Evaluation of a survey approach to estimating the prevalence of cattle carrying antimicrobial-resistant Escherichia coli.

A random survey of farms in the Highlands and Islands of Scotland provides estimated of the prevalence of calves, finishers and cows carrying ampicillin, apramycin and/or nalidixic acid resistant Escherichia coli. While the survey provides information on the geographical variation in risk, the results are of limited value for interpreting causality.

Dietary sulphur amino acid adequacy influences glutathione synthesis and glutathione-dependent enzymes during the inflammatory response to endotoxin and tumour necrosis factor-alpha in rats.

1. Glutathione concentrations in liver and lung fall when food intake or sulphur amino acid intake is inadequate. However, concentrations may be restored during inflammation, despite anorexia, provided that prior sulphur amino acid intake is adequate. 2. We studied the mechanisms of these changes by measuring the effect of sulphur amino acid and protein intake on hepatic glutathione synthesis and gamma-glutamylcysteine synthetase activity, hepatic and lung glutathione concentrations, glutathione reductase and glutathione peroxidase activities in young rats given an inflammatory challenge by intraperitoneal injection of tumour necrosis factor-alpha or endotoxin (lipopolysaccharide). 3. Diets containing 200 g of casein and 8 g of L-cysteine/kg (normal-protein diet), or 80 g of casein and 8 g of L-cysteine, or isonitrogenous amounts of L-methionine or L-alanine (low-protein diets) were fed ad libitum to young Wistar rats for 8 days. Dietary groups were subdivided into three: one subgroup continued feeding ad libitum, a second was given tumour necrosis factor or lipopolysaccharide and killed 24 h thereafter, while the third was pair-fed to the intakes of the second subgroup for 24 h before being killed. 4. Glutathione concentrations in liver and lung were reduced in rats fed the low-protein diet containing alanine, and in all dietary groups when food intake was restricted. The inflammatory challenges restored hepatic glutathione concentrations in all groups but the diet supplemented with alanine, which had an inadequate sulphur amino acid content. In lung, restoration occurred only in animals fed the normal-protein diet. 5. The activity of gamma-glutamylcysteine synthetase, which is rate limiting for glutathione synthesis, was unaffected by dietary or sulphur amino acid intake or by the inflammatory response. Substrate supply may therefore be a major determinant in glutathione synthesis in vivo. 6. Total hepatic glutathione synthesis was affected by food intake, the type and amount of sulphur amino acids in the diet and by inflammation. Total synthesis was 207, 137, 421 and 90 mumol/day for animals fed ad libitum the normal-protein diet, or low-protein diets supplemented with cysteine, methionine or alanine respectively, ad libitum. Pair-feeding resulted in values of 76, 31, 71, and 0 mumol/day respectively. After lipopolysaccharide injection, rates increased to 200, 117, 151 and 56 mumol/day respectively. 8. Reductase and peroxidase activities increased in liver and lung, when low-protein diets which contained supplemental methionine or alanine were consumed ad libitum. A reduction in food intake resulted in enzyme activity changes, which suggested that recycling of glutathione increased in lung and decreased in liver. Injection of tumour necrosis factor reversed this effect. 9. The restoration of glutathione concentrations in liver after an inflammatory challenge is closely associated with an enhanced rate of synthesis and increased recycling. The former is impaired when inadequate sulphur amino acid is consumed before the challenge. In lung, increased recycling of glutathione may help maintain concentrations when food intake is restricted, but not during inflammation.

Gp120 sequence variation in brain and in T-lymphocyte human immunodeficiency virus type 1 primary isolates.

OBJECTIVE: We sought to identify genetic determinants within human immunodeficiency virus type 1 (HIV-1) gp120 that differentiate viral species in brain from those in lymphocytes. STUDY DESIGN/METHODS: Polymerase chain reaction (PCR) was used to amplify gp120 genes from paired brain and T-cell isolates from two infants and two adults with dementia. Amplimers were molecularly cloned and sequenced. RESULTS: The degrees of amino acid divergence between brain and T-cell clones for the two adults were 7.7% and 8.6% and, for the two infants, 2.4% and 2.7%. For one adult and one infant, divergence was greater among brain cell clones than T-cell clones. In the adults, a 3-amino acid insertion, located similarly within V4 and encoding asparagine residues, was identified in the T-cell clones. CONCLUSIONS: HIV-1 genetic variation within brain cells is not necessarily restricted compared with that in blood lymphocytes. The species in brain cells can be distinguished from those in lymphocytes by determinants within V4. These differences suggest that immune-mediated selection ongoing within lymphoid cell compartments may not occur within brain.

3-Acetyl deoxynivalenol production in a strain ofFusarium culmorum insensitive to the fungicide difenoconazole.

The production of the mycotoxin, 3-acetyl deoxynivalenol (3-ADON), was investigated in a strain ofFusarium culmorum insensitive to the systemic fungicide, difenoconazole. On exposure to graded concentrations of the fungicide, the insensitive strain continued to synthesise 3-ADON when difenoconazole levels of 100 and 200µg/ ml media were used. In contrast, a control (sensitive) strain ceased production of 3-ADON at difenoconazole levels of 100 µg/ml. Differences between the two strains were also observed for 3-ADON production with time. Following incubation with fungicide at 0.1 µg/ ml, 3-ADON production occurred more rapidly in CS than in IS cultures. This is the first report of increased persistence and alteration of the pattern of production of a mycotoxin following the development of fungicide insensitivity in a fungal phytopathogen.

Accuracy of least squares designed spatial FIR filters for segmentation of images of fluorescence stained cell nuclei.

A method for accurate, real-time image segmentation is needed for the development of a fully automated image cytometer that combines the speed and case-of-use of flow cytometry with the detailed morphometry of imaging. Object intensity variation and inherent optical blur make real-time segmentation challenging. The best spatial finite impulse response (FIR) filter, implemented as a convolution, was tested for sharpening edges and creating the required contrast. The filter and threshold segmentation steps were treated as a two-category linear classifier. Best 3 x 3 through 25 x 25 filters were designed utilizing the perceptron criterion and nonlinear least squares, and tested on ten montage images of a combined 1,070 manually segmented DAPI stained cell nuclei. The resulting image contrast, or class separation, led to simple automatic thresholding via the histogram intermodal minimum. Image segmentation accuracy began to plateau at 7 x 7 filters and did not increase above 15 x 15. Little loss in accuracy occurred with application to the images not used for design. This segmentation method provides a systematic, fast and accurate means of creating binary object maps useful for subsequent measurement, processing and cell classification.

Cysteine and methionine supplementation modulate the effect of tumor necrosis factor alpha on protein synthesis, glutathione and zinc concentration of liver and lung in rats fed a low protein diet.

The synthesis of cysteine-rich compounds such as glutathione and metallothionein is an integral part of the response to cytokines. To examine the essentiality of an adequate supply of sulfur amino acids during a response to tumor necrosis factor alpha (TNF) we fed rats a low protein (8% casein) diet supplemented with either cysteine and alanine, methionine and alanine, or alanine alone, or a normal protein (20% casein) diet for 8 d before injection with TNF or saline. Those animals injected with saline were pair-fed the intake of their TNF-injected counterparts for the 24 h after injection. In a second experiment, control groups fed the same diets but receiving no treatment were also examined to establish baseline values. Although few significant differences between the non-injected animals consuming food ad libitum were apparent, TNF injection and pair-feeding resulted in differences between the dietary groups. Supplementation of the low protein diet with either cysteine or methionine improved growth and increased liver and lung glutathione concentration, zinc concentration, protein concentration and protein synthesis compared with results for the alanine-supplemented group. Lung polymorphonuclear cells were proportionally elevated in the TNF-treated, alanine-supplemented group compared with the other dietary groups treated with TNF. The changes in protein synthesis and glutathione concentration of the liver in response to TNF showed that sulfur amino acids may be partitioned to a greater extent into hepatic protein than into glutathione when sulfur amino acid intake is low. Consequently the adequacy of dietary sulfur amino acids will determine the extent to which antioxidant defenses are maintained during inflammation.

Adaptation of HIV-1 to pigtailed macaques.

In vitro infectivity experiments were performed to assess the susceptibility of cells from various monkey species to HIV-1. T lymphocytes from pigtailed macaques, but not those from rhesus or cynomolgus monkeys, were susceptible to infection, but virus expression was limited. The majority of HIV-1 isolates were unable to productively infect pigtailed macaque cells. Inoculation of autologous, HIV-1 expressing cells led to establishment of persistent infection in pigtailed macaques as evidenced by recovery of infectious virus and development of virus-specific antibody responses.

HIV-1 infection in pigtailed macaques.

Four pigtailed macaques were inoculated with autologous cells expressing low levels of human immunodeficiency virus type 1 (HIV-1). During the first 10 weeks, infectious virus was recovered from peripheral blood mononuclear cells (PBMCs) and lymph nodes from three of the animals. Subsequently, HIV-1 DNA was frequently detected in uncultured PBMCs from all three animals, and virus was isolated from one of them at weeks 38 and 61. The fourth animal, which was rechallenged at week 10 with cell-free virus isolated from one of the others, never became virus isolation positive, but harbored HIV-1 proviral genomes. These virus infections were accompanied by the development of varied HIV-1-specific humoral immune responses. Antibodies to gp160 were first apparent at week 8 in the three initially infected animals and persisted. The animal from whom virus was isolated at late times also developed persisting antibodies to HIV-1 p24 and gp120. Antibodies to gp120 and gp160 became apparent in the rechallenged animal at 1 week following reinoculation, but they waned with time. In vivo passage of the virus was attempted at week 6. One recipient pigtailed macaque and one recipient cynomolgus monkey failed to become detectably infected following transfusion of virus-positive blood and lymph node cells. The long-term presence of HIV-1-specific antibodies and proviral genomes in these animals, and the recovery of infectious virus more than 1 year following inoculation, are indicative of persistent infection, and confirm previous reports that pigtailed macaques are susceptible to HIV-1.

HTLV-III infection of EBV-genome-positive B-lymphoid cells with or without detectable T4 antigens.

The infection of a number of new and established B-cell lines by human T-cell lymphotropic virus III (HTLV-III) was investigated. The B lymphocytes differed in their expression of T4 antigens detected by specific monoclonal antibodies (MAbs) and the presence of Epstein Barr virus (EBV)-DNA or antigens. The presence of the EBV genome was the only requirement for infection of B-lymphocytes by HTLV-III, although its presence did not ensure infection. Two EBV genome and T4 antigen-positive B-cell lines, lacking EBV early antigens (EA) and viral capsid antigens (VCA), could be productively infected with no induction of known EBV antigens. Two other EBV genome-positive cell lines, lacking T4, EA, and VCA could also be infected. Another genome-positive cell line (P3HR-I) that was EBV-EA, VCA-positive and produced non-transforming EBV, could also be infected by HTLV-III. However, 3 EBV genome- and T4 antigen-negative B-cell lines could only be infected with HTLV-III after successful conversion to an EBV-genome-positive state by pre-infection with EBV. Five other EBV-genome-positive B-cell lines lacking T4 antigens were not infectible with HTLV-III even after super-infection with EBV. Incomplete inhibition of the HTLV-III infection of a T4-positive (LDV-7) and a T4-negative (Craig) was obtained by preadsorption with specific MAb to T4 (OKT4A and Leu 3A). From these observations, it is not clear whether the presence of T4 antigen on the cell surface is needed for the infection of B lymphoblastoid cells; however, successful infection does depend upon the presence of the EBV genome. The mechanism of interaction of HTLV-III and EBV-infected B-cell lines permitting this infection is not fully understood. Although the clinical implications of these observations remain to be determined, it is possible that infection of EBV-positive B-cells may contribute to aberrant humoral responses and/or increased frequency of B-cell malignancies observed in HTLV-III-infected individuals.

Presence of HTLV-III in tears and cells from the eyes of AIDS patients.

Tears, conjunctival epithelium, and corneoscleral tissue from AIDS patients were used for the isolation of HTLV-III and also for identifying cell types which support its replication. HTLV-III was isolated from tears of AIDS patients (66.6%) by cultivation of cells and fluid from patients' eyes with fresh human mononuclear cells. The cells from the conjunctival scrapings of these patients (33.3%) revealed HTLV-III antigens by indirect immunofluorescence (IF) using anti-P24 and P15 monoclonal antibodies. HTLV-III from the cell-free supernatant of the infected mononuclear cells from two patients' cocultures were further transmitted into fresh cells. The cells from right and left central cornea, as well as limbal cornea from an asymptomatic HTLV-III antibody-positive individual and one AIDS patient revealed HTLV-III upon cocultivation. HTLV-III P15 and P24 antigens were detected in cultured primary cornea epithelial cells. The tears and conjunctival cells from a control group were found to be free of HTLV-III. Although no documented cases of AIDS have been reported in corneal transplant recipients, serologic screening of donors prior to the use of the tissues for transplantation is advisable. Our data also raises important questions regarding possible transmission of virus during ophthalmologic examination by way of examiner's hands, through instruments and during contact lens fittings. Moreover, these findings indicate the need for testing various eye disinfectants for virus inactivation and/or inhibition.

Influence of time of feeding on time of parturition in beef cows.

The influence of time of feeding pregnant beef cows on the timing of parturition was investigated. The results suggest that the distribution of calvings between 'day' and 'night' can be altered by time of feeding. The calculations, based on a small quantity of data, suggest that feeding at 22.00 would result in 79 per cent of cows calving between 06.00 and 22.00. On the same basis of calculation the traditional 09.00 time of feeding would result in 57 per cent of cows calving during the 'day' period. There appears to be some evidence that by feeding pregnant beef cows in the evening better supervision of calving with a smaller proportion of cows calving at night can be achieved.

The 1979 revision of Z80.1--the American National Standard recommendations for prescription ophthalmic lenses--the AOA position.

Since April, 1972, the American National Standard Requirements for First-Quality Prescription Ophthalmic Lenses, Z80.1-1972, has been in the process of revision. The American National Standards Institute procedures are explained as well as details of the new revised standard. In addition, an explanation of the AOA position on the revision is given. Finally, what this standard means to the practicing optometrists is discussed.