Lipids in the inner membrane of dormant spores of Bacillus species are largely immobile

Bacterial spores of various Bacillus species are impermeable or exhibit low permeability to many compounds that readily penetrate germinated spores, including methylamine. We now show that a lipid probe in the inner membrane of dormant spores of Bacillus megaterium and Bacillus subtilis is largely immobile, as measured by fluorescence redistribution after photobleaching, but becomes free to diffuse laterally upon spore germination. The lipid immobility in and the slow permeation of methylamine through the inner membrane of dormant spores may be due to a significant (1.3- to 1.6-fold) apparent reduction of the membrane surface area in the dormant spore relative to that in the germinated spore, but is not due to the dormant spore's high levels of dipicolinic acid and divalent cations.     

A proinflammatory chemokine, CCL3, sensitizes the heat- and capsaicin-gated ion channel TRPV1

Pain, a critical component of host defense, is one hallmark of the inflammatory response. We therefore hypothesized that pain might be exacerbated by proinflammatory chemokines. To test this hypothesis, CCR1 was cotransfected into human embryonic kidney (HEK)293 cells together with transient receptor potential vanilloid 1 (TRPV1), a cation channel required for certain types of thermal hyperalgesia. In these cells, capsaicin and anandamide induced Ca(2+) influx mediated by TRPV1. When CCR1:TRPV1/HEK293 cells were pretreated with CCL3, the sensitivity of TRPV1, as indicated by the Ca(2+) influx, was increased approximately 3-fold. RT-PCR analysis showed that a spectrum of chemokine and cytokine receptors is expressed in rat dorsal root ganglia (DRG). Immunohistochemical staining of DRG showed that CCR1 is coexpressed with TRPV1 in >85% of small-diameter neurons. CCR1 on DRG neurons was functional, as demonstrated by CCL3-induced Ca(2+) ion influx and PKC activation. Pretreatment with CCL3 enhanced the response of DRG neurons to capsaicin or anandamide. This sensitization was inhibited by pertussis toxin, U73122, or chelerythrine chloride, inhibitors of Gi-protein, phospholipase C, and protein kinase C, respectively. Intraplantar injection of mice with CCL3 decreased their hot-plate response latency. That a proinflammatory chemokine, by interacting with its receptor on small-diameter neurons, sensitizes TRPV1 reveals a previously undescribed mechanism of receptor cross-sensitization that may contribute to hyperalgesia during inflammation.     

Decreased expression of phospholipase C-beta 2 isozyme in human platelets with impaired function

Platelets from a patient with a mild inherited bleeding disorder and abnormal platelet aggregation and secretion show reduced generation of inositol 1,4,5-trisphosphate, mobilization of intracellular Ca2+, and phosphorylation of pleckstrin in response to several G protein mediated agonists, suggesting a possible defect at the level of phospholipase C (PLC) activation (see accompanying report). A procedure was developed that allows quantitation of platelet PLC isozymes. After fractionation of platelet extracts by high-performance liquid chromatography, 7 out of 10 known PLC isoforms were detected by immunoblot analysis. The amount of these isoforms in normal platelets decreased in the order PLC- gamma 2 > PLC-beta 2 > PLC-beta 3 > PLC-beta 1 > PLC-gamma 1 > PLC- delta 1 > PLC-beta 4. Compared with normal platelets, platelets from the patient contained approximately one-third the amount of PLC-beta 2, whereas PLC-beta 4 was increased threefold. These results suggest that the impaired platelet function in the patient in response to multiple G protein mediated agonists is attributable to a deficiency of PLC-beta 2. They document for the first time a specific PLC isozyme deficiency in human platelets and provide an unique opportunity to understand the role of different PLC isozymes in normal platelet function.     

Possible effect of secretor locus on plasma concentration of factor VIII and von Willebrand factor

A significant fraction (30%) of the genetically determined variance in plasma concentration of the von Willebrand factor antigen (vWf:Ag) has been shown to be related to ABH determinants. Individuals with blood group O, who have the highest amounts of blood group H substance, have the lowest concentration of vWf:Ag. The Lewis substances, Le(a) and Le(b), are biochemically closely related to the ABH substances as both can be produced from the same precursor substance. We studied the effect of the presence of the Lewis antigens on the plasma concentration of vWf:Ag and factor VIII antigen (VIII:Ag) in 323 individuals of different ABO groups from a series of twins and in 58 blood donors of blood group O. Among persons belonging to blood group O, those with the Le(a) antigen had a higher concentration of both vWf:Ag and VIII:Ag than individuals lacking Le(a). Le(a+b-) people are nonsecretors and Le(a-b+) people are secretors of ABH substance. Thus, the lowest concentration of vWf:Ag and VIII:Ag was found in group O secretors. The effect is most likely due to an effect of the secretor locus. This finding may be of importance for the detection of carriers of hemophilia A and for the diagnosis of type I von Willebrand disease.     

Effect of withdrawal of somatostatin and growth hormone (GH)-releasing factor on GH release in vitro

The secretion of GH, in vivo, is pulsatile. We have proposed that the timing of the episodic bursts of GH secretion is set by somatostatin (SRIF) withdrawal, while the magnitude of the bursts is set by the amount of GH-releasing factor (GRF) impinging on the somatotrophs, before and during SRIF withdrawal. We have now used an in vitro model of perifused rat pars distalis cells to further examine the interaction between GRF and SRIF on the magnitude of the burst of GH release that follows SRIF withdrawal. We first characterized the GH response, with time, to constant perifusion with GRF. The initial burst, followed by a rapid decrease in GH release induced by constant perifusion is due to a loss of GRF bioactivity in the perifusion medium and not to a decreasing responsiveness of the somatotrophs. This was followed by studies on the interaction between GRF and SRIF. The burst of GH release after cessation of perifusion with SRIF (10(-9) M) plus GRF (10(-10) M) can be blocked by the administration of SRIF during the burst. Also, the magnitude of the burst is proportional to the concentration of GRF preceding the withdrawal of SRIF. It is likely that similar relations apply in vivo, where SRIF withdrawal sets the timing and duration of the episodic burst of GH release, while GRF sets the magnitude.     

Synergism between insulin and low concentrations of isoproterenol in the stimulation of leptin release by cultured human adipose tissue

The release of leptin by pieces of human adipose tissue incubated in primary culture for 24 or 48 hours in the presence of dexamethasone was reduced by isoproterenol. An inhibition of leptin release was observed at 24 hours in the presence of isoproterenol and was mediated by beta1-adrenergic receptors, since it was blocked by the specific beta1-adrenoceptor antagonist CGP-20712A. The inhibitory effect of 33 nmol/L isoproterenol on leptin release was reversed in the presence of 0.1 nmol/L insulin to a 2-fold stimulation of leptin release. These data suggest that the primary mechanism by which insulin stimulates leptin release is to blunt the inhibitory effects of beta1-adrenergic receptor agonists, and low concentrations of catecholamines actually enhance the stimulation of leptin release by insulin.     

Implementing rapid testing for tuberculosis in Mozambique

Problem

In Mozambique, pulmonary tuberculosis is primarily diagnosed with sputum smear microscopy. However this method has low sensitivity, especially in people infected with human immunodeficiency virus (HIV). Patients are seldom tested for drug-resistant tuberculosis.

Approach

The national tuberculosis programme and Health Alliance International introduced rapid testing of smear-negative sputum samples. Samples were tested using a polymerase-chain-reaction-based assay that detects Mycobacterium tuberculosis deoxyribonucleic acid and a mutation indicating rifampicin resistance; Xpert® MTB/RIF (Xpert®). Four machines were deployed in four public hospitals along with a sputum transportation system to transfer samples from selected health centres. Laboratory technicians were trained to operate the machines and clinicians taught to interpret the results.

Local setting

In 2012, Mozambique had an estimated 140 000 new tuberculosis cases, only 34% of which were diagnosed and treated. Of tuberculosis patients, 58% are HIV-infected.

Relevant changes

From 2012–2013, 1558 people were newly diagnosed with tuberculosis using sputum smears at intervention sites. Xpert® detected M. tuberculosis in an additional 1081 sputum smear-negative individuals, an increase of 69%. Rifampicin resistance was detected in 58/1081 (5%) of the samples. However, treatment was started in only 82% of patients diagnosed by microscopy and 67% of patients diagnosed with the rapid test. Twelve of 16 Xpert® modules failed calibration within 15 months of implementation.

Lessons learnt

Using rapid tests to diagnose tuberculosis is promising but logistically challenging. More affordable and durable platforms are needed. All patients diagnosed with tuberculosis need to start and complete treatment, including those who have drug resistant strains.     

Characterizing surf zone injuries from the five most populated beaches on the Atlantic-fronting Delaware coast

Introduction

Beaches are a popular destination for recreation activities. Surf zone injuries (SZI) can occur resulting from a variety of in-water activities. Little is known regarding the sustained injury types, or demographics of injured persons and activities leading to injuries.