Search PubMed⌕ Search

Biomedical subjects

G M Wyatt

Publications and source records attributed to G M Wyatt.

At least 19 recordsLinked to original sources
PubMed2004

A comparison of the traditional method of counting viable cells and a quick microplate method for monitoring the growth characteristics of Listeria monocytogenes.

AIMS: To determine: (i) the growth parameters (specific growth rate, lag time, asymptotic amount of growth, generation time and time for maximum growth rate) of Listeria monocytogenes in different broths by standard cultivation methods and (ii) whether a microplate method in conjunction with a standard nondedicated plate reader could be adapted to routine assay. METHODS AND RESULTS: Growth curves were determined from cell numbers in a standard tube method at 2 h intervals by serial dilution and plating, and in a microplate method by absorbance measurements. Growth curves were fitted with a modified Gompertz function. CONCLUSIONS: The microplate method was similar to the standard cultivation methods in accuracy, required less chemical reagents, and considerably reduced the time required for analyses. This work also illustrates that growth characteristics of bacteria are not necessarily constant, and depend on the methodology used. SIGNIFICANCE AND IMPACT OF THE STUDY: It is not the intended purpose of this paper to present all the data for the media tested but instead to illustrate the success of the microplate method for studying growth kinetics compared to a standard cultivation method and system precision. The method will be of considerable benefit to laboratories unable to afford dedicated workstations.

Bacteriological Techniques↗
PubMed2003

Principles of some novel rapid dipstick methods for detection and characterization of verotoxigenic Escherichia coli.

AIMS: The verotoxigenic Escherichia coli (VTEC) serotype most commonly associated with verotoxin (VT) production is O157:H7, but other serotypes have also been implicated in food-borne illness. These serotypes exhibit much greater genetic and biochemical diversity than E. coli O157:H7, making screening for all VTEC difficult. Here we describe development and testing of novel multi-analyte antibody-based dipstick methods for presumptive detection of VTEC cells and VTs, including non-O157 serotypes. METHODS AND RESULTS: The dipsticks are formatted as paddle-style and lateral flow devices. Test materials included raw milk, minced beef, apple juice and salami, spiked with VTEC. Prototype paddle dipsticks gave 47 of 48 E. coli O157-positive samples correct, and, simultaneously, 27 of 31 O26-positive samples correct, across the four food types. Prototype lateral flow dipsticks gave 12 of 12 E. coli O157-positive milk samples correct and, simultaneously, 28 of 28 positive VT samples correct. CONCLUSIONS: This work demonstrates that simple and rapid detection of more than one VTEC characteristic (toxin production and type, serogroup) is possible in a single dipstick test device, directly from a food enrichment culture. SIGNIFICANCE AND IMPACT OF THE STUDY: The development of simple easy-to-use rapid methods for simultaneous detection and preliminary characterization of VTEC will enable the risk presented by all VTEC to be more thoroughly assessed (e.g. in surveillance studies, outbreak investigations).

Antibodies, Bacterial↗
PubMed2001

Investigation of the interactions between beta-lactams and a metallo-beta-lactamase from bacillus cereus using a monoclonal antibody.

A monoclonal antibody recognizing the active site of a beta-lactamase from Bacillus cereus was identified and characterized. The binding of the monoclonal antibody to the active site was quantitatively inhibited by a broad spectrum of beta-lactam antibiotics. The levels of inhibition were found to be associated with particular structural features of the antibiotics and their ability to form stable enzyme/substrate complexes. A novel, broad specificity assay for beta-lactams was developed based on the inhibition of antibody binding of all the beta-lactams studied. The assay is applicable to detection of beta-lactams at or close to the MRL level and would be complementary to existing receptor-based assays. The approach described is relevant to the study of kinetic aspects of beta-lactamases and could prove a useful tool in future drug development.

Animals↗
PubMed2001

Development of an ELISA for sulfachlorpyridazine and investigation of matrix effects from different sample extraction procedures.

The development of an enzyme-linked immunosorbent assay (ELISA) for the detection of residues of sulfachlorpyridazine (SCP) is described for the first time. The assay is highly specific for SCP, is simple to perform and has a lower detection limit of 0.65 ng/ml in assay buffer. In potential application of the assay to detect residues of SCP at the 0.1 mg/kg level in eggs, milk, beef, lamb, pork, chicken, turkey, porcine kidney, porcine liver and pig feedstuffs is discussed with regard to the effects of sample extracts on the standard curves. The antibody exhibits a rare stability in assay buffers containing up to 30% methanol. It is concluded that the ELISA for SCP has the appropriate characteristics for development into a robust method for the detection of this sulphonamide in agri-food materials.

Animals↗
PubMed1996

Comparison of a microtitration plate ELISA with a standard cultural procedure for the detection of Salmonella spp. in chicken.

A rapid antibody-capture enzyme-linked immunosorbent assay (ELISA) detecting a wide range of Salmonella serotypes and employing only one culture stage was used to analyze the giblets and body cavity rinsings from frozen chickens. The results from the ELISA were compared with those obtained using a standard cultural procedure in current use in two laboratories, Norwich (N) and Ipswich (I), of the Public Health Laboratory Service (PHLS) in the UK. ELISAs were carried out on the same samples at each of two PHLS laboratories and at the Institute of Food Research with good agreement (94% and 90%). When compared with the cultural method there was 80% and 70% agreement with the ELISA with the PHLS(N) and PHLS(I) samples. The ELISA appeared to have a false-positive rate of 17% (samples from PHLS(N)) but on reculture of the "negative" samples this rate fell to 7%. The false-negative rate for the ELISA was 26% (samples from PHLS(N)) which appeared to be due to insufficient growth of the Salmonella spp. in the single cultural step employed in the ELISA rather than lack of recognition by the antibodies. The problem of false negatives with the cultural method is also discussed. These results are comparable to previously published studies relating immunoassays and the conventional procedure for Salmonella detection when analyzing similar samples. Suggestions are made as to how further increases in ELISA efficiency might be brought about.

Animals↗
PubMed1993

Further studies on the feasibility of one-day Salmonella detection by enzyme-linked immunosorbent assay.

A model system previously developed for the rapid detection of Salmonella typhimurium in foods was improved and extended to many other Salmonella serotypes. The original protocol, which consisted of an overnight nonselective culture followed by a specific enzyme-linked immunosorbent assay (ELISA), was modified and improved. A sandwich ELISA which used polyclonal antibodies for the capture stage and a cocktail of monoclonal antibodies for the detector stage was developed. The assay recognized a wide range of Salmonella serotypes; S. enteritidis, the most important serotype in the United Kingdom had a detection limit in the ELISA of about 4 x 10(2) cells ml-1. The cultural stage prior to the ELISA was either a single nonselective broth (incubated for 28 h) or a preenrichment broth (incubated for 7 h) plus a selective broth (incubated for 21 h). Antibodies which bind to cells grown in the unfavorable conditions of a selective medium were selected. It was concluded that, in the future, the shortened protocols for the detection of Salmonella spp. in foods described here will be of considerable value.

Animals↗
PubMed1990

Enzyme-linked immunosorbent assay for Salmonella typhimurium in food: feasibility of 1-day Salmonella detection.

A microtitration plate, antibody-capture, enzyme-linked immunosorbent assay was developed for detection of Salmonella typhimurium. The assay utilizes a monoclonal detector antibody which shows no cross-reactions with non-Salmonella species and only a slight cross-reaction with one other Salmonella serotype. By using only one cultural stage (in a nonselective, chemically defined medium) prior to the enzyme-linked immunosorbent assay, low numbers of cells in food (10 cells 25 g-1) were detected in 19 h. Non-Salmonella competing organisms did not interfere with detection of S. typhimurium even when present in the ratio of 10(6):1 (non-Salmonella/Salmonella spp.). The assay shows the feasibility of rapid, 1-day testing for Salmonella spp. with antibody technology.

Antibodies, Monoclonal↗
PubMed1988

Flatulence--causes, relation to diet and remedies.

In addition to causing embarrassment and unease, flatulence is linked to a variety of symptoms, some of which may be distressing. This review describes the origins of intestinal gas, its composition and methods which have been developed for its analysis. Emphasis is placed upon the effects of legumes in the diet in producing excessive intestinal gas and, particularly, on the role of raffinose-type oligosaccharides, containing alpha-galactosidic groupings. Suggestions for overcoming the problem are presented, including drug treatment, enzyme treatment, food processing and plant breeding. It is emphasised that removal of all raffinose-oligosaccharides from beans does not remove the problem of flatulence in animals and man; the compounds responsible--though assumed to be polysaccharides (or polysaccharide-derived oligomers formed by processing or cooking)--have yet to be characterised.

Animals↗
PubMed1988

Intestinal microflora and gastrointestinal adaptation in the rat in response to non-digestible dietary polysaccharides.

1. A comparison was made of the effect of a fibre-free diet and diets containing non-digestible polysaccharides on rat caecal and colonic physiology and microflora. 2. All polysaccharide-containing diets led to enlargement of the caecum and colon, associated with increased weight of contents, and of tissue. Carboxymethylcellulose (CMC) had the most marked effect and animals given this also had watery faeces. 3. The density of bacteria in the caecum and colon varied significantly with diet and the proportion of aerobic bacteria in the flora was increased by the CMC diet. 4. In vitro, CMC and hydroxypropylmethylcellulose were poorly fermented. 5. There was a high correlation (caecum r 0.93; colon r 0.94) between tissue weight and wet weight of organ contents but no correlation with bacterial density, number of bacteria per organ, moisture content or short-chain fatty acid content. 6. It is concluded that caecal and colonic enlargement is due to tissue hypertrophy in response to increased bulk of contents, irrespective of the nature of that bulk which varies with diet; it is unlikely that short-chain fatty acids or other microbial metabolites are the stimulus for the trophic response seen when non-digestible dietary polysaccharides are fed to rats.

Adaptation, Physiological↗
PubMed1988

Response of populations of human faecal bacteria to viscosity in vitro.

The effects of viscosity on cultures of human faecal bacteria in vitro were studied. The composition of the microbial populations which developed during in vitro incubation were affected by viscosity. The proportion of Bacteroides species was decreased at high viscosities and it is concluded that the 'viscotolerance' of a bacterial population contributes towards its composition. Viscosity alters the enzyme activity of cultures. Cell-associated total, and specific, beta-galactosidase activities of cultures of human faecal bacteria were decreased at high viscosity.

Bacteria↗
PubMed1987

Comparison of populations of human faecal bacteria before and after in vitro incubation with plant cell wall substrates.

Human faecal slurries were incubated anaerobically with larchwood xylan, oat spelt xylan, wheat bran, apple cell walls or sugar beet pulp as sole carbon sources. The populations which developed during incubation were different from the inoculum, the most marked changes being an increase in the number of Bacteroides species and a decrease in the number of Fusobacterium species for all carbon sources tested. With a water-soluble preparation of larchwood xylan the population was dominated by species able to ferment this substrate, in contrast to the population which developed with the insoluble substrates. The ability to use one plant cell wall substrate appeared to be related to the ability to use others. Strains capable of using plant cell wall substrates included Bacteroides spp., Clostridium clostridiiforme, Bifidobacterium longum, Fusobacterium spp. and Escherichia coli. When incubation with two contrasting substrates (bran and larchwood xylan) was replicated, the populations which developed were reproducibly different from the inoculum and from each other.

Bacteroides↗
PubMed1986

A change in human faecal flora in response to inclusion of gum arabic in the diet.

1. Gum arabic is a water-soluble polysaccharide resistant to human gut enzymes and thus can be described as dietary fibre. 2. Using a most-probable-number technique, estimates were made of total anaerobes and of gum-arabic fermenters in the faeces of a volunteer during a control period and during addition of 10 g gum arabic/d to the diet. Using an enrichment technique, the principal bacteria able to utilize gum arabic as the only carbohydrate source were isolated and characterized. 3. Faecal samples were analysed for undegraded gum arabic and, following acid-hydrolysis, for total sugars. 4. The proportion of the faecal flora able to degrade the gum arabic polymer rose from an initial level of 6.5% to more than 50% during gum-arabic ingestion, and subsequently returned to the control level after ingestion ceased. The principal gum-arabic fermenters were species of Bacteroides and Bifidobacterium. 5. Undegraded gum arabic was not detected in any faecal sample nor were there significant differences in the level of total sugars in acid-hydrolysed faeces between gum arabic and control periods. 6. The results presented indicate a direct and rapid change in faecal flora in response to a specific change in the diet of a human volunteer.

Adult↗
PubMed1986

Intestinal microflora, morphology and enzyme activity in zinc-deficient and Zn-supplemented rats.

1. Immature, male Wistar rats were given a low-zinc diet (2 mg/kg) for 22-24 d. Control groups received a similar diet supplemented with 58 mg Zn/kg either ad lib., or in amounts matched to the consumption of the Zn-deficient group. Food consumption, rate of growth and food conversion efficiency were markedly lower in the Zn-deficient group of rats compared with controls. Appetite, growth rate and food utilization improved dramatically over a subsequent 4 d period of Zn supplementation. 2. Morphological examination of samples of jejunum and ileum confirmed that Zn deficiency in the rat is accompanied by a reduction in villous dimensions and increase in villous density. After a short period of Zn supplementation, villous density and the basal width and maximum height of individual villi in the jejunum returned to normal. Similar changes occurred in the ileum but to a lesser extent. 3. Mucosal alkaline phosphatase (EC 3.1.3.1) activity was significantly lower in the small intestine of Zn-deficient rats compared with Zn-supplemented rats. Disaccharidase activities were lower in the Zn-deficient group, compared with their feed-restricted counterparts, but were similar to values for ad lib.-fed controls. Tissue alkaline phosphatase and disaccharidase activities were consistently higher after a 4 d period of Zn supplementation, compared with non-supplemented animals, but this increase was only significant for alkaline phosphatase. 4. Although there were striking similarities in the mucosal characteristics of gnotobiotic and Zn-deficient rats, there was no indication that even severe dietary Zn depletion reduced the numbers of viable bacteria present in either the small or large intestine.(ABSTRACT TRUNCATED AT 250 WORDS)

Alkaline Phosphatase↗
PubMed1979

Renal vein thrombosis: an underdiagnosed complication of multiple renal abnormalities.

Thirty-one cases of renal vein thrombosis (RVT) were reviewed retrospectively for clinical laboratory, and radiographic findings. An underlying renal disorder was present in 28 cases, absent in only 3. This supports other evidence that RVT is usually a complication of renal disease rather than a primary event, and that nephrotic syndrome may be due to renal disease rather than RVT. The findings also confirmed the large spectrum of urographic appearances in RVT, and were used as a basis for developing specific and liberal indications for renal venography.

Humans↗