Whole-blood gene expression has the potential to become an affordable alternative for the diagnosis of pulmonary tuberculosis (TB). Through an immune-based gene expression profile via the NanoString platform, we previously identified 12 genes from whole blood that distinguished active TB from other pulmonary diseases. Here, we performed a three-phase diagnostic test evaluation to determine the scientific validity and analytical and clinical performance of these 12 previously identified genes in accurately diagnosing active TB. We further employed binary classification machine learning models to assess and evaluate transcriptional signatures. Samples from 175 adults were included in the study. During phase one, we assessed the expression of 12 genes in 10 TB patients and 9 healthy controls. Elevated expression of FCGR1A, GBP5, PDCD1LG2, and IRAK3 was observed in TB patients compared with healthy controls (p value < 0.05). The analytical performance of these four genes was evaluated in a cohort of 100 symptomatic respiratory volunteers screened for TB, comprising 35 tuberculosis patients and 65 non-tuberculosis controls. The expression of FCGR1A, PDCD1LG2, and GBP5 consistently distinguished patients from both groups, with an area under the ROC curve (AUC) ≥ 0.75. These genes were combined into three transcriptional signatures (FCGB-2TB, FPG-3TB, and FCPD-2TB), which demonstrated good performance in a pilot clinical validation study phase (N = 56; sensitivity > 77%, specificity > 94%). The present analysis allowed the discovery of RT‒qPCR-based signatures for TB diagnosis. These signatures could serve as affordable alternatives for TB diagnosis at primary care centers.

The World Health Organization (WHO) has highlighted the need for new diagnostic tests for pulmonary tuberculosis (TB) that use easily obtainable samples, such as blood, to provide rapid and affordable results suitable for primary healthcare settings. To address this, we evaluated the diagnostic potential of 34 markers quantified by Luminex in supernatants of whole-blood cultures, either stimulated or not with an apolar lipid extract from Mycobacterium tuberculosis (Mtb). The study included 20 patients with pulmonary TB and 20 symptomatic respiratory (SR) non-TB individuals. In unstimulated cultures, eight biomarkers (IL-18, IL-1RA, IL-1β, IL-8, IP-10, MIP-1β, SDF-1α, and TNF-α) differentiated TB patients from SR - non-TB patients, with areas under the ROC curve (AUC) ranging from 0.71 to 0.82. Combinatorial analyses with four-marker panels, namely, IL-18 + IL-1β, IL-1RA + IL-18, IL-18 + IL-1β + IL-1RA and IL-18 + IL-1β + IL-1RA + MIP-1β, revealed AUCs of 0.84-0.90, specificities above 90 % and sensitivities between 70 % and 75 %. The addition of the lipid extract to the whole-blood culture did not improve the discriminatory power of the panels. Validation of the IL-1RA + IL-18 combination by ELISA in an independent group (21 TB patients and 33 SR patients) yielded an AUC of 0.76, a sensitivity of 62 %, a specificity of 88 %, and an accuracy of 78 %. The collective elevation of these cytokines suggests an interplay between pro- and anti-inflammatory pathways in the host response. Although the selected biomarker panels showed moderate diagnostic performance in the ELISA test, other combinations may be useful in helping to predict TB progression or monitor treatment outcomes.

Early diagnosis plays a pivotal role in breaking the epidemiological chain of Mycobacterium leprae transmission. Currently, diagnosis relies on clinical, dermato-neurological features, and histological/microbiological assessments. This prospective cross-sectional study investigated whether IgA, IgM, and IgG anti-lipid antibodies can be used to improve the diagnostic performance for leprosy-affected patients in a high-burden municipality in Brazil. Serum samples from 91 volunteers, including patients with leprosy (n=62), household contacts (n=21), and endemic controls (n=8) were screened by enzyme-linked immunosorbent assays (ELISA) for IgA, IgM, and total IgG against four lipids-namely, cardiolipin (CL), phosphatidylcholine (PTC), phosphatidylethanolamine (PE), and phosphatidylinositol (PI)-and a glycosphingolipid-sulfatide (SL)-found in the bacterial cell wall. Antibodies against all lipids were detected in the sera of patients with leprosy. Significantly higher levels of IgA anti-CL, anti-PE, and anti-PTC, IgM anti-CL, and total IgG anti-PTC were observed in these patients compared to household contacts and endemic controls (p < 0.0001). ROC curve analyses demonstrated high accuracy in discriminating patients with leprosy from the contacts, with moderate to high sensitivity and specificity, even in paucibacillary patients. Despite the small study population and the absence of patients with other dermatological lesions for differential diagnosis, these findings suggest the potential of anti-lipid antibodies as biomarkers for leprosy detection. This approach offers a promising method to improve early diagnosis in high-burden areas, such as the studied municipality in Brazil.

The capacity of Mycobacterium tuberculosis (Mtb) to establish long-term survival is attributed to its ability to subvert host defense mechanisms, especially macrophages. Although Mtb lipids are believed to play a role in this host-pathogen crosstalk, how mycobacterial lipids drive this complex interaction is poorly characterized. Here, we cultured macrophages with nonpolar cell wall Mtb lipids and applied high-throughput expression profiling (RNA sequencing), mass spectrometry-based targeted eicosanoid, and untargeted lipidomics analysis. This system-level analysis revealed that Mtb nonpolar lipid triggered the expression of phenotypic markers for classically and alternatively activated macrophages, a state previously referred as immunoregulatory. Specifically, under lipid stimulation, macrophages expressed high levels of proinflammatory markers, activated components of the interleukin-1 family, underwent an imbalance in lipid metabolism, and shifted the eicosanoid synthesis pathway toward the prostaglandin axis. Taken together, these results suggest an intricate mechanism of Mtb-driven macrophage immunomodulation that may favor its long-term survival.

This study assessed the diagnostic potential of nonpolar lipid extracts in enzyme-linked immunosorbent assays (ELISAs) for tuberculosis (TB) serodiagnosis. Nonpolar lipid extracts were harvested from Mycobacterium tuberculosis (Mtb) knockout in mce1 operon (∆mce1) and its parental wild type (WT) strains. IgM and IgG anti-nonpolar lipid serum levels were measured in TB patients (n=45), healthy individuals with positive (n=22) and negative (n=44) interferon-gamma release assay (IGRA) results, and symptomatic respiratory (SR) patients with negative TB tests (n=9). IgG anti-WT lipid distinguished TB patients from IGRA-positive individuals with 60% sensitivity and 77.3% specificity. Conversely, IgG anti-∆mce lipid levels didn't vary significantly across groups. Interestingly, most SR patients exhibited significantly higher IgM and IgG anti-WT lipid titers than the IGRA-positive and -nega groups. While the overall diagnostic potential of Mtb nonpolar lipids was limited, the impaired immunogenecity of Δmce1 lipid extract suggests that some missing lipid classes in this extract can potentially induce antibody production in TB patients.

A 22-year-old woman presented with an ulcer on her right earlobe 2 months ago, with inflammation and enlarged ipsilateral lymph nodes in her neck. She was treated with antibiotics without success and then was referred to an infectious disease specialist. She has a cat at home with sporotrichosis, but without direct contact with the lesion, she did not remember any scratching by the cat. She also mentioned wearing a semi-jewel earring. This is a rare and unusual case of sporotrichosis in the earlobe, probably caused by wearing an earring contaminated by the cat's fungus that was present in the home environment. The delay in diagnosis and treatment led to the worsening of the injury and loss of the earlobe.

Diabetes increases susceptibility to infections, including Leishmania braziliensis (Lb). Our group previously demonstrated that diabetic patients with cutaneous leishmaniasis (CL) take longer to heal lesions compared to non-diabetics. Since macrophages play a critical role in CL pathogenesis, we investigated how high glucose levels impact their response during Lb infection. Macrophages cultured in high glucose conditions showed increased parasite load than those in normal glucose conditions. The production of inflammatory mediators was similar between glucose conditions, but basal reactive oxygen species (ROS) production was elevated under high glucose conditions and remained unchanged after Lb infection, indicating glucose-induced oxidative stress does not control the parasite. In contrast, macrophages in normal glucose conditions, exhibited increased ROS production only after infection. Additionally, high glucose reduced TLR2 and TLR4 expression, which was also observed after Lb infection. TLR2/4 inhibition increased Lb infection in normal glucose conditions, mediated by TLR-dependent ROS production. However, this mechanism was absent under high glucose conditions, where elevated basal ROS production appeared TLR-independent. Biopsies from diabetic CL patients corroborated these findings, showing decreased TLR2 and TLR4 expression compared to non-diabetics. These findings suggest that high glucose levels induce oxidative stress and reduces TLR expression, impairing macrophage functions and rendering them less effective at controlling Lb infection.

IntroductionThe attenuation of BCG has led to the loss of not only immunogenic proteins but also lipid antigens.MethodsThus, we compared the macrophage and T-cell responses to nonpolar lipid extracts harvested from BCG and Mycobacterium tuberculosis (Mtb) to better understand the role of BCG lipids in the already known diminished responses of the vaccine strain.ResultsRelative to Mtb, nonpolar lipid extract from BCG presented a reduced capacity to trigger the expression of the genes encoding TNF, IL-1b, IL-6 and IL-10 in RAW 264.7 macrophages. Immunophenotyping of PBMCs isolated from healthy individuals revealed that lipids from both BCG and Mtb were able to induce an increased frequency of CD4+ and CD8+ T cells, but only the lipid extract from Mtb enhanced the frequency of CD4-CD8-double-negative, γσ+, CD4+HLA-DR+, and γσ+HLA-DR+ T cells relative to the nonstimulated control. Interestingly, only the Mtb lipid extract was able to increase the frequency of CD4+ memory (CD45RO+) T cells, whereas the BCG lipid extract induced a diminished frequency of CD4+ central memory (CD45RO+CCR7-) T cells after 48 h of culture compared to Mtb.DiscussionThese findings show that the nonpolar lipids of the BCG bacilli presented diminished ability to trigger both proinflammatory and memory responses and suggest a potential use of Mtb lipids as adjuvants to increase the BCG vaccine efficacy.

Cutaneous leishmaniasis (CL), caused by Leishmania braziliensis, is closely associated with a severe form of the disease, indicated by a positive Leishmania skin test (LST) that assesses and reflects the presence of immune T cells specific to Leishmania antigens. In this study, we compare the clinical, immunologic, and histopathologic features between Leishmania skin test-positive (LST+) and Leishmania skin test-negative (LST-) in CL. Compared to LST+ patients, LST- patients had larger lesions and had been sicker for longer, presented with more instances of therapeutic failure with meglumine antimonate, (MA) and the healing times were higher than LST+. While granulomas were less frequent and the parasite load was higher in LST-, there were more CD8+ T cells and an enhanced production of Granzyme B in the supernatants of biopsies from LST- subjects. This study shows that in LST-, an impairment in Th1 immune response is associated with a high parasite burden, and the pathology is mediated by CD8+ T cells and the enhanced production of Granzyme B. The abnormalities in the immunologic response in LST- patients lead to a more severe disease with a high rate of failure to therapy.

Leishmaniasis and leprosy comprise the group of granulomatous diseases. Although both diseases have a known occurrence in Brazil, their concurrent presence is rare, and few cases have been reported in the literature. We report the case of a 46-year-old male patient co-infected with leprosy and mucosal leishmaniasis. Examination revealed an ulcerated lesion on the nose and a lesion with a granulomatous surface on the palate, in addition to spots on the arms and legs. After diagnosis, the patient was treated for co-infection and has been under follow-up for 2 years without signs of recurrence. The diagnosis of leishmaniasis/leprosy co-infection is challenging because of the broad clinical spectrum. Few cases have been reported in the literature and the dentist may play an important role in its detection.

Hansen’s disease (HD) is an infectious, treatable, and chronic disease. It is the main cause of infectious peripheral neuropathy. Due to the current limitations of laboratory tests for the diagnosis of HD, early identification of infected contacts is an important factor that would allow us to control the magnitude of this disease in terms of world public health. Thus, a cross-sectional study was conducted in the Brazilian southeast with the objective of evaluating humoral immunity and describing the accuracy of the immunoassay based on IgA, IgM, and IgG antibodies against surface protein Mce1A of Mycobacterium, the predictive potential of these molecules, the clinical significance of positivity, and the ability to segregate new HD cases (NC; n = 200), contacts (HHC; n = 105), and healthy endemic controls (HEC; n = 100) as compared to α-PGL-I serology. α-Mce1A levels for all tested antibodies were significantly higher in NC and HHC than in HEC (p < 0.0001). The performance of the assay using IgA and IgM antibodies was rated as highly accurate (AUC > 0.85) for screening HD patients. Among HD patients (NC), positivity was 77.5% for IgA α-Mce1A ELISA, 76.5% for IgM, and 61.5% for IgG, while α-PGL-I serology showed only 28.0% positivity. Multivariate PLS-DA showed two defined clusters for the HEC and NC groups [accuracy = 0.95 (SD = 0.008)] and the HEC and HHC groups [accuracy = 0.93 (SD = 0.011)]. IgA was the antibody most responsible for clustering HHC as compared to NC and HEC, evidencing its usefulness for host mucosal immunity and as an immunological marker in laboratory tests. IgM is the key antibody for the clustering of NC patients. Positive results with high antibody levels indicate priority for screening, new clinical and laboratory evaluations, and monitoring of contacts, mainly with antibody indexes ≥2.0. In light of recent developments, the incorporation of new diagnostic technologies permits to eliminate the main gaps in the laboratory diagnosis of HD, with the implementation of tools of greater sensitivity and accuracy while maintaining satisfactory specificity.

This case-control study compared the clinical profile, parasite load, polymerase chain reaction positivity, and response to therapy in patients with recurrent cutaneous leishmaniasis (RCL) with primary cutaneous leishmaniasis (CL). The RCL patients had milder diseases with lower parasite loads, a lower number of lesions, and more self-healing diseases than primary CL patients.

Cutaneous leishmaniasis exhibits a spectrum of clinical presentations dependent upon the parasites' persistence and host immunopathologic responses. Although cytolytic CD8 T cells cannot control the parasites, they significantly contribute to pathologic responses. In a murine model of cutaneous leishmaniasis, we previously found that NKG2D plays a role in the ability of cytolytic CD8 T cells to promote disease in leishmanial lesions. Here, we investigated whether NKG2D plays a role in human disease. We found that NKG2D and its ligands were expressed within lesions from L. braziliensis-infected patients and that IL-15 and IL-1β were factors driving NKG2D and NKG2D ligand expression, respectively. Blocking NKG2D reduced degranulation by CD8 T cells in a subset of patients. Additionally, our transcriptional analysis of patients' lesions found that patients who failed the first round of treatment exhibited higher expression of KLRK1, the gene coding for NKG2D, than those who responded to treatment. These findings suggest that NKG2D may be a promising therapeutic target for ameliorating disease severity in cutaneous leishmaniasis caused by L. braziliensis infection.

Introdução/Objetivo: A COVID-19 trouxe a necessidade da compreensão dos mecanismos de agressão desta doença. Iniciado em 2020, o projeto COVID-19 Patologia Estrutural e Molecular (COVPEM), realizado em parceria entre Instituto Gonçalo Moniz (IGM)-FIOCRUZ-BA e Instituto Couto Maia-BA (ICOM) tem por objetivo estudar os casos fatais de COVID-19 através de Autopsias Minimamente Invasivas (AMIs). Posteriormente, esta parceria foi ampliada para outras doenças infectocontagiosas e realização de biópsias. Os casos estudados foram, periodicamente, utilizados para a realização de Sessões Anatomoclínicas (SACs). O objetivo deste trabalho é descrever as SACs como atividade acadêmica da Residência Médica/internato, bem como refletir sobre a necessidade desta temática na formação médica. Métodos: Uma equipe hospitalar multidisciplinar detecta os casos fatais de COVID-19 e outras doenças infecciosas no ICOM e solicita aos familiares, após aprovação da comissão de ética em pesquisa, a permissão das AMIs e assinatura de termos de consentimentos livres e esclarecidos. No internamento, o serviço social fornece aos responsáveis uma carta de apresentação, com apoio da diretoria da unidade e da Comissão de Ensino e Pesquisa, que menciona o histórico de trabalho desenvolvido no ICOM. Posteriormente, um patologista, realiza as AMIs/biópsias para as SACs, nas quais um residente do ICOM, um patologista do IGM e médicos especialistas/pesquisadores convidados de outras instituições brasileiras apresentam a história clínica, os aspectos anatomopatológicos e uma revisão/debates do tema, respectivamente. Resultados: Foram realizadas mais de 17 AMIs, sendo 13 de casos fatais de COVID-19. Em 2021 foram apresentadas as seguintes SACs: Clinica, Imagem e Patologia em casos de COVID-19 na Bahia, Coagulopatia Associada a COVID, Lesões Musculares na COVID-19, Lesão Renal Aguda na COVID-19. Em 2022 foi apresentado o tema Insuficiência Respiratória em Tempos de SARS-COV2. Dentre as biópsias realizadas os temas foram linfadenopatia a esclarecer e lesões de pele a esclarecer. Em 2023, das biópsias realizadas, os temas discutidos, até o momento, foram Pessoas Vivendo com HIV/AIDS apresentando lesões de pele e Sarcoma de Kaposi. Conclusão: As SACs promovem um ambiente de ensino e debate clínico e anatomopatológico, proporcionando aos residentes e internos uma vivência em áreas de conhecimentos da infectologia que contribuem para a formação médica baseada em evidências científicas.

Patients with cutaneous leishmaniasis (CL) present an exacerbated inflammatory response associated with tissue damage and ulcer development. In recent years, higher rates of failure to pentavalent antimoniate therapy have been observed, yet the underlying reason remains poorly understood. We hypothesize that the eicosanoid PGE2 favours the establishment of infection by L. braziliensis, which contributes to therapeutic failure. The aim of the present study was to investigate the influence of PGE2 on the survival of L. braziliensis in macrophages and rates of therapeutic failure in CL patients. PGE2, an eicosanoid derived from the metabolism of arachidonic acid by the COX-2 enzyme, plays several roles in immune response. We found that increased PGE2 decreases the microbicidal function of macrophages and is associated with disease severity and therapeutic failure. Additionally, the neutralization of COX-2 by NS398, a selective NSAID, increases the ability of macrophages to kill L. braziliensis and protects against the pathological inflammatory response. Our data suggest that NS398 may serve as an adjunct treatment for CL patients.

A esporotricose é uma micose subaguda ou crônica causada, na maioria das vezes, pela inoculação traumática do fungo dimórfico Sporothrix schenckii. É uma das micoses subcutâneas mais comuns na América Latina, com distribuição mundial. A doença atingiu recentemente proporções epidêmicas em algumas regiões do Brasil, como no Rio de Janeiro, onde o número de casos de transmissão zoonótica por gatos infectados aumentou significativamente. A forma tradicional de transmissão, porém, é a inoculação traumática do fungo na pele, por contato com solo, plantas ou substratos orgânicos contaminados. A apresentação mais comum é a cutânea e a doença tem sido classificada em três formas clínicas diferentes: cutânea, linfangítica e disseminada. Descrevemos um caso de esporotricose em um local único (o pavilhão auricular) com um modo de transmissão incomum. A demora no diagnóstico e tratamento resultou em maior morbidade, cicatrização inestética e perda do lóbulo da orelha. O relato consiste em um paciente de 22 anos, de Salvador, Bahia, com lesão cutânea ulcerada em lóbulo da orelha direita com duração aproximada de 2 meses, evoluindo com sinais inflamatórios locais e linfadenopatia cervical ipsilateral. Ela foi atendida por um dermatologista e antibióticos foram prescritos por 10 dias, sem melhora. Ao retornar ao dermatologista, foi encaminhada para avaliação por infectologista. Posteriormente, a paciente relatou a presença de um gato doente com esporotricose em sua residência, porém sem relato de mordida, arranhadura ou lambida no local da lesão. Ela se refere ao uso de brinco, sendo a composição uma semijoia. Na análise histopatológica da biopsia da lesão observou-se granulomas com neutrófilos células dendríticas ao centro, na coloração hematoxilina-eosina. Já na coloração de ácido periódico-schiff constatou-se a presença de Sporotrix, também observada na microscopia eletrônica. Também foi realizada cultivo de fungos na biopsia, comprovando a infecção. Após o tratamento com terbinafina a paciente foi curada da infecção e apresentou cicatrização inestética do lóbulo da orelha.

Dogs play an important role in transmission of Leishmania infantum, but epidemiologic and clinical studies of canine tegumentary leishmaniasis (CTL) are scarce. In an endemic area of human American tegumentary leishmaniasis (ATL) caused by Leishmania braziliensis, we determine the prevalence and incidence of both CTL and subclinical (SC) L. braziliensis infection in dogs and evaluated if the presence of dogs with CTL or SC L. braziliensis infection is associated with the occurrence of human ATL. SC infection in healthy animals and CTL in animals with ulcers were determined by PCR on biopsied healthy skin or on ulcers or by detecting antibodies against soluble leishmania antigen. We compared the occurrence of human ATL in homes with dogs with CTL or SC infection with control homes without dogs or with dogs without CTL or SC infection. The prevalence of SC infection was 35% and of CTL 31%. The incidence of SC infection in dogs was 4.6% and of CTL 9.3%. The frequency of ATL in humans was 50% in homes with infected dogs and 13% in homes without L. braziliensis infection in dogs. CTL and SC infection is highly prevalent, and dogs may participate in the transmission chain of L. braziliensis.

Hansen’s disease (HD) is an ancient disease, but more than 200,000 new cases were reported worldwide in 2019. Currently, there are not many satisfactory immunoassay methods for its diagnosis. We evaluated antibodies against Mce1A as a promising new serological biomarker. We collected plasma from new cases, contacts, and endemic controls in the city of Parnaíba and treated patients at Carpina, a former HD colony in Piauí state, northeastern Brazil. Receiver operating characteristic (ROC) curves were used to assess the assay thresholds, specificity and sensitivity of the IgA, IgM, and IgG antibodies against α-Mce1A by indirect ELISA and compared it with IgM anti-PGL-I and molecular diagnosis by quantitative polymerase chain reaction (qPCR). Venn diagrams were generated to represent the overlap in the antibody positivity pattern. Multivariate analysis was performed to assess the potential predictor of antibodies for the outcome of having an HD diagnosis. IgA and IgG were positive in 92.3 and 84% of patients, respectively. IgM was negative for all treated patients. IgG had a sensitivity and specificity of 94.7 and 100%, respectively. IgM-positive individuals had a 3.6 chance of being diagnosed with HD [OR = 3.6 (95% CI = 1.1–11.6); p = 0.028], while IgA-positive individuals had a 2.3 chance [OR = 2.3 (95% CI = 1.2–4.3); p = 0.005] compared to endemic controls. We found that the Mce1A antibody profile can be an excellent diagnostic method of HD. IgA is an ideal biomarker for confirming contact with the bacillus. IgM has potential in the detection of active disease. IgG antibodies confirm the performance of these serological markers in diagnosis and therapeutic follow-up.

The use of platelet rich plasma (PRP) for the treatment of hair loss has increased significantly in recent years. PRP treatment involves obtaining autologous PRP from a patient's blood sample and injecting it into the scalp. Although a promising treatment for alopecia, the role of platelet and growth factor (GF) concentration in PRP on hair growth is poorly understood. Herein, we present a case series of 3 patients (A, B and C) with androgenetic alopecia (AGA) who received PRP therapy containing platelet concentrations that were 15-55% of the baseline in the blood throughout the course of their treatment. Patients received 3 treatments over a 6-month period. Platelets in blood and PRP were counted using an automated hematology analyzer. An aliquot of the PRP was subjected to platelet activation using 20 mM calcium chloride to induce GF secretion and 11 different GFs were measured using commercially available kits. Clinical response was assessed prior to each treatment by a board-certified dermatologist. Additionally, an artificial intelligence photography device, HairMetrix® D200 EVO dermatoscope, was used to quantitate hair fibers in the frontal anterior, midscalp and vertex regions. Data generated 6-12 weeks after 3rd PRP treatment was compared to baseline. Patient A had the best response with a 20-50% increase in hair density and a 20% increase in average hair width across 3 scalp regions. Patient B improved slightly, while patient C's AGA remained stable. Platelet GF secretion profiles showed no fibroblast growth factor 2 (FGF2) in all of patient A samples, whereas the platelets of patient B and C secreted FGF2 upon activation. Secretion levels of brain-derived neurotropic factor was less than 0.5-fold in patient A's PRP samples compared to patients B and C. Our data suggests that high platelet numbers in PRP samples may not be necessary for hair growth and highlights key GFs that may have a negative effect on clinical response.