Detection of pathogenic Leptospira species associated with phyllostomid bats (Mammalia: Chiroptera) from Veracruz, Mexico.

The genus Leptospira encompass 22 species of spirochaetes, with ten pathogenic species that have been recorded in more than 160 mammals worldwide. In the last two decades, the numbers of records of these agents associated with bats have increased exponentially, particularly in America. Although order Chiroptera represents the second most diverse order of mammals in Mexico, and leptospirosis represents a human and veterinary problem in the country, few studies have been conducted to identify potential wildlife reservoirs. The aim of this study was to detect the presence and diversity of Leptospira sp. in communities of bats in an endemic state of leptospirosis in Mexico. During January to September 2016, 81 bats of ten species from three localities of Veracruz, Mexico, were collected with mist nets. Kidney samples were obtained from all specimens. For the detection of Leptospira sp., we amplified several genes using specific primers. Amplicons of the expected size were submitted to sequencing, and sequences recovered were compared with those of reference deposited in GenBank using the BLAST tool. To identify their phylogenetic position, we realized a reconstruction using maximum-likelihood (ML) method. Twenty-five samples from three bat species (Artibeus lituratus, Choeroniscus godmani and Desmodus rotundus) showed the presence of Leptospira DNA. Sequences recovered were close to Leptospira noguchii, Leptospira weilii and Leptospira interrogans. Our results include the first record of Leptospira in bats from Mexico and exhibit a high diversity of these pathogens circulating in the state. Due to the finding of a large number of positive wild animals, it is necessary to implement a surveillance system in populations of the positive bats as well as in related species, in order to understand their role as carriers of this bacterial genus.

Potential effects on Rhipicephalus microplus tick larvae fed on calves immunized with a Subolesin peptide predicted by epitope analysis.

The cattle tick Rhipicephalus microplus economically impacts cattle production in tropical and subtropical regions of the world. The recombinant R. microplus Subolesin antigen has been shown to protect cattle against tick infestations. In this study, we searched in silico protective epitopes in the subolesin gene and a recombinant peptide containing the predicted epitopes was expressed, and evaluated against a tick challenge. Two different epitope types, linear B-cells and conformational discontinuous epitopes, were predicted using bioinformatics strategies, in order to synthesize the recombinant peptide. Two Eightmonth-old calves European crossbred were immunized with two subcutaneous doses of the subolesin recombinant peptide, emulsified with Montanide ISA 50 V as an adjuvant, at 30-day intervals. The tick challenge was conducted with 5 000 R. microplus larvae/animal. ELISA test was used to evaluate the IgG immune response elicited against the peptide. After tick challenge, reduction in the number of engorged females (79%), and reduction in egg hatching (30%) was observed in tick population fed on immunized calves with regards to an untreated control group. The results showed a potential higher effect on tick reproduction for the recombinant peptide compared to other studies reported with Subolesin protein, demonstrating that the use of bioinformatics strategies to predict protective epitopes may lead to improve the immune response elicited against tick recombinant peptides and therefore to prevent cattle tick infestations.

Actin cytoskeleton role in the structural response of epithelial (MDCK) cells to low extracellular Ca2+.

Kinetic and stereometric assessment of the mechanical responses of epithelial cells to variations in the concentration of extracellular Ca2+ was carried out in vivo at the single cell level. Continuous monitoring of individual MDCK cells in subconfluent cultures attested to an intense, immediately relaxable, and cytochalasin D-sensitive contraction, equivalent to that seen in confluent monolayers following depletion of external Ca2+ (<0.1 mM). Increasingly greater and less readily reversible contractions were performed upon repeated stimulation with short-term cycles of alternating normal (30 min) and low Ca2+ (30 min) media. Constriction of a narrow horizontal girdle corresponding in position to the major ring-like bundle of actin filaments eventually develops into a deep lateral furrow in intensely contracted cells. Substantial membrane infolding in the contracted state is indicated also by stereometric estimates of apparent bounding surface area. Irrespective of the contracted or relaxed cell condition, rhodamine-phalloidin labeling showed a marginal position of the ring-like bundle of microfilaments and other components of the actin cytoskeleton. These results suggest, contrary to prevalent views, that the actin-myosin system stays associated to the cortex and retains contractile capability in epithelial cells deprived of external Ca2+. Hence, the mechanical responses to variations of Ca2+ may be an overstrained expression of a physiological mechanism.

Contraction of epithelial (MDCK) cells in response to low extracellular calcium is dependent on extracellular sodium.

Like other cells of epithelial origin, MDCK cells respond with a reversible structural transformation to a diminution in the concentration of extracellular Ca2+. Upon deprivation of Ca2+ in the medium the cells undergo an active contraction mediated by the actin-myosin cytoskeleton, in parallel to detachment of the intercellular contacts and appearance of free spaces in the epithelium or monolayer (Castillo et al., 1998). We now present results indicating that the decrease of external Ca2+ plays an indirect and non-specific role in activating contraction, probably by allowing an influx of Na+. The omission of external Ca2+ had no effect when it was replaced by Mg2+, Ba2+ or Hg2+, and the addition of any of these divalent cations induced relaxation of cells previously contracted by exposure to low Ca2+. A null or weak response was observed also when Ca2+ was lowered in a solution where Na+ was replaced by choline or in the presence of amiloride (30 microM), which reduces the permeability of the plasma membrane to Na+. Restitution of Na+ or removal of amiloride were followed by contraction in the same cultures. Li+ proved an able substitute of Na+ as requisite for cell contraction in response to Ca2+ depletion. Monensin (0.1 mM) -an ionophore selective for Na+- and to a lesser extent ouabain (0.1 mM) -an inhibitor of Na+ extrusion across the plasma membrane- , both stimulated contraction in the presence of the normal level of external Ca2+. Decreasing by half the normal concentration of external K+ facilitated cell contraction, but typical responses were observed when K+ was increased to 40 mM by partial substitution for Na+. These findings attest that cell contraction in response to low Ca2+ is likely due to an increase in the permeability of the plasma membrane to Na+, though not to membrane depolarization as such. Evidences from other motile systems suggest that Na+ influx might in turn cause an elevation of cytoplasmic Ca2+, which activates the actin-myosin cytoskeleton.

Myosin II-actin interaction in MDCK cells: role in cell shape changes in response to Ca2+ variations.

Cultured MDCK cell monolayers respond to a low level of extracellular calcium ([Ca2+]e < or = 5 microM) with a loss of transepithelial electrical resistance and transport function, and changes in position of a circumferential ring of actin filaments tethered to the plasma membrane at the zonula adhaerens. Keeping this cytoskeletal structure in place seems necessary to preserve the architecture of the tight junctions and therefore their sealing capacity. All three effects are reversible upon restituting normal [Ca2+]e. Recent work provided evidence of actin-myosin interactions at the filament ring, thus suggesting a contraction process involved in the alteration of the actin cytoskeleton. We now report that active contraction does occur and causes an extensive morphological transformation of MDCK cells. A marked increase in cell height simultaneous with a decrease in width and area of contact to the substratum was seen within 10 min of removal of [Ca2+]e; recovery began immediately after replacing calcium, although it took longer for completion. Conventional and confocal epifluorescence studies showed actin colocalized with myosin II at various planes of resting or contracted cells, in particular at the ring level. Electron-micrographs revealed the circumferential actin ring associated with the plasma membrane in a waist-like constriction where Ca2+ was removed from the cultures. Contraction, as well as relaxation, in response to [Ca2+]e variations were inhibited by cytochalasin-D (an actin-filament disrupting drug), by okadaic acid( an inhibitor of myosin light-chain dephosphorylation), and by 2,3-butanedione monoxime (a blocker of myosin II ATPase activity). Similarly, no response was observed in cells previously depleted of metabolic energy by 2,4-dinitrophenol and 2-deoxy-D-glucose preincubation. The actin-myosin mediated reversible structural transformation of MDCK cells in response to [Ca2+]3 poses new questions for the interpretation of in vitro experiments, as well as for the understanding of epithelial function.

[Differential changes in serum levels of N-acetylneuramic acid and L-fucose as manifestations of physiological and physiopathological adaptation in man. Pregnancy as an experimental model. The tumoral, vascular and hepatic diseases as possible pathological models]. / Cambios diferenciales en los niveles séricos del acido N-acetilneuramínico y de la L-fucosa como manifestaciones de adaptación fisiológica y fisiopatológica en el humano. La gestación como modelo experimental. Los padecimientos tumorales, vasculares y hepáticos como posibles modelos patológicos

In this paper we report on the modifications that normal pregnancy introduces on the blood maternal serum concentrations of both N-acetyl maternal serum concentrations of both N-acetyl-neuraminic acid which is elevated above normal level and L-fucose which on the contrary is diminished. The experiences were performed with women in the last trimester of normal gestations. The probable physiochemical meaning and biological significance of those such changes are discussed in connection both to the maternal physiological adaptation during pregnancy and to the possible metabolical changes in intracellular regulation that might take place in such a situation. Similarly it is emphasized the probability that numerous pathological conditions including malignancy vascular diseases and hepatic disorders which course with overt changes in plasma glucoproteins might be amenable to present notorious modifications in N-acetylneuraminic acid and L-fucose plasmatic levels.