Biomedical subjects

Stephen Forsythe

Publications and source records attributed to Stephen Forsythe.

5 recordsLinked to original sources

PubMed2006

The presence of endotoxin in powdered infant formula milk and the influence of endotoxin and Enterobacter sakazakii on bacterial translocation in the infant rat.

Lipopolysaccharide (LPS) is a heat stable endotoxin that persists during the processing of powdered infant formula milk (IFM). Upon ingestion it may increase the permeability of the neonatal intestinal epithelium and consequently bacterial translocation from the gut. To determine the level of endotoxin present in IFM, 75 samples were collected from seven countries (representing 31 brands) and analysed for endotoxin using the kinetic colorimetric Limulus amoebocyte lysate (LAL) assay. The endotoxin levels ranged from 40 to 5.5 x 10(4) endotoxin units (EU) per gram and did not correlate with the number of viable bacteria. The neonate rat model was used to address the risk of endotoxin-induced bacterial translocation from the gut. Purified Escherichia coli LPS was administered to rat pups followed by inoculation with Enterobacter sakazakii ATCC 12868. Bacteria were isolated from the mesentery, spleen, blood and cerebral spinal fluid (CSF) of endotoxin-treated rats due to enhanced gut and blood brain barrier penetration. Histological analysis of the colon showed marked distension of the mucosal and muscular layers. It is plausible that the risk of neonatal bacteraemia and endotoxemia, especially in neonates with immature innate immune systems, may be raised due to ingestion of IFM with high endotoxin levels.

Animals↗

PubMed2006

Identification of Enterobacter sakazakii from closely related species: the use of artificial neural networks in the analysis of biochemical and 16S rDNA data.

BACKGROUND: Enterobacter sakazakii is an emergent pathogen associated with ingestion of infant formula and accurate identification is important in both industrial and clinical settings. Bacterial species can be difficult to accurately characterise from complex biochemical datasets and computer algorithms can potentially simplify the process. RESULTS: Artificial Neural Networks were applied to biochemical and 16S rDNA data derived from 282 strains of Enterobacteriaceae, including 189 E. sakazakii isolates, in order to identify key characteristics which could improve the identification of E. sakazakii. The models developed resulted in a predictive performance for blind (validation) data of 99.3 % correct discrimination between E. sakazakii and closely related species for both phenotypic and genotypic data. Three main regions of the partial rDNA sequence were found to be key in discriminating the species. Comparison between E. sakazakii and other strains also constitutively positive for expression of the enzyme alpha-glucosidase resulted in a predictive performance of 98.7 % for 16S rDNA sequence data and 100% for phenotypic data. CONCLUSION: The computationally based methods developed here show a remarkable ability in reducing data dimensionality and complexity, in order to eliminate noise from the system in order to facilitate the speed and reliability of a potential strain identification system. Furthermore, the approaches described are also able to provide valuable information regarding the population structure and distribution of individual species thus providing the foundations for novel assays and diagnostic tests for rapid identification of pathogens.

Cronobacter sakazakii↗

PubMed2004

A selective differential medium for Enterobacter sakazakii, a preliminary study.

Enterobacter sakazakii can cause fatal invasive infection of neonates associated with the presence of this organism in powdered infant milk formula. A new chromogenic medium (Druggan-Forsythe-Iversen agar, DFI) is described for the selective detection of this emergent pathogen. The medium is based on the alpha-glucosidase reaction which is detected using 5-bromo-4-chloro-3-indolyl-alpha,D-glucopyranoside (XalphaGlc). Ent. sakazakii hydrolyses this substrate to an indigo pigment, producing blue-green colonies on this medium. DFI was compared with the current method of detection on violet red bile glucose agar (VRBGA) followed by pigment production on tryptone soy agar (TSA) after 48-72 h at 25 degrees C and subsequent biochemical profile determination using Biomerieux API20E. Ninety-five clinical and food strains of Ent. sakazakii were detected on the DFI chromogenic medium 2 days sooner than the alternative method. The characteristics of 148 strains representing 17 genera of non-Ent. sakazakii Enterobacteriaceae were compared using the two methods. Only 16/18 Escherichia vulneris strains, 2/3 strains of Pantoea spp. and 1/8 Citrobacter koseri strains gave false positive results on DFI agar. Eight alpha-glucosidase positive strains were identified as Pantoea using their API20E biochemical profile, but had higher percentage identification as Ent. sakazakii using ID32E. Therefore the DFI medium enables the detection of Ent. sakazakii within mixed cultures of Enterobacteriaceae, whereas the organism could be missed when using VRBGA since the latter is a general Enterobacteriaceae selective medium. In addition, the common use of API20E to check yellow pigmented colonies on TSA may lead to false negative results and consequently the acceptance of a batch of infant formula milk (IFM) that contains Ent. sakazakii.

Colony Count, Microbial↗

PubMed2004

Identification and phylogeny of Enterobacter sakazakii relative to Enterobacter and Citrobacter Species.

The phylogenetic relationships of Enterobacter sakazakii strains were investigated using 16S ribosomal DNA (rDNA) and hsp60 sequencing. Each analysis distributed E. sakazakii strains among four clusters, indicating substantial taxonomic heterogeneity. The E. sakazakii type strain 16S rDNA sequence was 97.8% similar to that of Citrobacter koseri but 97.0% similar to that of Enterobacter cloacae.

Chaperonin 60↗

PubMed2003

The toxicity of textile reactive azo dyes after hydrolysis and decolourisation.

The toxicity of C.I. Reactive Black 5 and three Procion dyes, as found in textile effluents, was determined using the bioluminescent bacterium Vibrio fischeri. Hydrolysed Reactive Black had a slightly greater toxicity than the parent form (EC(50) 11.4+/-3.68 and 27.5+/-4.01 mg l(-1), respectively). A baffled bioreactor with anaerobic and aerobic compartments was used to decolourise hydrolysed Reactive Black 5 in a synthetic effluent. Decolourisation of hydrolysed Reactive Black resulted in an increased toxicity (EC(50) 0.2+/-0.03 mg l(-1)). Toxicity was not detectable when decolourised Reactive Black 5 was metabolised under aerobic conditions. No genotoxicity was detected after the decolourisation of either the parent or the hydrolysed reactive dyes, either in vitro or in the bioreactor. The toxicity and genotoxicity of decolourised C.I. Acid Orange 7 was due to the production of 1-amino-2-naphthol (EC(50) 0.1+/-0.03 mg l(-1)).

Anaerobiosis↗