Non-target Strains Research Articles

Expanding vaccine efficacy estimation with dynamic models fitted to cross-sectional prevalence data post-licensure

The efficacy of vaccines is typically estimated prior to implementation, on the basis of randomized controlled trials. This does not preclude, however, subsequent assessment post-licensure, while mass-immunization and nonlinear transmission feedbacks are in place. In this paper we show how cross-sectional prevalence data post-vaccination can be interpreted in terms of pathogen transmission processes and vaccine parameters, using a dynamic epidemiological model. We advocate the use of such frameworks for model-based vaccine evaluation in the field, fitting trajectories of cross-sectional prevalence of pathogen strains before and after intervention. Using SI and SIS models, we illustrate how prevalence ratios in vaccinated and non-vaccinated hosts depend on true vaccine efficacy, the absolute and relative strength of competition between target and non-target strains, the time post follow-up, and transmission intensity. We argue that a mechanistic approach should be added to vaccine efficacy estimation against multi-type pathogens, because it naturally accounts for inter-strain competition and indirect effects, leading to a robust measure of individual protection per contact. Our study calls for systematic attention to epidemiological feedbacks when interpreting population level impact. At a broader level, our parameter estimation procedure provides a promising proof of principle for a generalizable framework to infer vaccine efficacy post-licensure.

A novel pentaplex real time (RT)- PCR high resolution melt curve assay for simultaneous detection of emetic and enterotoxin producing Bacillus cereus in food

Bacillus cereus causing emetic and diarrheal type of food poisoning is widely distributed in nature and is therefore considered a major foodborne pathogen. There is a growing demand for fast, accurate, reliable and economic detection of potentially toxigenic B. cereus. To improve differential diagnosis of toxigenic B. cereus, a highly sensitive pentaplex RT- PCR high resolution melt curve assay was developed for simultaneous detection of 4 major enterotoxim genes (cytK, entFM, hblD, nheA) and emetic toxin gene (ces). The average melting temperatures (Tm) of PCR products were 72.2 °C (ces), 74.23 °C (cytK), 76.55 °C (nheA), 78.42 °C (entFM) and 81.90 (hblD). The multiplex assay was evaluated using 71 bacterial strains including 17 emetic B. cereus reference strains, 9 enterotoxic B. cereus reference strains, 4 B. cereus group members, 23 wild B. cereus strains, 18 non-target strains, and was further tested on artificially inoculated foods. The detection limit in food samples was approximately 103 CFU/g without enrichment and 101 CFU/g was observed following 7 h enrichment. The DNA intercalating dye SYTO9 used in this study generated high resolution melt curve peaks for the target strains and genes in which the peaks were sharp and easily distinguishable from each other. Thus, the developed multiplex real-time (RT) PCR approach can be a reliable tool for the identification of emetic and enterotoxic strains of B. cereus present in food and food-related samples.

Rapid, specific and sensitive molecular detection assay for Alternaria helianthi that causes leaf blight disease in sunflower

The leaf blight disease of sunflower (Helianthus annuus) caused by Alternaria helianthi is a serious threat to its cultivation worldwide. Early and accurate detection of the pathogen is critical to efficient disease management and in avoiding further losses due to epidemics in sunflower. Conventional methods of detection and identification are time consuming, labour intensive, and lack specificity and sensitivity. A real-time PCR based TaqMan® probe assay was developed to target 156 bp Internal Transcribed Spacer (ITS) region of A. helianthi for its detection using fungal genomic DNA and infected plant tissues and seeds of sunflower. The specificity of the probe and primers was confirmed by testing their cross reactivity using genomic DNA of closely related Alternaria species isolated from 17 crop plants and 15 fungal species of other genera. No cross-reactivity could be detected with any of the other non-target fungal strains used in this study. The assay successfully detected as low as 1.0 pg fungal genomic DNA and up to 1 % infection in sunflower seed lots. To the best of our knowledge, this is the only probe based real-time PCR assay that enable high specificity and sensitivity for rapid detection of A. helianthi in infected seeds and plant tissues. The assay may also hold promise for application in effective bio-threat and risk mitigation program by early and accurate detection of the pathogen for effective management.

A cross-priming amplification assay coupled with vertical flow visualization for detection of Vibrio parahaemolyticus

Vibrio parahaemolyticus is a marine seafood-borne pathogen that causes gastrointestinal disorders in humans. In this study, we developed a cross-priming amplification (CPA) assay coupled with vertical flow (VF) visualization for rapid and sensitive detection of V. parahaemolyticus. This assay correctly detected all target strains (n = 13) and none of the non-target strains (n = 27). Small concentrations of V. parahaemolyticus (1.8 CFU/mL for pure cultures and 18 CFU/g for reconstituted samples) were detected within 1 h. CPA-VF can be applied at a large scale and can be used to detect V. parahaemolyticus strains rapidly in seafood and environmental samples, being especially useful in the field.

Development and evaluation of a loop-mediated isothermal amplification assay for rapid detection of bacterial blight pathogen (Xanthomonas axonopodis pv. dieffenbachiae) in anthurium

Anthurium bacterial blight caused by Xanthomonas axonopodis pv. dieffenbachiae (Xad) is an important and destructive disease worldwide, and no effective technique has been developed for its control. Detection of infection (latent) in anthurium plants is critical to evaluate disease progress and strengthening management to avoid a serious epidemic in the fields. In this paper, we presented a novel molecular method to detect Xad in anthurium using loop-mediated isothermal amplification (LAMP) technique (Xad-LAMP). The Xad-LAMP reaction could be finished by incubating at 61–65 °C for 1 h, and the amplificons were confirmed through gel electrophoresis, HpaII enzyme analysis, and visually inspected using SYBR Green I/calcein stain and lateral flow dipstick (LFD) assay. The specificity of Xad-LAMP primers set was widely validated on Xad and nontarget strains. In sensitivity testing, Xad-LAMP allowed detection as low as 1–10 f. pure genome DNA or 104 CFU/ml cells, 10–100 times more sensitive than conventional PCR. In addition, combining with the optimized DNA extraction, Xad-LAMP detections were successfully performed on both latent and disease samples, derived from artificially and naturally infected plants respectively. In all, this study provided a promising and practical molecular tool for Xad detecting, will facilitate the forecasting and control of anthurium blight disease.

Development and validation of a real-time quantitative PCR assay to detect Xanthomonas axonopodis pv. allii from onion seed

Bacterial blight of onion is an emerging disease threatening world onion production. The causal agent Xanthomonas axonopodis pv. allii is seed transmitted and a reliable and sensitive tool is needed to monitor seed exchanges. A triplex quantitative real-time PCR assay was developed targeting two X. axonopodis pv. allii-specific markers and an internal control chosen in 5.8S rRNA gene from Alliaceae. Amplification of at least one marker indicates the presence of the bacterium in seed extracts. This real-time PCR assay detected all the 79 X. axonopodis pv. allii strains tested and excluded 85.2% of the 135 non-target strains and particularly all 39 saprophytic and pathogenic bacteria associated with onion. Cross-reactions were mainly obtained for strains assigned to nine phylogenetically related X. axonopodis pathovars. The cycle cut-off was estimated statistically at 36.3 considering a risk of false positive of 1%. The limit of detection obtained in at least 95% of the time (LOD 95%) was 5×103CFU/g (colony forming unit/g). The sensitivity threshold was found to be 1 infected seed in 32,790 seeds. This real-time PCR assay should be useful for preventing the long-distance spread of X. axonopodis pv. allii via contaminated seed lots and determining the epidemiology of the bacterium.

Rapid detection of viable Bacillus cereus emetic and enterotoxic strains in food by coupling propidium monoazide and multiplex PCR (PMA-mPCR)

Bacillus cereus can cause emetic and diarrheal food poisoning. It is widespread in nature and therefore, considered a major foodborne pathogen. To develop a sensitive and reliable assay for detecting enterotoxin genes (nheA, entFM, hblD, cytK) and emetic toxin (ces), specific primers each targeting one individual gene were designed. Propidium monoazide (PMA) was coupled with the developed multiplex PCR (mPCR) for the detection of viable B. cereus. The inclusivity and exclusivity of the PMA-mPCR was confirmed using a panel of 44 strains including 17 emetic and 9 enterotoxic B. cereus reference strains and 18 non-target strains. The limit of detection (LOD) without PMA treatment in pure DNA was 2 pg/reaction tube. The LOD of mPCR assay in pure heat-killed dead bacteria was 4.0 × 102 CFU/mL. Also, the LOD on the viable bacteria with or without PMA treatment was similar (3.8 × 102 CFU/mL) showing that the PMA treatment did not significantly decrease sensitivity. Finally, the newly developed PMA-mPCR successfully detected 4.8 × 103 and 3.6 × 103 CFU/g of viable B. cereus F4810/72 (emetic) and B. cereus ATCC 12480 (enterotoxic) reference strains, respectively, in food samples. Hence, this study combines PMA and mPCR to detect viable B. cereus with a wide range of toxin detection (5 toxins). Thus, the novel PMA-mPCR assay developed in this study is a rapid and efficient diagnostic tool for the monitoring of viable B. cereus in food samples and potentially other samples via appropriate DNA extraction.

Development and evaluation of a loop-mediated isothermal amplification assay combined with enrichment culture for rapid detection of very low numbers of Vibrio parahaemolyticus in seafood samples.

The aim of this study was to develop and evaluate a rapid and effective method to detect Vibrio parahaemolyticus, a leading pathogen causing seafood-borne gastroenteritis. A newly designed loop-mediated isothermal amplification (LAMP) assay including a short enrichment period was optimized. This assay correctly detected all the target strains (n=61) but none of the non-target strains (n=34). Very low numbers of V. parahaemolyticus (2 colony forming unit (CFU) per gram of seafood) could be detected within 3 h and the minimum time of the whole assay was only 5 h. Comparative screening of various seafood samples (n=70) indicated that the LAMP assay is superior to polymerase chain reaction (PCR) and conventional culture methods because it is more rapid and less complex. This highly sensitive LAMP assay can be applicable as the method of choice in large-scale and rapid screening of seafood and environmental samples to detect V. parahaemolyticus strains.

The Possible Impact of Vaccination for Seasonal Influenza on Emergence of Pandemic Influenza via Reassortment.

BackgroundOne pathway through which pandemic influenza strains might emerge is reassortment from coinfection of different influenza A viruses. Seasonal influenza vaccines are designed to target the circulating strains, which intuitively decreases the prevalence of coinfection and the chance of pandemic emergence due to reassortment. However, individual-based analyses on 2009 pandemic influenza show that the previous seasonal vaccination may increase the risk of pandemic A(H1N1) pdm09 infection. In view of pandemic influenza preparedness, it is essential to understand the overall effect of seasonal vaccination on pandemic emergence via reassortment.Methods and FindingsIn a previous study we applied a population dynamics approach to investigate the effect of infection-induced cross-immunity on reducing such a pandemic risk. Here the model was extended by incorporating vaccination for seasonal influenza to assess its potential role on the pandemic emergence via reassortment and its effect in protecting humans if a pandemic does emerge. The vaccination is assumed to protect against the target strains but only partially against other strains. We find that a universal seasonal vaccine that provides full-spectrum cross-immunity substantially reduces the opportunity of pandemic emergence. However, our results show that such effectiveness depends on the strength of infection-induced cross-immunity against any novel reassortant strain. If it is weak, the vaccine that induces cross-immunity strongly against non-target resident strains but weakly against novel reassortant strains, can further depress the pandemic emergence; if it is very strong, the same kind of vaccine increases the probability of pandemic emergence.ConclusionsTwo types of vaccines are available: inactivated and live attenuated, only live attenuated vaccines can induce heterosubtypic immunity. Current vaccines are effective in controlling circulating strains; they cannot always help restrain pandemic emergence because of the uncertainty of the oncoming reassortant strains, however. This urges the development of universal vaccines for prevention of pandemic influenza.

Development of a multiplex real-time PCR method for simultaneous detection of Vibrio parahaemolyticus, Listeria monocytogenes and Salmonella spp. in raw shrimp

Vibrio parahaemolyticus, Listeria monocytogenes and Salmonella spp. are important pathogens contaminating seafood in China. In this study, we developed an efficient multiplex real-time PCR for the simultaneous detection of V. parahaemolyticus, L. monocytogenes and Salmonella spp. in raw shrimp without a prior enrichment step. In a test using 28 target and non-target strains only the targets were detected and two calibration curves, for pure cultures and artificially contaminated samples, were used to evaluate the efficiencies of this method. Amplification efficiencies of this multiplex real-time PCR were excellent in pure cultures and artificially contaminated shrimps. The limits of detection in artificially contaminated shrimps were 112 CFU/g for V. parahaemolyticus, 158 CFU/g for L. monocytogenes and 103 CFU/g for Salmonella spp. We validated this multiplex real-time PCR method on 48 commercial samples and the results were comparable to standard culture methods. This efficient multiplex real-time PCR, where each test takes only 50 min after DNA extraction, is a useful tool for high-throughput surveillance of V. parahaemolyticus, L. monocytogenes and Salmonella spp. in seafood products.

Detection of Listeria spp. using ACTERO listeria enrichment media.

ACTERO Listeria Enrichment Media (ACTERO Listeria) is a selective medium developed for a single-step recovery and enrichment of Listeria spp. from environmental samples. Robustness testing of the ACTERO Listeria medium demonstrated good performance when minor changes were introduced to the incubation temperature and time. All 54 Listeria strains tested, representing the most frequently isolated Listeria species from food (L. monocytogenes, L. ivanovii, L. seeligeri, L. welshimeri, and L. grayi), were successfully enriched in ACTERO Listeria. None of the 30 nontarget strains tested in the exclusivity study was recovered after incubation in ACTERO Listeria. Recovery of Listeria was consistent across three independently produced lots of the ACTERO Listeria, and the prepared medium was stable for 45 days when stored at 4 degrees C in the dark. Matrix studies performed with environmental sponge samples from plastic and stainless steel surfaces demonstrated similar recovery of Listeria spp. in a single-step enrichment using ACTERO Listeria from plastic, and significantly better recovery from stainless steel surfaces when compared to the U.S. Department of Agriculture-Food Safety and Inspection Service reference method. The results of this study prove that ACTERO Listeria Enrichment Media can be effectively used in replacement of the two-step enrichment suggested by the reference method without affecting the recovery of Listeria spp. from environmental samples.

Development of a Strain-Specific Real-Time PCR Assay for Enumeration of a Probiotic Lactobacillus reuteri in Chicken Feed and Intestine

A strain-specific real-time PCR assay was developed for quantification of a probiotic Lactobacillus reuteri (DSM 16350) in poultry feed and intestine. The specific primers were designed based on a genomic sequence of the strain derived from suppression subtractive hybridization with the type strain L. reuteri DSM 20016. Specificity was tested using a set of non-target strains from several sources. Applicability of the real-time PCR assay was evaluated in a controlled broiler feeding trial by using standard curves specific for feed and intestinal matrices. The amount of the probiotic L. reuteri was determined in feed from three feeding phases and in intestinal samples of the jejunum, ileum, and caecum of three, 14, and 39 day old birds. L. reuteri DSM 16350 cells were enumerated in all feeds supplemented with the probiotic close to the inclusion rate of 7.0×103 cfu/g, however, were not detected in L. reuteri DSM 16350 free feed. In three day old birds L. reuteri DSM 16350 was only detected in intestinal samples from probiotic fed animals ranging from 8.2±7.8×105 cfu/g in the jejunum, 1.0±1.1×107 cfu/g in the ileum, and 2.5±5.7×105 cfu/g in the caecum. Similar results were obtained for intestinal samples of older birds (14 and 39 days). With increasing age of the animals the amount of L. reuteri signals in the control animals, however, also increased, indicating the appearance of highly similar bacterial genomes in the gut microbiota. The L. reuteri DSM 16350 qPCR assay could be used in future for feeding trials to assure the accurate inclusion of the supplement to the feed and to monitor it's uptake into the GIT of young chicken.

Development and Application of a Loop‐Mediated Isothermal Amplification (LAMP) Method for Rapid and Sensitive Detection of Enterococcus faecalis in Drinking Water

AbstractThe genus Enterococcus comprises opportunistic pathogens such as Enterococcus faecalis and E. faecium, and is used to assess water quality. E. faecalis has become prominent in recent decades because of its ability to develop resistance to various antibiotics. A loop‐mediated isothermal amplification (LAMP) method for rapid and sensitive detection of E. faecalis in drinking water samples was developed and applied in this study. Four primers were specifically designed by targeting the mtlf gene. The mtlf‐based LAMP assay was able to specifically detect all of 54 E. faecalis isolates without amplification of 36 nontarget bacterial strains. The detection limit was 74 cfu/mL in pure culture, which was as high as 100‐fold more sensitive than that of the groES‐polymerase chain reaction (PCR) method. When applied to artificially contaminated water, the LAMP assay was able to detect E. faecalis with low concentration (3.2 cfu/250 mL), after 10 h enrichment culture. In the case of drinking water samples, 7.14% (7/98) of the samples were found to be positive by LAMP, which was in accordance with the results from the GB/T 8538‐2008 method. The LAMP assay established in this study may provide a candidate method for rapid and sensitive detection of E. faecalis in drinking water samples.Practical ApplicationsE. faecalis contamination in drinking water source and drinking water has been reported. In developing countries, such as China, the LAMP assay may prove as a useful tool for rapid assessment of water quality as it is more cost‐effective, simple and sensitive than PCR.

Evaluation of the effectiveness of Duddingtonia flagrans and Monacrosporium thaumasium in the biological control of gastrointestinal nematodes in female bovines bred in the semiarid region

Brazil has a herd of 212 million cattle and 171 million hectares of pastures that produce approximately 96% of Brazilian beef. The Brazilian production system enables animal infection by endoparasites, which are considered one of the main obstacles for the development of this industry and are responsible for considerable economic losses. The control of parasitic diseases is performed via the administration of antiparasitic drugs, but they leave residues of the products in the treated animal, affect non-target organisms and select resistant strains of the parasites. The species D. flagrans and M. thaumasium are promising and sustainable alternatives for controlling gastrointestinal helminths of ruminants and other herbivores. In this study, we evaluated the efficacy of isolates of these species, formulated in a sodium alginate matrix and administered twice a week, to reduce the number of environmental infective larvae of gastrointestinal nematodes that affect prepubescent zebu females. The treated animals presented fewer eggs and a lower number of infective larvae per gram of faeces (p < 0.05). The pastures occupied by treated animals showed a statistically significant reduction (p < 0.05) of the number of L3 and, furthermore, the genera Cooperia sp., Haemonchus sp., and Oesophagostomum sp. were the most prevalent. The average weight of the animals did not differ statistically (p > 0.05) among the treated and control groups. The use of sodium alginate pellets as vehicle for delivery of the fungus mycelia D. flagrans (isolate AC001) and M. thaumasium (isolate NF34A) proved effective in controlling trichostrongylids in prepubescent cows bred in the semi-arid region, with an effective reduction in the number of infective larvae in the pastures.

Real time loop-mediated isothermal amplification using a portable fluorescence scanner for rapid and simple detection of Vibrio parahaemolyticus

We evaluated the usefulness of the loop-mediated isothermal amplification (LAMP) using a portable ESE Quant tube scanner as a rapid and simple method for the detection of Vibrio parahaemolyticus, an important pathogen causing seafood-borne gastroenteritis. The real time LAMP (RT-LAMP) assay using a hemolysin gene (tlh/ldh)-specific primers was verified using V. parahaemolyticus strains (n = 91) from different countries and other non-target strains to check the utility of this method. Both the sensitivity and specificity of the RT-LAMP using 3 pairs of tlh/ldh-specific primers developed in this study were excellent (100%). The detection limit of the RT-LAMP was as low as 7 Colony forming unit per reaction and detection time was only 20 min. Comparative evaluation of the target bacterial strains with the RT-LAMP using ESE Quant tube scanner, API 20E system and conventional RT-PCR method revealed that the RT-LAMP assay developed in this study is simpler and more rapid than the latter two methods. Therefore, the RT-LAMP method using the easily portable ESE Quant tube scanner can be considered as an effective tool for the rapid screening of V. parahaemolyticus strains in environmental and clinical samples, especially, in remote areas of developing countries during epidemic periods.

Arcanobacterium pluranimalium leading to a bovine mastitis: Species identification by a newly developed pla gene based PCR

We are describing a clinical case of bovine mastitis due to Arcanobacterium pluranimalium in a Holstein-Friesian heifer, delivering bloody milk on the left hindquarter. Moreover, we report on the development and evaluation of PCR primers based on the pluranimaliumlysin (pla) gene for the identification of this species. With the primer pair PlaF/PlaR the A. pluranimalium type strain as well as the mastitis isolate 704 revealed a correctly sized amplification product (458 bp), whereas no amplification product was obtained for all non-target strains. The established PCR provides a new and convenient tool for the mastitis diagnostic to differentiate between A. pluranimalium and Trueperella pyogenes.

Inclusivity, exclusivity and limit of detection of commercially available real-time PCR assays for the detection of Salmonella

The traditional cultural detection of Salmonella spp. is both time- and labour-intensive. Salmonella is often a release criterion for the food industry and time to result is therefore an important factor. Storage of finished products and raw materials can be costly and may adversely impact available shelf-life. The application of real-time PCR for the detection of Salmonella spp. in food samples enables a potential time-saving of up to four days. The advancement of real-time PCR coupled with the development of commercially available systems in different formats has made this technology accessible for laboratories in an industrial environment. Ideally these systems are reliable and rapid as well as easy to use.The current study represents a comparative evaluation of seven commercial real-time PCR systems for the detection of Salmonella. Forty-nine target and twenty-nine non-target strains were included in the study to assess inclusivity and exclusivity. The limit of detection for each of the method was determined in four different food products. All systems evaluated were able to correctly identify the 49 Salmonella strains. Nevertheless, false positive results (Citrobacter spp.) were obtained with four of the seven systems. In milk powder and bouillon powder, the limit of detection was similar for all systems, suggesting a minimal matrix effect with these samples. Conversely, for black tea and cocoa powder some systems were prone to inhibition from matrix components. Up to 100% of the samples were inhibited using the proprietary extracts but inhibition could be reduced considerably by application of a DNA clean-up kit.

Rapid detection of emetic toxin producing Bacillus cereus strains using triple-primer polymerase chain reaction (PCR) assay

Emetic toxin producing Bacillus cereus can cause emetic food poisoning. In this study, an improved triple-primer polymerase chain reaction (PCR) assay was developed as a reliable and rapid identification method for the detection of emetic toxin producing B. cereus strains based on the unique gene sequences of the CER, ces and groEL. Specificity and sensitivity of the primers was confirmed using conventional PCR and gel electrophoresis. In addition, detection limit was determined in pure culture, rice and milk. In brief, sensitivity in pure culture was 10 fold or more higher than artificially inoculated food samples in improved triple-primer PCR detection limit assay. The primers did not react with genes from enterotoxin producing B. cereus or other non-target strains. The presented PCR assay is a useful tool for the rapid screening and diagnosis of emetic toxin producing B. cereus strains in food. Key words: Triple-primer polymerase chain reaction (PCR), Bacillus cereus, emetic toxin, food poisoning.

Molecular probes for the detection and identification of ichthyotoxic marine microalgae of the genus Pseudochattonella (Dictyochophyceae, Ochrophyta)

Phytoflagellates of the genus Pseudochattonella (Dictyochophyceae, Ochrophyta) form blooms in marine coastal waters in northern Europe, Japan, and New Zealand that at times cause fish kills with severe losses for the aquaculture industry. The aim of this study was to develop molecular probes for the detection and identification of Pseudochattonella at the genus and species level. A variety of probes were developed and applied to either dot blot hybridization, (q)PCR, or microarray format. In the dot blot hybridization assay, five different oligonucleotide probes targeting the small subunit (SSU) rDNA were tested against DNA from 18 microalgal strains and shown to be specific to the genus Pseudochattonella. A genus-specific PCR assay was developed by identifying an appropriate primer pair in the SSU-internal transcribed spacer 1 (ITS1) rDNA region. Its specificity was tested by screening against both target and non-target strains, and the assay was used to confirm the presence or absence of Pseudochattonella species in environmental samples. In order to distinguish between the two species of the genus, two PCR primer pairs each biased towards one of the species were designed in the large subunit (LSU) rDNA D1 domain and used for quantitative real-time PCR. Five selected probes (three SSU and two LSU rDNA) were adapted for the use on microarrays and included on a prototype multi-species microarray for the detection of harmful algae ( http://www.midtal.com ). Finally, microarrays and qPCR were used for the monthly monitoring of a sampling site in outer Oslofjorden during a 1-year period. Members of Pseudochattonella are difficult to identify by light microscopy in Lugol's preserved samples, and the two species Pseudochattonella verruculosa and Pseudochattonella farcimen can be morphologically distinguished only by transmission electron microscopy. The molecular probes designed in this study will be a valuable asset to microscopical detection methods in the monitoring of harmful algae and for biogeographical and ecological studies of this genus.

Samsung Salmonella Detection Kit

Salmonella, one of the most common causes of foodborne illness, is a significant public health concern worldwide. There is a need in the food industry for methods that are simple, rapid, and sensitive for the detection of foodborne pathogens. In this study, the Samsung Salmonella Detection Kit, a real-time PCR assay for the detection of Salmonella, was evaluated according to the current AOAC guidelines. The validation consisted of lot-to-lot consistency, stability, robustness, and inclusivity/exclusivity studies, as well as a method comparison of 10 different food matrixes. In the validation, the Samsung Salmonella Detection Kit was used in conjunction with the Applied Biosystems StepOnePlus PCR system and the Samsung Food Testing Software for the detection of Salmonella species. The performance of the assays was compared to the U.S. Department of Agriculture/Food Safety and Inspection Service-Microbiology Laboratory Guidebook (USDA/FSIS-MLG) 4.05: Isolation and Identification of Salmonella from Meat, Poultry, Pasteurized Egg, and Catfish and the and U.S. Food and Drug Administration/Bacteriological Analytical Manual (FDA/BAM) Chapter 5 Salmonella reference methods. The validation was conducted using an unpaired study design for detection of Salmonella spp. in raw ground beef, raw pork, raw ground pork, raw chicken wings, raw salmon, alfalfa sprouts, pasteurized orange juice, peanut butter, pasteurized whole milk, and shell eggs. The Samsung Salmonella Detection Kit demonstrated lot-to-lot consistency among three independent lots as well as ruggedness with minor modifications to changes in enrichment incubation time, enrichment incubation temperature, and DNA sample volume for PCR reaction. Stability was observed for 13 months at -20 degrees C and 3 months at 5 degrees C. For the inclusivity/exclusivity study, the Samsung Salmonella Detection Kit correctly identified 147 Salmonella species isolates out of 147 isolates tested from each of three different enrichment broths (a total of 441 isolates detected), and correctly excluded all 31 nontarget strains analyzed. For the method comparison, statistical analysis was conducted according to the Mantel-Haenszel Chi-square formula for unpaired test portions, and there was no significant difference in the number of positive samples detected between the Samsung Salmonella Detection Kit and the USDA/FSIS-MLG and FDA/BAM reference methods for all 10 food matrixes.