[Inflammatory gastrointestinal diseases in children : Radiological diagnosis]. / Entzündliche gastrointestinale Erkrankungen bei Kindern : Radiologische Diagnose.

BACKGROUND/CLINICAL ISSUE: Inflammatory bowel diseases are frequently seen in the clinical routine of a pediatric radiologist. The timely diagnosis of inflammatory bowel diseases in children is particularly important in acute cases. STANDARD RADIOLOGICAL METHODS/METHODOLOGICAL INNOVATIONS: This nonsystematic article intends to give an overview of the radiologic imaging methods for the diagnosis and work-up of pediatric patients with inflammatory bowel diseases. PERFORMANCE: Ultrasound imaging is an important basis tool in pediatric clinical practice. However, sensitivity and specificity depends on the experience of the operator. Cross-sectional imaging modalities in pediatric patients with inflammatory bowel disease are performed only in exceptional cases when clinically justified. Dedicated computed tomography (CT) protocols for children are indispensable to lower radiation dose. ACHIEVEMENTS: Knowledge about particularities in inflammatory bowel diseases in pediatric patients and a rational approach to the use of radiological investigations in order to prevent the harmful effects of ionizing radiation are indispensable in dedicated pediatric imaging departments. PRACTICAL RECOMMENDATIONS: From a radiation-hygiene point of view, the clinical application of ultrasound imaging should be favored in the work-up of pediatric patients with inflammatory bowel diseases. Knowledge about advanced imaging procedures is essential particularly in imaging departments specialized in pediatric radiology.

Evaluation of most-probable-number-PCR method with internal amplification control for the counting of total and pathogenic Vibrio parahaemolyticus in frozen shrimps.

The most-probable-number (MPN) method is often time-consuming for the isolation, detection, and quantification of Vibrio parahaemolyticus from natural sources. MPN counting of V. parahaemolyticus bacteria usually involves the isolation of typical V. parahaemolyticus colonies on selective medium, with subsequent confirmation by biochemical identification. In this study, we evaluated the use of a PCR on MPN enrichment cultures (MPN-PCR) for the direct detection of total and pathogenic V. parahaemolyticus cells in frozen shrimp. This reaction targeted the R72H, tdh, and trh sequences. An internal amplification control was added to the samples before R72H amplification. There was an excellent correlation between the results of the two methods for artificially inoculated and natural shrimp samples. Of 36 natural samples, 28 tested positive for the presence of V. parahaemolyticus, with an MPN value of 2 × 10(-1) to 9.2 × 10(1) per g. No pathogenic V. parahaemolyticus cells were detected. The test had a detection limit of one V. parahaemolyticus organism per g and was completed within two working days. These results support the use of the combination of PCR with MPN for the detection of total or potentially pathogenic V. parahaemolyticus cells in frozen shrimp.

A risk-based sampling plan for monitoring of histamine in fish products.

In 2008, the French Institute for Public Health Surveillance reported an increase in the number of histamine food poisoning outbreaks and cases in France. The aim of this study was to propose a new monitoring plan for characterizing consumers' exposure to histamine through fishery products. As fish products of concern are numerous, we proposed that the number of samples allocated for a fish category be chosen based on the risk associated with the category. Point risk estimates of histamine poisoning were assessed with the Risk Ranger tool. Fresh fish with high histidine content was found to contribute most to the number of cases. The (estimated) risks associated with the consumption of canned and deep-frozen fish appear marginal as compared with the risk associated with fresh fish with high histidine concentrations. Accordingly, we recommend excluding canned and deep-frozen fish from the monitoring plan, although these risk estimates can be biased. Within a category, samples were proportional to the relative food consumption of the different fishes. The spatial and seasonal consumption patterns were also taken into account for the design of the new monitoring plan. By testing appropriate numbers of samples from categories of fish products of concern, this plan will permit investigation of trends or comparison of product categories presenting risks of histamine poisoning.

Total and pathogenic Vibrio parahaemolyticus in shrimp: fast and reliable quantification by real-time PCR.

Vibrio parahaemolyticus is found in aquatic environments and is the leading cause of gastroenteritis due to seafood consumption worldwide. We evaluated a quantitative real-time PCR (Q-PCR) assay with hydrolysis probes, to determine whether this method could be used for the efficient counting of total, tdh and trh-positive V. parahaemolyticus in shrimps. We assessed the specificity of this assay, using 62 strains from 12 non target bacterial species of the Vibrio, Photobacterium, Shewanella and Aeromonas genera. Only V. parahaemolyticus with the appropriate target gene generated a fluorescent signal. We assessed the robustness of this assay, by analyzing spiked shrimps by Q-PCR and traditional culture methods, using most probable number (MPN)-PCR. After a 6h enrichment period, the assay successfully detected total and pathogenic V. parahaemolyticus in shrimps samples spiked with less than 5 V. parahaemolyticuscells/g of shrimp. The Q-PCR results obtained were compared with those obtained by most probable number (MPN)-PCR format. An excellent correlation between the two methods was observed in all cases (R² > 0.9742). The application of this Q-PCR assay to 85 natural shrimp samples also resulted in the successful quantification of V. parahaemolyticus in this matrix, and the counts obtained were correlated with those obtained by (MPN)-PCR (P=0.2598). Thus, this rapid and sensitive Q-PCR can be used to quantify V. parahaemolyticus in natural shrimp samples. This assay could be proposed, in response to the demands of the European Commission, as a tool for testing for the presence of vibrios in crustaceans, making it possible to legislate in this domain.

Modification of a virulence-associated phenotype after growth of Listeria monocytogenes on food.

AIM: To assess the effect of different foods, which have been implicated or not in cases of listeriosis, on the in vitro virulence-associated phenotype level of different Listeria monocytogenes strains. METHODS AND RESULTS: The virulence-associated phenotype level of L. monocytogenes was studied with the in vitro cell test based on a plaque-forming assay with a human adenocarcinoma cell line (HT-29) monolayer. Three strains of L. monocytogenes were grown in preparations (homogenate, 1-mum filtrate or 0.2-mum filtrate) of different food extracts ['rillettes' (potted minced pork), milk, raw salmon and cold-smoked salmon] or in a control medium, brain heart infusion (BHI). The bacterial suspensions grown in food extracts or in BHI at 37 degrees C were diluted with their growth medium (food extract or BHI) or with minimum essential medium before seeding on confluent HT-29 cell monolayers. Filtration of food extracts had no significant effect on the plaque numbers formed by the bacteria. A significant decrease in the plaque numbers was noted for the three strains when they grew in the rillettes extracts, compared with the other food extracts and BHI. The levels of in vitro virulence-associated phenotype of the strains after growth in the rillettes extract were similar to or lower than that of the hypovirulent internal reference strain L. monocytogenes 442. After growth in milk and cold-smoked salmon, the impact on virulence-associated phenotype depended on the strain. In contrast, plaque-forming assay indicated increased virulence-associated phenotype when the strains were switched from a nutrient-rich medium (food extract or BHI) to a minimum essential medium. CONCLUSIONS: In vitro virulence-associated phenotype level of the studied strains grown in BHI or cold-smoked salmon was the same as the control virulent strain EGD. In contrast, the nutrients present in rillettes may therefore substantially reduce the number of plaques but not the growth of L. monocytogenes. The utilization of minimum essential medium as diluent attenuates changes the effect of the food extract on virulence-associated phenotype in vitro. SIGNIFICANCE AND IMPACT OF THE STUDY: In the experimental design of this study, we showed that the nature of the food could affect the in vitro virulence-associated phenotype level of L. monocytogenes.

Determination of volatile compounds in whiting (Merlangius merlangus) using headspace-solid-phase microextraction-gas chromatography-mass spectrometry.

A method using headspace-solid-phase microextraction-gas chromatography-mass spectrometry to extract volatile compounds from whiting is developed. Several parameters such as sorption time, desorption time, fiber type, and matrix form are optimized to achieve better sensitivity in minimal analysis time. The efficiency of the method is determined by the linear range and repeatability; a mean relative standard deviation of approximately 7% is measured. It was possible to identify and quantitate 30 volatile compounds of interest present in spoiled whiting.

Use of PFGE typing for tracing contamination with Listeria monocytogenes in three cold-smoked salmon processing plants.

The sites of Listeria monocytogenes contamination in three cold-smoked salmon (Salmo salar) processing plants were detected by sampling salmon and the plant's environment and equipment at different production stages. Of the 141 samples collected from three processing plants, 59 (42%) were contaminated with L. monocytogenes. The rates of contamination varied as to the plant and the sample source. L. monocytogenes isolates from 17 various contaminated seafood products (fresh, frozen and smoked fishes, cooked mussels) were also studied. A total of 155 isolates from the three plants and the various seafoods were characterized by genomic macrorestriction using ApaI and SmaI with pulsed-field gel electrophoresis (PFGE) and 82 isolates were serotyped. Macrorestriction yielded 20 pulsotypes and serotyping yielded four serovars: 1/2a, 1/2b, 1/2c, 4b (or e), with 77 (93%) belonging to serovar 1/2a. One clone of L. monocvtogenes predominated and persisted in plant I and was the only pulsotype detected in the final product although it was not isolated from raw salmon. No L. monocytogenes was detected in the smoked skinned salmon processed in plant II, even though 87% of the raw salmon was contaminated. All the smoked salmon samples collected in plant III were contaminated with a unique clone of L. monocytogenes, which may have occurred during slicing. In the three plants, the contamination of final products did not seem to originate from the L. monocytogenes present on raw salmon, but from the processing environment.

Use of biogenic amines to evaluate spoilage in plaice (Pleuronectes platessa) and whiting (Merlangus merlangus).

Fish muscle decarboxylases generate biogenic amines during spoilage. We monitored spoilage in plaice and whiting by assaying biogenic amines represented by the amine index (AI), and by using expert assessors to determine a freshness index (FI) for each fish. First we characterized the assessor- and fish species-related effects on FI, and then we sought to correlate changes in AI and FI by statistical data analysis. We propose rejection limits on the basis of our findings.

Assay of biogenic amines involved in fish decomposition.

A liquid chromatographic (LC) method is described for quantitative determination of putrefaction amines: putrescine, cadaverine, histamine, spermidine, and spermine. These amines are extracted from fish by grinding with perchloric acid at -20 degrees C. The amines are reacted with dansyl chloride at alkaline pH, dried under a stream of nitrogen, placed in an automatic injector at -20 degrees C, and separated in a Kromasil C18 reversed-phase column at 25 degrees C. A new gradient elutes the amines in 22 min and regenerates the column in 30 min. Regression lines are obtained with high coefficients of correlation. This method is faster and more reproducible than other methods and shows that the quality index is a reliable, albeit species-specific, criterion of fish decomposition.

Cryptic Na+,K(+)-ATPase activity in rat liver canalicular plasma membranes: evidence for its basolateral origin.

Controversy exists concerning the localization of the enzyme Na+,K(+)-ATPase to canalicular membranes in hepatocytes. Most studies find enzyme activity only at the basolateral plasma membrane domain of the hepatocyte. However, Na+,K(+)-ATPase activity has been detected recently in a canalicular membrane fraction prepared by Mg++ precipitation, suggesting that differences in membrane domain fluidity account for these discrepancies. To reinvestigate this question, we used free-flow electrophoresis to further purify canalicular liver plasma membranes originally separated by sucrose density centrifugation. With this technique, canalicular membranes devoid of Na+,K(+)-ATPase activity by routine assay were separated into six subfractions. More than 80% of the activities of canalicular marker enzymes was recovered in two subfractions closest to the anode, which were totally devoid of Na+,K(+)-ATPase activity. However, Na+,K(+)-ATPase activity could now be detected in the four other fractions that contained only small amounts of canalicular marker enzymes. The basolateral marker enzyme, glucagon-stimulated adenyl cyclase, comigrated with this cryptic Na+,K(+)-ATPase activity. Furthermore, addition of 6 mumol/L [12-(2-methoxyethoxy)-ethyl-8-(cis-2-n-octylcyclopropyl)-octanoate ], a membrane-fluidizing agent, to the original canalicular membrane preparation and to all subfractions did not stimulate or unmask latent Na+,K(+)-ATPase activity. Finally, when canalicular membranes isolated by Mg++ precipitation were subjected to free-flow electrophoresis, they could not be separated from the more positively charged Na+,K(+)-ATPase-containing fractions, probably because of alterations in surface charge. Together these findings suggest that Na+,K(+)-ATPase is a basolateral enzyme, that represents a small contaminant when present in canalicular liver plasma membranes and that methodological differences may account for previous discrepancies.

Rapid Quantitative Determination of Trimethylamine using Steam Distillation.

A simple, rapid and inexpensive method is proposed for determination of trimethylamine (TMA) in fish muscle. This procedure includes a deproteinization step with trichloroacetic acid (TCA) followed by blocking of primary and secondary amines using formaldehyde at alkaline pH and finally steam distillation of TMA. No statistically significant differences were found between this new optimized procedure and either the Conway microdiffusion method or the colorimetric method. Using the technique proposed here it is possible to assay both the total volatile basic nitrogen (TVBN) and the TMA in less than 30 min.