The WHO multicenter study on Listeria monocytogenes subtyping: random amplification of polymorphic DNA (RAPD)

As part of a WHO multicenter study on Listeria monocytogenes subtyping methods the random amplification of polymorphic DNA (RAPD)-technique was evaluated. Six participants were asked to use a standard protocol to analyse a set of 80 L. monocytogenes strains. This set contained 22 groups of epidemiologically linked isolates and 11 pairs of duplicate strains. Using three different 10-mer primers the median reproducibility of the RAPD-results obtained by the six participants was 86.5% (range 0-100%). Failure in reproducibility was mainly due to results obtained with one particular primer. The number of epidemiological groups found to be homogeneous varied from 1-22 (median 16). However, for some groups an inhomogeneity was found by the majority of participants. The overall correlation between the results from the different participants ranged from 32 to 85%.     

Cystic prostatic disease associated with adrenocortical lesions in the ferret (Mustela putorius furo)

CONTEXT: In British Columbia, human immunodeficiency virus (HIV)-infected persons eligible for antiretroviral therapy may receive it free but the extent to which HIV-infected injection drug users access it is unknown. OBJECTIVE: To identify patient and physician characteristics associated with antiretroviral therapy utilization in HIV-infected injection drug users. DESIGN: Prospective cohort study with record linkage between survey data and data from a provincial HIV/AIDS (acquired immunodeficiency syndrome) drug treatment program. SETTING: British Columbia, where antiretroviral therapies are offered free to all persons with HIV infection with CD4 cell counts less than 0.50 x 10(9)/L (500/microL) and/or HIV-1 RNA levels higher than 5000 copies/mL. SUBJECTS: A total of 177 HIV-infected injection drug users eligible for antiretroviral therapy, recruited through the prospective cohort study since May 1996. MAIN OUTCOME MEASURES: Patient use of antiretroviral drugs through the provincial drug treatment program and physician experience treating HIV infection. RESULTS: After a median of 11 months after first eligibility, only 71 (40%) of 177 patients had received any antiretroviral drugs, primarily double combinations (47/71 [66%]). Both patient and physician characteristics were associated with use of antiretroviral drugs. After adjusting for CD4 cell count and HIV-1 RNA level at eligibility, odds of not receiving antiretrovirals were increased more than 2-fold for females (odds ratio [OR], 2.53; 95% confidence interval [CI], 1.08-5.93) and 3-fold for those not currently enrolled in drug or alcohol treatment programs (OR, 3.49; 95% CI, 1.45-8.40). Younger drug users were less likely to receive therapy (OR, 0.47/10-y increase; 95% CI, 0.28-0.80). Those with physicians having the least experience treating persons with HIV infection were more than 5 times less likely to receive therapy (OR, 5.55; 95% CI, 2.49-12.37). CONCLUSIONS: Despite free antiretroviral therapy, many HIV-infected injection drug users are not receiving it. Public health efforts should target younger and female drug users, and physicians with less experience treating HIV infection.     

Neuropsychological and motor functioning after unilateral anatomically guided posterior ventral pallidotomy. Preoperative performance and three-month follow-up

This study presents baseline and 3-month follow-up motor and neuropsychological data for 22 patients with Parkinson's disease (PD) who underwent anatomically guided unilateral posterior ventral pallidotomy (PVP). Postsurgical improvements were seen in psychomotor speed, fine motor accuracy, and dyskinesia, whereas grip strength decreased on the side contralateral to the surgery. No change was detected in overall level of cognitive functioning, nor were changes demonstrated in memory, language, or working memory when the entire sample of patients was evaluated. When the group was divided on the basis of side of surgery, patients with left-sided pallidotomies showed a decline in verbal fluency. Patients and caregivers reported improvement in psychosocial functioning. These initial findings of improved motor performance and largely unaffected cognitive functions are consistent with results obtained with functional PVP and provide support for the use of anatomically guided posterior ventral pallidotomy in the treatment of motor symptoms of PD.     

The proximal ligand variant His93Tyr of horse heart myoglobin

The spectroscopic and structural properties of the His93Tyr variant of horse heart myoglobin have been studied to assess the effects of replacing the proximal His residue of this protein with a tyrosyl residue as occurs in catalases from various sources. The variant in the ferric form exhibits electronic spectra that are independent of pH between pH 7 and 10, and it exhibits changes in absorption maxima and intensity that are consistent with a five-coordinate heme iron center at the active site. The EPR spectrum of the variant is that of a high-spin, rhombic system similar to that reported for bovine liver catalase. The 1D 1H-NMR spectrum of the variant confirms the five-coordinate nature of the heme iron center and exhibits a broad resonance at 112.5 ppm that is attributable to the meta protons of the phenolate ligand. This result indicates that the new Tyr ligand flips at a significant rate in this protein. The thermal stability of the Fe(III) derivative is unchanged from that of the wild-type protein (pH 8) while the midpoint reduction potential [-208 mV vs SHE (pH 8.0, 25 degrees C)] is about 250 mV lower. The three-dimensional structure of the variant determined by X-ray diffraction analysis confirms the five-coordinate nature of the heme iron center and establishes that the introduction of a proximal Tyr ligand is accommodated by a shift of the F helix (residues 88-99) in which this residue resides away from the heme pocket. Additional effects of this change are small shifts in the positions of Leu29, a heme propionate, and a heme vinyl group that are accompanied by altered hydrogen bonding interactions with the heme prosthetic group.(ABSTRACT TRUNCATED AT 250 WORDS)     

Kinetic parameters for the vesicular acetylcholine transporter: two protons are exchanged for one acetylcholine

The vesicular acetylcholine transporter (VAChT) mediates ACh storage in synaptic vesicles by exchanging cytoplasmic ACh with vesicular protons. This study sought to determine the stoichiometry of exchange by analysis of ligand binding and transport kinetics. The effects of different pH values inside and outside, external ACh concentrations, and electrical potential gradients on ACh transport by vesicles isolated from the electric organ of Torpedo were determined using a pH-jump protocol. The equilibrium binding of a high-affinity analogue of ACh is inhibited by protonation with a pKa of 7.4 +/- 0.3. A two-proton model fits the transport data much better than a one-proton model does, and uptake increases at more positive internal electrical potential, as expected for the two-proton model. Thus, the results support the two-proton model. The transport cycle begins with binding of external ACh to outwardly oriented site 2 (KACho = 20 mM) and protonation of inwardly oriented site 1 (pKa1 = 4.73 +/- 0.05). Loaded VAChT reorients quickly (73 000 min-1) and releases ACh to the inside (KAChi = 44 000 mM) and the proton to the outside. Unloaded, internally oriented site 2 binds a proton (pKa2 = 7.0), after which VAChT reorients (150 +/- 20 min-1) in the rate-limiting step and releases the proton to the outside to complete the cycle. Rate constants for the reverse direction also were estimated. Two protons provide a thermodynamic driving force beyond that utilized in vivo, which suggests that vesicular filling is regulated. Other phenomena related to VAChT, namely the time required to fill synaptic vesicles, the fractional orientation of the ACh binding site toward cytoplasm, orientational lifetimes, and the rate of nonquantal release of ACh from cholinergic nerve terminals, were computer-simulated, and the results are compared with physiological observations.