Subunit rotation in a single FoF1-ATP synthase in a living bacterium monitored by FRET

FoF1-ATP synthase is the ubiquitous membrane-bound enzyme in mitochondria, chloroplasts and bacteria which provides the 'chemical energy currency' adenosine triphosphate (ATP) for cellular processes. In Escherichia coli ATP synthesis is driven by a proton motive force (PMF) comprising a proton concentration difference {\Delta}pH plus an electric potential {\Delta}{\Psi} across the lipid membrane. Single-molecule in vitro experiments have confirmed that proton-driven subunit rotation within FoF1-ATP synthase is associated with ATP synthesis. Based on intramolecular distance measurements by single-molecule fluorescence resonance energy transfer (FRET) the kinetics of subunit rotation and the step sizes of the different rotor parts have been unraveled. However, these experiments were accomplished in the presence of a PMF consisting of a maximum {\Delta}pH ~ 4 and an unknown {\Delta}{\Psi}. In contrast, in living bacteria the maximum {\Delta}pH across the plasma membrane is likely 0.75, and {\Delta}{\Psi} has been measured between -80 and -140 mV. Thus the problem of in vivo catalytic turnover rates, or the in vivo rotational speed in single FoF1-ATP synthases, respectively, has to be solved. In addition, the absolute number of functional enzymes in a single bacterium required to maintain the high ATP levels has to be determined. We report our progress of measuring subunit rotation in single FoF1-ATP synthases in vitro and in vivo, which was enabled by a new labeling approach for single-molecule FRET measurements.Comment: 9 pages, 6 figure

Streptococcus uberis strains isolated from the bovine mammary gland evade immune recognition by mammary epithelial cells, but not of macrophages

Streptococcus uberis is frequently isolated from the mammary gland of dairy cattle. Infection with some strains can induce mild subclinical inflammation whilst others induce severe inflammation and clinical mastitis. We compared here the inflammatory response of primary cultures of bovine mammary epithelial cells (pbMEC) towards S. uberis strains collected from clinical or subclinical cases (seven strains each) of mastitis with the strong response elicited by Escherichia coli. Neither heat inactivated nor live S. uberis induced the expression of 10 key immune genes (including TNF, IL1B, IL6). The widely used virulent strain 0140J and the avirulent strain, EF20 elicited similar responses; as did mutants defective in capsule (hasA) or biofilm formation (sub0538 and sub0539). Streptococcus uberis failed to activate NF-κB in pbMEC or TLR2 in HEK293 cells, indicating that S. uberis particles did not induce any TLR-signaling in MEC. However, preparations of lipoteichoic acid (LTA) from two strains strongly induced immune gene expression and activated NF-κB in pbMEC, without the involvement of TLR2. The immune-stimulatory LTA must be arranged in the intact S. uberis such that it is unrecognizable by the relevant pathogen receptors of the MEC. The absence of immune recognition is specific for MEC, since the same S. uberis preparations strongly induced immune gene expression and NF-κB activity in the murine macrophage model cell RAW264.7. Hence, the sluggish immune response of MEC and not of professional immune cells to this pathogen may aid establishment of the often encountered belated and subclinical phenotype of S. uberis mastitis

Prevalence and Correlates of Food Insecurity among Palestinian Refugees in Lebanon: Data from a Household Survey.

Lebanon hosts the highest per capita refugee concentration worldwide. The Palestinian presence in Lebanon dates from 1948 and they remain a marginalized population. No information on their food security status has been reported previously. A survey of a representative sample of Palestinian refugee households in Lebanon (n = 2501) was conducted using a stratified two stage cluster sampling approach. We measured food insecurity using a modified USDA household food security module, locally validated. We collected data on household demographic, socioeconomic, health, housing, coping strategies and household intake of food groups and analysed these by food security status. About 41% (CI: 39-43) of households reported being food insecure and 20% (CI: 18-22) severely food insecure. Poor households were more likely to be severely food insecure (OR 1.41 (1.06-1.86)) while higher education of the head of household was significantly associated with protection against severe food insecurity (OR 0.66 (0.52-0.84)). Additionally, higher food expenditure and possession of food-related assets were significantly associated with food security (OR 0.93 (0.89-0.97) and OR 0.74 (0.59-0.92), respectively). After adjusting for confounders, households where at least one member suffered from an acute illness remained significantly more likely to be severely food insecure (OR 1.31(1.02-1.66)), as were households whose proxy respondent reported poor mental health (OR 2.64 (2.07-3.38)) and poor self-reported health (OR 1.62 (1.22-2.13). Severely food insecure households were more likely to eat cheaper foods when compared to non-severely food insecure households (p<0.001) and were more likely to rely on gifts (p<0.001) or welfare (p<0.001). They were also more likely to have exhausted all coping strategies, indicating significantly more frequently that they could not do anything (p = 0.0102). Food insecurity is a significant problem among Palestinian refugees in Lebanon and is likely to be exacerbated at this time when the Syrian crisis amplifies the problem

Characteristics of Palestinian refugees by levels of household (HH) food security.

<p>Estimates are weighted percentages or mean values and 95% confidence intervals.</p><p>^a Food-related assets represent the sum of fridge, freezer, oven, and microwave.</p><p>^b All health characteristics are self-reported physician diagnosed illnesses within the last six months.</p><p>Characteristics of Palestinian refugees by levels of household (HH) food security.</p

Mean dietary intake of food categories by Palestinian refugee households and by levels of food security.

<p>Estimates are weighted means and 95% confidence intervals. P-values are obtained using one-way analysis. The data on food category consumption was log transformed using the equation ln(x+1) and back transformed to achieve a normal distribution.</p><p>^a Significantly different from moderately and severely food insecure households (p<0.05).</p><p>^b Significantly different from food secure and severely food insecure households (p<0.05).</p><p>^c Significantly different from food secure and moderately food insecure households (p<0.05).</p><p>Mean dietary intake of food categories by Palestinian refugee households and by levels of food security.</p

Coping mechanisms employed by Palestinian refugee households who reported that the food that they had did not last, and they did not have money to buy more.

<p>Coping mechanisms employed by Palestinian refugee households who reported that the food that they had did not last, and they did not have money to buy more.</p

Logistic regression model for predictors of severe food insecurity among Palestinian refugee households in Lebanon.

<p>^a Estimates are weighted odds ratios generated from univariate logistic regression in relation to severe HH food insecurity.</p><p>^b Estimates from multivariate logistic regression model using stepwise selection (n = 2357) are weighted OR in relation to severe HH food insecurity. Model statistics include goodness-of-fit test: F (9, 1389) = 0.38, P = 0.9429.</p><p>Logistic regression model for predictors of severe food insecurity among Palestinian refugee households in Lebanon.</p