The structure of the transposable genetic element ISBsu2 in the cryptic plasmid p1516 from a soil Bacillus subtilis strain and the presence of homologues of this element in the chromosomes of various Bacillus subtilis strains

A cryptic plasmid from a soil strain of Bacillus subtilis was found to contain a sequence having features of IS element. Homologous sequences were also found in the chromosome of this strain and in the chromosomes of some other B. subtilis strains.     

TatC is a specificity determinant for protein secretion via the twin-arginine translocation pathway

The recent discovery of a ubiquitous translocation pathway, specifically required for proteins with a twin-arginine motif in their signal peptide, has focused interest on its membrane-bound components, one of which is known as TatC. Unlike most organisms of which the genome has been sequenced completely, the Gram-positive eubacterium Bacillus subtilis contains two tatC-like genes denoted tatCd and tatCy. The corresponding TatCd and TatCy proteins have the potential to be involved in the translocation of 27 proteins with putative twin-arginine signal peptides of which approximately 6-14 are likely to be secreted into the growth medium. Using a proteomic approach, we show that PhoD of B. subtilis, a phosphodiesterase belonging to a novel protein family of which all known members are synthesized with typical twin-arginine signal peptides, is secreted via the twin-arginine translocation pathway. Strikingly, TatCd is of major importance for the secretion of PhoD, whereas TatCy is not required for this process. Thus, TatC appears to be a specificity determinant for protein secretion via the Tat pathway. Based on our observations, we hypothesize that the TatC-determined pathway specificity is based on specific interactions between TatC-like proteins and other pathway components, such as TatA, of which three paralogues are present in B. subtilis.     

Avoiding False Positive Antigen Detection by Flow Cytometry on Blood Cell Derived Microparticles: The Importance of an Appropriate Negative Control

BackgroundMicroparticles (MPs), also called microvesicles (MVs) are plasma membrane-derived fragments with sizes ranging from 0.1 to 1μm. Characterization of these MPs is often performed by flow cytometry but there is no consensus on the appropriate negative control to use that can lead to false positive results.ResultsAnnexin-V positive events within a gate of 300-900nm were detected and defined as MPs. Our results confirmed that the characteristic antigens CD41/CD61 were found on platelet-derived-MPs validating our technique. However, for MPs derived from other cell types, we were unable to detect any antigen, although they were clearly expressed on the MP-producing cells in the contrary of several data published in the literature. Using the latex bead technique, we confirmed detection of CD41,61. However, the apparent expression of other antigens (already deemed positive in several studies) was determined to be false positive, indicated by negative controls (same labeling was used on MPs from different origins).ConclusionWe observed that mother cell antigens were not always detected on corresponding MPs by direct flow cytometry or latex bead cytometry. Our data highlighted that false positive results could be generated due to antibody aspecificity and that phenotypic characterization of MPs is a difficult field requiring the use of several negative controls.     

Cellular lysis in Bacillus subtilis; the affect of multiple extracellular protease deficiencies

Cellular lysis properties of strains of Bacillus subtilis deficient in the synthesis of extracellular proteases was investigated. In all cases, extracellular protease deficiency was found to increase the extent of cellular lysis of batch cultured strains following the transition to stationary phase, the time at which extracellular degradative enzymes are secreted in large quantities. The data indicates that the major extracellular proteases, NprE and AprE, are primarily responsible for the control of this autolytic activity in B. subtilis and has implications for the use of extracellular protease-deficient strains as hosts for the production of heterologous proteins.     

Chitosan and hot water treatments reduce postharvest green mould in ‘Murcott’ tangor

This study aims to evaluate the efficacy of hot water and chitosan treatments to control green mould caused by Penicillium digitatum in 'Murcott' tangor. P. digitatum conidial germination and mycelial growth were evaluated in assays in vitro to verify whether chitosan (0.5, 1 and 2%) or hot water (45, 50, and 55°C, for 30 s, 1, 2, and 5 min) acts directly on fungus development. In vivo assays consisted of inoculating the fruit with P. digitatum (10⁵ conidia ml⁻¹) 4 hr before the chitosan and hot water treatments. Subsequently, green mould incidence and severity were evaluated in fruits stored at 25°C/80% RH for 4 days. Also, treatments combining chitosan and hot water were investigated for controlling green mould and the effect on postharvest quality of fruit stored at 5°C/90% RH. The results showed that P. digitatum conidia germination and mycelial growth were significantly reduced by the hot water treatments especially at 50°C/5 min and 55°C/2 or 5 min in the first case and 50 and 55°C/5 min in the second. These two treatments, when applied alone, 1 min dipping in 2% chitosan or hot water at 50°C/5 min, significantly reduced green mould development in fruit kept at 25°C/80% RH or refrigerated. However, the hot water dip combined with chitosan did not improve green mould control on ‘Murcott’ tangor at room temperature or under refrigeration. Besides, chitosan and hot water did not impair fruit quality. Thus, chitosan and hot water could be an alternative to synthetic fungicides to control green mould in citrus while also contributing to a decrease in the postharvest losses of ‘Murcott’ tangor.     

An assessment of the use of drug and non-drug interventions in the treatment of Ichthyophthirius multifiliis Fouquet, 1876, a protozoan parasite of freshwater fish

Infection by the ciliate protozoan Ichthyophthirius multifiliis Fouquet, 1876 causes significant economic losses in freshwater aquaculture worldwide. Following the ban on the use of malachite green for treating food fish, there has been extensive research aimed at identifying suitable replacements. In this paper we critically assess drug and non-drug interventions, which have been tested for use or have been employed against this parasite and evaluate possibilities for their application in farm systems. Current treatments include the administration of formaldehyde, sodium chloride (salt), copper sulphate and potassium permanganate. However, purportedly more environmentally friendly drugs such as humic acid, potassium ferrate (VI), bronopol and the peracetic acid-based products have recently been tested and represent promising alternatives. Further investigation, is required to optimize the treatments and to establish precise protocols in order to minimize the quantity of drug employed whilst ensuring the most efficacious performance. At the same time, there needs to be a greater emphasis placed on the non-drug aspects of management strategies, including the use of non-chemical interventions focusing on the removal of free-swimming stages and tomocysts of I. multifiliis from farm culture systems. Use of such strategies provides the hope of more environmentally friendly alternatives for the control of I. multifiliis infections.