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J E Croft

Publications and source records attributed to J E Croft.

11 recordsLinked to original sources
PubMed2003

Indirect enzyme-linked immunosorbent assay for the detection of antibody against Rift Valley fever virus in domestic and wild ruminant sera.

An indirect enzyme-linked immunosorbent assay (I-ELISA) for the detection of specific IgG immunoglobulins against Rift Valley fever virus (RVFV) was validated in-house. A total of 3055 sera from sheep (n = 1159), goats (n = 636), cattle (n = 203), African buffalo (n = 928), and other wild ruminants (n = 129), including eland, kudu, and black wildebeest, was used. Sera from domestic ruminants were collected in West (n = 10), South (n = 1654) and East Africa (n = 334), and sera from wild ruminants (n = 1064) were collected in South Africa. In addition, 136 sera from eight experimentally RVFV-infected sheep, taken during a period of 28 days post infection (dpi), were used to study the kinetics of RVFV antibody production. Field sera were tested by the serum neutralization (VN) test and experimental sera by VN and haemagglutination-inhibition (HI) test. Based on VN test results, negative sera were regarded as reference controls from RVFV-free, and positive sera were regarded as reference controls from RVFV-infected subpopulations of animals. ELISA data were expressed as the percentage positivity (PP) of an internal high positive control. The two-graph receiver operating characteristics approach was used for the selection and optimization of I-ELISA cut-offs including the misclassification costs term and Youden index (J). In addition, cut-off values were determined as the mean plus two-fold standard deviation of the result observed with the RVFV-free subpopulations. Established optimal cut-offs were different for each of the data sets analyzed, and ranged from 1.65 PP (buffalo) to 9.1 PP (goats). At the cut-off giving the highest estimate of combined measure of diagnostic accuracy (highest J value), the I-ELISA test parameters were determined as follows: (1) Diagnostic sensitivity (%): cattle--84.31, buffalo--94.44, sheep--98.91, goats--99.18. (2) Diagnostic specificity (%): cattle--99.34, buffalo--98.28, sheep--99.16, goats--99.23 and other game ruminants--99.26. In the group of RVFV-experimentally infected sheep, seroconversion In all individuals was detected by VN on 4-6 dpi, by HI on 5-7 dpi, and by I-ELISA on 6-7 dpi. All tests showed the same kinetic pattern of immunological response. Antibody levels were low for a very short period before increasing to high titres, after which it was easily detectable by all tests. Compared to traditional tests, the lower sensitivity of I-ELISA in the detection of the earliest stage of immunological response may be practically insignificant, particularily when this assay is used in population-based, disease-surveillance programmes. The high sensitivity and specificity of I-ELISA established in this study, especially for the statistically more representative subpopulations of animals tested, seem to support this prediction. Test parameters determined in this study should, however, be regarded as in-house diagnostic decision limits, for which further updating is recommended, particularly for specimens from other countries, and preferably by applying a standardized method for sampling of new subpopulations of animals to be targeted by the assay.

Animals↗
PubMed1987

Expression of leucine genes from an extremely thermophilic bacterium in Escherichia coli.

The organisation of the leucine genes in Thermus thermophilus HB8 was analysed by examining the ability of recombinant DNAs to complement Escherichia coli mutations. The arrangement of the genes is different from that in the mesophilic bacteria E. coli and Salmonella typhimurium. The promoter responsible for the expression of the leuB, leuC and leuD genes of Thermus HB8 in E. coli was identified. The sequence of Thermus DNA containing this promoter revealed structural similarities to the promoter and attenuator regions of the E. coli leucine operon.

3-Isopropylmalate Dehydrogenase↗
PubMed1986

Unique properties of NADPH- and NADH-dependent metabolism of p-nitroanisole catalyzed by cytochrome P-450 isozyme 2 in pulmonary and hepatic microsomal preparations from rabbits.

Approximately 90% of the NADPH- and NADH-dependent O-demethylation of p-nitroanisole (PNA) in the hepatic microsomal fraction from phenobarbital (PB)-treated rabbits and in the pulmonary microsomal fraction from untreated rabbits is catalyzed by the same isozyme of cytochrome P-450. This isozyme of cytochrome P-450 catalyzes less than 60% of this reaction in the hepatic microsomal fraction from untreated rabbits. Antibodies to NADPH-cytochrome P-450 reductase inhibit NADPH-dependent metabolism of p-nitroanisole by about 90% but have no effect on NADH-dependent metabolism. Hepatic NADPH-dependent metabolism of pNA and reduction of cytochrome c are inhibited to the same extent with varying amounts of antibodies to NADPH cytochrome P-450 reductase. The same relationship between inhibition of monooxygenase and reductase activities is observed for the hepatic and pulmonary metabolism of benzphetamine and 7-ethoxycoumarin. In contrast, the relationship between inhibition of the pulmonary NADPH-dependent metabolism of pNA and reductase activity is biphasic; at 75% inhibition of reductase activity, metabolism of pNA is inhibited by less than 25%. For NADH-dependent metabolism of pNA, our results indicate that both electrons are transferred to cytochrome P-450 from cytochrome b5.

Animals↗
PubMed1985

Deacetylation to 2-aminofluorene as a major initial reaction in the microsomal metabolism of 2-acetylaminofluorene to mutagenic products in preparations from rabbit lung and liver.

The rabbit pulmonary and hepatic microsomal pathways for the metabolism of 2-acetylaminofluorene (AAF) and 2-aminofluorene (AF) to mutagenic products were investigated by means of high performance liquid chromatography and the Salmonella mutagenicity assay. Mutagenic activity approached a maximum with increasing concentrations of AAF incubated with hepatic microsomal preparations and Salmonella; with pulmonary microsomal preparations, mutagenic activity was proportional to the concentration of AAF over the range examined. The mutagenic activities of AF exhibited typical saturation kinetics with both hepatic and pulmonary microsomal preparations. Approximately 7 times more AF than N-hydroxy-2-acetylaminofluorene (N-hydroxy-AAF) was formed in incubations of AAF (0.5 mM) with hepatic microsomal preparations. When AAF was incubated with pulmonary microsomal preparations, formation of AF, but not N-hydroxy-AAF, was detected. The inclusion of paraoxon in the pulmonary incubations blocked the formation of AF but did not lead to the recovery of any N-hydroxy-AAF. We conclude that the metabolism of AAF to mutagenic products in pulmonary microsomal preparations from rabbits is initiated primarily, if not entirely, by deacetylation of AAF to AF. The mutagenic activity of AAF with the pulmonary microsomal preparations is limited by the deacetylase activity which, like mutagenic activity, exhibits a linear relationship with the concentration of AAF. On the basis of the rates of formation of AF and N-hydroxy-AAF and their mutagenic activities, we estimate that about 60% of the hepatic metabolism of AAF to mutagenic products is dependent upon deacetylation of AAF and subsequent oxidation of the AF formed.

2-Acetylaminofluorene↗
PubMed1983

A cryptic plasmid from Shigella sonnei.

pNZ500 is a 1.5 kb cryptic plasmid from a Shigella sonnei isolate. It was introduced into Escherichia coli by cotransformation, where it is maintained at about 30 copies per chromosome equivalent. Hybridization studies show that pNZ500 exhibits a high level of sequence similarity to other 1.5 kb plasmids found in different S. sonnei isolates but shares no homology with larger S. sonnei plasmids. pNZ500 shares a small degree of sequence homology with pBR322 and with pAC184. The homology with pBR322 is restricted to sequences close to the ori-bom region of this plasmid. Nevertheless, pNZ500 maintenance in E. coli is not dependent on DNA polymerase I activity, and does depend on continuing protein synthesis. pNZ500 encodes two polypeptide gene products whose monomer molecular weights are 24500 and 18000. The examination of host cells for the expression of possible plasmid phenotypes revealed no differences between cells bearing pNZ500 and plasmidless cells.

Bacterial Proteins↗
PubMed1983

The relationship between increases in the hepatic content of cytochrome P-450, form 5, and in the metabolism of aromatic amines to mutagenic products following treatment of rabbits with phenobarbital.

Treatment of rabbits with phenobarbital is followed by increases in the hepatic microsomal concentration of cytochrome P-450, form 5, and in the hepatic microsomal metabolism of aromatic amines to mutagenic products. Inhibition by antibodies to form 5 of the activation of 2-aminoanthracene and 2-aminofluorene demonstrates that these increases are directly related. The extent of the apparent induction of form 5 by phenobarbital is determined from single radial immunodiffusion, immunostaining of form 5 on nitrocellulose sheets containing microsomal proteins transferred from polyacrylamide gels, and the amount of antibody required for 50% of maximal inhibition of activity. The results indicate that phenobarbital increases the hepatic microsomal concentration of cytochrome P-450, form 5, to the same extent that it increases form 5-mediated metabolism of aromatic amines to mutagenic products: 10- to 12-fold. In contrast to the effects of phenobarbital, treatment of rabbits with beta-naphthoflavone decreases the hepatic microsomal concentration of cytochrome P-450, form 5, to less than detectable levels and has little effect on the metabolism of aromatic amines to mutagenic products. Our findings, along with the known effects of phenobarbital on cytochrome P-450, form 2, and the known catalytic activity of cytochrome P-450, form 4, lead to the following conclusions: (a) treatment of rabbits with phenobarbital results in increases in the hepatic microsomal concentrations of at least two structurally, immunochemically, and catalytically distinct isozymes of cytochrome P-450, forms 2 and 5; (b) the metabolism of aromatic amines to mutagenic products in rabbit hepatic microsomal preparations depends on the relative concentrations of at least two isozymes of cytochrome P-450, forms 4 and 5, that change in response to different inducers.

Amines↗