Molecular characterization of the gene encoding H antigen in Escherichia coli and development of a PCR-restriction fragment length polymorphism test for identification of E. coli O157:H7 and O157:NM.

Recent outbreaks of disease caused by Escherichia coli O157:H7 have focused much attention on this newly emerged pathogen. Identification of the H7 flagellar antigen is critical for the confirmation of E. coli O157:H7; however, clinical isolates are frequently nonmotile and do not produce detectable H antigen. To further characterize nonmotile isolates (designated NM), we developed a PCR-restriction fragment length polymorphism (PCR-RFLP) test to identify and characterize the gene encoding the H antigen (fliC) in E. coli. The entire coding sequence of fliC was amplified by PCR, the amplicon was restricted with RsaI, and the restriction fragment pattern was examined after gel electrophoresis. Two hundred eighty E. coli isolates representing serotypes O157:H7 and O157:NM, flagellar antigen H7 groups associated with other O serogroups, and all other flagellar antigen groups were analyzed. A single restriction pattern (pattern A) was identified for O157:H7 isolates, O157:NM isolates that produced Shiga toxin (formerly Shiga-like toxin or verotoxin), and 16 of 18 O55:H7 isolates. Flagellar antigen group H7 isolates of non-O157 serotypes had one of three banding patterns distinct from pattern A. A wide variety of patterns were found among isolates of the other 52 flagellar antigen groups; however, none was identical to the O157:H7 pattern. Thirteen of 15 nonmotile strains that did not produce the A pattern had patterns that matched those of other known H groups. The PCR-RFLP in conjunction with O serogroup determination will be useful in identifying E. coli O157:H7 and related strains that do not express immunoreactive H antigen and could be expanded to include other clinically important E. coli strains.     

Changes in serum vitamin B12 levels in patients with megaloblastic anaemia treated with folic acid

In 21 patients with megaloblastic anaemia associated with the tropical malabsorption syndrome serum vitamin B₁₂ levels have been measured serially before and during parenteral folic acid therapy.     

Domain communication in the dynamical structure of human immunodeficiency virus 1 protease.

A dynamical model for the structure of the human immunodeficiency virus 1 (HIV-1) protease dimer in aqueous solution has been developed on the basis of molecular dynamics simulation. The model provides an accurate account of the crystal geometry and also a prediction of the structural reorganization expected to occur in the protein in aqueous solution compared to the crystalline environment. Analysis of the results by means of dynamical cross-correlation coefficients for atomic displacements indicates that domain-domain communication is present in the protein in the form of a molecular "cantilever" and is likely to be involved in enzyme function at the molecular level. The dynamical structure also suggests information that may ultimately be useful in understanding and further development of specific inhibitors of HIV-1 protease.     

The Effects of 10 Weeks Military Training on Heel Ultrasound and Bone Turnover

To measure the physiological changes in bone in response to strenuous exercise we performed a prospective study of male army recruits over 10 weeks of basic training. Measurements performed at the start and completion of training consisted of ultrasound (US) measurements of the heel: velocity of sound (VOS in m/seconds) and broadband ultrasound attenuation (BUA in dB/MHz) and bone turnover markers; osteocalcin (OC), bone-specific alkaline phosphatase (BALP), and tartrate-resistant acid phosphatase (TRAP). Forty subjects were recruited for the study and 26 completed training. Over the 10-week study period there was a significant 1.7% fall in mean VOS [mean paired difference (mpd) 27.2 m/second, SEM 9.5 (95% CI 7.5–46.8) P= 0.009] and a nonsignificant 3.4% increase in BUA (P= 0.159). There were significant falls in markers of bone formation OC [11.6%, mpd 0.11 μg/liter (95% CI 0.07–0.14) P < 0.001] and BALP [13.3%, mpd 3.49 U/liter (CI 0.80–6.18) P= 0.013] and a nonsignificant 9.5% fall in TRAP a marker of bone resorption. The 10 recruits subsequently injured had a significantly lower VOS on entry [mean difference 24.2 m/seconds (95% CI 4.6–43.7) P= 0.017] and nonsignificantly raised BUA and baseline levels of all bone markers. The ultrasound changes may be accounted for by increase in trabecular separation and a fall in trabecular connectivity due to microfracture. The decrease in bone markers implies a fall in bone turnover. Received: 26 June 1997 / Accepted: 26 August 1998     

Allogeneic vaccination with a B7.1 HLA-A gene-modified adenocarcinoma cell line in patients with advanced non-small-cell lung cancer.

PURPOSE: To determine the safety, immunogenicity, and clinical response to an allogeneic tumor vaccine for non-small-cell lung cancer, we conducted a phase I trial in patients with advanced metastatic disease. PATIENTS AND METHODS: We treated 19 patients with a vaccine based on an adenocarcinoma line (AD100) transfected with B7.1 (CD80) and HLA A1 or A2. Patients were vaccinated intradermally with 5 x 10(7) cells once every 2 weeks. Three vaccinations represented one course of treatment. If patients had complete response, partial response, or stable disease, they continued with the vaccinations for up to three courses (nine vaccinations). Immune response was assessed by a change between pre-study and postvaccination enzyme-linked immunospot frequency of purified CD8 T-cells secreting interferon-gamma in response to in vitro challenge with AD100. RESULTS: Four patients experienced serious adverse events that were unrelated to vaccine. Another four patients experienced only minimal skin erythema. All but one patient had a measurable CD8 response after three immunizations. The immune response of six surviving, clinically responding patients shows that CD8 titers continue to be elevated up to 150 weeks, even after cessation of vaccination. Overall, one patient had a partial response, and five had stable disease. Median survival for all patients is 18 months (90% CI, 7 to 23 months), with corresponding estimates of 1-year, 2-year, and 3-year survival of 52%, 30%, and 30%, respectively. HLA matching of vaccine, age, sex, race, and pathology did not bear a significant relation to response. CONCLUSION: Minimal toxicity and good survival in this small population suggest clinical benefit from vaccination.