Conventional Culture Method For Detection Research Articles

Comparison of Legiolert and a Conventional Culture Method for Detection of Legionella pneumophila from Cooling Towers in Québec.

Background: Legionnaires' disease is a potentially lethal pneumonia contracted through inhalation of aerosolized water contaminated with Legionella bacteria. Detection and control of L. pneumophila, the primary species responsible for the disease, is critical to public health. In Québec, cooling towers and evaporative condensers are required to follow a maintenance and testing program to ensure L. pneumophila concentrations remain at acceptable levels. Objective: This study compared a new culture method based on the most probable number approach, Legiolert®, with the formal culture method used at EnvironeX for regulatory compliance testing to quantify L. pneumophila from cooling tower waters in Québec. Methods: A split-sample analysis was performed in which 401 samples from cooling towers in Québec were tested with both methods. Results: Results with 74 positive samples showed that Legiolert provided a significant increase in sensitivity for L. pneumophila compared with the agar plate method. Cooling tower samples often contain non-Legionella flora that necessitate multiple treatment and plating conditions to prevent interference with the test. Legiolert showed little to no impact from non-Legionella organisms in this study. Conclusions: Overall, Legiolert showed several advantages over the agar plate method, including increased sensitivity, reduced interference, a simplified test procedure, and an easy-to-read positive signal.

Comparison of OmpA Gene-Targeted Real-Time PCR with the Conventional Culture Method for Detection of Acinetobacter baumanii in Pneumonic BALB/c Mice

Acinetobacter baumannii is an important pathogen in health care and is responsible for severe nosocomial and community-acquired pneumonia. To design novel therapeutic agents, a mouse model for A. baumannii pneumonia is essential. We described a mouse model of A. baumannii using clinical and 19606R standard strains for developing a quantitative real-time PCR (qRT-PCR) for rapid identification of A. baumannii infection from lung tissues of BALB/c mice. To infect the mice, three doses of bacteria (0.5 × 108, 1 × 108, and 1.5 × 108 cfu/ml) were used. Lung tissues were cultured and compared with ompA gene. Clinical isolates had better positive results at day three with the highest dose than 19606 strain either in culture (4 versus 3) or in qRT-PCR (5 versus 4). However, qRT-PCR detection was 100%, the specificity was 70%, and the positive predictive value was 27%. The qRT-PCR detection of A. baumannii in the BALB/c mice model has a higher sensitivity than the culture method.

Immunochromatography versus Conventional Culture Method for Detection of Campylobacter Jejuni and Campylobacter Coli in Gastroenteritis

Background: Campylobacter is an important food-borne diarrheal disease in the whole world. The use of culture-independent diagnostic tests, such as stool antigen tests, for the detection of Campylobacter in stool is of particular interest.. Objectives: This study aimed to compare between conventional culture method and Immunochromatography (I.C) For Detection Of C.jejuni and C. coli in Gastroenteritisat Benha University hospitals, and determine their antibiotic susceptibility pattern. Methodology: This study was carried out on 100 patients who developed gastroenteritis like symptoms attending the Out Patient Clinic of Benha University Hospitals or admitted to the Pediatric and Internal medicine departments. Stool samples were cultured on modified charcoal cefoperazol dextrose agar (mCCDA) and were examined for Ag detection by I.C test using RIDA®QUICK kit. Antimicrobial susceptibility testing was performed for twelve different antibiotics by the Kirby–Bauer disc diffusion method. Results: Campylobacter was detected in 18% of samples by culture method.72.2% of them were C. jejuni, while the other 27.8% were C. coli. Out of 18 positive by culture, 16(88.9%) were positive by I.C. However, out of 82 negative by culture, 2(2.4%) were positive by I.C. The diagnostic accuracy values of the RidaQuick Campylobacter® versus culture were: sensitivity of 88.9%, specificity of97.6%, and positive and negative predictive values of 88.9% and 97.6%, respectively. Conclusion: RIDAQUICK Campylobacter®provide a rapid and reliable alternative for conventional culture in laboratory diagnosis of enteric infections with C. jejuni and C. coli.

A novel biomarker for detection of Listeria species in food processing factory

A biomarker-based PCR amplification method has been proved to be an effective tool for rapid and accurate detection of Listeria contamination in food factories. However, the current biomarkers (such as iap, hly genes) used for the detection of Listeria have a problem of detection efficiency for some Listeria species, including 7 recently recognized species. Therefore, in this study, a new comprehensive biomarker was developed for the effective detection of all Listeria species. The study employed a novel in-silico scheme to explore and characterize the alternative biomarkers. Biomarker BE-LisAll was identified against 34 Listeria and over 2700 other bacterial complete genomes by in-silico scheme. Specificity of biomarker BE-LisAll was then evaluated with 17 different Listeria species and 58 non-Listeria bacteria isolates. The result showed 100% specificity to Listeria species, and the biomarker could differentiate Listeria species from a variety of non-Listeria bacteria. Finally, the PCR amplification with BE-LisAll was compared with the conventional culture method for detection of Listeria spp. using 60 swab samples collected from a food-processing factory to verify the biomarker's applicability. The result demonstrated 100% correspondence to the results of the conventional culture method. The BE-LisAll biomarker-based PCR amplification is presented for rapid and comprehensive detection of all Listeria species with a high degree of accuracy and sensitivity. It is hoped that commercial food processing industries are able to employ this developed biomarker as a Listeria-detection tool in their factories to prevent or reduce economic losses due to Listeria contamination.

Identification of Vibrio parahaemolyticus in Seafood by Multiplex PCR

ABSTRACTVibrio parahaemolyticus is a human pathogen frequently found in seafood. Once the seafood is contaminated by V. parahaemolyticus, it can become a vehicle for foodborne illness. The conventional culture methods for detection of V. parahaemolyticus are time-consuming and cannot differentiate pathogenic strains from nonpathogenic ones. In this study, a multiplex polymerase chain reaction (PCR) technique was investigated for detecting tdh, chiA, and toxR of V. parahaemolyticus. The sensitivity of the multiplex PCR was determined by testing 28 strains of V. parahaemolyticus, 15 non-V. parahaemolyticus strains, and fresh seafood spiked with cells of V. parahaemolyticus. All the strains were analyzed for production of thermostable direct hemolysin (TDH) and chitinase. This study showed that both the chiA and toxR are excellent markers for detecting V. parahaemolyticus strains, and a multiplex PCR targeting chiA and tdh genes can be applied to simultaneously detect environmental and pathogenic V. parahaemolyticus.

Detection and genetic characterization of major pathogens isolated from bovine mastitis in Rajasthan

The present study was contemplated with an aim to detect and evaluate the major pathogens associated with clinical mastitis in bovines. The milk samples were obtained from the cows suffering from mastitis from different areas of Bikaner and veterinary clinical complex RAJUVAS, Rajasthan. After subjecting the samples to microbiological detection protocols, their 16S–23S rRNA gene internal transcribed spacer (ITS) regions were characterized and the feasibility of using ITS sequences to discriminate between five bacterial species was validated. The conventional bacteriological and PCR based examinations showed incidence of 20 (40%) isolates of E. coli, 16 (32%) isolates of S. aureus, 8 (16%) isolates of P. aeruginosa, and 1 (2%) isolate of Streptococcus spp. The sensitivity and specificity of the test was evaluated using standard cultures. PCR was found to be more sensitive than conventional culture methods for detection of common mastitic pathogens. The results suggest that the PCR assay could be used as an alternative method in routine diagnosis for rapid, sensitive and specific simultaneous detection of S. aureus, S. agalactiae, S. dysgalactiae, S. uberis, E. coli and P. aeruginosa in milk samples.

Comparison of Loop-Mediated Isothermal Amplification Assay and Conventional Culture Methods for Detection of Campylobacter jejuni and Campylobacter coli in Naturally Contaminated Chicken Meat Samples

We investigated the efficacy of a loop-mediated isothermal amplification (LAMP) assay for detection of chicken meat samples naturally contaminated with Campylobacter jejuni and Campylobacter coli. A total of 144 Preston enrichment broth cultures from chicken meat samples were assessed by using the LAMP assay and conventional culture methods, which consist of a combination of Preston enrichment culturing and plating onto Butzler and modified charcoal cefoperazone deoxycholate agars. Compared with C. jejuni-C. coli isolation using the conventional culture test, the LAMP results showed 98.5% (67/68) and 97.4% (74/76) sensitivity and specificity, respectively, and the positive and negative predictive values were 97.1% (67/69) and 98.7% (74/75), respectively. The conventional culture test required more than 3 to 4 days to isolate and identify C. jejuni and C. coli in the Preston enrichment cultures. In contrast, the LAMP assay was markedly faster, requiring less than 90 min from the beginning of DNA extraction to final detection and differentiation of C. jejuni and C. coli. In total, the LAMP assay required 23.5 to 25.5 h from the beginning of the enrichment culture to final determination. These results suggest that our LAMP assay is a powerful tool for rapid, sensitive, and practical detection of C. jejuni and C. coli which may facilitate surveillance and control of C. jejuni-C. coli contamination in chicken, as well as investigations of food poisoning incidents caused by these organisms. This is the first report of a highly sensitive and specific LAMP assay to detect and differentiate C. jejuni and C. coli in chicken meat samples.

Comparison of BD GeneOhm real-time polymerase chain reaction with chromogenic and conventional culture methods for detection of group B Streptococcus in clinical samples

A total of 200 antenatal high vaginal swabs were screened for the presence of group B Streptococcus (GBS) using a conventional culture method (recommended by Centers for Disease Control and Prevention). Screening was also performed by using a new chromogenic agar, chromID Strepto B, and by using the BD GeneOhm StrepB real-time polymerase chain reaction (PCR), which was performed directly on swabs without enrichment. Using a combination of all methods, we detected GBS in 101 samples. A total of 82 samples (81.2%) were positive using PCR, and 83 samples (82.2%) were confirmed as positive by culture (any method). PCR was more sensitive for detection of GBS than direct culture using any method ( P < 0.0005). PCR was also more sensitive than any single enrichment method, but this difference was not statistically significant. With culture as a “gold standard”, the PCR method showed a sensitivity of 77.1% and a positive predictive value of 79.3%. Of the culture-positive samples, significantly, more GBSs were detected by direct plating on chromID Strepto B than on selective sheep blood agar (67.5% versus 57% respectively, P < 0.02). After selective enrichment, 92.8% of GBS were isolated on chromID Strepto B compared with 89.2% isolated on sheep blood agar.

Growth inhibitory factors in bovine faeces impairs detection of Salmonella Dublin by conventional culture procedure

To analyse the relative importance of different biological and technical factors on the analytical sensitivity of conventional culture methods for detection of Salmonella Dublin in cattle faeces. Faeces samples collected from six adult bovines from different salmonella-negative herds were split into subpools and spiked with three strains of S. Dublin at a concentration level of c. 10 CFU g(-1) faeces. Each of the 18 strain-pools was divided into two sets of triplicates of four volumes of faecal matter (1, 5, 10 and 25 g). The two sets were pre-enriched with and without novobiocin, followed by combinations of culture media (three types) and selective media (two types). The sensitivity of each combination and sources of variation in detection were determined by a generalized linear mixed model using a split-plot design. Biological factors, such as faecal origin and S. Dublin strain influenced the sensitivity more than technical factors. Overall, the modified semi-solid Rappaport Vassiliadis (MSRV)-culture medium had the most reliable detection capability, whereas detection with selenite cystine broth and Mueller Kauffman tetrathionate broth combinations varied more in sensitivity and rarely reached the same level of detection as MSRV in this experiment. The study showed that for MSRV-culture medium and xylose lysine decarboxylase agar as the indicative medium, the sensitivity of the faecal culture method may be improved by focusing on the strain variations and the ecology of the faecal sample. Detailed investigation of the faecal flora (pathogens and normal flora) and the interaction with chemical factors may result in developing an improved method for detection of S. Dublin.

Modified Colorimetric DNA Hybridization Method and Conventional Culture Method for Detection of Salmonella in Foods: Comparison of Methods

Abstract A new, improved colorimetric DNA hybridization method for detection of Salmonella in food was developed. A new modified Colorimetric GENETRAK ® Salmonella Assay (m-DNAH) is similar to the current Colorimetric GENE-TRAK Salmonella Assay (c-DNAH) except that the number of assay steps has been reduced, indicator dyes have been added to some reagents to facilitate the tracking of reagent additions, and the probe set has been modified so that subgenus V Salmonella can be detected. The new probe set detected 453/453 Salmonella strains representing at least 239 serovars and 50 somatic antigen groups, including 8 strains of subgenus V Salmonella that were not detected by the current probe set. In addition, a total of 225 nonsalmonellae representing highly homologous organisms such as Enterobacter agglomerans, Escherichia coli, and Citrobacter freundii were all nonreactive with the modified probe set. Overall, a comparison of m-DNAH to the conventional Bacteriological Analytical Manual/AOAC culture method using 240 inoculated food samples and 80 naturally contaminated raw poultry samples showed that both methods were equally effective for detecting Salmonella in food products. The method has been adopted first action by AOAC International to replace method 990.13.

Comparison of immunofluorescence and culture for the detection of Actinomyces israelii in wearers of intra-uterine contraceptive devices

A direct immunofluorescence (IF) method was compared with traditional culture methods for the detection of Actinomyces israelii in endocervical and intra-uterine-device (IUD) smears from 124 IUD wearers. Of 11 specimens that gave positive results by IF, only one was positive by culture. Of the 10 patients with positive IF specimens, three (30%) had signs and symptoms suggestive of pelvic infection and no other pathogen was detected. Direct IF of cervical smears offers a simple, relatively cheap method to screen IUD wearers for A. israelii. Clinical management of such cases is discussed.

Evaluation of a Colorimetric DNA Hybridization Test for Detection of Salmonellae in Meat and Poultry Products

A commercially available colorimetric DNA hybridization test was compared with the USDA/FSIS conventional culture method for detection of salmonellae in naturally contaminated meat and poultry products and inoculated ground beef samples. All samples which were Salmonella-positive by the culture method were also positive by the DNA probe assay. There were no false-negative or false-positive results by the colorimetric DNA hybridization test, which was slightly more sensitive than the culture method.

Comparative Study of Colorimetric DNA Hybridization Method and Conventional Culture Procedure for Detection of Salmonella in Foods

A second generation nucleic acid hybridization assay has been developed and evaluated against the conventional culture method for detection of salmonellae in foods. The assay involves a liquid hybridization with Salmonella-specific oligonucleotide probes, capture of probe:target hybrids onto a solid support (plastic dipstick), and a colorimetric end point detection. The assay can be completed in 2.5 h, following approximately 44 h of culture enrichment. One thousand samples representing 20 food types were analyzed in parallel by both methods. Samples included uninoculated test product, and product inoculated with Salmonella at 2 levels. Eighteen Salmonella serotypes were used as inocula. The data demonstrate that the colorimetric hybridization method and the conventional culture method are equivalent in their ability to detect Salmonella contamination of foods.