Adjuvant ArtinM favored the host immunity against Cryptococcus gattii infection in C57BL/6 mice.

Aim: Cryptococcus gattii causes a severe fungal infection with high mortality rate among immunosuppressed and immunocompetent individuals. Due to limitation of current antifungal treatment, new immunotherapeutic approaches are explored. Methods: This study investigated an immunization strategy utilizing heat-inactivated C. gattii with ArtinM as an adjuvant. C57BL/6 mice were intranasally immunized with heat-killed C. gattii and ArtinM was administrated either before immunization or along with HK-C. gattii. Mice were infected with C. gattii and the efficacy of the immunization protocol was evaluated. Results: Mice that received ArtinM exhibited increased levels of IL-10 and relative expression of IL-23 in the lungs, reduced fungal burden and preserved tissue integrity post-infection. Conclusion: Adjuvant ArtinM improved immunization against C. gattii infection in C57BL/6 mice.

Cryptococcus gattii is a fungus that can make lungs sick. Right now, there are no good treatments for it, so scientists are trying to find new ways to fight it. In a recent study, they tested a type of immunotherapy called ArtinM to see if it could help. When they gave ArtinM to mice, the mice got healthier and had less fungus in their lungs. This means ArtinM might be able to help fight this fungus.

Adjuvant Pam3CSk4 does not improve the immunization against Cryptococcus gattii infection in C57BL/6 mice.

Background: Cryptococcosis is a relevant invasive fungal infection that affects immunocompromised and immunocompetent individuals when caused by Cryptococcus gattii. Host innate and adaptive immune responses can be subverted by C. gattii, that blocks the differentiation of T helper (Th) 1 and Th17 cells, which are involved in the protection against cryptococcosis. Moreover, the macrophage polarization is modulated by C. gattii infection that requires a balance in the macrophage subsets to control the C. gattii infection. Toll-like receptor (TLR) 2 agonists are important immunomodulators favoring a pro-inflammatory response with potential fungicidal activity, and TLR2 agonists have been used as adjuvants in vaccines against infections caused by bacteria or viruses. Therefore, this work aimed to evaluate the immunomodulatory effect of the tripalmitoyl lipopeptide S-glycerol cysteine (Pam3CSK4 or P3C4), a TLR2 agonist, as an adjuvant in the vaccination against C. gattii infection. Methods and Results: C57BL/6 mice were immunized with 2 × 107 inactivated yeasts of C. gattii via intranasal route on day 1, 14 and 28 (Immunized group). Immunization was associated with 1µg or 10µg of adjuvant P3C4 (Immunized+P3C4-1µg or Immunized+P3C4-10 µg), followed by C. gattii infection on day 42 after the immunization protocol. Immunized+P3C4-1 µg group had reduced levels of IgG1, IgG2a and IgA and no significant difference in the IgG and IgM anti-GXM antibody titer was detected, compared to the Immunized group. High levels of IL-17 and IL-1ß in lung tissue of mice from the Immunized+P3C4-1µg group did not promote a predominance of Th17 cells, in contrast, the frequency of TLR2+ cells was increased in immunized mice that received 1 µg of P3C4. The reduction in the relative expression of T-bet and high levels of Foxp3 detected in the lungs of the Immunized+P3C4-1µg group suggest a prevalence of regulatory T cells in the tissue, which did not contribute to the control of C. gattii infection. The immunization protocol associated with 10 µg of adjuvant P3C4 induced high levels of IL-17 in the lung tissue, whereas the levels of pro-inflammatory cytokines were downregulated. To evaluate the effect of adjuvant P3C4 in the control of C. gattii infection, quantification of the fungal burden in the lungs was performed by the CFU assay, and the groups with adjuvant P3C4 showed a pulmonary C. gattii burden that was not significantly altered when compared with the immunized group. The mice that received 1 µg of adjuvant P3C4 had a lower percentage of inflammatory infiltrate in the lungs. Conclusion: The immunomodulatory effect of P3C4, associated with the immunization protocol, plays an imbalance between pro- and anti-inflammatory response in the lungs that did not favor a protection against C. gattii infection, which is related to the immune response characterized by a suppressive/regulatory profile in the pulmonary microenvironment after C. gattii infection.

Molecular analysis of the prevalence of Acinetobacter baumannii in hospitals and the surrounding environments: a cross-sectional study

Acinetobacter baumannii is widely recognized in clinical environments due to its infectious capacity, antimicrobial adaptability, and lethality. Analyzing the prevalence of this agent in intra- and extra-hospital environments may reveal target indicators for appropriate management interventions. In this observational cross-sectional study, we evaluated the prevalence of A. baumannii within hospitals with intensive care units and in their external surroundings in a macro-health region of Brazil. Samples of Columba livia (pigeon) droppings from the external environment of four hospitals (n = 40), from floor surfaces (n = 20), and door handles (n = 20) of different hospital wards were collected based on random sampling, all of which were evaluated for the presence of A. baumannii using polymerase chain reactions (PCR). The sensitivity and specificity of the technique was verified after the collected samples were contaminated with clinical samples positive for A. baumannii. We detected a significantly higher A. baumannii prevalence (87.50%, CI = 71.29­100.00) in samples collected within the hospital environment compared with those obtained from the external environment (12.50%, CI = 0.00­28. 71) (p = 0.003). In addition, samples collected from floor surfaces contained bacterial densities (181.3 ± 11.58) that exceeded those in environmental (93.32 ± 1.56) and door handle (142.70 ± 17.14) samples by 94% and 78.71%, respectively. The findings of this study will enhance our understanding of the spatial distribution of A. baumannii and additionally, validate the efficiency of PCR for diagnosis of this infectious agent.

In vitro antifungal activity of Morinda citrifolia (noni) extract against Candida albicans.

INTRODUCTION: Candida albicans is the main agent of the most common fungal infection, Candidiasis. It is an opportunistic and dangerous pathogen, especially in immunosuppressed patients. The biological properties of Morinda citrifolia (noni) make it a potent antifungal. In this study, antifungal effect of M. citrifolia was evaluated to verify its effect on human cells. METHODOLOGY: Extract of M. citrifolia was used against strains of C. albicans (cEC 1291). Glucose consumption in C. albicans biofilm was determined at different concentrations of M. citrifolia, and germ tube formation was evaluated in the presence and absence of M. citrifolia. Fungicidal activity was determined by the kinetics of fungal cell death. THP-1 and HeLa cells were used for cell viability and apoptosis, and cell proliferation assays, respectively. RESULTS: Cells treated with M. citrifolia maintained higher concentration of glucose than the control group (p < 0.05). Germ tube formation was inhibited in cells treated with M. citrifolia (p < 0.05). M. citrifolia exerted a cytotoxic effect on C. albicans cells with 99.99% lethality after 6.82 h (1:1 and 1:2), and reduced the viability of THP-1 cells by 25% and 67% after 12 and 36 h, respectively. Annexin V expression in THP-1 increased in groups that received higher concentrations of M. citrifolia (p < 0.05), reducing the proliferation of THP-1 and HeLa cells (2.8-fold). A greater cytotoxic effect was observed in fungal cells. CONCLUSIONS: These results indicate that M. citrifolia exerts biological activity against C. albicans and reduces the viability and proliferation of human cells.

Adjuvant Curdlan Contributes to Immunization against Cryptococcus gattii Infection in a Mouse Strain-Specific Manner.

The low efficacy and side effects associated with antifungal agents have highlighted the importance of developing immunotherapeutic approaches to treat Cryptococcus gattii infection. We developed an immunization strategy that uses selective Dectin-1 agonist as an adjuvant. BALB/c or C57BL/6 mice received curdlan or ß-glucan peptide (BGP) before immunization with heat-killed C. gattii, and the mice were infected with viable C. gattii on day 14 post immunization and euthanized 14 days after infection. Adjuvant curdlan restored pulmonary tumor necrosis factor- α (TNF-α) levels, as induced by immunization with heat-killed C. gattii. The average area and relative frequency of C. gattii titan cells in the lungs of curdlan-treated BALB/c mice were reduced. However, this did not reduce the pulmonary fungal burden or decrease the i0,nflammatory infiltrate in the pulmonary parenchyma of BALB/c mice. Conversely, adjuvant curdlan induced high levels of interferon-γ (IFN-γ) and interleukin (IL)-10 and decreased the C. gattii burden in the lungs of C57BL/6 mice, which was not replicated in ß-glucan peptide-treated mice. The adjuvant curdlan favors the control of C. gattii infection depending on the immune response profile of the mouse strain. This study will have implications for developing new immunotherapeutic approaches to treat C. gattii infection.

Predicting Blood Parasite Load and Influence of Expression of iNOS on the Effect Size of Clinical Laboratory Parameters in Acute Trypanosoma cruzi Infection With Different Inoculum Concentrations in C57BL/6 Mice.

In Chagas disease, the initial responses of phagocyte-mediated innate immunity are strongly associated with the control of Trypanosoma cruzi and are mediated by various signaling pathways, including the inducible nitric oxide synthetase (iNOS) pathway. The clinical and laboratory manifestations of Chagas disease depend on the parasite-host relationship, i.e., the responsive capacity of the host immune system and the immunogenicity of the parasite. Here, we evaluated effect sizes in clinical and laboratory parameters mediated by acute infection with different concentrations of T. cruzi inoculum in mice immunosuppressed via iNOS pathway inactivation. Infection was induced in C57BL/6 wild-type and iNOS-/- mice with the "Y" strain of T. cruzi at three inoculum concentrations (3 × 102, 3 × 103, and 3 × 104). Parasitemia and mortality in both mouse strains were monitored. Immunohistochemistry was performed to quantify amastigotes in cardiac tissues and cardiac musculature cells. Biochemical parameters, such as blood urea nitrogen, sodium, albumin, and globulin concentrations, among others, were measured, and cytokine concentrations were also measured. Effect sizes were determined by the eta squared formula. Compared with that in wild-type animals, mice with an absence of iNOS expression demonstrated a greater parasite load, with earlier infection and a delayed parasitemia peak. Inoculum concentration was positively related to death in the immunosuppressed subgroup. Nineteen parameters (hematological, biochemical, cytokine-related, and histopathological) in the immunocompetent subgroup and four in the immunosuppressed subgroup were associated with parasitemia. Parasitemia, biochemical parameters, and hematological parameters were found to be predictors in the knockout group. The impact of effect sizes on the markers evaluated based on T. cruzi inoculum concentration was notably high in the immunocompetent group (Cohen's d = 88.50%; p <.001). These findings contribute to the understanding of physiopathogenic mechanisms underlying T. cruzi infection and also indicate the influence of the concentration of T. cruzi during infection and the immunosuppression through the iNOS pathway in clinical laboratory heterogeneity reported in acute Chagas disease.

Corrigendum to "scFv against HSP60 of Strongyloides sp. and Its Application in the Evaluation of Parasite Frequency in the Elderly".

[This corrects the article DOI: 10.1155/2020/4086929.].

Administration of artinm lectin reduces the severity of the acute phase infection with Trypanosoma cruzi.

The acute phase of experimental Trypanosoma cruzi infection is associated with a strong inflammatory reaction, physiological changes, amastigote nests in tissues, and hematological alterations. ArtinM, a lectin extracted from Artocarpus heterophyllus seeds, is a homotetramer exhibiting immunomodulatory properties that promotes Th1 immune responses against intracellular pathogens, including the induction of neutrophil migration and increase in IL-12 production. This study aimed to evaluate the effects of ArtinM on experimental Chagas disease in mice. We evaluated mouse survival curves, parasitemia, hematological parameters including quantification of inflammatory infiltrates, and amastigote nests in cardiac tissue during infection. The results showed a reduced number of parasites in the blood, an increase in animal survival, improvements in hematological parameters, and decrease in inflammatory infiltrates and amastigote nests in the group treated with ArtinM. Collectively, these data suggest that the administration of ArtinM can lower the number of parasites in peak parasitemia caused by the Colombian strain of T. cruzi and can increase survival of infected mice. The observed reduction in cardiac tissue injury may be due to fewer T. cruzi amastigote nests and lower levels of inflammatory infiltrates. This study highlights the need for further investigation into the use of ArtinM as a potential alternative therapeutic for treating Chagas disease.

Macrophage migration inhibitory factor (MIF) and pregnancy may impact the balance of intestinal cytokines and the development of intestinal pathology caused by Toxoplasma gondii infection.

Toxoplasma gondii (T. gondii) is an intracellular parasite responsible for causing toxoplasmosis. When infection occurs during pregnancy, it can produce severe congenital infection with ocular and neurologic damage to the infant. From the oral infection parasite reaches the intestine, causing inflammatory response, damage in tissue architecture and systemic dissemination. Macrophage migration inhibition factor (MIF) is a cytokine secreted from both immune and non-immune cells, including gut epithelial cells. MIF is described to promote inflammatory responses, to be associated in colitis pathogenesis and also to play role in maintaining the intestinal barrier. The aim of the present study was to evaluate the influence of the pregnancy and MIF deficiency on T. gondii infection in the intestinal microenvironment and to address how these factors can impact on the intestinal architecture and local cytokine profile. For this purpose, small intestine of pregnant and non-pregnant C57BL/6 MIF deficient mice (MIF-/-) and Wild-type (WT) orally infected with 5 cysts of ME-49 strain of T. gondii were collected on day 8th of infection. Intestines were processed for morphological and morphometric analyses, parasite quantification and for cytokines mensuration. Our results showed that the absence of MIF and pregnancy caused an increase in T. gondii infection index. T. gondii immunolocalization demonstrated that segments preferentially infected with T. gondii were duodenum and ileum. The infection caused a reduction in the size of the intestinal villi, whereas, infection associated with pregnancy caused an increase in villi size due to edema caused by the infection. Also, the goblet cell number was increased in the ileum of MIF-/- mice, when compared to the corresponding WT group. Analyses of cytokine production in the small intestine showed that MIF was up regulated in the gut of pregnant WT mice due to infection. Also, infection provoked an intense Th1 response that was more exacerbated in pregnant MIF-/- mice. We also detected that the Th2/Treg response was more pronounced in MIF-/- mice. Altogether, our results demonstrated that pregnancy and MIF deficiency interferes in the balance of the intestinal cytokines and favors a Th1-immflamatory profile, which in turn, impact in the development of pathology caused by T. gondii infection in the intestinal microenvironment.

scFv against HSP60 of Strongyloides sp. and Its Application in the Evaluation of Parasite Frequency in the Elderly.

The present study is aimed at evaluating serological method using scFv anti-Strongyloides sp. and reporting the frequencies of the results with conventional parasitological technique (faeces) in elderly individuals. Among 112 elderly individuals (≥60 years of age), 14.28% were positive for at least one enteroparasite, with one individual positive for S. stercoralis. Sera were evaluated for the presence of anti-Strongyloides sp. antibodies using total or detergent fraction extracts of Strongyloides venezuelensis, which presented positivity rates of 19.64% and 10.71%, respectively. An anti-HSP60 single-chain variable fragment from Strongyloides sp. was used to detect parasite antigens, with 5.36% (6 individuals) of ELISA-positive individuals returning a positive result. While the serological test indicates previous or recent infection and may be limited by antigen purification, the anti-HSP60 method reflects the presence of Strongyloides sp. immune complexes and exhibits greater sensitivity and specificity. Our results demonstrate the variable occurrence of enteroparasites in elderly individuals residing in long-term nursing homes and validate a novel epidemiological tool to describe infection cases by Strongyloides sp.

Interleukin-6, tumor necrosis factor-α, C-reactive protein, and hematological parameters in experimental periodontal disease after ß-adrenergic blockade.

BACKGROUND: Changes in the levels of C-reactive protein (CRP), tumor necrosis factor α (TNF-α), and interleukin 6 (IL-6) observed during periodontal disease were linked with vascular manifestations. Recent studies showed that the beta-blocker propranolol reduces the pathological parameters associated with certain molecules at sites of bone injury. Hence, in this study, we evaluated the activity of propranolol on hematological parameters and systemic concentrations of inflammatory proteins in a model of experimental periodontitis. MATERIALS AND METHODS: Periodontal disease was induced in rats. After euthanasia, the number of inflammatory cells in each rat was quantified using histopathological assays. In addition, hematological parameters were quantitated using automated analysers, cytokine levels were determined using an enzyme-linked immunosorbent assay, and CRP levels were determined using a high-sensitivity immunoturbidimetric assay. RESULTS: Low doses of propranolol suppressed the systemic production of CRP, TNF-α, and IL-6; however, the hematological parameters were not affected. CONCLUSIONS: ß-adrenergic activation indirectly contributes to the pattern of systemic inflammatory molecules observed in periodontal disease. These molecules may initiate cardiovascular diseases as a consequence of periodontitis.

Systemic effects in naïve mice injected with immunomodulatory lectin ArtinM.

Toll-like receptors (TLR) contain N-glycans, which are important glycotargets for plant lectins, to induce immunomodulation. The lectin ArtinM obtained from Artocarpus heterophyllus interacts with TLR2 N-glycans to stimulate IL-12 production by antigen-presenting cells and to drive the immune response toward the Th1 axis, conferring resistance against intracellular pathogens. This immunomodulatory effect was demonstrated by subcutaneously injecting (s.c.) ArtinM (0.5 µg) in infected mice. In this study, we evaluated the systemic implications of ArtinM administration in naïve BALB/c mice. The mice were s.c. injected twice (7 days interval) with ArtinM (0.5, 1.0, 2.5, or 5.0 µg), LPS (positive control), or PBS (negative control) and euthanized after three days. None of the ArtinM-injected mice exhibited change in body weight, whereas the relative mass of the heart and lungs diminished in mice injected with the highest ArtinM dose (5.0 µg). Few and discrete inflammatory foci were detected in the heart, lung, and liver of mice receiving ArtinM at doses ≥2.5 µg. Moreover, the highest dose of ArtinM was associated with increased serum levels of creatine kinase MB isoenzyme (CK-MB) and globulins as well as an augmented presence of neutrophils in the heart and lung. IL-12, IFN-γ, TNF-α, and IL-10 measurements in the liver, kidney, spleen, heart, and lung homogenates revealed decreased IL-10 level in the heart and lung of mice injected with 5.0 µg ArtinM. We also found an augmented frequency of T helper and B cells in the spleen of all ArtinM-injected naïve mice, whereas the relative expressions of T-bet, GATA-3, and ROR-γt were similar to those in PBS-injected animals. Our study demonstrates that s.c. injection of high doses of ArtinM in naïve mice promotes mild inflammatory lesions and that a low immunomodulatory dose is innocuous to naïve mice.

Morinda citrifolia (Noni) Fruit Juice Reduces Inflammatory Cytokines Expression and Contributes to the Maintenance of Intestinal Mucosal Integrity in DSS Experimental Colitis.

Morinda citrifolia L. (noni) has been shown to treat different disorders. However, data concerning its role in the treatment of intestinal inflammation still require clarification. In the current study, we investigated the effects of noni fruit juice (NFJ) in the treatment of C57BL/6 mice, which were continuously exposed to dextran sulfate sodium (DSS) for 9 consecutive days. NFJ consumption had no impact on the reduction of the clinical signs of the disease or on weight loss. Nonetheless, when a dilution of 1 : 10 was used, the intestinal architecture of the mice was preserved, accompanied by a reduction in the inflammatory infiltrate. Regardless of the concentration of NFJ, a decrease in both the activity of myeloperoxidase and the key inflammatory cytokines, TNF-α and IFN-γ, was also observed in the intestine. Furthermore, when NFJ was diluted 1 : 10 and 1 : 100, a reduction in the production of nitric oxide and IL-17 was detected in gut homogenates. Overall, the treatment with NFJ was effective in different aspects associated with disease progression and worsening. These results may point to noni fruit as an important source of anti-inflammatory molecules with a great potential to inhibit the progression of inflammatory diseases, such as inflammatory bowel disease.

Antibiotic Resistance of Bacteria Involved in Urinary Infections in Brazil: A Cross-Sectional and Retrospective Study.

Empirical and prolonged antimicrobial treatment of urinary tract infections caused by Escherichia coli is associated with the emergence of bacterial resistance, and not all countries have strict policies against the indiscriminate use of drugs in order to prevent resistance. This cross-sectional and retrospective study (2010-2015) aimed to evaluate the sensitivity and resistance of patient-derived E. coli to different drugs broadly used to treat urinary infections in Brazil: ampicillin + sulbactam, cephalothin, ciprofloxacin, norfloxacin, and nitrofurantoin. We obtained 1654 E. coli samples from ambulatory patients with disease symptoms of the urinary tract from a Brazilian public hospital. While all antibiotics were effective in killing E. coli to a large degree, nitrofurantoin was the most effective, with fewer samples exhibiting antibiotic resistance. We assessed the costs of generic and brand name versions of each antibiotic. Nitrofurantoin, the most effective antibiotic, was the cheapest, followed by the fluoroquinolones (ciprofloxacin and norfloxacin), ampicillin + sulbactam and, lastly, cephalothin. Finally, assessment of antibiotic resistance to fluoroquinolones over the study period and extrapolation of the data led to the conclusion that these antibiotics could no longer be effective against E. coli-based urinary infections in approximately 20 years if their indiscriminate use in empirical treatment continues.

Establishing standards for studying renal function in mice through measurements of body size-adjusted creatinine and urea levels.

Strategies for obtaining reliable results are increasingly implemented in order to reduce errors in the analysis of human and veterinary samples; however, further data are required for murine samples. Here, we determined an average factor from the murine body surface area for the calculation of biochemical renal parameters, assessed the effects of storage and freeze-thawing of C57BL/6 mouse samples on plasmatic and urinary urea, and evaluated the effects of using two different urea-measurement techniques. After obtaining 24 h urine samples, blood was collected, and body weight and length were established. The samples were evaluated after collection or stored at -20°C and -70°C. At different time points (0, 4, and 90 days), these samples were thawed, the creatinine and/or urea concentrations were analyzed, and samples were restored at these temperatures for further measurements. We show that creatinine clearance measurements should be adjusted according to the body surface area, which was calculated based on the weight and length of the animal. Repeated freeze-thawing cycles negatively affected the urea concentration; the urea concentration was more reproducible when using the modified Berthelot reaction rather than the ultraviolet method. Our findings will facilitate standardization and optimization of methodology as well as understanding of renal and other biochemical data obtained from mice.

Anti-inflammatory effects of ellagic acid on acute lung injury induced by acid in mice.

Acute lung injury (ALI) is characterized by alveolar edema and uncontrolled neutrophil migration to the lung, and no specific therapy is still available. Ellagic acid, a compound present in several fruits and medicinal plants, has shown anti-inflammatory activity in several experimental disease models. We used the nonlethal acid aspiration model of ALI in mice to determine whether preventive or therapeutic administration of ellagic acid (10 mg/kg; oral route) could interfere with the development or establishment of ALI inflammation. Dexamethasone (1 mg/kg; subcutaneous route) was used as a positive control. In both preventive and therapeutic treatments, ellagic acid reduced the vascular permeability changes and neutrophil recruitment to the bronchoalveolar lavage fluid (BALF) and to lung compared to the vehicle. In addition, the ellagic acid accelerated the resolution for lung neutrophilia. Moreover, ellagic acid reduced the COX-2-induced exacerbation of inflammation. These results were similar to the dexamethasone. However, while the anti-inflammatory effects of dexamethasone treatment were due to the reduced activation of NF- κ B and AP-1, the ellagic acid treatment led to reduced BALF levels of IL-6 and increased levels of IL-10. In addition, dexamethasone treatment reduced IL-1 ß. Together, these findings identify ellagic acid as a potential therapeutic agent for ALI-associated inflammation.