Biomedical subjects
Publications and source records attributed to J G Morris.
A rapid protocol for subtyping vancomycin-resistant enterococci by pulsed-field gel electrophoresis is reported. The procedure is simple and potentially cost-effective and allows reproducible subtyping of the strains in approximately 1 day.
There have been no previous three-dimensional volumetric studies of regional brain atrophy in patients with pathologically confirmed progressive supranuclear palsy (PSP). Postmortem cortical and subcortical volumes were compared with neuropathology in 9 patients with PSP, 15 patients with Parkinson's disease, 10 patients with dementia with Lewy bodies, and 23 controls. Cases with the neuritic pathology of Alzheimer's disease were excluded. The topography of brain atrophy differed according to clinicopathological phenotype. Patients with Parkinson's disease had atrophy confined to the amygdala. Atrophy of the frontal lobe was found in both PSP and dementia with Lewy bodies and correlated with increasing neurofibrillary tangle or Lewy body densities, respectively. Patients with PSP could be differentiated by their marked atrophy of the internal globus pallidus. Further analysis of variance revealed that trends for greater frontal lobe atrophy correlated with clinical dementia in PSP, whereas both greater frontal and hippocampal atrophy and higher densities of Lewy bodies and Lewy neurites correlated with clinical dementia in cases with Lewy bodies. The present study provides evidence for selective regional atrophy that correlates with the underlying pathology of PSP and Lewy body disease.
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The gait and other clinical features of 22 patients presenting to our hospital over the last 10 years are shown on video. In 12 patients, a diagnosis of psychogenic gait was made; in the remainder, the gait abnormality was the result of a neurologic disease. Psychogenic gaits are compared and contrasted with "organic" gaits. In one patient, the psychogenic gait occurred in the setting of a neurologic disease. The "traditional" approach to psychogenic gait, attempting to exclude underlying neurologic and psychiatric disease and seeking evidence for primary and secondary gain, was found to be of limited value. More useful were the features of the gait itself, in particular, exaggerated effort, extreme slowness, variability throughout the day, unusual or uneconomic postures, collapses, convulsive tremors, and distractibility; certain aspects of the history were also helpful. A list of comments is provided. The diagnosis of psychogenic gait, particularly in the elderly, remains fraught with hazard, and a balance has to be sought between subjecting an anxious patient to needless investigations and yet not losing sight of the fact that the patient may be elaborating on symptoms of genuine disease. The bizarre gait of some neurologic disorders, particularly dystonia and chorea, may be a pitfall for the unwary.
The higher copy number of pUC19, compared to its parent plasmid pBR322, is known to be due to deletion of rop, also known as rom, and to an ori mutation that impedes RNAI:RNAII interaction. pUC19, unlike pBR322, fails to transform E. coli rho mutant rho026 cells. Here we identify two features of pUC19 that contribute to this transformation defect. (1) The pUCori mutation is involved because replacing the pUCori with that of pBR322 restored transformation. (2) Transcription from the lac promoter in pUC19 is important, since deletion or inversion of the promoter or insertion of a transcription terminator (lambdat0) downstream of it restored transformation. Host RNase E activity is responsible for the transformation defect because introduction of an rne-1 allele into rho026 cells suppressed this defect, indicating that RNAI instability due to RNase E is aggravated in the rho026 strain. We suggest that in rho026 cells pUC19 RNAI:RNAII interaction is more impeded than in rho+ cells and Rop/Rom may confer stability by protecting RNAI against RNase E activity because expression of a rom gene inserted into pUC19 restored transformation.
Mutations in the brain specific P/Q type Ca2+ channel alpha1 subunit gene, CACNA1A, have been identified in three clinically distinct disorders, viz. episodic ataxia type 2 (EA-2), familial hemiplegic migraine (FHM) and spinocerebellar ataxia 6 (SCA6). For individuals with EA-2, the mutations described thus far are presumed to result in a truncated protein product. Several different missense mutations have been identified in patients with FHM. At least two of these mutations have been identified on two different chromosome 19p13 haplotypes and thus represent recurrent mutations. In the present study, we have screened several individuals for mutations in all 47 exons in the CACNA1A gene by single-strand conformation analysis. We have characterised a novel missense mutation, G5260A, in exon 32 in a family segregating for EA-2. The consequence of this mutation is an amino acid substitution at a highly conserved position within the CACNA1A gene. This represents the first point mutation not resulting in a proposed truncated protein. Furthermore, this mutation has been detected in a family member with mild clinical signs including only migraine. Additionally, a second previously identified recurrent muta tion, C2272T, in exon 16 has been discovered in a patient with FHM.
BACKGROUND: Gram-positive organisms, including vancomycin-resistant enterococci (VRE), have emerged as major pathogens on the organ transplant service at our institution. We hypothesized that our use of vancomycin as part of routine surgical prophylaxis increased the risk of VRE colonization and infection; conversely, there was concern that failure to use vancomycin prophylaxis would increase peri-operative morbidity due to gram-positive organisms. METHODS: Renal transplant recipients (n = 88) were randomized to receive either a) vancomycin/ceftriaxone or b) cefazolin; and pancreas transplants (n = 24) to receive either a) vancomycin/gentamicin or b) cefazolin/gentamicin. Stool samples or rectal swabs were obtained for culture for enterococci within 24 h of transplantation and weekly while hospitalized. RESULTS: Enterococci were isolated on stool culture from 38 (34%) of 102 patients at the time of transplantation; 4 (11%) of the isolates were VRE. The percentage of patients who subsequently acquired VRE was low (1-7% per wk) but remained constant during hospitalization. There was no association between new VRE detection and vancomycin use for either prophylactic or therapeutic purposes. Forty-four patients (39%) had a post-operative infection with 46% of these infections due to gram-positive organisms; rates were unaffected by prophylactic vancomycin use. Pancreas transplant patients who did not receive vancomycin prophylaxis had a significantly longer initial hospitalization (p = 0.03); however, differences were not statistically significant when total length of stay (LOS) within the first 90 d of transplantation was compared. CONCLUSIONS: Vancomycin surgical prophylaxis does not appear to have an effect on VRE colonization or infection, or on rates of infection with gram-positive bacteria. Elimination of vancomycin prophylaxis in renal transplant patients may be a reasonable part of an overall program to limit vancomycin usage, although as a single measure, its impact may be minimal. Vancomycin surgical prophylaxis may be of greater importance in pancreas transplants.
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The minimal chlorine (chloride) requirement of kittens for growth was determined in a study in which kittens were given purified diets containing 0.1, 0.4, 0.7, 1.0 or 1.3 g of chloride (Cl) as KCl/kg diet. Each dietary group contained six (three males and three females) specific-pathogen-free kittens; the diets were fed for 30 d. Kittens receiving diets with <0.7 g Cl/kg became alkalotic as evidenced by an increase in blood pH, blood bicarbonate, standard bicarbonate, actual base excess, standard base excess, partial pressure of carbon dioxide (pCO(2)) and total CO(2). In addition, kittens were hypochloremic and hypokalemic; they had decreased serum ionized calcium and a negative chloride balance. On the basis of the chloride balance measurements, a broken-line nonlinear least-square analysis predicted a Cl requirement as 0.89 g Cl/kg diet (22 kJ/g diet). Because the dietary Cl concentration closest to 0.89 g/kg that we tested was 1.0 g Cl/kg, we recommend a minimum chloride requirement of 1.0 g Cl/kg diet for growing kittens. This value is considerably less than the recommended chloride requirement of the National Research Council of 1.9 g Cl/kg diet, or the allowance of the Association of American Feed Control Officials of 3.0 g Cl/kg diet. Because the bioavailability of chloride is high, the previous estimates appear excessive.
The objective of this study was to determine the effect of dietary protein and taurine on cysteine desulfhydration in various kitten tissues. Cysteine desulfhydration was assessed in liver, kidney, skeletal muscle, heart, spleen, brain and jejunum of kittens fed one of the following diets for 5 wk: 20% protein, 0% taurine diet (LP0T); 20% protein, 0.15% taurine diet (LPNT); 60% protein, 0% taurine diet (HP0T); and 60% protein, 0.15% taurine diet (HPNT). Cats fed LP0T and HP0T had been fed a taurine-free diet for 10 wk before the 5-wk experiment. The activity of cysteine desulfhydration was determined by measuring the production of H(2)(35)S from (35)S-cysteine in the presence and absence of alpha-ketoglutarate (alphaKG) in the incubation medium. Liver and kidney had the highest total activities among the tissues tested (P < 0.01). Total hepatic desulfhydration activities [micromol H(2)S/(min. kg body wt)] in cats fed LP0T, LPNT, HP0T and HPNT were (mean +/- SEM) 117 +/- 6, 135 +/- 10, 137 +/- 10 and 190 +/- 9, respectively. Dietary taurine had a significant effect on activity when expressed per gram liver (P < 0.01), per gram protein (P < 0.05) and per kilogram body weight (P < 0.001). Dietary protein had a significant effect (P < 0.001) only when activity was expressed relative to body weight because of the significant effect of protein on relative liver weight. The direct pathway via cysteine desulfhydrase appears to be the major route of cysteine desulfhydration in kitten liver because the values obtained in the absence of alphaKG were 81-88% of those obtained in the presence of alphaKG.
The sodium requirement of adult cats for maintenance was determined using a randomized block design of eight dietary sodium treatments (0.1, 0.4, 0.5, 0.66, 0.8, 1.2, 1.6 or 2.0 g Na/kg in a casein-lactalbumin-based purified diet) administered for periods of 4 wk. A total of 35 adult specific-pathogen-free domestic shorthaired cats (26 males and 9 females, 1.5-3 y of age) was given an equilibration diet (2 g Na/kg) for 14 d before assignment (or reassignment) to the treatments. A total of 12 cats (8 males, 4 females) was randomly assigned to the lowest six levels of sodium, and four cats to the highest two sodium levels. Cats consuming the diet containing 0.1 g Na/kg had significantly elevated aldosterone concentration in plasma, and packed cell volume. In addition, these cats exhibited anorexia, body weight loss, reduced urinary specific gravity and sodium excretion, and had a negative sodium balance. However, adult cats did not develop polydypsia and polyuria reported in sodium-deficient kittens. Cats given the diet containing 0.66 g Na/kg did not have an increased packed cell volume, but aldosterone concentration in the plasma was significantly elevated. However, cats given diets containing >/=0.8 g Na/kg had plasma aldosterone concentrations </=0.7 nmol/L (reference value for sodium-replete cats) and normal packed-cell volumes. A minimal sodium requirement of adult cats for maintenance of 0.8 g Na/kg diet (energy density = 22 kJ/g diet) or 0.4 mmol Na. kg body weight-1. d-1 is proposed.
Changes in plasma 25-hydroxyvitamin D (25-OHD) were used as an index of vitamin D status of cats. Plasma 25-OHD concentration of kittens given a purified vitamin D-free diet and exposed to direct summer sun for 15 h/wk declined at a similar rate as kittens given the same diet kept indoors. Similarly, plasma 25-OHD of kittens exposed to ultraviolet (UV) lamps declined at a similar rate as kittens not exposed, and these kittens developed clinical signs of vitamin D deficiency. Eight weaned kittens were given the vitamin D-free purified diet until their plasma concentrations of 25-OHD were < 5 nmol/L. They then had the hair on their backs clipped at weekly intervals and were paired on the basis of skin color and exposed to UV light for 2 h/d. One member of each pair was given an inhibitor of 7-dehydrocholesterol (5, 7-cholestradien-3beta-ol)-delta7-reductase (EC 1.3.1.21) in the diet. Cats receiving the inhibitor had a progressive increase in 25-OHD concentration of plasma with time to 91 +/- 22 nmol/L (mean +/- SEM), whereas cats not receiving the inhibitor had plasma 25-OHD concentrations that were not detectable (P < 0.001). Biopsy samples of skin from cats receiving the inhibitor had more than five times the concentration of 7-dehydrocholesterol (P < 0.001) than the skin of control cats. Low concentration of 7-dehydrocholesterol (presumably due to high activity of the reductase) in the skin of cats is the major impediment to effective vitamin D synthesis. Analysis of wild caught potential prey of cats indicated that these animals could supply adequate vitamin D to meet the requirement of growing kittens.
Vitamin D synthesis by growing kittens exposed to ultraviolet light is ineffective. Concentration of 25-hydroxyvitamin D (25-OHD) in plasma (the most useful index of vitamin D status) was measured in six groups each of seven kittens given a purified diet (12 g calcium and 8 g phosphorus/kg, calculated metabolizable energy = 20 kJ/g) that contained either 0.0, 3.125, 6.25, 12.5, 18.75 or 25 microg of cholecalciferol/kg diet. All kittens received these diets from 9 to 22 wk of age, and the two groups given the 0.0 and 3.125 microg cholecalciferol/kg treatments continued to receive the diets until they were 34 wk old. Total and ionizable calcium and phosphorus in plasma were not affected by treatments. No adverse clinical changes were observed or found on radiographic examination of the kittens at 22 or 34 wk of age. Plasma concentration of 25-OHD was linearly related (r2 = 0.99, P < 0.001) to dietary intake of cholecalciferol. Plasma concentration of 25-OHD in kittens given the diet without added vitamin D was significantly less at 22 wk than at 9 wk, whereas kittens receiving the diet containing 3.125 microg cholecalciferol/kg had significantly higher 25-OHD concentrations at 22 and 34 wk than at 9 wk of age. Kittens given the 6.25 microg cholecalciferol/kg diet had plasma 25-OHD concentrations at 22 wk > 50 nmol/L which is considered replete for humans. An allowance of 6. 25 microg (250 IU) of cholecalciferol/kg diet is suggested to provide a margin of safety.
We previously demonstrated that a purified diet containing 3.125 microg of cholecalciferol/kg was adequate to maintain plasma concentrations of 25-hydroxyvitamin D in growing kittens. With the use of this concentration of cholecalciferol, the response of growing kittens to varying levels of calcium in purified diets was measured. Five groups (treatments 1-5), each comprised of seven weaned kittens, were given diets containing 3.8, 5.0, 6.0, 7.2 or 8. 1 g calcium/kg diet (Ca:P ratio of 1:1.25) from 9 to 18 wk of age. Two further groups of kittens (treatments 6 and 7) received similar diets containing 6.0 g Ca/kg diet, with Ca:P ratios of 1:1.55 and 1:2.61, respectively. No clinical signs of calcium deficiency were observed, i.e., growth rate, energy intake and plasma total calcium were not affected by the treatments. However, ionized calcium was significantly lower in kittens in treatment 7. Plasma phosphorus was lower in kittens in treatment 7 than in kittens in treatments 1, 2, 3 and 4, and there was a negative relationship between dietary and plasma phosphorus concentrations. Kittens in treatment 7 had a significantly higher alkaline phosphatase concentration in plasma than kittens in treatments 1, 2, 3 and 5. Kittens in treatment 1 had a lower percentage of bone minerals measured by dual-energy X-ray absorptiometry than kittens in treatments 2-6. These results indicate that the calcium requirement of growing kittens is not >6.0 g/kg diet, (calculated metabolizable energy approximately 20 kJ/g) and that kittens are not very sensitive to inverse Ca:P ratios up to 1:1.55.
The DNA sequence of the O-antigen biosynthesis cluster (wbf) of a recently emergent pathogen, Vibrio cholerae serogroup O139, has been determined. Here we report the sequence of the genes downstream of the O139 wbfX gene and analysis of the genes flanking the wbf gene cluster in other serogroups. The gene downstream of wbfX, designated rjg (right junction gene), is predicted to be not required for O-antigen biosynthesis but appears to be a hot spot for DNA rearrangements. Several variants of the rjg gene (three different insertions and a deletion) have been found in other serogroups. DNA dot blot analysis of 106 V. cholerae strains showed the presence of the left and right junction genes, gmhD and rjg, respectively, in all strains. Further, these genes mapped to a single I-CeuI fragment in all 21 strains analyzed by pulsed-field gel electrophoresis, indicating a close linkage. The insertion sequence element IS1358, found in both O1 and O139 wb* regions, is present in 61% of the strains tested; interestingly, where present, it is predominantly linked to the wb* region. These results indicated a cassette-like organization of the wb* region, with the conserved genes (gmhD and rjg) flanking the divergent, serogroup-specific wb* genes and IS1358. A similar organization of the wb* region in other serogroups raises the possibility of the emergence of new pathogens by homologous recombination via the junction genes.
Yersinia bercovieri, a recently identified Y. enterocolitica-like species, produces a heat-stable enterotoxin (designated YbST) which has biologic activity in infant mice and increases short circuit current in Ussing chambers. Although YbST has some properties in common with the heat-stable enterotoxins of Y. enterocolitica (YST I and YST II), it appears to be a novel toxin because (i) it was not neutralized by anti-YST I antiserum, (ii) YbST-neutralizing antiserum did not neutralize YST I, and (iii) Y. bercovieri strains did not hybridize with genetic probes for yst I, yst II, and other known enterotoxins.
Vibrio vulnificus is a human pathogen whose virulence has been associated with the expression of capsular polysaccharide (CPS). Multiple CPS types have been described; however, virulence does not appear to correlate with a particular CPS composition. Reversible-phase variation for opaque and translucent colony morphologies is characterized by changes in CPS expression, as suggested by electron microscopy of cells stained nonspecifically with ruthenium red. Isolates with opaque colony morphologies are virulent and appear to be more thickly encapsulated than naturally occurring translucent-phase variants, which have reduced, patchy, or absent CPS. Previously, we have shown that the virulence of translucent-phase variants was intermediate between opaque-phase variants and acapsular transposon mutants, suggesting a correlation between virulence and the amount of CPS expressed. In the present study, CPS expression of phase variants and genetically defined mutants of V. vulnificus M06-24/O was examined by using a CPS-specific monoclonal antibody with an enzyme-linked immunosorbent assay, flow cytometry, and immunoelectron microscopy. Semiquantitative analyses of CPS expression correlated well among these assays, confirming that the translucent-phase variant was intermediate in CPS expression and retained type I CPS-specific epitopes. Cell surface expression of CPS varied with the growth phase, increasing during logarithmic growth and declining in stationary culture. Significantly greater CPS expression (P = 0.026) was observed for cells grown at 30 degrees C than for those at 37 degrees C. These studies confirm that phase variation and virulence in V. vulnificus correlate with the amount of CPS expressed and demonstrate the fluidity of bacterial polysaccharide expression in response to environmental conditions.