Luiz Antonio Custodio

Protective effect of Artin M from extract of Artocarpus integrifolia seeds by Th1 and Th17 immune response on the course of infection by Candida albicans.

The immunoregulatory effect of Artin M and jacalin from extract of Artocarpus integrifolia seeds (jack extract) against infection with Candida albicans was investigated. Swiss mice received jack extract containing 500 μg protein/ml PBS intraperitoneally (i.p.) or PBS alone and after 72 h were infected i.p. with C. albicans CR15 (10(7)) and sacrificed after 30 min, 2, 6, 24, and 72 h. ELISA analysis revealed that in jack extract-treated mice IFN-γ was predominantly produced versus IL-10 in control mice. These results suggest that jack extract induced a protective immune response, since C. albicans clearance was complete at 72 h postinfection. Jack extract presents two lectins (Artin M and jacalin) with distinct biological properties. Artin M was able to induce IL-12 production by macrophages. Also, Artin M in different concentrations, associated with jacalin or in jack extract induced both IFN-γ and IL-17 production. As a consequence, phagocytic and candidacidal activity increased significantly. Alanine aminotransferase activity (ALT) was used as parameter for damage of the liver. The activity of ALT correlated with inoculum size that increased significantly in control group, however, mice pretreated with jack extract 3 days before infection presented normal ALT. Mice pretreated with jack extract that received a lethal inoculum of Candida presented 90% survival versus 20% among controls or mice pretreated with jacalin. Thus, the results suggest that Artin M by itself, associated with jacalin or present in jack extract is able to induce protective Th1 and Th17 immune responses against Candida albicans infection.

Increased tumour necrosis factor-α production, higher mannose receptor activity and ability to kill Candida by concanavalin-A-activated macrophages

In a previous study, our group verified that mice pretreated with concanavalin-A (Con-A) produced more tumour necrosis factor (TNF)-alpha and presented greater Candida clearance from the peritoneal cavity, liver and spleen, which yielded a higher survival rate than control animals. In this work, the hypothesis that macrophages were of crucial importance in overcoming the infection was tested. Thus, peritoneal macrophages from mice pretreated for 3 days with Con-A or phosphate-buffered saline (PBS) were coincubated with CR1, CR15 and 577 isolates of Candida albicans for 0.5, 1 and 2 h. The ability of Con-activated macrophages to produce TNF-alpha, ingest via mannose receptors and kill all the isolates was significantly greater compared with PBS-treated macrophages, and activated macrophages exhibited a lower incidence of apoptosis, verified by binding to annexin V-fluorescein isothiocyanate. The transition of yeast cells to filamentous forms during coincubation for 2 h with control macrophages was about 73-80%, whereas in the presence of Con-A-activated macrophages, it was 35-40%. Our results suggest that a greater clearance of C. albicans infection through treatment with Con-A is probably due to the activation of macrophages, which produce more TNF-alpha, express more mannose receptors and are better endowed to kill ingested C. albicans.

Artin M enhances TNF-α production and phagocytosis of Candida albicans mediated by dectin-1 and mannose receptors.

The activities of dectin-1 and mannose receptors on phagocytosis of Candida albicans and the production of TNF-α by macrophages from mice pretreated for 3 days with extract of Artocarpus intergrifolia seeds (jack extract), Artin M or jacalin were studied. Macrophages from these mice were coincubated with C. albicans CR15 (yeast), in the presence of mannose (50mM) plus mannan (100 μg) or laminarin (1mg). Phagocytosis was significantly enhanced to 52% in macrophages from mice pretreated intraperitoneally for 3 days with jack extract (500 μg/250 μl PBS). Reduction in phagocytosis from 52% to 34% (P<0.05) occurred in the presence of mannose receptor inhibitors and from 52% to 16% (P<0.01) in the presence of dectin-1 inhibitor laminarin, whereas only 20% of control macrophages phagocytosed blastoconidia. Similar results were verified for pretreatment of mice with Artin M (2.5 μg/250 μl PBS), but not for jacalin (25 μg/250 μl PBS). Macrophages from mice pretreated 3 days previously with jack extract or Artin M and then coincubated for 2h with C. albicans presented a significant increase in TNF-α production, correlating with significantly less transition of yeast to filamentous forms compared to pretreatment with jacalin. These results suggest that Artin M, but not jacalin present in jack extract significantly increased TNF-α production and the activity of mannose and dectin-1 receptors.

Detection of parasite-specific IgG and IgA in paired serum and saliva samples for diagnosis of human strongyloidiasis in northern Paraná state, Brazil

Human strongyloidiasis is an infection caused by the helminth Strongyloides stercoralis that can be fatal, especially in immunosuppressed patients. The aim of this study is to evaluate parasite-specific IgG and IgA levels using S. venezuelensis third-stage (L3) infective larvae alkaline extract as a heterologous antigen by ELISA in paired serum and saliva samples with improved sensitivity and specificity. Individuals from northern Paraná state, Brazil were divided into three groups: 30 patients copropositive for S. stercoralis (Group I); 30 clinically healthy individuals (Group II); and 30 patients copropositive for other parasites (Group III). The area under ROC curve (AUC), an overall index of diagnostic accuracy, and Kappa index were calculated. Data were analyzed using analysis of variance (ANOVA) followed by a Kruskal-Wallis test. Probability (p) values of <0.05 were regarded as significant. In Group I, IgG was detected in 96.7% serum and in 6.7% saliva samples. IgG was not detected in Group II. In Group III, cross-reactivity was observed for serum IgG in 26.7% and in 6.7% for saliva samples. In Group I, IgA was detected in 76.7% serum and 56.7% saliva samples. In Group II, 3.3% were positive for IgA in serum, whereas IgA was not detected in any saliva samples. Group III showed 6.7% serum and 26.7% saliva-positive samples. The sensitivity values for detection of IgG and IgA in serum samples were 96.7% and 76.7%, respectively. In saliva samples, the sensitivity values for detection of IgG and IgA were 6.7% and 56.7%, respectively. The specificity value was 100% for the detection of IgG in serum and for detection of IgG and IgA in saliva, and 96.7% for detection of IgA in serum samples. The proper choice of immunological diagnosis to supplement parasitological methods is essential to estimate the true prevalence of the parasite, and will permit analysis of population immune response profiles, particularly in northern Paraná state, where there are no previous reports.

Brazilian propolis promotes immunomodulation on human cells from American Tegumentar Leishmaniasis patients and healthy donors infected with L. braziliensis

American Tegumentar Leishmaniasis (ATL) is an infectious disease caused by Leishmania parasites with ineffective treatment. The properties of propolis have been studied in different experimental studies, however, few works have investigated the effects of propolis on human-derived peripheral blood mononuclear cells (PBMC) in leishmaniasis models. Thus, we investigate the immunomodulatory effects of propolis treatment on PBMC from ATL patients and on PBMC from healthy donors infected with Leishmania braziliensis. Our data demonstrate that propolis pretreatment shows immunomodulatory effects on both healthy donors and ATL patients adherent cells, increasing IL-4 and IL-17 and decreasing IL-10, in either the presence or absence of the L. braziliensis infection, demonstrating that propolis contributes with the decrease of the inflammation and could also contribute with parasite control.

Concanavalin‐A induces IL‐17 production during the course of Candida albicans infection

In a previous study, our group verified that 100% of mice survived to a lethal dose of Candida albicans following pretreatment with concanavalin-A (Con-A) for 3 days. This work proposed to investigate whether treatment could mediate an adaptative immune response involving T(H) 17 cells. A significant increase in IL-17 levels at 6 h postinfection was observed and was maintained up to 18 h in the Con-A group, whereas in control mice, a reduction in this cytokine was verified. In addition, T(H) 17 cells develop in the presence of TGF-β, IL-1 β, and IL-6 that were increased significantly 2 h postinfection in Con-A-treated mice. Macrophages were involved in the process, engulfing greater numbers of yeast cells, and were activated through TNF-α and interferon-γ produced at significant levels at 2 h postinfection. A significant increase in IL-12 levels was also observed at 2 h postinfection. Thus, activated macrophages were probably more capable of killing and processing Candida antigens, signalizing an adaptative immune response. Macrophages from controls did not prevent yeast-to-hyphae transition and were partially destroyed, as shown in scanning microscopy. These results suggest that treatment with Con-A facilitated the triggering of T(H) 17 and T(H) 1 responses via IL-17 and IFN-γ production, leading to the resolution of C. albicans infection.

Detection of Lsr2 gene of Mycobacterium leprae in nasal mucus

In the present study, nasal mucus from patients with leprosy were analyzed by PCR using specific primers for Lsr2 gene of Mycobacterium leprae. The presence of Lsr2 gene in the nasal mucus was detected in 25.80% of patients with paucibacillari leprosy, and 23.07% of contacts. Despite the absence of clinical features in the contact individuals, it was possible to detect the presence of Lsr2 gene in the nasal mucus of these individuals. Therefore, PCR detection of M. leprae targeting Lsr2 gene using nasal mucus samples could contribute to early diagnosis of leprosy.

INQUÉRITOS COPROPARASITOLÓGICOS DE PARASITOS INTESTINAIS NA CIDADE DE LONDRINA-PR: UMA ANÁLISE RETROSPECTIVA

Intestinal parasites are a major public health problem. It is important to inform and educate the public about these infections, especially where such data are scarce. This study aimed to determine the prevalence of intestinal parasites from the analysis of medical records of individuals of the city of Londrina. We analyzed 11,641 fecal reports from February 2009 to December 2012. Data were cataloged after the completion of parasitological testing of Hoffmann, Pons & Janer, Faust and Kato-Katz. From 11,641 reports, 19.1% were positive for intestinal parasites. Among those, 52.1% pertained to females and 47.9% to males, with predominance of positivity of 27.1% among children 0-10 years. For the regions studied, the northern region stood out with 35.4% of cases and prevalence of 6.8%. Among the pathogenic protozoa, reports of Giardia lamblia comprised 19.1% of positivity, while hookworms were the most frequent among helminths, comprising 7.8% of positive cases. It follows that poor conditions of basic sanitation contribute to the dissemination of these parasites. Early diagnosis is a determinant of successful treatment. Additionally, epidemiological data may be used to study the risk factors for transmission and may result in measures applicable to improving living conditions in the community