Randomized controlled trial comparing Moviprep® and Phosphoral® as bowel cleansing agents in patients undergoing colonoscopy.

BACKGROUND: Introducing colonoscopy as part of colorectal cancer screening on a national level, we aimed at evaluating the efficacy of the two most common bowel cleansing agents, Moviprep(®) and Phosphoral(®). Secondly, we evaluated the benefit for the patient and society in terms of sick leave and discomfort. METHODS: In a single-blinded randomized equivalence trial, Phosphoral(®) (NaP) was compared with Moviprep(®) (2 l polyethylene glycol + ascorbic acid) in patients undergoing colonoscopy due to suspicion of cancer. Patients filled out questionnaires concerning stool consistency, discomfort and number of sick days due to bowel cleansing. Blinded colonoscopists estimated the efficiency of the cleansing using the validated Harefield Cleansing Scale. RESULTS: Two hundred and sixty-six patients were included 250 of whom underwent full colonoscopy. There was no difference in the percentage of acceptable bowel cleansings in the two groups; however, a significantly higher number of A scores were observed in the Moviprep(®) group (p = 0.028). We found no correlation between stool consistency and outcome of the cleansing and no difference in subjective discomfort during cleansing. Vomiting during cleansing occurred more often in the Phosphoral(®) group (p = 0.002). There was a trend toward a smaller number of sick days in patients who used Moviprep(®) compared with Phosphoral(®). CONCLUSIONS: Moviprep(®) and Phosphoral(®) provided equally efficient bowel cleansing in 90 % of patients, but Moviprep(®) provided a higher quality of cleansings graded as successful. The two agents were equally tolerated, and no difference was found in the related number of sick days.

Nucleic acid aptamers against proteases.

Proteases are potential or realized therapeutic targets in a wide variety of pathological conditions. Moreover, proteases are classical subjects for studies of enzymatic and regulatory mechanisms. We here review the literature on nucleic acid aptamers selected with proteases as targets. Designing small molecule protease inhibitors of sufficient specificity has proved a daunting task. Aptamers seem to represent a promising alternative. In our review, we concentrate on biochemical mechanisms of aptamer selection, protein-aptamer recognition, protease inhibition, and advantages of aptamers for pharmacological intervention with pathophysiological functions of proteases. Aptamers can be selected so that they bind their targets highly specifically and with affinities corresponding to KD values in the nM range. Aptamers can be selected so that they recognize their targets conformation-specifically, for instance with vastly different affinities to zymogen and active enzyme forms. Furthermore, aptamers can be selected to inhibit the enzyme activity of the target proteases, but also to inhibit functionally important exosite interactions, for instance cofactor binding. Several protease-inhibiting aptamers, directed against blood coagulation factors, are in clinical trials as anticoagulant drugs. Several of the studies on protease-binding aptamers have been pioneering and trend-setting in the field. The work with protease-binding aptamers also demonstrates many interesting examples of non-standard selection strategies and of new principles for regulating the activity of the inhibitory action of aptamers of general interest to researchers working with nucleic acid aptamers.

Mass screening for antimicrobial resistant Escherichia coli in dairy cows in northern New South Wales.

OBJECTIVE: To describe aspects of the epidemiology of antimicrobial resistance in Escherichia coli shed in the faeces of milking cows in a dairying region of New South Wales. DESIGN: A survey based on multi-stage sampling with repeated measures made within herds for estimating within-herd correlation of resistance status, and with repeated measures made on identical specimens for estimating test-retest reliability. PROCEDURE: From a population of 110 dairy herds, 30 were selected at random and from each herd between 5 and 10 faecal specimens were obtained from fresh manure pats. E coli from faecal specimens were grown on hydrophobic grid membrane filters (HGMF) and replicated onto chromogenic agar and agar containing antimicrobials (gentamicin, ampicillin, tetracycline and sulfamethoxazole). Image analysis was used to assess colony growth. Data were analysed descriptively, by generalised linear mixed models and by Taylor series linearisation to account for attributes of the survey design. RESULTS: Of the 10,279 E coli isolates assessed, 91% expressed no resistance, 7.3% were resistant to sulfamethoxazole, 3.6% to tetracycline, 2.2% to ampicillin and 0.09% to gentamicin. The most common multiple resistance phenotype was ampicillin-tetracycline-sulfamethoxazole (1.8% of isolates). Most multiple resistant isolates appeared clustered within particular herds but were too rare to obtain valid estimates of variance, confidence intervals or intra-herd correlation. The estimated proportion of isolates in the population that were susceptible to all four antimicrobials was 97% (95% CI: 91% to 100%) and 55% of cows had no resistance detected in faecal E coli (95% CI: 27% to 83%). Within-herd correlation of shedding status (any resistance pattern) was absent and test-retest reliability of the measurement system was estimated to be at the lower end of good (0.40) but increased to excellent (0.89) after excluding sulfamethoxazole resistance, which had a greater measurement error. CONCLUSION: Antimicrobial resistance was uncommon in E coli in the population of dairy cows studied. HGMF and image analysis is an effective tool for detecting rare forms of resistant E coli that are not uniformly distributed in livestock populations.

The relationship between concentration of a dual marker strain of Salmonella Typhimurium in bovine faeces and its probability of detection by immunomagnetic separation and culture.

AIMS: To modify a strain of Salmonella serotype Typhimurium to express unique marker traits and then define how the concentration of the marker in bovine faeces affects the probability of its detection by culture preceded by immunomagnetic separation (IMS). METHODS AND RESULTS: DNA encoding for the production of green fluorescent protein (gfp) and resistance to kanamycin was inserted into the bacterial chromosome of Salm. Typhimurium. Transposon insertion was demonstrated by Southern blot hybridization. Varying amounts of one electroporant (gfpSal-1) were inoculated into suspensions of bovine faeces and attempts made to isolate gfpSal-1 using a protocol based on pre-enrichment incubation, IMS and enrichment in selective media. Isolates of gfpSal-1 were differentiated from wild strains of Salmonella using fluorescence under u.v. light and expression of kanamycin resistance. A logistic and Gompertz function each derived from the dose-response data partially explained the observations with the fit of the Gompertz function judged to be superior. The 10, 50 and 90% limits of detection from the Gompertz function were estimated to be 1.92, 2.03 and 2.27 CFU g(-1) respectively. CONCLUSIONS: Reliance on the traditional concept of 'limit of detection' could introduce unacceptable errors in the interpretation of test findings when the concentration of Salm. Typhimurium in bovine faeces (pooled or individual) is below ca 3 CFU g(-1) of faeces. SIGNIFICANCE AND IMPACT OF THE STUDY: The dose-response curve can be used to aid the design of protocols for detecting Salmonella in individual and pooled faecal specimens. The experiments demonstrate that both reporter genes in tandem are useful for studying the performance of culture-based methods for detecting pathogens in faeces.

Isolation and initial characterization of the BRCA2 promoter.

The hereditary breast cancer susceptibility gene, BRCA2, is considered to be a tumor suppressor gene that may be involved in the cellular response to DNA damage. The transcript for this gene is cell cycle regulated with mRNA levels reaching a peak just before the onset of DNA synthesis. In order to define the mechanisms by which BRCA2 is transcriptionally regulated, we have begun to study upstream regulatory sequences. In this report, we define a minimal promoter region that has strong activity in human breast epithelial cells. Deletions of this sequence narrowed the strong basal activity to a region extending from -66 to +129 with respect to the BRCA2 transcriptional start site. This sequence demonstrated cell cycle regulated activity with kinetics similar to the endogenous transcript. Examination of the sequence revealed several consensus binding sites for transcription factors including an E-box, E2F and Ets recognition motifs. Electrohoretic mobility shift assays revealed specific protein binding to two sequences upstream of the start site; the palindromic E-box and an Ets/E2F site. Site-directed mutagenesis of either of these sites reduced both the basal activity in log phase cells and the cell cycle regulated activity of the promoter. Mutational inactivation of both sites within the same construct effectively eliminated promoter activity. Antibodies to candidate transcription factors used in super shift experiments revealed specific interactions between the BRCA2 promoter and the basic region/helix - loop - helix containing USF-1 and 2 proteins and Elf-1, an Ets domain protein. Binding of these factors depended upon the presence of intact recognition sequences. The USF factors were shown to bind predominantly as a heterodimeric complex of USF-1 and 2 while Elf-1 bound the promoter when it was not occupied by USF. Co-transfection studies with USF proteins and the varicella zoster IE62 protein provide evidence for the involvement of endogenous and exogenous USF in the activation of the BRCA2 promoter. We propose that interactions between USF-1, USF-2 and Elf-1 play an important role in the transcriptional regulation of the BRCA2 gene.

Utilization of ileal digestible amino acids by growing pigs: isoleucine.

Two experiments were conducted to determine the utilization of ileal digestible isoleucine by growing pigs. In the first, the apparent ileal digestibility of amino acids in cottonseed meal, lupin-seed meal and soya-bean meal was determined in pigs fitted with 'T'-shaped cannulas. In the second, three isoleucine-deficient diets were formulated to 0.23 g ileal digestible isoleucine/MJ digestible energy (DE) with the three protein concentrates contributing the only source of isoleucine in sucrose-based diets. An additional three diets were formulated with supplements of isoleucine to confirm that isoleucine was limiting in the first three diets. The growth performance and retention of isoleucine by pigs given the six diets over the 20-45 kg growth phase were then determined. The apparent ileal digestibility of isoleucine in the three protein concentrates (proportion of total) was: cottonseed meal 0.68, lupin-seed meal 0.86, soya-bean meal 0.86. There were no significant differences (P > 0.05) in growth rates (g/d) and crude protein deposition rates (g/d) of the pigs given the three diets formulated to 0.23 g ileal digestible isoleucine/MJ DE: cottonseed meal 590, 84; lupin-seed meal 613, 87; soya-bean meal 594, 91 (SEM 13.0, 2.9) respectively. The response of pigs to the addition of isoleucine confirmed that isoleucine was limiting in these diets. The proportion of ileal digestible isoleucine retained by pigs given the cottonseed meal (0.65) was slightly lower than that retained by pigs given soya-bean meal (0.73; P < 0.05). These results indicate that values for the ileal digestibility of isoleucine in protein concentrates more closely reflect the proportion of isoleucine that can be utilized by the pig than occurs for other amino acids such as lysine, threonine and methionine.

Utilization of ileal digestible amino acids by growing pigs: tryptophan.

The ileal digestibility of tryptophan for growing pigs was determined for cottonseed, meat-and-bone and soya-bean meals. Tryptophan in the food and digesta was measured by two analytical procedures (NaOH hydrolysis and colorimetric estimation (method 1) and LiOH hydrolysis and HPLC determination (method 2)). The results were respectively: cottonseed meal 0.46, 0.81; meat-and-bone meal 0.55, 0.65; soya-bean meal 0.74, 0.90. In the first experiment the values for method 1 were shown to be inapplicable to pigs. In a second experiment three tryptophan-deficient diets (0.05 g ileal digestible tryptophan/MJ digestible energy (DE)) were formulated using values from method 2 for cottonseed meal, meat-and-bone meal plus L-tryptophan and soya-bean meal respectively as the only sources of tryptophan in the diets. This experiment was terminated after 28 d as overall growth performance of the pigs was very low. A third experiment was conducted in a similar manner to Expt 2 except that the diets were formulated to 0.065 g ileal digestible tryptophan/MJ DE and growth responses and tryptophan retention were assessed over the 20-45 kg growth phase. Growth rates (g/d) of the pigs given the three diets were significantly different (P < 0.01): cottonseed meal 393, meat-and-bone meal plus L-tryptophan 531, soya-bean meal 437 (SED 39.0). Tryptophan retention (as a proportion of ileal digestible tryptophan intake) was significantly different (P < 0.05): cottonseed meal 0.51, meat-and-bone meal plus L-tryptophan 0.49, soya-bean meal 0.41. These results indicate (1) that the colorimetric technique for assessing tryptophan was inapplicable and (2) that ileal digestible values for tryptophan were not suitable for formulating diets containing heat-processed proteins, possibly due to absorption of some of the tryptophan in a form that was non-utilizable, and/or to underestimation of total tryptophan in the protein concentrates.

Utilization of ileal digestible amino acids by growing pigs: methionine.

An experiment was conducted to determine the utilization of ileal digestible methionine by growing pigs. Three methionine-deficient diets (0.09 g ileal digestible methionine/MJ digestible energy (DE)) were formulated using cottonseed meal, meat-and-bone meal and soya-bean meal respectively as the only source of methionine in the diet. An additional three diets were formulated with supplements of methionine to confirm that methionine was limiting in the first three diets. The growth performance and retention of methionine by pigs given the six diets over the 20-45 kg growth phase was then determined. Growth rates (g/d) of pigs given the three diets formulated to 0.09 g ileal digestible methionine/MJ DE were significantly different (P < 0.01): cottonseed meal 411, meat-and-bone meal 442, soya-bean meal 496 (SED 24.6). The response of pigs to the addition of methionine confirmed that methionine was limiting in these diets. Crude protein (N x 6.25) deposited by the pigs (g/d) was significantly higher (P < 0.05) for those given soya-bean meal (61) and meat-and-bone meal (57) relative to cottonseed meal (47; SED 3.3). The proportion of ileal digestible methionine retained by pigs given the three protein concentrates was: cottonseed meal 0.39, meat-and-bone meal 0.45, soya-bean meal 0.47 (SED 0.019). These results indicate that values for the ileal digestibility of methionine in protein concentrates do not reflect the proportion of methionine that can be utilized by the pig. It appears that, with heat-processed meals, a considerable proportion of the methionine is absorbed in a form(s) that is (are) inefficiently utilized.

A comparison of the availability and ileal digestibility of lysine in cottonseed and soya-bean meals for grower/finisher pigs.

The availability of lysine and the ileal digestibility of amino acids in three cottonseed meals and a soya-bean meal for grower/finisher pigs were determined. The usefulness of the availability estimates for formulating diets was assessed. The availability of lysine, as assessed with a slope-ratio assay, was (proportion of total): cottonseed meal no. 1, 0.27; no. 2, 0.30; no. 3, 0.29; soya-bean meal, 0.90. Ileal digestibility of lysine in the meals (proportion of total) was: cottonseed meal no. 1, 0.58; no. 2, 0.68; no. 3, 0.72; soya-bean meal, 0.89. Pigs given diets formulated to the same available lysine concentration grew at similar rates and retained the same amount of lysine in the carcasses. The results indicate that, for meals of high availability (soya-bean meal), reduced ileal digestibility appears to be the main reason for reduced availability. However, in meals of low availability (cottonseed meal), reduced ileal digestibility only accounts for part of the reduced availability. Thus, the ileal digestibility of lysine is not a reliable indicator of lysine availability.

Utilization of ileal digestible amino acids by pigs: lysine.

Two experiments were conducted to determine the utilization of ileal digestible lysine by pigs. In the first, the apparent ileal digestibility of amino acids in cottonseed meal, meat-and-bone meal and soya-bean meal was determined in pigs fitted with 'T'-shaped cannulas. In the second experiment, three lysine-deficient diets were formulated to 0.36 g ileal digestible lysine/MJ digestible energy (DE), with lysine contributed from the three protein concentrates as the only source of lysine in sugar-based diets. An additional three diets were formulated with supplements of lysine to verify that lysine was limiting in the first three diets. The growth performance and retention of lysine by pigs given the six diets over the 20-45 kg growth phase were then determined. The apparent ileal digestibility of lysine in the three protein concentrates (proportion of total) was: cottonseed meal 0.74, meat-and-bone meal 0.78, soya-bean meal 0.89. Growth rates (g/d) of the pigs given the three diets formulated to 0.36 g ileal digestible lysine/MJ DE were significantly different (P less than 0.001): cottonseed meal 377, meat-and-bone meal 492, soya-bean meal 541. The response of pigs to the addition of lysine confirmed that lysine was limiting in these diets. Crude protein (nitrogen x 6.25) deposited by the pigs was significantly higher (P less than 0.001) for those given soya-bean meal (77 g/d), relative to meat-and-bone meal (66 g/d) and cottonseed meal (38 g/d). The proportion of ileal digestible lysine retained by pigs given the three protein concentrates was: cottonseed meal 0.36, meat-and-bone meal 0.60, soya-bean meal 0.75. The results indicate that values for the ileal digestibility of lysine in protein concentrates are unsuitable in dietary formulations as the assay does not reflect the proportion of lysine that can be utilized by the pig. It appears that, with heat-processed meals, a considerable proportion of the lysine is absorbed in a form(s) that is (are) inefficiently utilized.

Utilization of ileal digestible amino acids by growing pigs: effect of dietary lysine concentration on efficiency of lysine retention.

Diets were formulated using sugar, soya-bean meal and free amino acids to contain 0.1-0.8 g lysine/MJ digestible energy (DE) and offered at three times maintenance to male and female pigs from 20 to 45 kg live weight. Growth responses and retentions of protein, fat, energy and lysine were assessed. Increasing the dietary lysine concentration resulted in significant (P less than 0.001) linear and curvilinear increases in growth rates and decreases in food conversion ratios. There was only a small effect of lysine concentration on total energy retention, but a substantial effect on the partitioning of energy deposition, with increases in the rate of protein deposition and decreases in fat retention. There was no difference in the efficiency of protein deposition between male and female pigs but males responded more to higher lysine concentrations than females (estimated 0.93 and 0.74 g lysine/MJ DE for males and females respectively). Lysine concentration in the protein deposited by the pigs increased linearly and curvilinearly (P less than 0.01) from 5.8 to 6.6 g lysine/16 g N with increasing dietary lysine concentration. There was a linear and quadratic response (P less than 0.001) in retention of ileal digestible lysine, with the minimum retention of 0.16 occurring at 0.1 g lysine/MJ DE and increasing to a maximum retention of 0.73 at a dietary concentration of 0.47 g lysine/MJ DE. The efficiency of lysine retained/ileal digestible lysine intake was 0.86 and the endogenous lysine loss was estimated at 0.94 g/d.

Nutritional value of lupin (Lupinus albus)-seed meal for growing pigs: availability of lysine, effect of autoclaving and net energy content.

1. Two experiments were conducted to assess the nutritional value of lupin (Lupinus albus)-seed meal for growing pigs. In the first, the availability of lysine was assessed using slope-ratio analysis. In the second, the effects of autoclaving lupin seeds and formulating the diets on the basis of estimated digestible or net energy were assessed. 2. In the first experiment, the availability of lysine in three samples of lupin-seed meal was compared with that in meat-and-bone meal and soya-bean meal. Availability of lysine in the five protein concentrates, using food conversion efficiency on a carcass basis as the criterion of response, was (proportion of total): lupin-seed meal no. 1 0.44, no. 2 0.57, no. 3 0.53, meat-and-bone meal 0.42, soya-bean meal 0.80. 3. Availability estimates, based on protein deposited:food intake, were: lupin-seed meal no. 1 0.82, no. 2 0.73, no. 3 0.70, meat-and-bone meal 0.27, soya-bean meal 0.77. These estimates had higher standard deviations than those based on carcass response. 4. Regressing the measures of response v. lysine intake resulted in estimates of availability similar to, or higher than, the slope-ratio analysis but was associated with greater statistical invalidity and higher standard deviations. 5. The proportion of energy retained in the carcasses was unaffected by the inclusion levels of lysine or soya-bean meal. Energy retention was depressed (P less than 0.05) with the three lupin-seed meals and the meat-and-bone meal. 6. In the second experiment, the response of pigs given a diet containing lupin-seed meal was inferior, on a carcass basis (P less than 0.05), to that of pigs given a diet containing soya-bean meal formulated to similar total lysine and digestible energy contents. 7. The addition of soya-bean oil to the diet containing lupin-seed meal, to equalize the estimated net energy of the diet to that of the diet containing soya-bean meal, depressed protein deposition (P less than 0.05) and increased fat deposition (P less than 0.05), indicating that energy was not limiting the growth of pigs given the lupin-seed-meal diet. 8. Autoclaving the lupin-seed at 121 degrees for 5 min had no effect on the growth of pigs, indicating that the low availability of lysine was not due to the presence of heat-labile anti-nutritional factors.

Effect of heat on the nutritional value of lupin (Lupinus angustifolius)--seed meal for growing pigs.

1. Two experiments were conducted to determine the effects of heat on the nutritional value of lupin (Lupinus angustifolius cv. Uniharvest and Unicrop)-seed meal, relative to soya-bean meal, for growing pigs 2. In both experiments, values for carcass gain/d and food conversion ratio (FCR) on a carcass basis of pigs fed on the diets containing lupin-seed meal were inferior (P less than 0.05) to those produced by pigs fed on soya-bean meal. 3. In the first experiment, heating lupin seed at temperatures from 105 to 150 degrees for 15 min resulted in a linear depression in carcass gain/d, a quadratic increase in carcass FCR, a linear decrease in lean in the ham and a linear increase in backfat thickness. In the second experiment, autoclaving lupin seed from 5 to 45 min at 121 degrees resulted in a linear depression in carcass gain/d and a linear increase in carcass FCR. 4. The addition of L-lysine to the diets containing lupin-seed meal verified that lysine was limiting in both experiments. The additions of L-lysine did not overcome the differences in carcass gains/d of pigs fed on lupin-seed meal relative to those fed on diets containing soya-bean meal. 5. It is concluded that the low lysine availability in lupin-seed meal for pigs is not due to the presence of heat-labile anti-nutritional factors in the seed.

Phenoxyethanol as a nontoxic substitute for formaldehyde in long-term preservation of human anatomical specimens for dissection and demonstration purposes.

Formaldehyde has recently been declared a potential carcinogen. Occupational health authorities throughout the world are therefore likely to put stricter regulations to its use also within anatomical disciplines. We have been able to reduce the atmospheric concentration of formaldehyde in our dissection rooms to below the detection limit of a conventional Dräger tube multigas analyzer (i.e., below 0.5 ppm or 0.6 mg formaldehyde/m3 air), by extracting previously formaldehyde-fixed material for more than 3 months in 1% phenoxyethanol in tap water. In this fluid our material has remained soft and flexible with a consistency and color retention suitable for dissection and demonstration purposes for up to 10 years. Fungal attacks are rare and we have been unable to raise bacteria from such specimens. Even the microscopical structure of most tissues remains satisfactory after 5 years in 1% phenoxyethanol. The unpleasant and irritating smell traditionally felt in dissection rooms is almost absent in our facilities, but some of our students still mention slight odor, headache, drowsiness, and mild eye, nose, and throat irritation during their dissection practice periods.

Availability of lysine in vegetable protein concentrates as determined by the slope-ratio assay with growing pigs and rats and by chemical techniques.

The availability of lysine in nine vegetable-protein concentrates was assessed using the slope-ratio assay for growing pigs and rats. Diets were equalized for crude fibre using solka floc to minimize any possible effects of variation in fibre content on availability estimates. The availability of lysine in the nine proteins for pigs, using food conversion efficiency (FCE) on a carcass basis as the criterion of response were (proportion of total): cottonseed meal 0.39, lupin (Lupinus angustifolius) seed meal no. 1 0.37, no. 2 0.65, no. 3 0.54, no. 4 0.54, field peas (Pisum sativum) 0.93, peanut (groundnut) meal 0.57, soya-bean meal no. 1 0.98, no. 2 0.89. Estimates of available lysine for rats as assessed by the slope-ratio assay using FCE on a carcass basis were in close agreement with the pig estimates for cottonseed meal (0.35) and soya-bean meal no. 1 (0.91) and no. 2 (0.89), higher for lupin-seed meals (range 0.70-0.94 with a mean of 0.81) and peanut meal (0.76) and lower for field peas (0.76). The differences in available lysine were not detected by the chemical Silcock available-lysine test (Roach et al. 1967) or by the direct 1-fluoro-2,4-dinitrobenzene procedure (Carpenter, 1960).