A comparison between the one-step and the two-step copper thiocholine procedure for the cytochemical localization of cholinesterases

Summary A comparison between the one-step and the two-step copper thiocholine procedure for the subcellular localization of cholinesterases was under-taken. The results indicate that only under experimental conditions with short incubation times and precise control of the conversion into sulphide is the localization of the primary precipitate and that of the secondary precipitate identical. It was concluded that the conversion of the primary precipitate into Cu-sulphide is not necessary and can lead to artefacts.     

Molecular detection of<Emphasis Type="Italic">Ureaplasma urealyticum</Emphasis> infection from clinical urogenital swabs

A simple nucleic acid amplification test (NAAT) was developed for detection of Ureaplasma urealyticum infection based on the PCR amplification of the urease gene (UU1/UU2 Test). DNA was extracted from urogenital swabs and a 225-bp long DNA fragment was amplified by PCR. NAAT was compared to the commercial amplification kit for sexually transmitted disease reference assay. The sensitivity and specificity of the UU1/UU2 Test were determined to be 100 and 98.9%, respectively. The overall prevalence rate in this group of patients was found to be about 236 per 1000 (283 and 166 per 1000 in females and males, respectively). These data demonstrate that UU1/UU2 Test is suitable for effective epidemiological screening and/or diagnostic practice.     

Forest above-ground volume assessments with terrestrial laser scanning: a ground-truth validation experiment in temperate,managed forests

Background and AimsQuantifying the Earth’s forest above-ground biomass (AGB) is indispensable for effective climate action and developing forest policy. Yet, current allometric scaling models (ASMs) to estimate AGB suffer several drawbacks related to model selection and uncertainties about calibration data traceability. Terrestrial laser scanning (TLS) offers a promising non-destructive alternative. Tree volume is reconstructed from TLS point clouds with quantitative structure models (QSMs) and converted to AGB with wood basic density. Earlier studies have found overall TLS-derived forest volume estimates to be accurate, but highlighted problems for reconstructing finer branches. Our objective was to evaluate TLS for estimating tree volumes by comparison with reference volumes and volumes from ASMs.MethodsWe quantified the woody volume of 65 trees in Belgium (from 77 to 2800 L; Pinus sylvestris, Fagus sylvatica, Larix decidua, and Fraxinus excelsior) with QSMs and destructive reference measurements. We tested a volume expansion factor (VEF) approach by multiplying the solid and merchantable volume from QSMs by literature VEF values.Key ResultsStem volume was reliably estimated with TLS. Total volume was overestimated by +21 % using original QSMs, by +9 % and –12 % using two sets of VEF-augmented QSMs, and by –7.3 % using best-available ASMs. The most accurate method differed per site, and the prediction errors for each method varied considerably between sites.ConclusionsVEF-augmented QSMs were only slightly better than original QSMs for estimating tree volume for common species in temperate forests. Despite satisfying estimates with ASMs, the model choice was a large source of uncertainty, and species-specific models did not always exist. Therefore, we advocate for further improving tree volume reconstructions with QSMs, especially for fine branches, instead of collecting more ground-truth data to calibrate VEF and allometric models. Promising developments such as improved co-registration and smarter filtering approaches are ongoing to further constrain volumetric errors in TLS-derived estimates.     

Subcellular localization of Type VI secretion system assembly in response to cell–cell contact

Bacteria require a number of systems, including the type VI secretion system (T6SS), for interbacterial competition and pathogenesis. The T6SS is a large nanomachine that can deliver toxins directly across membranes of proximal target cells. Since major reassembly of T6SS is necessary after each secretion event, accurate timing and localization of T6SS assembly can lower the cost of protein translocation. Although critically important, mechanisms underlying spatiotemporal regulation of T6SS assembly remain poorly understood. Here, we used super‐resolution live‐cell imaging to show that while Acinetobacter and Burkholderia thailandensis can assemble T6SS at any site, a significant subset of T6SS assemblies localizes precisely to the site of contact between neighboring bacteria. We identified a class of diverse, previously uncharacterized, periplasmic proteins required for this dynamic localization of T6SS to cell–cell contact (TslA). This precise localization is also dependent on the outer membrane porin OmpA. Our analysis links transmembrane communication to accurate timing and localization of T6SS assembly as well as uncovers a pathway allowing bacterial cells to respond to cell–cell contact during interbacterial competition.     

CHOLINESTERASE ACTIVITY OF NODAL AND INTERNODAL REGIONS OF MYELINATED NERVE FIBERS OF FROG

The distribution of cholinesterase (Ch-esterase) in isolated myelinated fibers of the frog has been investigated. Quantitative microgasometric measurements have confirmed the previous histochemical observations. Both approaches indicate that in frog nerve fibers acetylcholinesterase (ACh-esterase) is the only or the predominant enzyme when selective inhibitors and different substrates are used: acetylcholine (ACh), butyrylcholine, and acetyl-B-methylcholine (Mecholyl). By means of the microgasometric technique, a significant difference in ACh-esterase activity between axons isolated from ventral (37.2 ± 1.7 µmole x 10^-5 ACh/mm²/hr) and dorsal roots (2.0 ± 0.9 µmole x 10^-5 ACh/mm²/hr) was found. In the region of the node of Ranvier the enzyme activity (50.4 ± 4.4 µmole x 10^-5 ACh/mm²/hr) appears to be considerably higher than in the internodal area (36.6 ± 2.1 µmole x 10^-5 ACh/mm²/hr). The findings are discussed in relation to the theory of saltatory conduction and the ACh system.     

Cystatin, a protein inhibitor of cysteine proteases alters viral protein cleavages in infected human cells

Chicken cystatin is a previously described small protein which has the property of inhibiting cysteine active site proteases. The protein, when added to cultured human cells, alters the intracellular proteolytic processing of poliovirus proteins, and causes a reduction in virus yield. It is suggested that the cystatin is able to penetrate to the cellular cytoplasm and inhibit the action of the poliovirus protease.     

Survival outcomes and effect of early vs. deferred cART among HIV-infected patients diagnosed at the time of an AIDS-defining event: a cohort analysis

Objectives

We analyzed clinical progression among persons diagnosed with HIV at the time of an AIDS-defining event, and assessed the impact on outcome of timing of combined antiretroviral treatment (cART).

Methods

Retrospective, European and Canadian multicohort study.. Patients were diagnosed with HIV from 1997–2004 and had clinical AIDS from 30 days before to 14 days after diagnosis. Clinical progression (new AIDS event, death) was described using Kaplan-Meier analysis stratifying by type of AIDS event. Factors associated with progression were identified with multivariable Cox regression. Progression rates were compared between those starting early (<30 days after AIDS event) or deferred (30–270 days after AIDS event) cART.

Results

The median (interquartile range) CD4 count and viral load (VL) at diagnosis of the 584 patients were 42 (16, 119) cells/µL and 5.2 (4.5, 5.7) log₁₀ copies/mL. Clinical progression was observed in 165 (28.3%) patients. Older age, a higher VL at diagnosis, and a diagnosis of non-Hodgkin lymphoma (NHL) (vs. other AIDS events) were independently associated with disease progression. Of 366 patients with an opportunistic infection, 178 (48.6%) received early cART. There was no significant difference in clinical progression between those initiating cART early and those deferring treatment (adjusted hazard ratio 1.32 [95% confidence interval 0.87, 2.00], p = 0.20).

Conclusions

Older patients and patients with high VL or NHL at diagnosis had a worse outcome. Our data suggest that earlier initiation of cART may be beneficial among HIV-infected patients diagnosed with clinical AIDS in our setting.