Genetic elements associated with antimicrobial resistance in enteropathogenic <Emphasis Type="Italic">Escherichia coli</Emphasis> (EPEC) from Brazil

Background  

We recently observed an association of resistance with a certain enteropathogenic Escherichia coli (EPEC) serotypes and identified a conjugative plasmid, similar to plasmid pED208, that was conserved among archival O111:H2/NM and O119:H2 strains of diverse geographical origin. In this study, we sought to determine the prevalence and distribution of this plasmid among a collection of EPEC isolates from Brazil, as well as to study the susceptibilities of these isolates to antimicrobial agents.     

DA Forum: <Emphasis Type="Italic">De eso que llaman Anthropología Mexicana</Emphasis>

De eso que llaman Antropología Mexicana (of that known as Mexican Anthropology) first appeared in 1969 in Mexico at critical national and international conjunctures in an effort to both take the pulse of Mexican anthropology and move the discipline in new directions. As the text approaches its 50th anniversary, Dialectical Anthropology organized a forum, soliciting six critical evaluations of the book’s historical role.     

Immunohistochemical localization of TLR2 and CD14 in gingival tissue of healthy individuals and patients with chronic periodontitis

We used immunohistochemistry to quantify and compare the expression of Toll-like receptor 2 (TLR2) and cluster of differentiation 14 (CD14) in gingival tissues of both healthy individuals and patients with chronic periodontitis. We also correlated the expression of TLR2 and CD14 with the histological grades of chronic periodontitis. We examined 30 gingival specimens from chronic periodontitis patients and 10 from healthy individuals. Tissues from both groups were immunostained with antibodies against TLR2 and CD14. TLR2 and CD14 were expressed by endothelial cells, fibroblasts, lymphocytes and plasma cells. The immunohistochemical expression of TLR2 and CD14 was significantly greater in inflammatory cells of the chronic periodontitis group than in healthy individuals. Expression of these molecules was greater in the inflammatory cells of connective tissue adjacent to pocket epithelium in both groups. The expression of TLR2 and CD14 was greatest in the periodontitis group that was classified as severe grade, followed by moderate and mild grades, which suggests a role of TLR2 and CD14 in the pathogenesis of chronic periodontitis. The positive correlation of TLR2 and CD14 expression levels with the severity grades of chronic periodontitis suggests that they are correlated also with disease severity; therefore, they may be useful for predicting disease progression. Our findings are consistent with the possibility that CD14 acts as a co-receptor for TLR2.     

Variable Substrate Preference among Phospholipase D Toxins from Sicariid Spiders

Venoms of the sicariid spiders contain phospholipase D enzyme toxins that can cause severe dermonecrosis and even death in humans. These enzymes convert sphingolipid and lysolipid substrates to cyclic phosphates by activating a hydroxyl nucleophile present in both classes of lipid. The most medically relevant substrates are thought to be sphingomyelin and/or lysophosphatidylcholine. To better understand the substrate preference of these toxins, we used ³¹P NMR to compare the activity of three related but phylogenetically diverse sicariid toxins against a diverse panel of sphingolipid and lysolipid substrates. Two of the three showed significantly faster turnover of sphingolipids over lysolipids, and all three showed a strong preference for positively charged (choline and/or ethanolamine) over neutral (glycerol and serine) headgroups. Strikingly, however, the enzymes vary widely in their preference for choline, the headgroup of both sphingomyelin and lysophosphatidylcholine, versus ethanolamine. An enzyme from Sicarius terrosus showed a strong preference for ethanolamine over choline, whereas two paralogous enzymes from Loxosceles arizonica either preferred choline or showed no significant preference. Intrigued by the novel substrate preference of the Sicarius enzyme, we solved its crystal structure at 2.1 Å resolution. The evolution of variable substrate specificity may help explain the reduced dermonecrotic potential of some natural toxin variants, because mammalian sphingolipids use primarily choline as a positively charged headgroup; it may also be relevant for sicariid predatory behavior, because ethanolamine-containing sphingolipids are common in insect prey.     

Calorimetric determination of the partition coefficient for chlorpromazine hydrochloride in aqueous suspensions of dimyristoylphosphatidylcholine vesicles

Lipid suspensions containing from 0.1 to 0.2% by weight dimyristoylphosphatidylcholine were mixed in a flow calorimeter with equal volumes of chlorpromazine hydrochloride at concentrations ranging from 6×10^?5 to 1.2×10^?4 M. The vesicle bilayer volume fraction of the suspension was determined by density measurements. Linear relationships were obtained between heat production per ml suspension and chlorpromazine concentration at each level of lipid volume. Using phase partitioning as a model, the values of the partition coefficient and the enthalpy change were found to be K_c′=1300 and ΔH=?30 kJ·mol^?1 at 25°C.Heat outputs at slightly higher concentrations of chlorpromazine increased less than linearly because of repulsive forces between neighboring chlorpromazine cations absorbed in the bilayer phase. At still higher concentrations the slope increased again but partition coefficients became variable, which indicated a change in the nature of the interactions.In batch calorimeter titrations at higher concentrations a sharp increase in heat output was observed at the critical micelle concentrations of chloropromazine (4 mM) and a final levelling off at 6 mM. Enthalpies of dilution of chlorpromazine obtained in separate experiments were large and endothermic, but no break in the curve could be detected at the critical micelle concentrations.     

Tissue distribution of a plasmid DNA encoding Hsp65 gene is dependent on the dose administered through intramuscular delivery

In order to assess a new strategy of DNA vaccine for a more complete understanding of its action in immune response, it is important to determine the in vivo biodistribution fate and antigen expression. In previous studies, our group focused on the prophylactic and therapeutic use of a plasmid DNA encoding the Mycobacterium leprae 65-kDa heat shock protein (Hsp65) and achieved an efficient immune response induction as well as protection against virulent M. tuberculosis challenge. In the present study, we examined in vivo tissue distribution of naked DNA-Hsp65 vaccine, the Hsp65 message, genome integration and methylation status of plasmid DNA. The DNA-Hsp65 was detectable in several tissue types, indicating that DNA-Hsp65 disseminates widely throughout the body. The biodistribution was dose-dependent. In contrast, RT-PCR detected the Hsp65 message for at least 15 days in muscle or liver tissue from immunized mice. We also analyzed the methylation status and integration of the injected plasmid DNA into the host cellular genome. The bacterial methylation pattern persisted for at least 6 months, indicating that the plasmid DNA-Hsp65 does not replicate in mammalian tissue, and Southern blot analysis showed that plasmid DNA was not integrated. These results have important implications for the use of DNA-Hsp65 vaccine in a clinical setting and open new perspectives for DNA vaccines and new considerations about the inoculation site and delivery system.     

Foraging behavior of the communal spider,Philoponella republicana (Araneae: Uloboridae)

The communal orb-weaving spider, Philoponella republicana,was observed in the subtropical moist forest of Southeast Peru. These spiders live in colonies of conspecifics whose individual orbs are connected by silk. The wrapping of a prey prior to feeding is a large component of the prey capture process because P. republicanahas no venom with which to kill an insect. Wrapping time was the only aspect of prey capture that was strongly correlated with the size of the insect captured. Occasionally we observed several individuals working together to wrap a prey item. These joint efforts were more frequent on prey larger than the capturing spider. Although group captures accounted for only 5.5% of captures, they represented 14.7% of the biomass obtained. A comparison of the relationship between wrapping time and prey size for solitary and group efforts suggested that, by working together, the spiders reduced their total handling time. In most cases only one spider fed on the captured prey.     

Hemolysis induced by Prymnesium parvum toxin calorimetric studies

The relationship between binding of the hemolytic toxin (prymnesin) to bovine erythrocytes and the amount of heat liberated was examined as a function of pH using a flow microcalorimeter and ³H-labelled toxin isolated from the euryhaline alga Prymnesium parvum. A high degree of correlation (correlation coefficient = 0.986) was found between the amount of heat generated and the quantity of toxin that was allowed to interact with the erythrocytes. No significant binding of toxin was observed at pH 7 but it increased linearly as the pH was reduced to 5.5. Maximum heat and binding occured at a pH range 4.5–5.5. The same pattern was followed in terms of the amount of heat liberated and the hemolytic activity of the toxin. The differences in the maximum binding and heat production as a function of pH was independent of the average red cell volume which remained constant at pH 5.5 and 6.2 (102.4 and 102.6 μm³, respectively).