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Riparins are alkamides naturally found in the fruits of Aniba riparia (Nees) Mez, but currently synthetic molecules as Riparin E (Rip-E) can be obtained. Potential biological of Rip-E as schistosomicidal agent against Schistosoma mansoni worms, as well as against Staphylococcus aureus strains has already been described. However, the mechanism of action related to antimicrobial activity of Rip-E against bacterial or fungi species has not yet been reported. This study had as objective to evaluate the Rip-E antimicrobial activity against Gram-positive and Gram-negative bacteria, as well as against yeast species of clinical importance. Minimal inhibitory concentrations of the compound against bacterial and yeast strains were determined by microdilution method. To verify if a possible lethal effect caused by Rip-E were related to plasma membrane damage, microbial cells treated with Rip-E were stained with 7-aminoactinomycin D (7-AAD) and analyzed by flow cytometry. Rip-E showed a bactericide effect against Gram-positive species S. aureus and S. epidermidis, as well as, against Gram-negative species Escherichia coli and Salmonella enterica Typhimurium, but was inactive against Pseudomonas aeruginosa. Moreover, Rip-E showed activity against fungi species Candida albicans and C. tropicalis. S. aureus, E. coli and C. albicans cells treated with Rip-E were marked with 7-aminoactinomycin D (7-AAD) indicating that Rip-E can cause plasma membrane damage, acting as a potential microbicide agent for prevention or treatment of infectious diseases.

The aim of this study was to investigate intrinsic antimicrobial activity of three monoterpenes nerol, dimethyl octanol and estragole, against bacteria and yeast strains, as well as, investigate if these compounds are able to inhibit the NorA efflux pump related to fluoroquinolone resistance in Staphylococcus aureus. Minimal inhibitory concentrations (MICs) of the monoterpenes against Staphylococcus aureus, Escherichia coli and Candida albicans strains were determined by micro-dilution assay. MICs of the norfloxacin against a S. aureus strain overexpressing the NorA protein were determined in the absence or in the presence of the monoterpenes at subinhibitory concentrations, aiming to verify the ability of this compounds act as efflux pump inhibitors. The monoterpenes were inactive against S. aureus however the nerol was active against E. coli and C. albicans. The addition of the compounds to growth media at sub-inhibitory concentrations enhanced the activity of norfloxacin against S. aureus SA1199-B. This result shows that bioactives tested, especially the nerol, are able to inhibit NorA efflux pump indicating a potential use as adjuvants of norfloxacin for therapy of infections caused by multi-drug resistant S. aureus strains.

Objective. This in vitro study investigated the effect of Fluoride (F), Chlorhexidine (CHX), and their association on the viability of Streptococcus mutans using a biofilm model. Materials and Methods. Biofilms were anaerobically grown on glass slides that were vertically suspended in 24-well plates for 5 days. After 48 h of initial growth, biofilms were treated for the next 72 h, 2x/day with CHX at 0.12% and 2%, F as NaF at 0.08% and 0.4% and their association. Results. CHX treatment decreased the bacteria counts either alone or in association with both F concentrations, when compared with control group and the F treatments alone (p

Summary The gene Ldccys1 enconding a cysteine proteinase of 30 kDa from Leishmania (Leishmania) chagasi , as well as the recombinant cysteine proteinase rLdccys1, obtained by cloning and expression of the Ldccys1 gene in the pHIS vector, were used to evaluate their ability to induce immune protective responses in BALB/c mice against L. (L.) chagasi infection. Mice were immunized subcutaneously with rLdccys1 plus Bacille Calmette Guerin (BCG) or Propionibacterium acnes as adjuvants or intramuscularly with a plasmid carrying the Ldccys1 gene ( Ldccys1 /pcDNA3) and CpG ODN as the adjuvant, followed by a booster with rLdccys1 plus CpG ODN. Two weeks after immunization the animals were challenged with 1 × 10 7 amastigotes of L. (L.) chagasi . Both immunization protocols induced significant protection against L. (L.) chagasi infection as shown by a very low parasite load in the spleen of immunized mice compared to the non-immunized controls. However, DNA immunization was 10-fold more protective than immunization with the recombinant protein. Whereas rLdccys1 induced a significant secretion of IFN-γ and nitric oxide (NO), animals immunized with the Ldccys1 gene increased the production of IgG2a antibodies, IFN-γ and NO. These results indicated that protection triggered by the two immunization protocols was correlated to a predominant Th1 response.

Objective: This study aimed to test the calcium hydroxide associated with vehicles Aloe vera, chlorhexidine digluconate (2%) and saline, in inhibiting bacterial growth on Mueller-Hinton agar plates. Methods: The species Staphiloccocus aureus, Streptococcus pyogenes, Escherichia coli and Enterecoccus faecalis were isolated and inoculated in 3 mL of BHI (Brain Heart Infusion). Holes (5 mm diameter) were made in the plates and filled with the test materials. After incubation, readings were taken with a hand lens and a caliper with 0.1 mm accuracy to determine the diameter of the inhibition zone, after 24 and 48 hours. Each experiment was repeated six times, and the average values were obtained. Results: Chlorhexidine digluconate without calcium hydroxide resulted in better inhibition of bacterial growth, followed by the hydroxide pastes evaluated. E. coli strains were the most resistant to the tested compounds, followed by S. pyogenes and S. aureus. There was no statistically significant interaction between the variables. Conclusion: Chlorhexidine digluconate (2%) alone showed the best antimicrobial effectiveness. Aloe vera is a promising vehicle for the calcium hydroxide but more studies should be conducted on herbal medicines in dentistry.

A 500 bp fragment encoding an isoform of cysteine proteinase from Leishmania (Leishmania) amazonensis was subcloned and expressed in the pHis vector, resulting in a recombinant protein of 24 kDa, rLacys24. In Western blots of L. (L.) amazonensis extracts, antibodies directed to rLacys24 recognized a cysteine proteinase isoform of 30 kDa. Analysis by fluorescence-activated cell sorter showed a significantly higher expression of CD8(+) lymphocytes in animals immunized with rLacys24 plus CFA, whereas a low expression of CD4(+) lymphocytes was observed in these animals. The cytotoxicity of lymphocytes isolated from mice immunized with rLacys24 plus CFA on L. (L.) amazonensis-infected macrophages was significantly higher than that observed in the presence of lymphocytes from control animals. Immunization of BALB/c mice with rLacys24 plus CFA resulted in a low but significant decrease of foot lesions after challenge with L. (L.) amazonensis compared to those exhibited by control mice.

A recombinant protein, rLdccys1, produced by expression of the gene encoding a 30 kDa cysteine proteinase from Leishmania (Leishmania) chagasi, was used to detect specific antibodies in serum by enzyme-linked immunosorbent assays and to test for reactivity in delayed-type hypersensitivity (DTH) responses of dogs from an endemic region of visceral leishmaniasis (VL), Teresina, Piaui State, Brazil. Amastigote or promastigote extracts were also assayed for comparison. The sensitivity for detection of specific antibodies to L. (L.) chagasi using rLdccys1, lysates from L. (L.) chagasi promastigotes and amastigotes was 96%, 68%, and 69%, respectively. No cross-reactivity between rLdccys1 and Chagas disease was observed, and little reactivity was found with sera from dogs with babesiosis and ehrlichiosis. Among 106 sera from symptomatic dogs and 22 from non-infected controls, no false negatives and only two false positive sera were found for rLdccys1. In contrast, amastigote lysates yielded 11 false positives and 13 false negatives, whereas the corresponding numbers for promastigote lysates were 17 and 16. DTH responses were determined after intradermal injection of rLdccys1 or amastigote extract and the induration area was measured at 24, 48 and 72 h after injection. All asymptomatic dogs showed a positive intradermal response to rLdccys1 (>10 mm) which peaked at 48 h, whereas no significant reactivity to the recombinant antigen was found in the symptomatic group. Histological analysis of the intradermal induration showed a predominance of necrotic and hemorrhagic areas in sections from asymptomatic dogs injected with L. (L.) chagasi amastigote extract, whereas a typical granulomatous reaction mediated by mononuclear cells was observed in sections from asymptomatic animals injected with rLdccys1. Grouping data from ELISA and DTH assays with rLdccys1 and L. (L.) chagasi amastigote extracts showed that humoral and cellular responses were inversely correlated during the development of canine VL. Overall, these findings indicate that L. (L.) chagasi recombinant cysteine proteinase is potentially useful for diagnosis of canine VL, as well as for the discrimination of clinical and subclinical forms of the disease.

Background A recombinant cysteine proteinase from Leishmania (Leishmania) infantum chagasi (rLdccys1) was previously shown to induce protective immune responses against murine and canine visceral leishmaniasis. These findings encouraged us to use rLdccys1 in the immunotherapy of naturally infected dogs from Teresina, Piauí, a region of high incidence of visceral leishmaniasis in Brazil. Methodology/Principal Findings Thirty naturally infected mongrel dogs displaying clinical signs of visceral leishmaniasis were randomly divided in three groups: one group received three doses of rLdccys1 in combination with the adjuvant Propionibacterium acnes at one month interval between each dose; a second group received three doses of P. acnes alone; a third group received saline. The main findings were: 1) dogs that received rLdccys1 with P. acnes did not display increase of the following clinical signs: weight loss, alopecia, onychogryphosis, cachexia, anorexia, apathy, skin lesions, hyperkeratosis, ocular secretion, and enlarged lymph nodes; they also exhibited a significant reduction in the spleen parasite load in comparison to the control dogs; 2) rLdccys1-treated dogs exhibited a significant delayed type cutaneous hypersensitivity elicited by the recombinant antigen, as well as high IgG2 serum titers and low IgG1 serum titers; sera from rLdccys1-treated dogs also contained high IFN-γ and low IL-10 concentrations; 3) control dogs exhibited all of the clinical signs of visceral leishmaniasis and had low serum IgG2 and IFN-γ levels and high concentrations of IgG1 and IL-10; 4) all of the dogs treated with rLdccys1 were alive 12 months after treatment, whereas dogs which received either saline or P. acnes alone died within 3 to 7 months. Conclusions/Significance These findings illustrate the potential use of rLdccys1 as an additional tool for the immunotherapy of canine visceral leishmaniasis and support further studies designed to improve the efficacy of this recombinant antigen for the treatment of this neglected disease., Author Summary Visceral leishmaniasis (VL) is an important public health problem and dogs are the main domestic reservoirs of zoonotic VL which has resulted in an annual incidence of 40,100–75,500 new human cases. Because canine VL chemotherapy is limited by the low efficacy of drugs currently used for human VL treatment, immunotherapy may provide a viable alternative. We used a recombinant cysteine proteinase from L. (L.) infantum chagasi, rLdccys1, in combination with the adjuvant P. acnes for the treatment of naturally infected mongrel dogs from Teresina, Pauí a state in Brazil that has a high incidence of VL. Dogs treated with rLdccys1 showed a significant delayed type hypersensitivity reaction against the recombinant antigen and displayed high serum concentrations of IgG2 and IFN-γ and low concentrations of IgG1 and IL-10. Immunotherapy with rLdccys1 resulted in no increase of the clinical signs of canine VL and an extensive reduction of spleen parasite load. Furthermore, all of the dogs treated with rLdccys1 survived for at least 12 months after treatment, whereas those that received either saline or P. acnes alone died within 3 to 7 months. These findings support the potential of rLdccys1 immunotherapy as an additional option for the treatment of canine VL.