Bacterial biofilms with emphasis on coagulase-negative staphylococci

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Cytokine production in experimental paracoccidioidomycosis of murine selected lineages for acute inflammatory response (air)

Paracoccidioidomycosis is a systemic human mycosis caused by Paracoccidioides brasiliensis (P. brasiliensis), an imperfect dimorphic fungus whose conidia are its infective form. Mice genetically selected for maximum (AIRmax) and minimum (AIRmin) acute inflammatory response were used as experimental paracoccidioidomycosis models. The animals were intraperitoneally inoculated with P. brasiliensis (strain 18) and killed 6, 12 and 24 hours or 3, 7 and 14 days after infection. In these periods, fragments from their spleen, liver and lung were removed for evaluation of the infection level by fungal cells, assessment of macrophagic activity by peritoneal and splenic macrophages - through the determination of nitric oxide (NO) concentrations and production of pro- and anti-inflammatory cytokines of lung and spleen homogenate supernatants. In the present study, it was observed that AIRmax lineages presented greater control of the infectious process than the AIRmin ones. Regarding NO production, AIRmax animals produced more metabolites in late periods, what may help control the infectious process. Concerning cytokine production, it was observed that the production of INF-gamma, TNF-alpha, IL-1, IL-6, IL-8 and IL-12 were increased in AIRmax lineages in most analyzed organs and periods, thus contributing to the greater resistance exhibited by such lineages against infection, except for IL-4 and IL-10 that showed decreased production in AIRmax lineage, reproducing its suppressive biological effect. From these results, it was observed that the AIRmax lineage was more effective in controlling the infectious process, with an important involvement of the analyzed cytokines. These findings are probably related to the genetically selected factors involved in the acute inflammatory response.

ROLE of TLR2 and TLR4 on human neutrophil functions against Paracoccidioides brasiliensis

In paracoccidioidomycosis, a systemic mycosis caused by the fungus Paracoccidioides brasiliensis (Pb), studies have focused on the role of neutrophils that are involved in the primary response to the fungus. Neutrophil functions are regulated by pro- and anti-inflammatory cytokines. Molecular mechanisms involved in this process are not fully understood, but there are strong evidences about the involvement of toll-like receptors (TLRs). We aimed at evaluating TLR2 and TLR4 expression on human neutrophils activated by GM-CSF, IL-15, TNF-alpha or IFNgamma and challenged with a virulent strain of P. brasiliensis (Pb18). Moreover, we asked if these receptors have a role on fungicidal activity, H(2)O(2) and IL-6, IL-8, TNFalpha and IL-10 production, by activating and challenging cells. All cytokines increased TLR2 and TLR4 expression. Pb18 also increased TLR2 expression, inducing an additional cytokine effect. on the contrary, it inhibited TLR4 expression. All cytokines increased neutrophil fungicidal activity and H(2)O(2) production; however, this process was not associated with TLR2 or TLR4. Neutrophil activation by GMCSF and TNF-alpha resulted in a significant increase of IL-8 production, while IL-15 and IFN-alpha have no effect. Pb18 also augmented IL-8 expression, inducing an additional effect to that of cytokines. None of the cytokines activated neutrophils by releasing IL-10. This cytokine was only detected after Pb18 challenge. Interestingly, IL-8 and IL-10 production involved TLR2 and mainly TLR4 modulation. The present results suggest that Pb18 interaction with neutrophils through TLR2 and TLR4 with consequent IL-8 and IL-10 production may be considered a pathogenic mechanism in paracoccidioidomycosis.

Biological properties of medicinal plants: a review of their antimicrobial activity

Plants have been used for thousands of years to flavor and conserve food, to treat health disorders and to prevent diseases including epidemics. The knowledge of their healing properties has been transmitted over the centuries within and among human communities. Active compounds produced during secondary vegetal metabolism are usually responsible for the biological properties of some plant species used throughout the globe for various purposes, including treatment of infectious diseases. Currently, data on the antimicrobial activity of numerous plants, so far considered empirical, have been scientifically confirmed, concomitantly with the increasing number of reports on pathogenic microorganisms resistant to antimicrobials. Products derived from plants may potentially control microbial growth in diverse situations and in the specific case of disease treatment, numerous studies have aimed to describe the chemical composition of these plant antimicrobials and the mechanisms involved in microbial growth inhibition, either separately or associated with conventional antimicrobials. Thus, in the present work, medicinal plants with emphasis on their antimicrobial properties are reviewed.

Anti-coagulase-negative Staphylococcus activity of ethanolic extracts of propolis from two Brazilian regions and synergism with antimicrobial drugs by the E-test method

Propolis is a natural resinous substance collected by bees from vegetal sources and its therapeutic properties have been investigated. In this work, we evaluated the inhibitory activity of ethanolic extracts of propolis (EEP) from the Southeast and South of Brazil on coagulase-negative Staphylococcus (CNS) growth as well as the EEP in vitro synergism with antimicrobial drugs by using the diffusion method (E-test). The EEP chemical characteristics (dry weight, pH, flavonoid and phenolic compounds) were determined. Seven drugs were tested, and synergism was observed between three drugs and Southeast EEP, six drugs and South EEP, and one drug and ethanol control. Ethanolic extracts of propolis from the South of Brazil presented the greatest flavonoid content and synergism rate, while EEP from the Southeast presented the greatest anti-CNS activity and phenolic compound content. Results showed the correlation among anti-CNS activity, synergism rate and chemical characteristics of propolis.

Evalution of methods for the detection of MRSA in patients of Botucatu Medical School Hospital, São Paulo, Brazil

Oxacillin is the main drug of choice for the treatment of S. aureus infections. However, S. aureus resistance to oxacillin has become a major problem in the recent decades. The study aimed assess the rates of oxacillin resistance in S. aureus samples obtained at the Botucatu Medical School Hospital, UNESP, and to compare phenotypic techniques for the detection of MRSA against the gold standard method (mecA gene detection) in these samples. A total of 102 samples, previously isolated between 2002 and 2006, and kept at the Culture Collection of the Department of Microbiology and Immunology, in the Botucatu Biosciences Institute, UNESP, were included. Oxacillin resistance was assessed by oxacillin and cefoxitin disk diffusion and agar dilution tests, screening tests using Mueller-Hinton agar with 6 mu g/mL of oxacillin and 4% NaCl, E-test, and mecA gene detection. of the samples analyzed, 46 (45.1%) were mecA-positive. Oxacillin disk sensitivity and specificity were 86.9% and 91.1%, respectively. Cefoxitin disk sensitivity and specificity were respectively 91.3% and 91.1%. The screening test with the cefoxitin disk showed almost the same level of sensitivity (91.3%) and specificity (91.1%). With E-test strips, sensitivity was higher (97.8%) and specificity was comparable to that found with the other methods (91.1%). Ninety-three percent of the samples produced beta-lactamase and five of them were mecA-negative. There was a gradual increase in the number of oxacillin-resistant S. aureus samples between 2002 and 2004. However, from 2004 to 2006, the number of resistant samples dropped from 55% of MRSA in 2004, to 45% in 2005 and 34.6% in 2006. The data obtained reveal that, among phenotypic methods, the E-test yielded the best results, with higher sensitivity levels when compared to the other methods. The decreased resistance rate observed over the most recent years may be explained by the rational use of antimicrobial agents associated with good practices in the control of hospital infection, or may be related to the diminished use of oxacillin as a treatment option.

Impact of malnutrition on immunity and infection

Malnutrition may be a consequence of energy deficit or micronutrient deficiency. It is considered the most relevant risk factor for illness and death, particularly in developing countries. In this review we described the magnitude of this problem, as well as its direct effect on the immune system and how it results in higher susceptibility to infections. A special emphasis was given to experimental models used to investigate the relationship between undernutrition and immunity. Malnutrition is obviously a challenge that must be addressed to health authorities and the scientific community.

Candidemia in a Brazilian tertiary hospital: microbiological and clinical features over a six-year period

Yeasts are becoming a common cause of nosocomial fungal infections in immunocompromised patients. Such infections often develop into sepsis with high mortality rates. The aim of this study was to evaluate some of the numerous factors associated with the development of candidemia. Medical records were retrospectively analyzed of 98 Candida spp. patients. Results showed that the most prevalent risk factors for developing candidemia were: antibiotics and antifungal agents (93.9% and 79.6%, respectively); the use of central venous catheter (93.9%); mechanical ventilation (73.5%); and parenteral nutrition (60.2%). The main species of Candida found were: C. parapsilosis (37.76%), C. albicans (33.67%); and others (28.57%). C. glabrata showed the highest mortality rate (75%), followed by C. tropicalis (57.1%) and C. albicans (54.5%). The elevated mortality rate found in this study indicates that preventive measures against candidemia must be emphasized in hospitals.

Identification and antimicrobial susceptibility of Staphylococcus from home-treated peritoneal dialysis patients

Staphylococcus aureus is the main agent of infections during peritoneal dialysis (PD). The presence of S. aureus in the nasal cavity has been extensively studied and suggested as a risk factor of dialysis-related infections, whereas coagulase-negative Staphylococcus (CNS) species are frequently considered part of the normal human microbiota. The aim of this study was to identify Staphylococcus in the nasal cavity, pericatheter skin and peritoneal effluent from PD patients, as well as to evaluate the antimicrobial activity evolution in vitro. Thirty-two chronic PD patients were observed during 12 months and had nasal and pericatheter skin samples collected for culture. When peritonitis was detected, samples were also collected from the peritoneal effluent for culture. The activity of several antimicrobial drugs (penicillin G, oxacillin, cephalothin, ofloxacin, netilmicin and vancomycin) against different Staphylococcus species was measured by using the agar drug diffusion assay (Kirby-Bauer method). Staphylococcus was separated into S. aureus, S. epidermidis and other CNS species in order to determine the in vitro resistance level. S. epidermidis resistance to oxacillin progressively increased during the study period (p < 0.05). Resistance to ofloxacin was inexpressive, whereas resistance to netilmicin and vancomycin was not detected. of the oxacillin-resistant species (n = 74), 83% were S. epidermidis, 13% other CNS and 4% S. aureus (p < 0.05). Regarding multidrug resistant strains (n = 45), 82% were S. epidermidis, 13% other CNS, and 5% S. aureus (p < 0.05). This study shows the relevance of resistance to oxacillin and CNS multi-drug resistance, particularly concerning S. epidermidis, in PD patients.

Kinetics of venom and antivenom serum levels, clinical evaluation and therapeutic effectiveness in dogs inoculated with Crotalus durissus terrificus venom

This work evaluated the clinical and therapeutic aspects as well as serum levels of venom and antivenom IgG by enzyme-linked immunosorbent assay (ELISA) in experimental envenomation of dogs with Crotalus durissus terrificus venom. Twenty-eight mixed breed adult dogs were divided into four groups of seven animals each, Group I: only venom; Group II, venom + 50 ml of anti-bothropic-crotalic serum (50mg) + fluid therapy; Group III, venom + 50 ml of anti-bothropic-crotalic serum + fluid therapy + urine alkalination; Group IV, 50 ml of anti-bothropic-crotalic serum. The lyophilized venom of Crotalus durissus terrificus was reconstituted in saline solution and subcutaneously inoculated at the dose of 1mg/kg body weight. The dogs presented clinical signs of local pain, weakness, mandibular ptosis, mydriasis, emesis and salivation. The venom levels detected by ELISA ranged from 0 to 90ng/ml, according to the severity of the clinical signs. Serum antivenom ranged from 0 to 3ug/ml and was detected for up to 138h after treatment. ELISA results showed the effectiveness of the serum therapy for the venom neutralization.

Morphological, morphometric, and molecular characterization of Hepatozoon spp. (Apicomplexa, Hepatozoidae) from naturally infected Caudisona durissa terrifica (Serpentes, Viperidae)

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Description of gamontogonic and sporogonic stages of Hepatozoon spp. (Apicomplexa, Hepatozoidae) from Caudisoma durissa terrifica (Serpentes, Viperidae)

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Morphological and molecular characterization of Strongyloides ophidiae (Nematoda, Strongyloididae)

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Seasonal variation of Hepatozoon spp. (Apicomplexa, Hepatozoidae) parasitemia from Boa constrictor amarali (Serpentes, Boidae) and Hydrodynastes gigas (Serpentes, Colubridae)

The aim of this study was to evaluate the parasitemia variation of three Hepatozoon species in Brazilian snakes. This study was conducted between 2001 and 2003 and included Hepatozoon terzii from Boa constrictor amarali, and Hepatozoon migonei and Hepatozoon cyclagrasi from Hydrodynastes gigas. It was observed that the parasitemia tended to decrease in all three Hepatozoon species but the parasites were not eliminated. This data suggest that Hepatozoon infection may be similar to Toxoplasma gondii infection, in that it persists throughout host life.

Natural history of the South American water snake Helicops leopardinus (Colubridae : Hydropsini) in the pantanal, Central Brazil

The South American water snake Helicops leopardinus is very abundant in the Pantanal wetlands of Central Brazil. We studied the biology of this species based on specimens collected by local workers in the Paraguay River. Females attained greater body lengths and had larger heads than males, but the latter had longer tails. Helicops leopardinus fed on a wide variety of fishes (70% of the total prey items) and frogs, with fishes of the order Gymnotiformes (33.34%) and frogs of the family Hylidae (23.34%) representing the most common prey items. There was relatively lows diet overlap between the sexes and between juveniles and adults. Mean litter size was 6.6 and ranged from 4-11. The reproductive cycle was seasonal, with birth occurring late in the wet season (simultaneous with the flooding of the Paraguay River), facilitating dispersal of young snakes by floating vegetation. Floating vegetation may be a critical habitat for H. leopardinus, and management plans for their maintenance need to be developed.