Extracellular enzymatic activities of Bipolaris sorokiniana isolates

Several enzymatic activities were investigated in six isolates of the fungus Bipolaris sorokiniana, originating from different areas of Brazil. Among the glycosidases studied, beta-glucosidase, beta-N-acetylglucosaminidase, beta-xylosidase, cellobiohydrolase, and chitobiohydrolase were the major activities. In some isolates, beta-glucuronidase, beta-galactosidase, and alpha-mannosidase activities were also present. Polysaccharide-hydrolyzing enzymes, such as pectin lyase and carboxymethyl cellulase were detected in significant amounts, and their activities were variable among the different isolates. Other enzymes, namely phosphatases, proteinases and phenol oxidase, were also examined, showing variable amounts depending on the isolate. The pH dependence of all enzymes tested was investigated. Endoproteinase, carboxymethyl cellulase, and phenoloxidase had maximum activity in the pH range of 6-8, whilst all other enzymes showed maximum activity at pH 4-6.     

Mode of action of antimicrobial peptide P45 on Listeria monocytogenes

The mode of action of an antimicrobial peptide produced by Bacillus sp. P45 isolated from the intestine of the Amazonian basin fish Piaractus mesopotamicus was investigated. The antimicrobial peptide was purified from culture supernatants by precipitation with ammonium sulfate and gel filtration chromatography. The peptide has an EC(50) of 300 AU (activity units) ml(-1) and kills all viable cells of Listeria monocytogenes with a concentration of 800 AU ml(-1). A decrease in OD was observed when L. monocytogenes cultures were treated with the peptide, suggesting that cells were lysed. Transmission electron microscopy showed damage of the cell envelope and loss of protoplasmic material. The peptide P45 was bactericidal and bacteriolytic to L. monocytogenes. There is evidence that the mode of action is interfering at cell membranes and the cell wall. The knowledge of the mode of action of antimicrobial peptides is an essential step to consider their utilization in food or clinic.     

Characterization of human sperm N-acetylglucosaminidase

N-acetylglucosaminidase (NAG) is particularly active in mammalian spermatozoa and appears to be involved in fertilization. Although it is assumed that this enzyme is acrosomal, previous results from our laboratory suggest the presence of NAG at the sperm plasma membrane level. The present study attempted to analyse the subcellular distribution of this enzyme in human spermatozoa. Sperm were incubated under different conditions and NAG activity measured in the soluble extracts and cell pellets using a specific fluorometric substrate. A significant proportion of NAG activity was released when sperm were incubated in culture medium, suggesting a weak association with the plasma membrane. This location was confirmed by western blot analysis of plasma membrane fractions and immunofluorescence on non-permeabilized sperm, which showed a positive signal mainly on the acrosomal domain. The distribution of NAG activity between plasma membrane and acrosome was analysed after cell disruption by freezing and thawing. Triton X-100 stimulated sperm and epididymal NAG activity but not the enzyme obtained from other sources. In addition, biotinylated human recombinant NAG was able to bind to human sperm. Finally, after sperm incubation under capacitating conditions, NAG total activity increased and the sperm enzyme lost its ability to be stimulated by Triton X-100. The possible connection of these results with sperm maturation, capacitation and NAG participation in primary binding to the zona pellucida, was discussed.     

Andrology: A new predictive test for in-vitro fertilization based on the induction of sperm acrosome reaction by N-acetylglucosamine-neoglycoprotein

Neoglycoproteins with N-acetylglucosamine residues (BSA-GIcNAc)induced specifically the acrosome reaction (AR) in human spermatozoa.Our objective was to investigate the relationship between thisphenomenon and the in-vitro fertilization (IVF) rate. Spermsuspensions from IVF protocols were incubated with BSA-GlcNAc(t), using calcium ionophore (i) or medium alone (c) as positiveor negative controls. When the normalized AR percentage ratio(STIM) (%ARt-%ARc): (%ARi-%ARc) was compared with fertilizationrate in 31 couples from our IVF programme, a positive correlationwas found (r = 0.46, P < 0.01). The fertilization rate inpatients with STIM 0.2 was higher than in non-responders (STIM< 0.2); 72 ± 7% compared with 5 ± 3%. The overallpredictive value of this test for adequate fertilization rate(>30%) was 87%, sensitivity 91% and specificity 78%. Falsepositives were 9% and false negatives 22%. For successful fertilizationrates (>60%), the results were: overall predictive value,84%; sensitivity 100%; specificity 64%. False positives were23% and no false negatives were found. The results indicatedthat the induction of AR in human spermatozoa by GlcNAc-neoglycoproteinscould be used to predict their fertilizing ability in vitro.     

Prevalence of Gender Dysphoria in Southern Brazil: A Retrospective Study

Archives of Sexual Behavior - To ensure that public health services provide comprehensive and inclusive health care to the general population, it is important for countries to estimate how many of...     

Bacteriocin-like substance from Bacillus amyloliquefaciens shows remarkable inhibition of Acanthamoeba polyphaga

The effectiveness of a bacteriocin-like substance (BLS) produced by Bacillus amyloliquefaciens was tested against Acanthamoeba polyphaga strains, and its cytotoxic potential on Vero cells was investigated. Amebicidal activity of the purified BLS was tested by plate bioassays with concentrations ranging from 12.5 to 6,400 AU mL⁻¹. Damage to A. pholyphaga cells was monitored using an inverted microscope and counted in a Fuchs–Rosenthal chamber after 24, 48, and 72 h. According to the results obtained, the BLS showed remarkable amebicidal and amebostatic effect on A. polyphaga and showed no cytotoxicity on the Vero cells. These results may have great relevance in the development of new acanthamoebicidal compounds.     

Indigenous traditional medicine: in vitro anti-giardial activity of plants used in the treatment of diarrhea

The ethnopharmacology for treatment of parasitic infections facilitates and directs the search for new chemical entities. In this direction, this study evaluated the cytotoxicity in vitro, against trophozoites of Giardia lamblia, of aqueous extracts of leaves Achyrocline satureioides (Lam.) DC., barks of Eugenia uniflora L., aerial parts of Foeniculum vulgare Miller, and barks of Psidium guajava L. These plants are traditionally used for the treatment of diarrhea by the indigenous population Mbyá-Guarani, located at the Lomba do Pinheiro, Porto Alegre, Rio Grande do Sul, Brazil. The in vitro susceptibility qualitative analysis of G. lamblia to aqueous extracts was performed in serial dilutions from 2.5 to 0.02 mg/mL. Results revealed the minimal inhibitory concentrations: 0.313 mg/mL for A. satureioides and E. uniflora, 0.02 mg/mL for P. guajava, and F. vulgare did not present any cytotoxic effect. Quantitative assays of viable trophozoites, showed that A. satureioides presented the highest cytotoxic effect (93.5%), followed by P. guajava (82.2%), and E. uniflora (67.3%). Indigenous Guarani use mainly A. satureioides for the treatment of diarrhea, revealing the conformity with results obtained in vitro. Bioguided assays are necessary to identify the compounds responsible for the activity of the aqueous extract of A. satureioides.