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Objective: To evaluate the tracking of children's fruit and vegetable intake over 2 years., Design: Children in the control group from the Gimme 5 nutrition education intervention trial completed three 7-day food diaries once a year in 1994, 1995, and 1996, beginning when they were in third grade., Subjects: Five hundred sixty-one students from public elementary schools., Statistical Analyses Performed: Tracking was examined through correlation of fruit and vegetable servings over the 2 years as well as the percent of subjects remaining in the same or adjacent quintiles of fruit and vegetable intake, using the kappa statistic., Results: Tracking of fruit and vegetable intake was fair to moderate over the 2 years. One-year correlations for total fruit and vegetable servings were .37 for boys and .46 for girls; 2-year correlations were .48 for boys and .40 for girls. kappa values for remaining in the same quintile generally exceeded chance rates only for the top and bottom quintiles. No values exceeded .50. APPLICATION/CONCLUSION: The drift in ranking may have been a result of true changes in fruit and vegetable intake or measurement error (i.e., inability to accurately assess actual intake). If this drift reflects true change, it may be difficult for nutrition education programs targeting this age group to demonstrate long-term behavioral effects on fruit and vegetable intake. If the drift is the result of measurement limitations, improved dietary assessment may yield stronger tracking.

The impact of school health education programs is often attenuated by inadequate teacher implementation. Using data from a school-based nutrition education program delivered in a sample of fifth graders, this study examines the discriminant and predictive validity of three measures of curriculum implementation: class-room observation of fidelity, and two measures of completeness, teacher self-report questionnaire and post-implementation interview. A fourth measure, obtained during teacher observations, that assessed student and teacher interaction and student receptivity to the curriculum (labeled Rapport) was also obtained. Predictive validity was determined by examining the association of implementation measures with three study outcomes; health knowledge, asking behaviors related to fruit and vegetables, and fruit and vegetable intake, assessed by 7-day diary. Of the 37 teachers observed, 21 were observed for two sessions and 16 were observed once. Implementation measures were moderately correlated, an indication of discriminant validity. Predictive validity analyses indicated that the observed fidelity, Rapport and interview measures were significantly correlated with post-test student knowledge. The association between health knowledge and observed fidelity (based on dual observation only), Rapport and interview measures remained significant after adjustment for pre-test knowledge values. None of the implementation variables were significantly associated with student fruit and vegetable intake or asking behaviors controlling for pre-test values. These results indicate that the teacher self-report questionnaire was not a valid measure of implementation completeness in this study. Post-implementation completeness interviews and dual observations of fidelity and Rapport appear to be more valid, and largely independent methods of implementation assessment.

A high-performance liquid chromatographic method with diode-array detection for the determination of nicotine and its metabolites, cotinine and caffeine, in meconium is described. This method is suitable to assess foetus exposure to tobacco smoke. The analytes were extracted by solid-phase extraction before chromatography. From among 30 meconium samples 11 were positive for cotinine (20-86 ng/g) and 27 for caffeine (10-45 ng/g). No nicotine was present in the samples because of its rapid metabolism into cotinine.

Objective: This study examined whether providing a school-based teacher wellness program enhances the impact of a health curriculum on student outcomes and improves cognitive, behavioral, and physiological outcomes among participating teachers., Methods: Thirty-two elementary schools were randomly assigned to experimental or comparison conditions. Comparison group schools received the Gimme-5 program, a curriculum designed to increase fourth and fifty graders' consumption of fruits and vegetables. Experimental schools received Gimme-5 and the teacher wellness program, which included 54 workshops over 2 years, along with several schoolwide health activities. Physiological, behavioral, and cognitive outcomes were assessed in teachers and students., Results: There was no evidence that the intervention favorably modified any student or teacher end points; nor did intervention teachers deliver the Gimme-5 program with greater fidelity than comparison teachers., Conclusion: Confidence in the null results is bolstered by the randomized design, baseline sample equivalence, appropriate mixed-model analyses, and lack of selective or differential attrition. Insufficient participation in the wellness program appears a likely explanation for the lack of teacher and student effects. Factors specific to the school setting and intervention may have diminished participation and, thus, intervention effects.

We performed a comparative evaluation of anatomical and functional changes in kidneys after urinary tract reconstructive procedures and in patients with history of urinary tract infections using renal scintigraphy and other imaging techniques. Statistically significant differences in the prevalence of pathological findings in imaging between these groups were shown. When children after the RVU are taken into account renal scintigraphy supplies us with objective and precise data about the renal scarring degree. In this group repeated renal scintigraphy should be warranted. After the reconstructive procedures due to hydronephrosis the images from ultrasonography and renal scintigraphy correlate well. Ultrasonography is a golden standard for screening these patients. Statistically significant higher degree of renal scarring in renal scintigraphy in patients after urinary tract infections without urinary tract defect proves the important role of infections in the pathogenesis of these changes. Children with recurrent symptomatic urinary tract infections should be submitted to renal scintigraphy imaging on regularly basis, as a risk group of higher chronic pyelonephritis prevalence.

Objective: To assess differences in children's consumption of fruit and vegetables (F&V) by day of the week and meal of the day., Design: Baseline data from two school based nutrition education studies were combined for analysis., Subjects/setting: 2984 third grade students from 48 participating elementary schools in three school districts in the metropolitan Atlanta area., Measures of Outcome: The frequency of consumption of F&V abstracted by trained registered dietitians from prompted 7-day food records. STATISTICAL-ANALYSES PERFORMED: Mixed model analysis with meals and days as terms, controlling for the within school correlation, gender and ethnic group., Results: F&V were most frequently consumed at weekday lunch, and second most frequently at dinner. Participation in school lunch accounted for a substantial proportion of F&Vs consumed at lunch. Few F&Vs were consumed at breakfast or snack., Conclusions: School lunch makes an important contribution to elementary school students' F&V consumption. Dietary change programs should target parents to increase F&V consumption at dinner, and target students for the meals over which they assert the most control: breakfast and snacks.

Social-cognitive theory (SCT) was used to explain the fruit and vegetable intake of 1,398 3rd graders. SCT variables assessed included self-efficacy, outcome expectations, preferences, social norms, asking skills, and knowledge. Fruit and vegetable intake was assessed with 7-day records. Bivariate correlations with fruit and vegetable intake ranged from .17 for asking skills to .29 for fruit and vegetable preferences. In analyses controlling for school-level clustering, only preferences and positive outcome expectations remained significantly associated with fruit and vegetable intake, accounting for approximately 10%-11% of the variance. Limitations in the conceptualization, scope, and measurement of the variables assessed may have contributed to the weak associations observed. Models incorporating factors other than individual-level social-cognitive variables may be required to more fully explain children's dietary behavior.

Two novel series of 2-pyridones were synthesized by transposition of the nitrogen of 4-quinolones to the bridgehead position. This subtle interchange of the nitrogen atom with a carbon atom yielded two novel heterocyclic nuclei, pyrido[1,2-alpha]pyrimidine and quinolizine, which had not previously been evaluated as antibacterial agents and were found to be potent inhibitors of DNA gyrase. Quinolizines with a methyl group at the 9-position such as (S)-45a (ABT-719) demonstrate exceptional broad spectrum antibacterial activity. Most notably, they are active against resistant bacteria such as methicillin-resistant Staphylococcus aureus, vancomycin-resistant strains of enterococci, and ciprofloxacin-resistant organisms. In addition, 2-pyridones also possess favorable physiochemical and pharmacokinetic properties. These 2-pyridones were synthesized from the commercially available starting materials by 10-17 linear transformations. The structure of an adduct yielded by this sequence, (S)-45a (ABT-719), was determined by X-ray crystallographic analysis.

A TLC method for the separation of heavy metals and their complexes with dithizone, 4-(2-pyridylazo)resorcinol and ethylenediaminetetraacetic acid was devised and conditions for the solid-phase extraction of heavy metal ions in human bones, placenta and milk after microwave mineralization and in air after alkaline absorption were elaborated.

Objective: We developed, pilot-tested, and field-applied a stages of change questionnaire regarding fruit and vegetable (F&V) consumption among fourth- and fifth-grade students., Methods: The design included cross-sectional assessment of internal consistency and construct validity, and 2-week and 7-week longitudinal assessment of test-retest reliability. Subjects included 134 students from one school for pilot-testing and 252 from two schools for field application. Thirty-two questions from McConnaughy et al's stages of change questionnaire were adapted and pilot-tested; minor revisions were made for field application. Statistical analyses included principal components analysis to identify subscales; cluster analysis to identify subgroups within students; Cronbach's alpha coefficient to assess internal consistencies; Pearson product-moment correlations to assess test-retest reliabilities; and oneway ANOVA's by F&V stages of change clusters with actual F&V consumption, F&V self-efficacy subscales, F&V preferences, and F&V outcome expectations subscales to determine construct validity., Results: Principal components analysis from the field application indicated two subscales (precontemplation and beyond precontemplation) accounting for 39.5% of variance. Cluster analysis indicated 6 interpretable clusters; 2 (n = 63) provided responses inconsistent with the stages of change theory and 4 (n = 189) provided responses consistent with the theory. Internal consistencies and test-retest reliabilities were acceptable. Students in the "beyond precontemplation" clusters had higher levels of self-efficacy and outcome expectations regarding eating F&V., Conclusions: Measuring stages of change other than the precontemplation stage in F&V consumption among elementary school children is problematic. Perhaps the theoretical concept does not apply to children, or elementary school children lack the ability to comprehend the questions measuring the concept, or the approach used was not entirely appropriate.

Objective: Due to the increased emphasis on fruit and vegetable (F+V) consumption and a desire to use the simplest dietary assessment method appropriate to determine F+V intake, we assessed the reliability and validity of weekly and monthly fruit and vegetable food frequency questionnaires (F+V FFQ) among fourth and fifth grade students by comparing them to food records previously validated through school lunch observations., Methods: The multiethnic sample (primarily African-American and Caucasian) included predominantly lower-middle socioeconomic students from 10 classes at one elementary school. F+V FFQ were printed on optical scanning forms and classroom administered on two occasions. Students completed preprinted food record forms at school and received new forms weekly. To allow comparison between the F+V FFQ and food records, a dietitian abstracted information from the records according to a written protocol and transferred it to F+V FFQ forms covering respective time periods. F+V FFQ were converted to estimated daily servings and aggregated to create five F+V indices plus three summary indices. Statistical analyses included Spearman correlations and paired t-tests., Results: Students demonstrated acceptable reliability in completing food records as well as weekly and monthly F+V FFQ; however, validity was unacceptable for both F+V FFQ versions due to significant overreporting., Conclusion: Of the two methods studied, the food record procedure is preferred over the FFQ procedure for collecting self-reported F+V consumption data among fourth and fifth grade students.

Fourth- and fifth-grade students' food records completed by using a labor-intensive approach (with daily monitoring) or a less labor-intensive approach (with weekly monitoring) were compared with school-lunch observations. Records were completed at three elementary schools for 14-23 d by students in 26 classes. Record pages were printed forms with areas for recording items consumed, number of servings, and locations of meals. Students were randomly selected for lunch observation. Kappa coefficients and percent of agreement for interobserver reliability for both approaches combined ranged from 0.43 to 0.91 and 72.6% to 95.5%, respectively, on 24 observations. Pearson correlations for 117 record/observation matches were significant for eight and three of nine meal items for the daily and weekly monitoring approaches, respectively. Thus, students were reasonably accurate in keeping records and more accurate with the more labor-intensive approach. Researchers need to select an approach based on the degree of accuracy desired and available finances.

Various nutrition guidelines recommend increased consumption of fruits and vegetables (F+V); a Year 2000 goal targets five or more daily servings of F+V. Decisions are needed regarding how to define F+V to facilitate measurement of goal achievement. Four alternative definitions (narrow and broad, each with and without legumes) were developed and applied to food frequencies from 133 children and 211 parents. Results varied by definition. Mean intake met the goal of five or more daily servings regardless of definition and exceeded it for both broad definitions by children and both broad definitions and the narrow definition with legumes for adults. Median intakes met the goal for all definitions except for children with the narrow definition without legumes, and exceeded it for children with both broad definitions and for adults with the broad definition with legumes. These differences by definition indicate that a clear definition of F+V is needed.

Background: Achievement of the nation's Year 2000 fruit and vegetable goal of five or more daily servings may relate to children's preferences for fruits and vegetables. There are, however, no easily administered instruments for measuring preferences for fruits and vegetables among children., Methods: An initial questionnaire systematically varied preparation methods after focus group discussions had indicated that this had an impact on fruit and vegetable preferences. The three-part questionnaire (fruits, vegetables, fruit and/or vegetable snacks) was classroom administered. Internal consistencies and test-retest reliabilities were acceptable. Principal components analyses, however, revealed food-specific rather than preparation factors; thus, preparation was eliminated from a revised questionnaire., Results: Repeat classroom administration of the revised questionnaire yielded acceptable internal consistencies and test-retest reliabilities. Preferences were higher for fruits than vegetables. Principal components analyses indicated single fruit and vegetable factors and a two-factor solution for snacks (vegetables, fruits). Pearson correlations between preferences and consumption were significant, but accounted for only a small fraction of the variance. Correlations between fruit exposure and preferences, and between vegetable exposure and preferences were 0.55 and 0.51, respectively., Conclusion: The revised questionnaire provides a reliable, valid, and simple tool for assessing fruit and vegetable preferences of 4th- and 5th-grade students. The relationship between these preferences and actual consumption is weak; however, making fruits and vegetables more available may enhance preferences.

The level of physical activity of 3- and 4-year-old children was assessed in alternative physical locations by month and time of day and by age, gender, and ethnicity. Physical activity was assessed by observation with the Children's Activity Rating Scale (CARS) for up to 12 hours from 7:00 am to 7:00 pm. A sample of 191 three- and four-year-old children was observed for up to four times in the course of a year. The sample was tri-ethnic. Boys were significantly more active than girls. Activity was consistently higher outside than inside. There were significant differences in the amount of time children in this age group spent inside versus outside by time of year; the activity levels of boys and girls differed by time of year, particularly when outside. A model including gender, month, and location terms accounted for 75% of the variance in physical activity. These data further documented gender differences in physical activity among very young children using measures not subject to self-report biases but did not explain or clarify the gender differences. The substantial differences by physical location and time of year deserve future attention, but more refined methods will be needed to balance data by location and important seasonal times. An inference from these results is that activity levels among young children may be increased by encouraging them to spend more time outdoors.

DNA topoisomerases, a class of enzymes that change the topological structure of DNA, have been shown to be the target of many therapeutic agents, including antibacterial agents (quinolones) and anticancer agents. These drugs inhibit the enzyme in a unique way so that the enzyme is converted into a cellular poison. Candida albicans and Aspergillus niger are two major opportunistic fungal pathogens. Our results show that these fungi have high levels of both type I and type II topoisomerases (with a minimum of 5 x 10(5) ATP-independent relaxation units and 2 x 10(5) P-4 unknotting units per liter of wild-type C. albicans). The ATP-dependent type II topoisomerase (termed C. albicans topoisomerase II) was purified by approximately 2,000-fold from C. albicans cells by using a simple isolation scheme that consists of three column procedures: hydroxylapatite, phosphocellulose, and heparin-agarose chromatographies. The responses of the Candida and the calf thymus topoisomerase II to some known topoisomerase II inhibitors were measured. Etoposide and 4'-(9-acridinylamino)methanesulfon-m-anisidide, compounds known to inhibit catalysis and to enhance DNA breakage by mammalian topoisomerase II, and A-80198, an etoposide derivative, enhanced cleavage by both enzymes at similar concentrations of these compounds, with the response of the calf thymus topoisomerase II from slightly to fourfold higher in magnitude than the response of the Candida enzyme in the same concentration range. In contrast, A-75272 (a cytotoxic tricyclic quinolone) shows a slightly stronger DNA cleavage enhancement effect with the Candida enzyme than with the mammalian counterpart. The abundance of the enzyme in cells and the different drug responses of the host enzyme and the fungal enzyme suggest that the fungal topoisomerase may serve as a target for the discovery of effective and safe antifungal agents.

Barker recently hypothesized that factors affecting prenatal and infant growth are related to adult blood pressure and CVD mortality. Predictions from Barker's hypothesis in regard to infant feeding were tested among a sample of 3 or 4 year old children. The relationship of infant-feeding characteristics (duration of breast-feeding, times of introduction of high fat, high carbohydrate, high potassium foods and table salt) to indicators of cardiovascular functioning (resting blood pressures and heart rates, and heart rate response to graded activity) while controlling for anthropometric (height, sum of seven skinfolds, BMI) and demographic (ethnicity, gender, social status) characteristics revealed that infant-feeding practices were not related to CV functioning in the predicted directions among this sample of 3 or 4 year old children. Furthermore, the positive relationship between height and systolic blood pressure was inconsistent with the Barker hypothesis.

We assessed the effects of socioeconomic status and amount of time mother and child were together during the day on the accuracy of the mother's 24-hour recall report of her 3- or 4-year-old child's dietary intake. Accuracy of the mother's report was compared for nutrients and foods with dietary intake recorded by an observer. Neither socioeconomic status nor amount of time mother and child were together were related to any of the nutrient indicators of accuracy. Although socioeconomic status and amount of time mother and child were together were not related to the indicators of food accuracy, only 64.8% agreement was obtained between mothers and observers on foods consumed. Mothers were more likely to underreport (17.7%) than overreport (10.4%) foods. The low mean difference scores for nutrient indicators supported the use of the 24-hour recall method when collecting group data. The substantial disagreement on foods, however, suggests that previous studies assessing agreement on nutrients have overestimated the accuracy of the 24-hour recall method.

It is proposed to identify histologically indeterminate human cancers by comparing their qualitative content of stained proteins, separated by two-dimensional electrophoresis, with patterns of proteins from a series of identified human cancers of comparable or different stem cell origin. The hypothesis to be tested is that in cancer, biochemical ontogeny, reflected in the distribution of cellular and tissue proteins, will recapitulate phylogeny, albeit in a manner less organized than normal. Patterns of protein from cancer cells, whether histologically identified or not but originating from embryologically similar stem cells, are expected to exhibit greater homology, compared with protein profiles of cancer cells derived from stem cells of different embryologic origin. Normal or cancer cell proteins common or unique to one or more cell sources can be identified, and subsets of proteins that characterize cancer cells of specific stem cell origin identified. There are reasons to believe that cancer cells rarely, if ever express properties inconsistent with the basic cellular commitment of their stem cell of origin. Photo-optical scanning and computer-assisted analysis will greatly facilitate collating the many hundreds of discrete proteins, among which those used to characterize different cancer cells are to be found.

The distribution of Coomassie blue-stained proteins from uninvolved regions of 4 human livers, from 1 hepatocellular carcinoma, and from 4 samples each of uninvolved colon, primary adenocarcinoma of the colon, and colon cancer metastatic to the liver was analyzed by two-dimensional protein electrophoresis. From a comparison of acidic proteins between pI 3.5 and 6.5, we conclude (1) that the majority (66 of 82) of denoted acidic proteins from 4 normal liver samples were represented in the hepatocellular carcinoma. Fifty-one of 58 proteins denoted in the 4 colon samples were detected in each of the 4 primary colon cancers; (2) that the "normograms" of proteins from normal colon and normal liver differed in many details, and their dissimilar patterns identified the source of the sample; (3) that hepatoma and primary adenocarcinoma of the colon were easily distinguished by their distribution of proteins; (4) that colon cancer metastatic to the liver contained a majority (50/58) of acidic proteins enumerated in primary colon cancer. These results indicate that uninvolved liver and colon and their primary or secondary cancers can be identified by their distribution of electrophoresed acidic proteins.

The distribution of proteins in samples from 8 prostate and 4 colon adenocarcinomas and 1 hepatoma was analyzed by 2-dimensional protein electrophoresis. Composite "normograms" based upon their distribution of proteins were developed from these patterns. Particular attention was paid to acidic proteins between pI 3.5 and 5.9. Of 161 proteins enumerated, 23 of the first 135 were present in prostate cancers, compared with 68 in colon cancer and 85 in the hepatoma. The 26 proteins denoted from nos. 135 to 161 were prostate associated, and none was evident in the colon or hepatoma samples. Twenty-seven prostate, 20 colon, and 48 liver cancer proteins were "unique" to each of the 3 cancers, respectively. The patterns of protein associated with each type of cancer were so dissimilar that with this technique no difficulty should be experienced in distinguishing these carcinomas originating from 3 different types of "stem" cells without obligatory recourse to microscopy.

Proteins from nine normal rat organs, including heart, lung thymus, liver, spleen, kidney, prostate, abdominal muscle, and brain, were solubilized, separated by electrophoresis according to their different isoelectric points and molecular weights, and stained with Coomassie blue. Patterns of major proteins unique to each organ were identified. Nine "unknown" samples, chosen from the same sources and submitted for analysis in a single blind study, were easily identified by comparing their protein profiles against the nine "reference" patterns. The ability to identify the origin of a tissue sample without recourse to microscopy, by comparing the pattern of its electrophoresed proteins with a "catalogue" of identified protein profiles, provides a prototype for the identification of histologically indeterminate normal and abnormal cells, tissues, and organs. Application of this technique to problems in human pathology and forensic medicine could prove to be very useful.

Samples from nine normal prostates (N), ten benign hypertrophic prostates (BPH), and eight adenocarcinomas of the prostate (ACP), obtained from the National Prostatic Cancer Project were analyzed by two-dimensional (2D)-protein electrophoresis. Qualitative analysis of Coomassie-blue-stained acidic proteins between pI 3.5 and 6.8 yielded these results: The total number of enumerated proteins was 136 (N), 107 (BPH), and 147 (ACP). Ninety-two denoted proteins were deemed common to the three kinds of samples, three were considered present in N and BPH, 7 in BPH and ACP, and 28 in N and ACP. Thirteen, five, and 20 proteins were detected only in N, BPH, or ACP. These proteins cannot be considered "unique" to their respective samples, since amounts of protein that do not exceed the minimum sensitivity of the dye may be present in the other samples. The similarity in the patterns of proteins from these three types of prostate samples is consistent with the following hypothesis: The distribution of proteins from histologically ambiguous prostate samples will correlate more closely with these protein profiles than with those associated with cancers originating from other organs.

Urocanic acid production was studied in 166 bacterial cultures isolated from mahimahi which had been stored for 14 days on ice in seawater. After 4 days of incubation at 10 degrees C in histidine-supplemented Trypticase soy broth (BBL Microbiology Systems, Cockeysville, Md.), urocanic acid was measured by thin-layer chromatography. Fifty-eight of the cultures were positive for urocanic acid, with the eight most active isolates producing ca. 10 mg/100 ml. These observations, coupled with a report that urocanic acid may be the predominant histidine metabolite in fish held at low temperatures, all suggest that urocanic acid may be a useful alternative to histamine as a spoilage index in scombroid and other fish that are rich in endogenous histidine.

Alkyl esters of six hydroxycinnamic acids, shown to be active antimicrobial agents when tested against Pseudomonas fluorescens, were further investigated for their effects against this organism. There was no statistically significant adaptation by this organism to either of the methyl esters of caffeic, rho-coumaric, cinnamic, or rho-hydroxybenzoic acids. Mixtures of these compounds taken two at a time gave at least additive effects, with some mixtures showing synergism. Preliminary work was also performed to determine the mode of inhibitory action for these compounds. The inhibition of oxygen utilization by the methyl esters correlated well with growth inhibition. Short-term lethality studies showed that none of the alkyl esters or methyl or propyl paraben produced any bacteriocidal effects. Oil-water partition coefficients were determined for these compounds and were shown to have no correlation with growth inhibitions. These all point to a specific mode of action, based in part on cellular energy depletion, rather than the nonspecific membrane-disrupting effects of other phenolic antimicrobial agents.

Six obese patients undergoing end-to-side jejunoileal bypass had gastric acid output and serum gastrin measurements performed preoperatively and within three months postoperatively. Meat extract stimulated acid output was higher in obese than in normal patients. Following the bypass procedure, basal and meat extract stimulated acid output significantly decreased. Basal and meat extract stimulated serum gastrin levels did not change following bypass. These studies demonstrate that after a small intestinal bypass, acid output decreases. Gastrin plays no role in this change.

The isolator 1.5 Microbial Tube (E. I. du Pont de Nemours & Co., Inc., Wilmington, Del.) is a new blood culture system for detection of bacteremia in children which involves lysis of blood cells and direct plating of blood lysate on agar media. Haemophilus influenzae type b is a major pediatric pathogen and may be associated with low-density bacteremia (less than or equal to 50 CFU per ml), yet the clinical data on the efficacy of this system for recovery of H. influenzae b are limited. We used a rat model of H. influenzae b bacteremia and fresh, whole human blood inoculated in vitro with H. influenzae b to compare the 1.5 Microbial Tube and conventional Trypticase soy broth (BBL Microbiology Systems, Cockeysville, Md.) for the detection of low-density H. influenzae b bacteremia. There was a significantly lower recovery of H. influenzae b from blood of bacteremic rats and from in vitro-inoculated human blood by using the 1.5 Microbial Tube than by using Trypticase soy broth. Studies on the mechanism of this lower recovery suggested that the contents of the 1.5 Microbial Tube are toxic to H. influenzae b in the presence of human blood. Additional clinical data are necessary before the 1.5 Microbial Tube can be recommended for detection of low-density H. influenzae b bacteremia.

We examined the incorporation of radioactive amino acids into nuclear proteins occurring at nonpermissive conditions in tsH1 Chinese hamster ovary cells with a temperature-sensitive defect in cytosol nonmitochondrial protein synthesis. In leucine-free medium at 40 degrees C, total cellular protein synthesis declined by 1-1.5%/min. As reported by others, preincubating these cells at 42 degrees C for 5-10 min sharply increased the rate of decline. The synthesis of acidic nuclear proteins at nonpermissive conditions (40 degrees C + 300 micrograms/ml cycloheximide) was demonstrated by the nuclear incorporation of 3H-tryptophan. Radioactivity, seen by autoradiography to be associated with these isolated Triton-X-100-washed nuclei, was released after incubating labelled nuclei with proteolytic enzymes. During incubation of tsH1 cells at nonpermissive conditions, pulse/chase experiments were consistent with the loss of some nuclear radioactivity into the cytoplasm. The distribution of cytosol and nuclear proteins, labelled at permissive or nonpermissive conditions and separated by isoelectric focusing, differed quantitatively and probably qualitatively, confirming the residual synthesis of acidic nuclear proteins at 40 degrees C in the presence of cycloheximide. Most newly synthesized acidic proteins retained by nuclei from cells labeled at nonpermissive conditions were present in a transciptionally active chromatin fraction. Although under these conditions the apparent rate of cellular RNA synthesis was unchanged, inhibiting residual cycloheximide-resistant nuclear protein synthesis with puromycin proportionately reduced RNA synthesis. Preincubating cells with 20 micrograms/ml of actinomycin D did not inhibit residual labelling of nuclear proteins; effects on residual nuclear labelling of impaired mitochondrial respiration were ambiguous. Nuclear proteins labelled under nonpermissive conditions probably included some of the 'prompt' heat shock proteins recently described. Provided certain assumptions are correct, our results are consistent with very limited protein synthesis associated with and even intrinsic to cell nuclei. They also suggest that this residual cycloheximide-resistant protein synthesis could be concerned with optimum synthesis or processing of certain nuclear RNA species.

Although the functional target of quinolone antibacterials such as nalidixic acid and norfloxacin has been identified as the enzyme DNA gyrase, the direct binding site of the drug is the DNA molecule [Shen, L. L., & Pernet, A. G. (1985) Proc. Natl. Acad. Sci. U.S.A. 82, 307-311]. As described in this paper, binding specificity and cooperativity of quinolones to DNA were further investigated with the use of a variety of DNA species of different structures and different base compositions. Results show that the drug binding specificity is controlled and determined largely by the DNA structure. The drug binds weakly and demonstrates no base preference when DNA strands are paired. The drug binds with much greater affinity when the strands are separated, and consequently, binding preference emerges: it binds better to poly(G) and poly(dG) over their counterparts including poly(dI). The results suggest that the drug binds to unpaired bases via hydrogen bonding and not via ring stacking with DNA bases. The weak binding to relaxed double-stranded DNA and the stronger binding to single-stranded DNA are both nonspecific as they do not demonstrate binding saturation and cooperativity. The specific type of binding, initially demonstrated in our previous publication with the supercoiled DNA and more recently with complex formed between linear DNA and DNA gyrase [Shen, L. L., Kohlbrenner, W. E., Weigl, D., & Baranowski, J. (1989) J. Biol. Chem. (in press)], occurs near the drug's supercoiling inhibition concentration. As shown in this paper, binding saturation curves of this type are highly cooperative (with Hill constant greater than 4).(ABSTRACT TRUNCATED AT 250 WORDS)

Retention of reference memory was evaluated in an animal model of cerebral ischemia. Rats were trained for 70 daily trials on an 8-arm radial maze with 5 arms baited, subjected to 30 minutes of forebrain ischemia, allowed to recover for 30 days, and then tested for an additional 50 trials. Post ischemic (PI) rats demonstrated normal retention of reference memory (p less than 0.05). Working memory was significantly impaired postoperatively (p less than 0.05). Morphologic analysis showed that PI rats had primary loss of pyramidal neurons in the CA1 region of hippocampus.

As a means of gaining additional information on the topoisomerase-mediated cytotoxicity induced by a variety of antibacterial and antitumor compounds we have examined the interaction of the quinolone anti-bacterial agent, norfloxacin, with the bacterial topoisomerase, DNA gyrase. Membrane filtration and spin-column techniques were used to study the binding of [3H]norfloxacin to purified plasmid DNA, DNA gyrase, and complexes formed by adding gyrase to different forms of plasmid DNA. Consistent with previous results (Shen, L. L., and Pernet, A. G. (1985) Proc. Natl. Acad. Sci. U.S.A. 82, 301-311) little [3H]norfloxacin binds to reconstituted gyrase, but significant levels of drug bind nonspecifically to relaxed DNA. However, when DNA and gyrase are incubated together additional norfloxacin binding sites are detectable. These complex-dependent sites are distinguishable from those sites involved in nonspecific DNA binding in that the complex-dependent sites are saturable and they retain bound norfloxacin after centrifuging the complex through a spin column. In addition, extent of binding is influenced by the topological state of DNA used to form the complex. The complex-dependent norfloxacin binding sites are likely involved in the inhibition of the enzyme since saturation of these sites occurs in the same norfloxacin concentration range as the inhibition of DNA supercoiling activity. Moreover, there is a close correlation of norfloxacin-induced DNA breakage with levels of norfloxacin bound to complexes of gyrase and relaxed DNA. These findings provide the first direct correlation of quinolone binding with inhibition of enzyme activity and induction of DNA breakage, and they suggest that the inhibition of DNA gyrase by norfloxacin occurs as a result of binding to a site which appears after the formation of a gyrase-DNA complex.

Plasma fibronectin concentration was measured in neonates of 2 to 5 days of age. Although breast fed and formula fed infants were similar in demographic characteristics, the mean (SD) plasma concentration of fibronectin in 26 breast fed infants, 237 (117) mg/l, was significantly higher than in 27 formula fed infants (171 (91) mg/l). Fibronectin was detected in five colostrum specimens (mean concentration 13.4 mg/l). Similar bands were detected after gel electrophoresis of purified adult plasma fibronectin and whole plasma from breast fed and formula fed neonates after staining or immunoblotting. Fibronectin isolated from breast milk also appeared similar to purified plasma fibronectin. It is possible, although unlikely, that fibronectin is absorbed intact from ingested colostrum. Alternatively, a factor(s) might be present in colostrum that contributes to the regulation of plasma fibronectin concentration.