 1.01 (95% confidence interval [CI], 0.97 to 1.04) for cervical intraepithelial neoplasia grade 2 or worse (CIN2+) and 1.01 (95% CI, 0.97 to 1.06) for CIN3+ and relative specificity of 1.02 (95% CI, 1.00 to 1.03) for a CIN grade of ≤1. The clinical sensitivity of Anyplex for CIN2+ and CIN3+ was comparable to that of hc2 (P values for McNemar test [pMcN] of 0.655 and 0.564, respectively), but its specificity was significantly higher (pMcN = 0.008). The sensitivity and specificity of Anyplex were also noninferior to those of cobas, with relative sensitivity of 1.01 (95% CI, 0.98 to 1.04) for CIN2+ and 1.01 (95% CI, 0.99 to 1.04) for CIN3+ and relative specificity of 1.00 (95% CI, 0.99 to 1.01) (pMcN value of >0.05 in all cases). Regardless of the clinical outcome (CIN2+ or CIN3+), age restriction (women ≥30 years old), or comparator test used, Anyplex consistently showed excellent clinical performance and can be considered validated for primary cervical cancer screening.

Multiplex Quantitative PCR Assays for the Detection and Quantification of the Six Major Non-O157 Escherichia coli Serogroups in Cattle Feces†

2016 ◽  
Vol 79 (1) ◽  
pp. 66-74 ◽  
Author(s):  
P. B. SHRIDHAR ◽  
L. W. NOLL ◽  
X. SHI ◽  
B. AN ◽  
N. CERNICCHIARO ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Quantitative Pcr ◽  
Foodborne Pathogens ◽  
Virulence Genes ◽  
Target Genes ◽  
Culture Methods ◽  
Fecal Samples ◽  
E Coli ◽  
Cattle Feces ◽  
Before And After

ABSTRACT Shiga toxin–producing Escherichia coli (STEC) serogroups O26, O45, O103, O111, O121, and O145, called non-O157 STEC, are important foodborne pathogens. Cattle, a major reservoir, harbor the organisms in the hindgut and shed them in the feces. Although limited data exist on fecal shedding, concentrations of non-O157 STEC in feces have not been reported. The objectives of our study were (i) to develop and validate two multiplex quantitative PCR (mqPCR) assays, targeting O-antigen genes of O26, O103, and O111 (mqPCR-1) and O45, O121, and O145 (mqPCR-2); (ii) to utilize the two assays, together with a previously developed four-plex qPCR assay (mqPCR-3) targeting the O157 antigen and three virulence genes (stx1, stx2, and eae), to quantify seven serogroups and three virulence genes in cattle feces; and (iii) to compare the three mqPCR assays to a 10-plex conventional PCR (cPCR) targeting seven serogroups and three virulence genes and culture methods to detect seven E. coli serogroups in cattle feces. The two mqPCR assays (1 and 2) were shown to be specific to the target genes, and the detection limits were 4 and 2 log CFU/g of pure culture–spiked fecal samples, before and after enrichment, respectively. A total of 576 fecal samples collected from a feedlot were enriched in E. coli broth and were subjected to quantification (before enrichment) and detection (after enrichment). Of the 576 fecal samples subjected, before enrichment, to three mqPCR assays for quantification, 175 (30.4%) were quantifiable (≥4 log CFU/g) for at least one of the seven serogroups, with O157 being the most common serogroup. The three mqPCR assays detected higher proportions of postenriched fecal samples (P < 0.01) as positive for one or more serogroups compared with cPCR and culture methods. This is the first study to assess the applicability of qPCR assays to detect and quantify six non-O157 serogroups in cattle feces and to generate data on fecal concentration of the six serogroups.

scholarly journals Sensitivity and specificity of a rapid diagnostic test for chronic Chagas disease at a referral center in Brazil - can it be included as a standard serological diagnostic test in the clinical practice of a referral center?

2021 ◽  
Vol 1 ◽  
pp. e1236
Author(s):  
Alejandro M. Hasslocher-Moreno ◽  
Ingebourg Georg ◽  
Luiz H. C. Sangenis ◽  
Mauro F. F. Mediano
Keyword(s):  
Clinical Practice ◽  
Diagnostic Test ◽  
Chagas Disease ◽  
Sensitivity And Specificity ◽  
Diagnostic Testing ◽  
Chronic Phase ◽  
Neglected Tropical Disease ◽  
Rapid Diagnostic Testing ◽  
Referral Center ◽  
Chronic Chagas Disease

Introduction: Chagas disease (CD) is a neglected tropical disease. In the chronic phase of CD, the diagnosis is essentially serologic. Conventional reactions are currently in use. More recently, the use of rapid diagnostic testing (RDT) is indicated when conventional techniques are not available. Objective: To evaluate the sensitivity and specificity of RDTs for chronic CD diagnosis. Methodology: Individuals under suspicion of CD were evaluated using ELISA, Chemiluminescence (ChLIA) and RDT tests. Results: The RDT showed 95.1% sensitivity and 96.7% specificity, respectively. Conclusion: The findings of the present study showed that RDT used in the diagnosis of CD at a referral center in Brazil were not able to detect all CD cases when compared to Elisa and ChLIA.

scholarly journals Evaluation of a vanA-Specific PCR Assay for Detection of Vancomycin-Resistant Enterococcus faecium during a Hospital Outbreak

1999 ◽  
Vol 37 (10) ◽  
pp. 3348-3349 ◽  
Author(s):  
Michel Roger ◽  
Marie-Claude Faucher ◽  
Pierre Forest ◽  
Pierre St-Antoine ◽  
François Coutlée
Keyword(s):  
Enterococcus Faecium ◽  
Pcr Assay ◽  
Culture Methods ◽  
Agar Culture ◽  
Enrichment Broth ◽  
Specific Pcr ◽  
Hospital Outbreak ◽  
Vancomycin Resistant Enterococci ◽  
Vancomycin Resistant ◽  
Specific Pcr Assay

We investigated the use of PCR as an alternative to culture of fecal samples for detection of vanA-containingEnterococcus faecium during a recent hospital outbreak. Rectal swabs collected consecutively from 223 patients were analyzed by culture with and without enrichment broth and byvanA-specific PCR of enrichment broth samples. Fifty-five specimens were positive for vanA-containing E. faecium by at least one method. The sensitivities of thevanA-specific PCR assay and agar culture with and without enrichment broth were 94.5, 98, and 89%, respectively. All three methods were 100% specific. Final results were obtained much more rapidly by PCR (within 24 to 30 h of specimen submission) than by the culture methods (4 to 5 days). Thus, PCR is an accurate and rapid alternative to culture for detection of vancomycin-resistant enterococci during hospital outbreaks.

Direct detection ofCampylobacter jejuniin human stool samples by real-time PCR

2008 ◽  
Vol 54 (9) ◽  
pp. 742-747 ◽  
Author(s):  
Shiyong Lin ◽  
Xinying Wang ◽  
Haoxuan Zheng ◽  
Zhengguo Mao ◽  
Yong Sun ◽  
...  
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Direct Detection ◽  
Culture Method ◽  
Turnaround Time ◽  
Culture Methods ◽  
Time Saving ◽  
Whole Process ◽  
Pure Cultures ◽  
Stool Samples

Our purpose was to establish a quick and accurate real-time PCR (rtPCR) method to detect Campylobacter jejuni directly from human diarrheal stool as an alternative to traditional culture methods. To determine the consistency of rtPCR and culture method, 256 clinical diarrheal stool samples and 50 normal stool samples from healthy individuals were examined, and the whole process was double-blinded. Our data showed that the sensitivity of rtPCR in pure cultures and stool was 102CFU·mL–1and 103CFU·g–1, respectively. Of the 256 diarrheal samples, 10 specimens were successfully detected by both methods, whereas two specimens were PCR positive but culture negative. No positive results were found by these two methods in 50 normal specimens. Our data suggested that rtPCR was convenient in operation and time-saving (turnaround time 3.5–4 h), so it could be used for clinical diagnostic and epidemiological purposes.

scholarly journals The urban and rural capybaras (Hydrochoerus hydrochaeris) as reservoir of Salmonella in the western Amazon, Brazil

2019 ◽  
Vol 39 (1) ◽  
pp. 66-69 ◽  
Author(s):  
Itacir O. Farikoski ◽  
Luciana S. Medeiros ◽  
Yuri K. Carvalho ◽  
David A. Ashford ◽  
Eduardo E. S. Figueiredo ◽  
...  
Keyword(s):  
Urban Area ◽  
Rural Area ◽  
Salmonella Spp ◽  
Convenience Sample ◽  
Enrichment Broth ◽  
Fecal Samples ◽  
Zoonotic Infections ◽  
Hydrochoerus Hydrochaeris ◽  
Pcr Targeting

ABSTRACT: The capybara (Hydrochoerus hydrochaeris) is the largest rodent in the world. In the state of Acre, Brazil, populations of capybaras have been increasing significantly. The role of capybaras in the transmission of certain bacterial zoonotic infections is not well understood, including bacteria of the genus Salmonella. Salmonella spp. generally cause enteritis or septicemia in mammals, however many mammalian species can carry the bacteria asymptomatically and shed it in their feces. To better understand the possible role of capybaras as reservoirs of Salmonella spp., we conducted a study of Salmonella within fecal samples from capybara in Acre. In a convenience sample, 54 capybaras from two urban and two rural areas of Acre were captured and kept for three to four days for sampling. None of the animals were symptomatic of any intestinal illness. Three separate fecal samples were collected from each animal, during their stays in captivity. Each sample was cultured for the presence of Salmonella spp. at the bacteriology laboratory of the Veterinary College of the Federal University of Acre. Samples were seeded in tetrationate pre-enrichment broth and in pre-enrichment broth peptone. After a 24 hour of incubation all samples were streaked on MacConkey Agar (MC) and Salmonella-Shigella Agar (SS). Suggestive colonies were submitted to biochemical analysis. Salmonella compatible colonies according to biochemical profile were submitted to serotyping (Sorokit for Salmonella - Probac do Brasil). In addition, the first sample from each of the 54 capybara was tested for Salmonella spp. using PCR targeting gene hilA. Eight (5%) of the 162 samples examined by bacterial culture were positive for Salmonella spp., while four (7%) of the 54 examined by PCR were positive. From the eight positive animals on culture, five were from urban area and three from rural area. On PCR, only one positive animal was from urban area and four were from rural area. Overall, by either test, one of the 54 animals was positive. All samples were collected in free - living animals with no apparent clinical signs of salmonellosis, indicating the potential of capybara as reservoir on this ecosystem.

Novel Campylobacter Isolation Method Using Hydrophobic Grid Membrane Filter and Semisolid Medium†

2007 ◽  
Vol 70 (2) ◽  
pp. 355-362 ◽  
Author(s):  
ALFONSO VALDIVIESO-GARCIA ◽  
KATHLEEN HARRIS ◽  
EDWARD RICHE ◽  
STEPHANIE CAMPBELL ◽  
ANNE JARVIE ◽  
...  
Keyword(s):  
Membrane Filter ◽  
Simple Procedure ◽  
Meat Products ◽  
Variable Region ◽  
Initial Study ◽  
Epidemiological Studies ◽  
Campylobacter Coli ◽  
Semisolid Medium ◽  
Enrichment Broth ◽  
Pure Cultures

Culture procedures for isolation of thermophilic campylobacters from food matrices are complex, labor intensive, and time-consuming. Most available methods include the use of antibiotics as selective agents to prevent the growth of competing microflora. A simple procedure for isolation of thermophilic campylobacters after enrichment in Rosef's enrichment broth was developed using a hydrophobic grid membrane filter (HGMF) on semisolid medium (SSM). SSM contains no antibiotics, and the HGMF physically separates Campylobacter from the enrichment broth, allowing isolation based on differential motility. The HGMF-SSM method was compared to the Agriculture and Agri-Food Canada Food Safety Procedures Manual (FSPM-10) method (Isolation of Thermophilic Campylobacters from Fresh Pork, Beef, Veal, Poultry and Ready-to-Eat Meat Products), which includes the use of selective antibiotics. During the initial study, after enrichment the HGMF-SSM method yielded pure cultures of campylobacters after 16 to 18 h (overnight) compared with 48 h for the FSPM-10 method. Ninety-four turkey samples collected at local retail stores and 38 frozen pig fecal samples were processed by both methods. Thirty-five samples (26.5%) were positive by the HGMF-SSM method; 24 (18.2%) of these positive samples contained Campylobacter jejuni and 11 (8.3%) contained Campylobacter coli. With the FSPM-10 method, 25 samples (18.9%) were positive: 21 (15.9%) with C. jejuni and 4 (3%) with C. coli. For a subsequent field study, only the HGMF-SSM method was used to isolate Campylobacter from 1,200 chicken samples and 454 turkey samples sold at retail. Analysis of five subisolates from various samples indicated that only one type of Campylobacter was recovered by the HGMF-SSM method, as ascertained by MICs for 10 antimicrobials, sequencing of the short variable region of the flaA gene, and fingerprinting based on amplified fragment length polymorphism. The absence of antibiotics in the SSM may explain the higher recovery of thermophilic campylobacters. The HGMF-SSM method resulted in improved isolation of campylobacters and is simpler, faster, cheaper, and less labor intensive than the FSPM-10 method. The recovery of one type of Campylobacter from the chicken samples may have important implications, particularly in epidemiological studies.

scholarly journals Comparison of 2 ELISAs for detecting exposure to Brucella ovis

2020 ◽  
Vol 32 (5) ◽  
pp. 700-705
Author(s):  
Molly J. Elderbrook ◽  
Brant A. Schumaker ◽  
Massaro W. Ueti ◽  
Meila Bastos de Almeida ◽  
Thallitha S. W. J. Vieira ◽  
...  
Keyword(s):  
Sensitivity And Specificity ◽  
Operating Characteristic ◽  
Relative Sensitivity ◽  
The United States ◽  
Characteristic Analysis ◽  
Cutoff Value ◽  
Brucella Ovis ◽  
Cutoff Values ◽  
Test Kit ◽  
Significant Difference

Control of Brucella ovis infection in sheep flocks in the United States depends on early detection of B. ovis antibodies via serologic testing. We used 2,276 sheep sera and various cutoff values to compare seroprevalence and agreement between 2 ELISAs: the National Veterinary Services Laboratories (NVSL) B. ovis indirect ELISA and the IDEXX B. ovis ELISA kit. A subset of 295 sera was used to compare agreement and evaluate relative sensitivity and specificity of the 2 ELISAs with an agar gel immunodiffusion (AGID) test kit. There was no significant difference in B. ovis seroprevalence between the ELISAs; however, there was poor agreement between them. When the AGID test was used as the reference test, the IDEXX ELISA with a moderate cutoff value (S/P ratio = 45%) had the highest relative sensitivity of 38.1% and specificity of 92.0%. The NVSL ELISA with a lax cutoff value (S/P ratio = 0.75) had relative sensitivity of 19.1% and specificity of 94.6%. Receiver operating characteristic analysis revealed that optimal cutoff values for the NVSL and IDEXX ELISAs were 0.091 and 16.5%, respectively. This results in sensitivity and specificity of 85.7% and 31.8% for the NVSL ELISA, and sensitivity and specificity of 81.0% and 53.6% for the IDEXX ELISA, respectively.

scholarly journals A survey of feline trichomonosis suggests a low incidence of Tritrichomonas blagburni among cats in the Czech Republic

2017 ◽  
Vol 62 (No. 5) ◽  
pp. 269-273 ◽  
Author(s):  
V. Ceplecha ◽  
V. Svobodova ◽  
C. Lendon ◽  
R. Husnik ◽  
K. Horackova ◽  
...  
Keyword(s):  
Czech Republic ◽  
Single Case ◽  
Risk Groups ◽  
Species Differentiation ◽  
Culture Methods ◽  
The Czech Republic ◽  
Pcr Screening ◽  
Low Incidence ◽  
Medium Method

Tritrichomonas blagburni (previously called T. foetus) has been implicated as an aetiological agent of long-term large-bowel diarrhoea in cats in many countries worldwide. The aim of this study was to determine the presence of, and risk factors for T. blagburni among a cohort of cats living in different conditions in the Czech Republic. Samples were collected from 170 cats living in different environments. The InPouch™ TF-Feline medium method was used for diagnosis of feline trichomonosis. A single case (0.6%) with motile trichomonads identified as Pentatrichomonas hominis was found in a cat from a multi-cat household. Our study suggests that trichomonads and in particular, T. blagburni, infection may be much less common in the Czech Republic than in neighbouring countries, despite the inclusion of cats that were likely to be from higher-risk groups. A review of studies of the association of trichomonads and feline diarrhoea carried out in different countries revealed variation in the frequency of trichomonads detected. Different combinations of PCR or culture methods for screening or confirmation have been utilised, with or without species differentiation; however, this could not solely account for the variation in the occurrence between countries. From those studies where differentiation was performed, we calculated from the combined studies that T. blagburni occurred in six cats without diarrhoea (1.1%) and 47 cases with diarrhoea (5%). This finding supports an association with diarrhoea as well as the occurrence of asymptomatic cases. We note that in many studies, including our own, the occurrence of T. blagburni may well be underestimated and suggest that future studies use a combination of PCR screening of both faeces and faecal cultures, with differentiation of trichomonad species.

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