A new multiplex PCR for easy screening of methicillin-resistant Staphylococcus aureus SCCmec types I–V

A multiplex PCR with four primer-pairs was designed to identify the five main known SCCmec types. A clear and easily discriminated band pattern was obtained for all five types. The SCCmec type was identified for 98% of 312 clinical isolates of methicillin-resistant Staphylococcus aureus (MRSA). SCCmec type IV was by far the most common SCCmec type among both hospital- and community-acquired MRSA isolates in Denmark.     

Isolation and preliminary evaluation of a low-molecular-mass antigen preparation for improved detection of Helicobacter pylori immunoglobulin G antibodies.

Previously, immunoglobulin G (IgG) antibodies to five antigens with a relative molecular mass of between 15 and 30 kDa from Helicobacter pylori were found to be significantly more frequent in H. pylori-infected patients than in noninfected patients. In this study, these specific low-molecular-mass (LMW) antigens were separated by ultrafiltration of whole-cell sonicates. The LMW antigen preparation was evaluated by enzyme-linked immunosorbent assay with serum samples from 76 children with abdominal symptoms and 151 adults with dyspeptic symptoms. H. pylori was cultured or seen in 40 (53%) children and 83 (55%) adults. Increased antibody levels to H. pylori were found in serum from 35 (46%) children and 88 (58%) adults. Values for sensitivity, specificity, and predictive value of positive and negative results of the test were higher with LMW antigens than with the heat-stable antigen previously described. The low specificity and predictive value of a positive result were due to seropositive results for 21 persons with a negative culture for H. pylori and negative microscopy results for Helicobacter-like organisms in biopsies from gastric mucosa. Histologically, chronic gastritis was demonstrated in 43% of these persons, and 19% had peptic ulcer, indicating that they have or have had H. pylori infection. Specific antibodies to H. pylori were confirmed in all 21 patients by the Western immunoblot technique. Use of the LMW antigen improved the IgG antibody detection in patients with H. pylori infection, even though the results reflect the difficulties in establishing a true gold standard for diagnosis of H. pylori infection.     

Nucleotide sequence of the geminivirus chloris striate mosaic virus

The genome of chloris striate mosaic virus (CSMV) comprises a single circular DNA as determined by analyses on virion single-stranded (ss) DNA and virus-specific covalently closed circular (ccc) DNA isolated from infected plants. The nucleotide sequence of CSMV DNA was determined from cccDNA and the data were accommodated into one DNA circle of 2750 nucleotides. Comparison of the nucleotide sequence with those of maize streak virus (MSV), wheat dwarf virus (WDV), and digitaria streak virus (DSV) showed 49, 47, and 48% DNA homology, respectively. The sequence has four potential open reading frames for proteins of greater than 10,000 mol wt, two in the viral (+) sense and two in the complementary (-) sense. Three of these potential coding regions have homologous counterparts, by comparison of the amino acid sequences, among the open reading frames reported for MSV, WDV, and DSV. CSMV encapasidates primer molecules able to prime the synthesis in vitro of a complementary strand to virion DNA, initiating this reaction at one site on the genome. The CSMV primer comprising approximately 88 nucleotides was located within the smaller of two intergenic or noncoding regions.     

Morbidity in a Danish national cohort of 472 IVF/ICSI twins, 1132 non-IVF/ICSI twins and 634 IVF/ICSI singletons: health-related and social implications for the children and their families

BACKGROUND: There is a lack of knowledge on child health as well as family well-being in IVF/ICSI twins. METHODS: These data originated from questionnaires completed by mothers taking part in a national cohort study of twin and singleton births occurring in Denmark in 1997. The overall response rate was 83%. The three cohorts consisted of all IVF/ICSI twin children (n = 472), all IVF/ICSI singletons (n = 634) and all non-IVF/ICSI twin children (n = 1132) born in Denmark in 1997. RESULTS: No major differences in physical health were observed between IVF/ICSI twins and non-IVF/ICSI twins. Compared with IVF/ICSI singletons, more IVF/ICSI twins were admitted to a neonatal intensive care unit (NICU) (P < 0.01) and more had surgical interventions (P = 0.03) and special needs (P = 0.02), moreover they had poorer speech development (P < 0.01). Correspondingly, IVF/ICSI twin mothers rated their infant's general health poorer than IVF/ICSI singleton mothers did. All discrepancies between IVF/ICSI twins and singletons disappeared after stratification for birthweight except for NICU admissions and speech development. Multiple logistic regression analyses showed that both IVF/ICSI and non-IVF/ICSI twin parents experienced more marital stress [odds ratio (OR) 2.9, 95% CI 2.2-3.8] and that twins had more impact on the mother's life (OR 1.7, 95% CI 1.2-2.4) compared with singletons. Nevertheless, the only predictor of low divorce/separation risk was IVF/ICSI treatment. CONCLUSION: Our study indicates that physical health of IVF/ICSI twins is comparable with that of non-IVF/ICSI twins. However, physical health of IVF/ICSI twins is poorer and the implications for the families stronger compared with IVF/ICSI singletons.     

Genomic approach to identification of Mycobacterium bovis diagnostic antigens in cattle

Differential delayed-type hypersensitivity skin testing with tuberculin purified protein derivatives from Mycobacterium bovis and M. avium is the standard for diagnosing bovine tuberculosis. However, improved tests based on defined, specific antigens are urgently needed. In the present study, a combination of bioinformatics, molecular biology, and bovine models of infection were used to screen mycobacterial proteins for their potential as diagnostic reagents which could be used in a whole-blood assay for diagnosis of tuberculosis. Initial screening of 28 proteins selected in silico and expressed as recombinants in Escherichia coli indicated that CFP-10, ESAT-6, TB27.4, TB16.2, TB15.8, and TB10.4 induced strong gamma interferon responses in experimentally infected cattle. A more thorough investigation over time in two groups of animals infected with a high (10(6) CFU) and a low (10(4) CFU) dose of M. bovis revealed that, for both groups, the strength of the in vitro response to individual antigens varied greatly over time. However, combining the results for ESAT-6, CFP-10, and TB27.4, possibly supplemented with TB10.4, gave sensitivities at different infection stages close to those obtained with M. bovis purified protein derivative. Importantly, while responsiveness to ESAT-6 and CFP-10 correlated strongly for individual samples, the same was not the case for ESAT-6 and TB27.4 responsiveness. The results suggest that combinations of specific antigens such as these have great potential in development of optimized diagnostic systems for bovine tuberculosis.     

Characterization of monoclonal antibodies to fimbria-associated adhesins of Bacteroides loescheii PK1295.

Bacteroides loescheii PK1295 fimbriae, which mediate the lactose-sensitive coaggregation with Streptococcus sanguis 34 and the lactose-insensitive coaggregation with Actinomyces israelii PK14, were injected into mice to raise adhesin-specific monoclonal antibodies (MAbs). Supernatants of hybridomas were screened for the capacity to inhibit coaggregation and agglutinate intact bacteria. Of the 10 MAbs that were isolated, 4 were specific and potent inhibitors of the coaggregation between B. loescheii and S. sanguis and two other MAbs specifically inhibited the B. loescheii-A. israelii interaction. None of the six MAbs which inhibited adherence were capable of agglutinating whole cells of B. loescheii, whereas the four remaining MAbs agglutinated whole cells but had no effect on coaggregation. Fab fragments of two MAbs, one that inhibited the coaggregation with S. sanguis and another that inhibited the interaction with A. israelii, also were shown to inhibit the respective coaggregation interactions, suggesting that each of the immunoglobulins recognized its adhesin molecule at or near the active sites. By immunoblotting or immunoprecipitation, the S. sanguis adhesin-specific MAbs reacted with a 75-kilodalton polypeptide present in fimbria-enriched preparations, whereas the A. israelii adhesin-specific MAbs recognized a 45-kilodalton polypeptide in the same preparations. By screening hybridoma supernatants directly for their capacity to block coaggregation, we isolated MAbs which were used to establish that the B. loescheii-S. sanguis and the B. loescheii-A. israelii interactions were mediated by different adhesins.     

Problematic Methods in the Assessment of Scholarly Productivity in Clinical PhD Programs

We review the methods in the ranking of clinical psychology doctoral programs provided by Stewart, Roberts, and Roy (2007). Using our own program as an example, we identify several areas of concern (e.g., authorship credits, criteria applied, faculty attrition). The inaccuracies identified for our program ranking, in combination with methodological concerns highlighted by previous commentaries, suggest that the validity of the rankings can be called into question.     

Effect of omeprazole and cimetidine on duodenal ulcer. A double-blind comparative trial

We conducted a double-blind randomized study of 132 patients to determine whether the new, investigational proton-pump inhibitor, omeprazole (30 mg per day), would accelerate healing and pain relief, as compared with cimetidine (1 g per day), in patients with duodenal ulcer. After two weeks of treatment, which was completed by all patients, the healing rates were 73 per cent in the omeprazole group and 46 per cent in the cimetidine group (P less than 0.01). After four weeks of treatment, which was completed by 118 patients, the corresponding figures were 92 and 74 per cent (P less than 0.05). In the omeprazole group 55 per cent of the patients were free of pain after the first week, as compared with 40 per cent of those treated with cimetidine (P greater than 0.05). No major clinical or biochemical side effects of omeprazole or cimetidine were noted. A six-month follow-up study revealed no significant difference between the recurrence rates after omeprazole and after cimetidine treatment. In May 1984 clinical trials with omeprazole were temporarily suspended, since a study of long-term toxicity in rats had shown the development of gastric carcinoid tumors.