DltX of Bacillus thuringiensis Is Essential for D-Alanylation of Teichoic Acids and Resistance to Antimicrobial Response in Insects.

The dlt operon of Gram-positive bacteria is required for the incorporation of D-alanine esters into cell wall-associated teichoic acids (TAs). Addition of D-alanine to TAs reduces the negative charge of the cell envelope thereby preventing cationic antimicrobial peptides (CAMPs) from reaching their target of action on the bacterial surface. In most gram-positive bacteria, this operon consists of five genes dltXABCD but the involvement of the first ORF (dltX) encoding a small protein of unknown function, has never been investigated. The aim of this study was to establish whether this protein is involved in the D-alanylation process in Bacillus thuringiensis. We, therefore constructed an in frame deletion mutant of dltX, without affecting the expression of the other genes of the operon. The growth characteristics of the dltX mutant and those of the wild type strain were similar under standard in vitro conditions. However, disruption of dltX drastically impaired the resistance of B. thuringiensis to CAMPs and significantly attenuated its virulence in two insect species. Moreover, high-performance liquid chromatography studies showed that the dltX mutant was devoid of D-alanine, and electrophoretic mobility measurements indicated that the cells carried a higher negative surface charge. Scanning electron microscopy experiments showed morphological alterations of these mutant bacteria, suggesting that depletion of D-alanine from TAs affects cell wall structure. Our findings suggest that DltX is essential for the incorporation of D-alanyl esters into TAs. Therefore, DltX plays a direct role in the resistance to CAMPs, thus contributing to the survival of B. thuringiensis in insects. To our knowledge, this work is the first report examining the involvement of dltX in the D-alanylation of TAs.

Influence of the hand squeeze and mask distensibility on tidal volume measurements during neonatal mask ventilation.

BACKGROUND: During mask ventilation, the mask volume can vary as it is pressurized or when it is squeezed. The change in volume of the mask may affect tidal volumes delivered and difference in inspired (Vti) and expired tidal volumes (Vte). OBJECTIVES: To investigate whether hand squeeze and distensibility of the mask during ventilation influences tidal volume measurements. METHODS: For both experiments, we ventilated a leak-free mask ventilation model using pressures of 25/5 cm H2O through a t-piece. Vti and Vte were measured. (A) Two consultants performed mask ventilation with (1) consistent hand squeeze, (2) release during inflation and squeeze during expiration, (3) squeeze during inflation, release during expiration, and (4) gentle squeeze. RESULTS: (B) Thirty caregivers performed mask ventilation. Experiment A: Vti was different during consistent hold (1) 8.1 ml (0.4) and loose grip (4) 8.2 ml (0.3), compared to squeezing during inflation (2) 18.9 ml (1.9), or expiration (3) 6.4 ml (3.5). Variance in difference between Vti and Vte occurred only when the mask was squeezed during inflation (-47.4% (101.5)). Experiment B: volumes measured were consistent (intraindividual CV 3-5%, interindividual CV 9-10%). When comparing gas flow rate of 6-10 l/min, volumes increased by approximately 8%, differences in Vti and Vte were small with both flow settings (-0.9% (-3.9-1.4) and -0.6% (-3.3-1.8); n.s.). CONCLUSION: Variation in mask hold during mask ventilation can influence volume measurement, but this hardly occurs when testing caregivers.

Pathogenic potential of Bacillus cereus strains as revealed by phenotypic analysis.

The Bacillus cereus pathogenic spectrum ranges from strains used as probiotics to human-lethal strains. However, prediction of the pathogenic potential of a strain remains difficult. Here, we show that food poisoning and clinical strains can be differentiated from harmless strains on the basis of host colonization phenotypes.