Streptomyces ansochromogenes Tur-10 produces a substance with antifungal bioactivity.

The increased incidence of fungal infections and the development of drug resistance have led to the search for microorganisms capable of producing bioactive metabolites with antifungal activity. Among these microorganisms, Streptomyces spp are distinguished mainly owing to their potential to secrete bioactive molecules. The aim of this study was to evaluate the production of secondary metabolites by Streptomyces sp TUR-10 against 12 fungal clinical isolates (yeast and filamentous fungi). In the preliminary screening, Streptomyces sp TUR-10 showed activity against 75% of the clinical isolates, and was selected for fermentation. In this assay, we tested three different media (MPE, M1, and ISP-4) for 96 h at pH 7.0 and 30°C for the production of bioactive metabolites. Increased production of bioactive compounds was observed when using the MPE medium for 48 h, with good activity against Candida pelliculosa. The minimum inhibitory concentration showed significant antifungal activity values ranging from 15.6 to 250 µg/mL. The actinobacterium was characterized by 16S rRNA analysis and the pattern suggested that the isolate studied belonged to the species Streptomyces ansochromogenes. The biotechnological potential of this strain was also demonstrated by the detection of the nrps and pks genes. These results indicate the production of secondary metabolites of biotechnological interest by actinobacteria from the rhizosphere, suggesting great potential for further research.

Activity and in vivo tracking of Amphotericin B loaded PLGA nanoparticles.

The development of biocompatible polymeric nanoparticles has become an important strategy for optimizing the therapeutic efficacy of many classical drugs, as it may expand their activities, reduce their toxicity, increase their bioactivity and improve biodistribution. In this study, nanoparticles of Amphotericin B entrapped within poly (lactic-co-glycolic) acid and incorporated with dimercaptosuccinic acid (NANO-D-AMB) as a target molecule were evaluated for their physic-chemical characteristics, pharmacokinetics, biocompatibility and antifungal activity. We found high plasma concentrations of Amphotericin B upon treatment with NANO-D-AMB and a high uptake of nanoparticles in the lungs, liver and spleen. NANO-D-AMB exhibited antifungal efficacy against Paracoccidioides brasiliensis and induced much lower cytotoxicity levels compared to D-AMB formulation in vivo and in vitro. Together, these results confirm that NANO-D-AMB improves Amphotericin B delivery and suggest this delivery system as a potential alternative to the use of Amphotericin B sodium deoxycholate.

Nanobiotechnological approaches to delivery of DNA vaccine against fungal infection.

Vaccines play an essential role in keeping humans healthy. Innovative approaches to their use include the utilization of plasmid DNA encoding sequences to express foreign antigens. DNAhsp65 from Mycobacterium leprae is suitable for this purpose due to its ability to elicit a powerful immune response. Controlled release systems represent a promising approach to delivering vaccines. In this work, we used liposomes or PLGA systems to deliver DNAhsp65 to treat the pulmonary fungal infection Paracoccidioidomycosis. Both formulations modulated a protective immune response and reduced the pulmonary fungal burden even in the groups receiving less than four times the amount of the DNAhps65 entrapped within the nanoparticles. Although both systems had the same effective therapeutic results, the advantage of the liposome formulation was that it was administered intranasally, which may be more easily accepted by patients. These systems are a great alternative to be considered as adjuvant vaccine therapy for systemic mycosis.

Immunogenicity and antigenic properties of Pf332-C231, a fragment of a non-repeat region of the Plasmodium falciparum antigen Pf332.

Antigen Pf332, a megadalton protein has been shown to be associated with the membrane of infected erythrocytes. Detailed functional studies on the antigen have remained hampered by the cross-reactive nature of antibodies generated to Pf332. Pf332-C231, identified in the C-terminal region of Pf332 was cloned and antibodies against the C231 fragment were shown to react with intact Pf332 antigen by both immunofluorescence and immunoblotting analyses. Antibodies to C231 inhibited in vitro Plasmodium falciparum growth efficiently. In addition, human sera from malaria-exposed individuals reacted with recombinant C231. We show that Pf332-C231 represents a functional domain and is expected to facilitate further studies on Pf332 as a potential target for protective immune responses and the function of the antigen.

Antibody responses to a C-terminal fragment of the Plasmodium falciparum blood-stage antigen Pf332 in Senegalese individuals naturally primed to the parasite.

Previous studies have shown that antibodies from humans exposed continuously to malaria recognize the Plasmodium falciparum asexual blood-stage antigen Pf332. Here we analysed the antibody responses to a C-terminal fragment of Pf332, designated C231, in individuals from Senegal, by measuring the serum levels of immunoglobulin M (IgM), IgG class and subclass and IgE antibodies. IgG antibody reactivity with crude P. falciparum antigen was detected in all the donors, while many of the children lacked or had low levels of such antibodies against C231. The antibody levels increased significantly with age for both crude P. falciparum antigen and C231, and in the older age groups most of the donors displayed antibodies to C231. This was also true for IgM, IgE and IgG subclass reactivity against C231. Moreover, the ratio of IgG1/IgG2 was considerably lower for C231 than for crude P. falciparum antigen, and in age groups 10-14 and 15-19 years the levels of IgG2 against C231 even exceeded that of IgG1. The IgG2/IgG3 ratios suggest that C231 gives similar levels of IgG2 and IgG3, except for children aged 4-9 years, where IgG3 was higher. Raw IgM, IgG class and subclass and IgE antibody levels to C231 tended to be higher in those who did not experience a malaria attack, but following linear multivariate analysis the trends were not significant.

Comparative immunization study using RNA and DNA constructs encoding a part of the Plasmodium falciparum antigen Pf332.

Development of nucleic acid-based vaccines against parasitic diseases shows great promise, although certain concerns about safety aspects of conventional DNA vaccines have been raised. This study presents a comparison of antibody responses induced in mice by DNA and RNA-based immunization with vectors encoding a part of the P. falciparum antigen Pf332. Two types of plasmids were used, one conventional DNA plasmid containing a cytomegalovirus promoter and one suicidal DNA plasmid encoding the Semliki Forest virus (SFV) replicase. RNA, encoding the SFV replicase and the relevant antigen, was delivered either as naked RNA or packaged in SFV suicide particles. In general, the antibody responses induced by the DNA plasmids were low and peaking after three injections, the conventional plasmid giving the highest responses. Also the RNA delivered in SFV particles consistently induced antibody responses, although comparatively low. Analyses of the ratio of immunoglobulin (Ig)G1/IgG2a subclasses in the responses indicated that all plasmids resulted in a bias for a Th2-type of response, while the SFV-particles elicited a Th1 type of response. Importantly, all these immunogens induced an immunological memory, which could be efficiently activated by a booster injection with the corresponding protein, with unchanged patterns of IgG subclasses.