ABSTRACT
Secondary infections are one of the complications in COVID-19 patients. We aimed to analyze the antimicrobial prescriptions and their influence on drug resistance in fungi and bacteria isolated from severely ill COVID-19 patients. Seventy-nine severely ill COVID-19 hospitalized patients with secondary bacterial or fungal infections were included. We analyzed the prescribed antimicrobial regimen for these patients and the resistance profiles of bacterial and fungal isolates. In addition, the association between drug resistance and patients' outcome was analyzed using correlation tests. The most prescribed antibacterial were ceftriaxone (90.7% of patients), vancomycin (86.0%), polymyxin B (74.4%), azithromycin (69.8%), and meropenem (67.4%). Micafungin and fluconazole were used by 22.2 and 11.1% of patients, respectively. Multidrug-resistant (MDR) infections were a common complication in severely ill COVID-19 patients in our cohort since resistant bacteria strains were isolated from 76.7% of the patients. Oxacillin resistance was observed in most Gram-positive bacteria, whereas carbapenem and cephalosporin resistance was detected in most Gram-negative strains. Azole resistance was identified among C. glabrata and C. tropicalis isolates. Patients who used more antimicrobials stayed hospitalized longer than the others. The patient's age and the number of antibacterial agents used were associated with the resistance phenotype. The susceptibility profile of isolates obtained from severely ill COVID-19 patients highlighted the importance of taking microbial resistance into account when managing these patients. The continuous surveillance of resistant/MDR infection and the rational use of antimicrobials are of utmost importance, especially for long-term hospitalized patients with COVID-19.
Subject(s)
COVID-19 Drug Treatment , Microbial Sensitivity Tests , Anti-Bacterial Agents/pharmacology , Anti-Bacterial Agents/therapeutic use , Bacteria , Fungi , Prescriptions , Drug ResistanceABSTRACT
The co-infection between visceral leishmaniasis (VL) and human immunodeficiency virus (HIV) has increased in several countries in the world. The current serological tests are not suitable since they present low sensitivity to detect the most of VL/HIV cases, and a more precise diagnosis should be performed. In this context, in the present study, an immunoproteomics approach was performed using Leishmania infantum antigenic extracts and VL, HIV and VL/HIV patients sera, besides healthy subjects samples; aiming to identify antigenic markers for these clinical conditions. Results showed that 43 spots were recognized by antibodies in VL and VL/HIV sera, and 26 proteins were identified by mass spectrometry. Between them, ß-tubulin was expressed, purified and tested in ELISA experiments as a proof of concept for validation of our immunoproteomics findings and results showed high sensitivity and specificity values to detect VL and VL/HIV patients. In conclusion, the identified proteins in the present work could be considered as candidates for future studies aiming to improvement of the diagnosis of VL and VL/HIV co-infection.
Subject(s)
Coinfection/diagnosis , HIV Infections/diagnosis , Leishmania infantum/isolation & purification , Leishmaniasis, Visceral/diagnosis , Proteomics/methods , Protozoan Proteins/analysis , Adult , Brazil , Female , Humans , Male , Middle AgedABSTRACT
The laboratorial diagnosis of leishmaniasis is based on parasitological methods, which are invasive, present high cost, require laboratorial infrastructure and/or trained professionals; as well as by immunological methods, which usually present variable sensitivity and/or specificity, such as when they are applied to identify asymptomatic cases and/or mammalian hosts presenting low levels of antileishmanial antibodies. As consequence, new studies aiming to identify more refined antigens to diagnose visceral (VL) and tegumentary (TL) leishmaniasis are urgently necessary. In the present work, the Leishmania eukaryotic elongation factor-1 beta (EF1b) protein, which was identified in L. infantum protein extracts by antibodies in VL patients' sera, was cloned and its recombinant version (rEF1b) was expressed, purified and tested as a diagnostic marker for VL and TL. The post-therapeutic serological follow-up was also evaluated in treated and untreated VL and TL patients, when anti-rEF1b antibody levels were measured before and after treatment. Results showed that rEF1b was highly sensitive and specific to diagnose symptomatic and asymptomatic canine VL, as well as human TL and VL. In addition, low cross-reactivity was observed when sera from healthy subjects or leishmaniasis-related diseases patients were tested. The serological follow-up showed also that rEF1b-specific antibodies declined significantly after treatment, suggesting that this protein could be also evaluated as a prognostic marker for human leishmaniasis.
Subject(s)
Dog Diseases/parasitology , Eukaryotic Initiation Factor-1/immunology , Leishmania infantum/immunology , Leishmaniasis, Visceral/parasitology , Leishmaniasis, Visceral/veterinary , Protozoan Proteins/immunology , Adult , Animals , Antibodies, Protozoan/immunology , Antigens, Protozoan/genetics , Antigens, Protozoan/immunology , Cross Reactions , Dog Diseases/diagnosis , Dog Diseases/immunology , Dogs , Enzyme-Linked Immunosorbent Assay , Eukaryotic Initiation Factor-1/genetics , Female , Humans , Leishmania infantum/genetics , Leishmania infantum/isolation & purification , Leishmaniasis/diagnosis , Leishmaniasis/immunology , Leishmaniasis/parasitology , Leishmaniasis/veterinary , Leishmaniasis, Visceral/diagnosis , Leishmaniasis, Visceral/immunology , Male , Middle Aged , Protozoan Proteins/genetics , Serologic TestsABSTRACT
The measures for leishmaniasis control include the precise diagnosis of disease. However, although several recombinant antigens have been tested with this biotechnological purpose, no effective product exists, which could detects patients with the active disease, as well as differentiates them from cured and treated patients. In this study, a conserved Leishmania hypothetical protein, which was identified in Leishmania infantum parasites, but evaluated to presents high homology in the amino acid sequences between distinct parasite species, was evaluated for the diagnosis of tegumentary and visceral leishmaniasis. In addition, PBMCs collected from treated and untreated mucosal leishmaniasis (ML) and visceral leishmaniasis (VL) patients, as well as in healthy subjects living in endemic region of disease, were in vitro stimulated, when IFN-γ, IL-4 and IL-10 levels were evaluated in the cell supernatant. Regarding the serological analyses, ELISA experiments using the recombinant protein (rLiHyL) and a human serological panel revealed high sensitivity and specificity values to detect both diseases, while control antigens showed worst results. Regarding the cellular response, results showed that rLiHyL-stimulated cells produced higher IFN-γ and lower IL-4 and IL-10 levels in the supernatants. Also, the anti-protein antibody production was evaluated in these patients, and data showed higher IgG2 and lower IgG1 levels found in the treated patients and healthy controls, demonstrating the stimulation of a Th1-type response induced by the rLiHyL protein. In conclusion, this hypothetical protein can be considered as antigenic in TL and VL, as well as a vaccine candidate to be tested in future studies to protect against disease.
Subject(s)
Antigens, Protozoan/immunology , Leishmaniasis, Visceral/immunology , Protozoan Proteins/immunology , Recombinant Proteins , Adult , Antibodies, Protozoan/immunology , Antigens, Protozoan/genetics , Biomarkers , Case-Control Studies , Cytokines/metabolism , Enzyme-Linked Immunosorbent Assay , Female , Humans , Immunoglobulin G/immunology , Leishmaniasis, Visceral/diagnosis , Leishmaniasis, Visceral/parasitology , Male , Middle Aged , Protozoan Proteins/genetics , Serologic TestsABSTRACT
The diagnosis of visceral leishmaniasis (VL) presents problems due to the toxicity and/or high cost of drugs. In addition, no vaccine exists to protect against human disease. In this study, the antigenicity and immunogenicity of amastin protein were evaluated in L. infantum-infected dogs and humans. For the diagnosis, besides the recombinant protein, 1 linear B-cell epitope was synthetized and evaluated in serological assays. Results showed high sensitivity and specificity values to detect the disease when both antigens were employed against a canine and human serological panel. By contrast, when using rA2 and a soluble Leishmania antigenic preparation, sensitivity and specificity values proved to be lower. A preliminary immunogenicity study showed that the amastin protein induced high IFN-γ and low IL-10 production in stimulated PBMC derived from treated VL patients and healthy subjects, thus suggesting a potential use of this protein as an immunogen to protect against human disease.
Subject(s)
Antigens, Protozoan/immunology , Leishmania infantum/isolation & purification , Leishmaniasis, Visceral/diagnosis , Leishmaniasis, Visceral/veterinary , Protozoan Proteins/immunology , Animals , Antibodies, Protozoan/blood , Antigens, Protozoan/chemistry , Antigens, Protozoan/genetics , Cells, Cultured , Cytokines/metabolism , Dog Diseases/blood , Dog Diseases/diagnosis , Dog Diseases/parasitology , Dogs , Epitopes, B-Lymphocyte/immunology , Humans , Immunogenicity, Vaccine , Leishmania infantum/immunology , Leishmaniasis, Visceral/blood , Leishmaniasis, Visceral/parasitology , Leukocytes, Mononuclear/immunology , Protozoan Proteins/chemistry , Protozoan Proteins/genetics , Recombinant Proteins/chemistry , Recombinant Proteins/immunology , Sensitivity and Specificity , Serologic TestsABSTRACT
There is no suitable vaccine against human visceral leishmaniasis (VL) and available drugs are toxic and/or present high cost. In this context, diagnostic tools should be improved for clinical management and epidemiological evaluation of disease. However, the variable sensitivity and/or specificity of the used antigens are limitations, showing the necessity to identify new molecules to be tested in a more sensitive and specific serology. In the present study, an immunoproteomics approach was performed in Leishmania infantum promastigotes and amastigotes employing sera samples from VL patients. Aiming to avoid undesired cross-reactivity in the serological assays, sera from Chagas disease patients and healthy subjects living in the endemic region of disease were also used in immunoblottings. The most reactive spots for VL samples were selected, and 29 and 21 proteins were identified in the promastigote and amastigote extracts, respectively. Two of them, endonuclease III and GTP-binding protein, were cloned, expressed, purified and tested in ELISA experiments against a large serological panel, and results showed high sensitivity and specificity values for the diagnosis of disease. In conclusion, the identified proteins could be considered in future studies as candidate antigens for the serodiagnosis of human VL.
Subject(s)
Antigens, Protozoan/immunology , Leishmania infantum/physiology , Leishmaniasis, Visceral/immunology , Protozoan Proteins/immunology , Adult , Female , Humans , Male , Middle Aged , ProteomicsABSTRACT
Serological tests are important tools for the diagnosis of Leishmania infection. However, they are not effective markers to diagnose asymptomatic cases of visceral leishmaniasis (VL) and patients developing tegumentary leishmaniasis (TL), since antileishmanial antibodies can be encountered in low levels resulting in false-negative results in the serological trials. In this context, antigens able to be recognized by antibodies in sera from both VL and TL patients will be desirable to be employed in a more sensitivity and specific diagnosis of disease. In the present study, a conserved Leishmania protein, small myristoylated protein-3 (SMP-3), which was showed to be conserved in different Leishmania species and an effective vaccine candidate against Leishmania infantum infection in a murine model, was cloned and the recombinant protein was evaluated as a serological marker for the diagnosis of human TL and canine VL. In addition, a linear B cell-specific epitope (MQKDEESGEFKCEL) was identified, synthetized and also investigated as a serological marker. As antigen controls, rA2 protein and antigenic Leishmania extracts (SLA) were used. Results showed that ELISA-rSMP-3 and ELISA-Peptide presented sensitivity and specificity values higher than 90% in both diseases in humans and canids, having identified all asymptomatic cases and did not present cross-reaction with cross-reactivity diseases in both mammalian hosts. On the other hand, sensitivity and specificity values were worst when rA2 or SLA were used as antigens in humans and dogs. In conclusion, results showed the efficacy and Leishmania SMP-3 protein, employed as a recombinant antigen or a B cell epitope, for the improvement of the serodiagnosis of human TL and canine VL. This candidate can be tested in other diagnostic platforms, such as rapid immunochromatographic dipstick tests, aiming its use in epidemiological studies in remote areas where laboratories are not readily accessible for conventional assays.
Subject(s)
Antigens, Protozoan/immunology , Dog Diseases/diagnosis , Epitopes, B-Lymphocyte/immunology , Leishmania/physiology , Leishmaniasis, Visceral/diagnosis , Recombinant Proteins/immunology , Zoonoses/diagnosis , Adult , Aged , Animals , Antibodies, Protozoan/blood , Antigens, Protozoan/genetics , Cross Reactions , Dogs , Epitopes, B-Lymphocyte/genetics , Female , Humans , Male , Middle Aged , Recombinant Proteins/genetics , Sensitivity and Specificity , Serologic Tests , Young AdultABSTRACT
In the present study, a conserved Leishmania hypothetical protein, LiHyE, was evaluated for the serodiagnosis of leishmaniasis. Results showed that it presented high sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) to serologically identify visceral leishmaniasis (VL) dogs when 40 positive sera and 95 cross-reactive samples were used. rLiHyE also showed the best results of sensitivity, specificity, PPV, and NPV to identify tegumentary leishmaniasis (TL) and VL patients when 45 leishmaniasis patients' sera and 90 cross-reactive samples were used. Results were better in comparison to those obtained when rA2 or Leishmania antigenic extract was employed as controls. The posttreatment follow-up showed that rLiHyE-specific antibodies declined significantly after the end of treatments, and a predominance of the IgG2 subclass was found in comparison to IgG1 levels in both TL and VL patients. In conclusion, rLiHyE can be considered a candidate for the serodiagnosis of canine and human leishmaniasis.
Subject(s)
Biomarkers/blood , Dog Diseases/diagnosis , Leishmaniasis, Visceral/diagnosis , Leishmaniasis/diagnosis , Protozoan Proteins , Serologic Tests , Animals , Antibodies, Protozoan , Antigens, Protozoan , Dog Diseases/drug therapy , Dog Diseases/parasitology , Dogs , Enzyme-Linked Immunosorbent Assay/methods , Follow-Up Studies , Humans , Immunoglobulin G/blood , Immunoglobulin G/immunology , Leishmaniasis/drug therapy , Leishmaniasis/parasitology , Leishmaniasis, Visceral/drug therapy , Leishmaniasis, Visceral/parasitology , Protozoan Proteins/immunology , Recombinant Proteins/immunology , Sensitivity and Specificity , Serologic Tests/methodsABSTRACT
New candidates for serological markers against leishmaniasis are required to be identified, since the presence of high titers of anti-Leishmania antibodies remain detected in sera of treated and cured patients, when current antigens have being employed. In this study, the diagnostic performance of a conserved Leishmania hypothetical protein was evaluated against a human and canine serological panel. The serological follow-up of the patients was also evaluated, using this recombinant antigen (rLiHyS) in ELISA assays. In the results, high sensitivity and specificity values were found when rLiHyS was used in the serological tests, while when the recombinant A2 (rA2) protein or an antigenic Leishmania preparation were used as controls, low sensitivity and specificity were found. Regarding the serological follow-up of the patients, significant reductions in the anti-rLiHyS antibody levels were found and, one year after the treatments, the anti-protein IgG production was similar to this found in the non-infected groups, reflecting a drop of the anti-rLiHyS antibody production. In conclusion, the present study shows for the first time a new recombinant antigen used to identify tegumentary and visceral leishmaniasis, as well as being able to serologically distinguish treated and cured patients from those developing active disease.
Subject(s)
Leishmania braziliensis/immunology , Leishmania infantum/immunology , Leishmaniasis, Cutaneous/diagnosis , Leishmaniasis, Visceral/diagnosis , Protozoan Proteins/immunology , Adult , Amino Acid Sequence , Animals , Antigens, Helminth/genetics , Antigens, Helminth/immunology , Biomarkers/blood , Chagas Disease/diagnosis , Chagas Disease/immunology , Dog Diseases/blood , Dog Diseases/immunology , Dog Diseases/parasitology , Dogs , Enzyme-Linked Immunosorbent Assay , Female , Humans , Leishmania braziliensis/chemistry , Leishmania infantum/chemistry , Leishmaniasis, Cutaneous/immunology , Leishmaniasis, Visceral/diet therapy , Leishmaniasis, Visceral/immunology , Male , Middle Aged , Prognosis , Protozoan Proteins/genetics , Recombinant Proteins/immunology , Sensitivity and Specificity , Serologic Tests/methods , Young AdultABSTRACT
Visceral leishmaniasis (VL) is a potentially fatal disease, in which the treatment based on chemotherapy is considered toxic. The cure of disease is associated with the life-long Th1-type immunity against the infection. The Th1-related cytokines production by peripheral blood mononuclear cells (PBMCs) seems to be crucial for host control of parasite load and clinical cure. In the current study, we used five proteins (IgE-dependent histamine-releasing factor [HRF], LiHyD, LiHyV, LiHyT and LiHyp6) recently shown to be antigenic and/or immunogenic in the canine VL, aiming to evaluate the antigen-specific antibody levels and cytokine production in PBMCs culture supernatants collected from VL patients before and after anti-VL treatment. In the results, when PBMCs were exposed to rHRF, rLiHyD and rLiHyT, higher IFN-γ and lower IL-10 levels were observed in all patients that were treated and clinically cured. Analysis of specific antibody subclasses was in line with in vitro cellular response, since a higher IgG2 production was found in the treated and cured patients, when compared to the IgG1 subclass levels. In addition, evaluating the diagnostic efficacy of the recombinant molecules, the rHRF, rLiHyD and rLiHyT proteins showed the best results in the serology assays to identify all VL patients, as well as these antigens were not recognized by antibodies in sera from non-infected subjects or those with leishmaniasis-related diseases. Our results corroborate the view that clinical cure of VL is associated with a sustained Th1-related response, and indicate the potential use of rHRF, rLiHyD and rLiHyT as immune biomarkers of VL treatment.
Subject(s)
Antibodies, Protozoan/blood , Biomarkers, Pharmacological/blood , Leishmania infantum/physiology , Leishmaniasis, Visceral/diagnosis , Leukocytes, Mononuclear/immunology , Th1 Cells/immunology , Adult , Animals , Antigens, Protozoan/genetics , Antigens, Protozoan/immunology , Cells, Cultured , Disease Progression , Dogs , Female , Humans , Interferon-gamma/metabolism , Interleukin-10/metabolism , Leishmaniasis, Visceral/therapy , Leukocytes, Mononuclear/parasitology , Lymphocyte Activation , Male , Middle Aged , Recombinant Proteins/genetics , Recombinant Proteins/immunology , Young AdultABSTRACT
Visceral leishmaniasis (VL) represents a serious public health problem, as Leishmania infantum is one of main disease causative agents in the Americas. In a previous immunoproteomic study, the prohibitin (PHB) protein was identified in L. infantum promastigote and amastigote extracts by antibodies in asymptomatic and symptomatic VL dog sera. This protein was found to be highly conserved between different Leishmania spp., but it presented a low identity with amino acid sequences of other organisms. The aim of the present study was to evaluate the cellular response induced by the recombinant PHB (rPHB) protein in BALB/c mice, as well as in PBMCs purified from untreated and treated VL patients, as well as to evaluate its protective efficacy against an infection by L. infantum promastigotes. Our data showed that there was a Th1 cellular response to rPHB, based on high levels of IFN-γ, IL-12, and GM-CSF in the immunized animals, as well as a proliferative response specific to the protein and higher IFN-γ levels induced in PBMCs from individuals who had recovered from the disease. The protection was represented by significant reductions in the parasite load in the animals' spleen, liver, bone marrow, and draining lymph nodes, as compared to results found in the control groups. In addition, an anti-rPHB serology, using a canine and human serological panel, showed a high performance of this protein when diagnosing VL based on high sensitivity and specificity values, as compared to results found for the rA2 antigen and the soluble Leishmania antigenic extract. Our data suggest that PHB has a potential application for the diagnosis of canine and human VL through antibody detection, as well as an application as a vaccine candidate to protect against disease.
Subject(s)
Leishmaniasis, Visceral/diagnosis , Leishmaniasis, Visceral/immunology , Repressor Proteins/immunology , Animals , Antigens, Protozoan/immunology , Dogs , Humans , Leishmania infantum/immunology , Leishmania infantum/metabolism , Leishmaniasis, Visceral/metabolism , Mice , Mice, Inbred BALB C , Prohibitins , Recombinant Proteins/metabolism , Repressor Proteins/metabolism , Th1 Cells/immunology , Vaccines/metabolismABSTRACT
Different Leishmania proteins have been evaluated in order to find a potential vaccine candidate or diagnostic marker capable of providing long lasting protection against infection or helping to identify infected mammalian hosts, respectively. However, just few molecules have fulfilled all the requirements to be evaluated. In the current study, we evaluated the prophylactic and diagnostic value against visceral leishmaniasis (VL) of a small glutamine-rich tetratricopeptide repeat-containing (SGT) protein from Leishmania infantum species. In a first step, the immune response elicited by the immunization using the recombinant protein (rSGT) plus saponin was evaluated in BALB/c mice. Immunized animals had a low parasitism in all evaluated organs. They developed a specific Th1 immune response, which was based on protein-specific production of IFN-γ, IL-12 and GM-CSF, and a humoral response dominated by antibodies of the IgG2a isotype. Both CD4+ and CD8+ T cells contributed to the IFN-γ production, showing that both T cell subtypes contribute to the resistance against infection. Regarding its value as a diagnostic marker, rSGT showed maximum sensitivity and specificity to serologically identify L. infantum-infected dog and human sera. No cross-reactivity with sera from humans or dogs that had other diseases was found. Although further studies are necessary to validate these findings, data showed here suggest immunogenicity of rSGT and its protective effect against murine VL, as well as its potential for the serodiagnosis of human and canine VL.
Subject(s)
Leishmania infantum/immunology , Leishmaniasis Vaccines/immunology , Leishmaniasis, Visceral/diagnosis , Leishmaniasis, Visceral/prevention & control , Protozoan Proteins/immunology , Animals , Antibodies, Protozoan/immunology , CD8-Positive T-Lymphocytes/immunology , CD8-Positive T-Lymphocytes/pathology , Cross Reactions , Cytokines/immunology , Dogs , Female , Humans , Immunoglobulin G/immunology , Leishmania infantum/genetics , Leishmaniasis Vaccines/genetics , Leishmaniasis, Visceral/genetics , Leishmaniasis, Visceral/immunology , Mice, Inbred BALB C , Protozoan Proteins/genetics , Protozoan Proteins/pharmacology , Recombinant Proteins/genetics , Recombinant Proteins/immunology , Recombinant Proteins/pharmacology , Th1 Cells/immunology , Th1 Cells/pathologyABSTRACT
BACKGROUND: Propolis has been used as a natural health product mainly due to the presence of polyphenols, flavonoids, phenolic aldehydes, amino acids, vitamins and others bioactive constituents. To this natural substance are attributed different biological and pharmacological properties which are influenced by its chemical composition and organoleptic properties. The aim of this work was to evaluate the physicochemical properties and parameters of green propolis collected during a period of six years (2008-2013) in the state of Minas Gerais, located at the southeastern region of Brazil. METHODS: The methodology were in accordance with Brazilian legislation on the identity and quality standards of propolis. The evaluated parameters of hydroalcoholic from green propolis were total flavonoids, antioxidant activity - DPPH method, oxidation index, wax content, humidity and insoluble impurities. RESULTS: Propolis samples collected in different seasons during the years 2008 to 2013 presented mean values of total flavonoids (3.4 ± 0.11 mg/g), antioxidant activity DPPH (4.76 ± 0.16 µg/mL), oxidation index (3, 4 ± 0.33 seconds) and wax (15.14 ± 0.78% m/m), which are in accordance with Brazilian legislation. CONCLUSION: Green propolis did not show abrupt seasonal changes during the six years of investigation, and may be considered as an adequate functional ingredient.
Subject(s)
Propolis/chemistry , Antioxidants/chemistry , Baccharis , Biphenyl Compounds/chemistry , Brazil , Flavonoids/analysis , Phenols/analysis , Picrates/chemistry , Seasons , Waxes/analysisABSTRACT
BACKGROUND: Paracoccidioidomycosis (PCM) is a systemic mycosis caused by dimorphic fungi Paracoccidioides brasiliensis and Paracoccidioides lutzii. It is prevalent in Latin American, mainly in Brazil. Therefore, PCM has fundamental impact on the Brazilian global economy, especially in public health system, since it is affecting economical active population in different country regions. OBJECTIVE: The present study aimed to standardize the Real Time-Polymerase Chain Reaction (rt-PCR) for an efficient and safe PCM diagnosis amplifying the recombinant protein PB27 gene, only expressed by specimens of Paracoccidioides genus. METHODS: To standardize a methodology of rt-PCR using species-specific primers and probe designed for annealing in this specific region of the fungi´s genome, amplifying the recombinant protein PB27 gene, only expressed by specimens of Paracoccidioides genus. Followed by design in silico, experiments were performed in vitro to determine rt-PCR specificity, efficiency and genome detection limit. RESULTS: The primers and probe sequences were deposited in Brazilian Coordination of Technological Innovation and Transfer (CTIT), under patent reference number BR1020160078830. The present study demonstrated the rt-PCR applicability for support on diagnosis of paracoccidioidomycosis, presenting low cost, which makes it affordable for public health services in developing countries as Brazil. It is noteworthy that it is necessary to validate this methodology using clinical samples before to use as a safe method of diagnosis. A review of all patents related to this topic was performed and it was shown that, to date, there are no records of patent on kits for paracoccidioidomycosis´s diagnostic. Indeed, there is still a lot to go to reach this goal. CONCLUSION: The reaction developed was standardized and patented, opening perspectives to molecular diagnosis development for paracoccidioidomycosis, since rt-PCR can be applied to a broad spectrum of infectious diseases. It would need to be tested in biological samples in order to validate this method and then generate a diagnostic kit for Paracoccidioidomycosis.
Subject(s)
Fungal Proteins/genetics , Paracoccidioides/isolation & purification , Paracoccidioidomycosis/diagnosis , Real-Time Polymerase Chain Reaction/methods , Brazil , Computer Simulation , Genome, Fungal , Humans , Paracoccidioides/genetics , Paracoccidioidomycosis/microbiology , Patents as Topic , Real-Time Polymerase Chain Reaction/economics , Sensitivity and Specificity , Species SpecificityABSTRACT
BACKGROUND: Visceral leishmaniasis (VL) is a zooanthroponosis affecting both rural and periurban areas, and can also spread into urban areas. VL has emerged in many countries in the world, presenting new cases in new countries of occurrence. Thus, studies concerning epidemiological aspects in different world regions are very meaningful. METHODS: With this purpose, this study analyzed 89 cases of VL, treated between June 2006 and June 2014 at Eduardo de Menezes Hospital (HEM), a Reference Center of Infectious Diseases situated in Belo Horizonte, in Minas Gerais state, Brazil. RESULTS: According to the results, it was observed that males are mostly infected (84%/n=75) and the most affected age range was 20-49 years old (83%/n=74). The treatment liposomal amphotericin B (33%/n=29) was mostly used. Recurrences were more frequent in patients treated with Glucantime® (17%/n=9). No side effects were reported among the 29 patients treated with liposomal amphotericin B. On the other hand, there were 23 cases related to the occurrence of acute renal failure (ARF) and the use of conventional amphotericin B, both when it was administered alone or in combination with other drugs. Additionally, we observed a close relationship between the VL and HIV infection, observing a coinfection rate of 28.1% (n=25). CONCLUSION: From the survey data, it was possible to conclude that the majority of VL patient treated at HEM is male, classified as brown racial group, economically active, and may be drug addicts, chronic alcoholics and/or smokers. They may present some Non-communicable diseases (NCD), such as hypertension, diabetes mellitus, dyslipidemia and/or obesity and, predominantly present a great chance of being a carrier of the Human Immunodeficiency Virus, associated or not with tuberculosis. As symptoms, these patients possibly will present hepatosplenomegaly, fever and pronounced weight loss.
Subject(s)
Amphotericin B/therapeutic use , Antiprotozoal Agents/therapeutic use , Leishmaniasis, Visceral/drug therapy , Meglumine/therapeutic use , Organometallic Compounds/therapeutic use , Adolescent , Adult , Amphotericin B/administration & dosage , Antiprotozoal Agents/administration & dosage , Brazil/epidemiology , Child , Child, Preschool , Coinfection , Female , HIV Infections/epidemiology , Humans , Infant , Leishmaniasis, Visceral/complications , Leishmaniasis, Visceral/epidemiology , Male , Meglumine/administration & dosage , Meglumine Antimoniate , Middle Aged , Organometallic Compounds/administration & dosage , Recurrence , Retrospective Studies , Sex Distribution , Young AdultABSTRACT
Sulforaphane (SFN) is a molecule within the isothiocyanate (ITC) group of organosulfur compounds. SFN is a phytochemical commonly found in cruciferous vegetables such as broccoli, brussels sprouts and cabbages. It has been widely studied in order to evaluate its chemopreventive properties and some of those have already been established by means of animal and human models. The SFN induces Phase I and II enzymes involved in detoxification processes of chemical carcinogens in order to prevent the start of carcinogenesis. It also presents anti-tumor action at post-initiation Phase, suggesting supplementary roles in cancer prevention. In a dose dependent manner, ITC inhibits the viability of human cancer cells, modifies epigenetic events that occur in cancer cells and present antiinflammatory effect acting during the initial of uncontrolled cell proliferation. This protective effect may be due to its antioxidant status, its recognized capacity to induce the expression and/or activity of of different cytoprotective proteins involved in the activating "Nuclear factor erythroid-derived 2-like 2" (Nrf2). Nevertheless, the effects on health and the possible connections among different diet constituents in humans must be carefully studied as there are limitations in the current data in order to better understand the molecular mechanisms responsible for those effects. This survey also includes relevant patents on the use of SFN, like its use in skin cancer treatment (US2015038580); and as an adjuvant in anti-cancer treatment (US2014228419). The use of SFN as an antioxidant dietary supplement, methods for compositions that promote glutathione production (WO2015002279) and methods for extracting and purifying SFN from broccoli seeds (CN104086469) are also included in this review.
Subject(s)
Antineoplastic Agents, Phytogenic/therapeutic use , Antioxidants/therapeutic use , Dietary Supplements , Isothiocyanates/therapeutic use , Neoplasms/drug therapy , Animals , Antineoplastic Agents, Phytogenic/adverse effects , Antioxidants/adverse effects , Antioxidants/chemistry , Apoptosis/drug effects , Dietary Supplements/adverse effects , Drug Design , Epigenesis, Genetic/drug effects , Humans , Isothiocyanates/adverse effects , Isothiocyanates/chemistry , Molecular Structure , Neoplasms/genetics , Neoplasms/metabolism , Neoplasms/pathology , Patents as Topic , Signal Transduction/drug effects , Structure-Activity Relationship , SulfoxidesABSTRACT
The diagnosis of visceral leishmaniasis (VL) is a challenging issue and several studies worldwide have evaluated the different tools to reach a diagnostic solution. The polymerase chain reaction (PCR) has proven to be effective in detecting the genome of Leishmania species in different biological samples. In this study, we compared the conventional PCR and real-time PCR using the Sybr Green system and their application in molecular diagnosis of visceral leishmaniasis in peripheral blood as a biological sample. The genus-specific conserved region of kinetoplast DNA (kDNA) was the target of amplification. We studied 30 samples from patients with suspect of visceral leishmaniasis who were treated by the Medical Clinic of Santa Casa de Belo Horizonte Hospital, Brazil. Among the samples studied, 19 had a confirmed diagnosis for VL by serology and/or by clinical findings. Among these 19 samples, 63% (n = 12) presented positive results for serology and 79% (n = 15) positive results in both PCR methodologies. This fact suggests that the PCR technique can assist in the diagnosis of visceral leishmaniasis in patients who do not have detectable antibodies by serology but can present the genome of the parasite circulating in whole blood. Also, it was possible to observe that there was conformity between the results of the techniques of cPCR and qPCR using the Sybr Green system in 100% of samples analyzed. These data suggest that both PCR techniques were equally effective for detection of the genome of the parasite in the patient's blood.
Subject(s)
Leishmaniasis, Visceral/diagnosis , Real-Time Polymerase Chain Reaction/methods , DNA, Kinetoplast/genetics , Humans , Leishmaniasis, Visceral/parasitologyABSTRACT
We describe the history of a 24-year-old immunocompetent man with an expansive lesion in the brainstem that, after many misdiagnoses, was found to be caused by a Candida albicans abscess. One year after surgery and 3 months of fluconazole treatment, the patient was asymptomatic and all image and laboratory tests were normal.
ABSTRACT
In this study, we report four cases of Histoplasma capsulatum infection in eight biologists who made an expedition to determine the prevalence of this fungus in a cave localized in the northwest of Minas Gerais state, Brazil. This case report demonstrates the importance of evaluating the H. capsulatum presence in Brazilian caves before opening to public visitations.
ABSTRACT
Cruciferous vegetables, such as broccoli and watercress, have been studied extensively aiming to evaluate their chemopreventive properties. Some of them have already been established using animal models. The ITCs induce Phase II enzymes related to detoxification processes of chemical carcinogens to prevent the start of carcinogenesis. They also exhibit antitumor activity at post-initiation phase, suggesting their additional role(s) in cancer prevention. Sulforaphane is the most extensively studied isothiocyanate, focused in its anti-tumoral activity and it is mainly found in great amounts in broccoli and other cruciferous. In a dose dependent manner, ITCs inhibit the cell viability of human cervical cancer cells, human pancreatic cancer cells, human hepatocellular carcinoma cells, human ovarian cancer cells, and have antiinflammatory properties in the treatment of human T-cell leukemia cells. This protective effect may be due to improved antioxidant status. Although the health effects of diet in humans are generally considered promising, there are definite challenges and limitations of the current data in better understanding of the molecular mechanisms responsible for this effect, together with the possible interactions between different dietary constituents. The survey of relevant patents on the use of isothiocyanates such as sulforaphane for cancer and cardiovascular diseases treatments is also included in this review.