Recommended Immunological Strategies to Screen for Botulinum Neurotoxin-Containing Samples

Botulinum neurotoxins (BoNTs) cause the life-threatening neurological illness botulism in humans and animals and are divided into seven serotypes (BoNT/A–G), of which serotypes A, B, E, and F cause the disease in humans. BoNTs are classified as “category A” bioterrorism threat agents and are relevant in the context of the Biological Weapons Convention. An international proficiency test (PT) was conducted to evaluate detection, quantification and discrimination capabilities of 23 expert laboratories from the health, food and security areas. Here we describe three immunological strategies that proved to be successful for the detection and quantification of BoNT/A, B, and E considering the restricted sample volume (1 mL) distributed. To analyze the samples qualitatively and quantitatively, the first strategy was based on sensitive immunoenzymatic and immunochromatographic assays for fast qualitative and quantitative analyses. In the second approach, a bead-based suspension array was used for screening followed by conventional ELISA for quantification. In the third approach, an ELISA plate format assay was used for serotype specific immunodetection of BoNT-cleaved substrates, detecting the activity of the light chain, rather than the toxin protein. The results provide guidance for further steps in quality assurance and highlight problems to address in the future.     

Magnitude of body-cell-mass depletion and the timing of death from wasting in AIDS

The impact of malnutrition on survival in AIDS was evaluated by examining the magnitude of body-cell-mass depletion as a function of time from death. Body cell mass was estimated as total body-potassium content and determined by whole-body counting. There was progressive depletion of body cell mass as patients neared death. The extrapolated and observed values for body cell mass at death were 54% of normal. Body weight had a similar relationship to death, with a projected body weight at death of 66% of ideal. We conclude that death from wasting in AIDS is related to the magnitude of tissue depletion and is independent of the underlying cause of wasting. The degree of wasting seen in this study is similar to historical reports of semistarvation, with or without associated infections. This observation suggests that successful attempts to maintain body mass could prolong survival in patients with AIDS.     

Beta adrenoceptor control of the microvascular reserve in rabbit myocardium

This study was performed to determine if the unperfused microvascular reserve in the rabbit heart can be controlled by beta adrenoceptors. Anesthetized, open chest rabbits (N = 54) were subjected to saline i.v., 0.7 mg/kg of atenolol i.v., 1 mg/kg of practolol, 0.1 microgram/kg of salbutamol, 1 microgram/kg of salbutamol or 0.7 mg/kg of atenolol + 1 microgram/kg of salbutamol treatments. In half these animals coronary flows were determined before and after treatment using radioactive microspheres. The others were given 100 mg/kg of fluorescein isothiocyanate-dextran and the hearts were removed and analyzed for perfused and total microvascular morphology. The fluorescence marked the perfused microvessels and the slides were stained to show all vessels. Control group blood flow was 213 +/- 25 ml/min/100 g. Flows decreased 25% with atenolol and practolol whereas no change was seen with salbutamol or atenolol + salbutamol. Total capillary volume fraction ranged from 0.15 to 0.20 mm3/mm3 with 60% of this perfused in controls. Atenolol significantly reduced this to 47% whereas practolol (90%), high-dose salbutamol (97%) and salbutamol + atenolol (99%) resulted in a significant mobilization. Low-dose salbutamol resulted in a mobilization of capillaries to a degree intermediate (79%) between controls and high-dose salbutamol. Total arteriolar volume fraction ranged from 0.002 to 0.004 mm3/mm3 and in controls 65% of this was perfused. This percentage was reduced with atenolol and significantly increased with all other treatments except practolol. Thus, blockade of beta-1 adrenoceptors results in decreases in the percentage of perfused microvessels whereas stimulation of beta-2 adrenoceptors results in an increase.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of age and gender on CNS serotonergic responsivity in normal adults

The effects of age and gender on central nervous system (CNS) serotonergic responsivity were assessed with a neuroendocrine challenge test in 30 normal adults. Subjects greater than or equal to 30 years of age, compared with younger subjects, exhibited decreased prolactin secretion in response to a 60-mg oral dose of dl-fenfluramine hydrochloride, an indirect serotonin agonist. Furthermore, women had greater prolactin responses than men. As prolactin secretory capacity appears to be stable through midlife, the age-associated decrease in fenfluramine-induced prolactin release suggests a decline in CNS serotonergic responsivity. In contrast, the finding of greater prolactin release in women than in men probably reflects the effects of nonserotonergic modulatory influences at the level of the lactotroph. Age and gender effects must be considered in studies of the CNS serotonergic system.     

Effect of medial arch-heel support in inserts on reducing ankle eversion: a biomechanics study

Background  

Excessive pronation (or eversion) at ankle joint in heel-toe running correlated with lower extremity overuse injuries. Orthotics and inserts are often prescribed to limit the pronation range to tackle the problem. Previous studies revealed that the effect is product-specific. This study investigated the effect of medial arch-heel support in inserts on reducing ankle eversion in standing, walking and running.     

A threonine residue in the M2 region of the beta1 subunit is needed for expression of functional alpha1beta1 GABA(A) receptors.

Although there is a high degree of homology in the M2 transmembrane segments of alpha1 and beta1 subunits, subunit-specific effects were observed in alpha1beta1 GABA(A) receptors expressed in Spodoptera frugipedra (Sf9) cells when the conserved 13' threonine residue in the M2 transmembrane region was mutated to alanine. When threonine 263 (13') was mutated to alanine in the beta1 subunit, high-affinity muscimol binding and the response to GABA were abolished. This did not occur when the threonine 263 (13') was mutated to alanine in the alpha1 subunit, but the rate of desensitisation increased and the effect of bicuculline, a competitive inhibitor, was reduced. The results show differential effects of subunits on receptor function and support a role for M2 in desensitisation.     

Laparoscopic pelvic lymph node dissection.

Traditionally, cancer of the prostate has been staged by digital exam, ultrasound, CT scan, bone scan, prostatic acid phosphatase (PAP), and prostate specific antigen (PSA) determinations. These methods commonly lead to understaging, resulting in surgical or radiation therapy of questionable benefit. Pathologic staging, even though reliable and accurate, requires laparotomy with its associated morbidity and lengthy hospitalization/recovery period. Following national trends, we have recently introduced the technique of Laparoscopic Pelvic Lymph Node Dissection (LPND) at our institution. In July 1990 we performed the first LPND at CAMC (Memorial Division). This report details our experience with the first three patients treated in this manner and suggest that the procedure can be performed safely, effectively, and with a significant reduction in morbidity, thus allowing the surgeon to obtain an adequate specimen for pathologic staging. Possible cost containment, minimal discomfort, and little scarring are other advantages that appeal to both patients and surgeons alike.     

Chronic toxicity and carcinogenicity studies of olestra in Fischer 344 rats

Two 2-year feeding studies were carried out in Fischer 344 rats with olestra, a mixture of the hexa-, hepta- and octaesters of sucrose formed with long-chain fatty acids. Olestra was fed at 0, 0.99, 4.76 or 9.09% (w/w) of the diet in the first study, and at 0 or 9.09% (w/w) of the diet in the second. Daily observations, feed consumption and body weights, ophthalmoscopic examinations, organ weights, serum chemistry, haematology, urinalysis and histopathological evaluations revealed no evidence of any adverse effects associated with olestra ingestion. Relative to controls, there was a higher incidence of basophilic liver foci in olestra-fed female rats at 12 months. At 24 months, foci were observed in most animals in all groups but were more numerous in olestra-fed females. The foci were not associated with hepatic tumours, alterations in liver function, or increases in liver weight and therefore not considered to represent a toxic response to olestra. Isolated statistically significant differences in mortality, mononuclear cell leukaemia, and pituitary adenomas were observed but were not considered to be related to olestra ingestion since they were not reproducible across the two studies, generally not dose responsive, not consistent between sexes, and the incidences were within the ranges for historical and contemporary laboratory controls. The results of the two studies show that olestra was not toxic or carcinogenic when fed to rats at up to 9% of the diet for 24 months.