Search Results (376)

Background and Objectives: Laboratory parameters associated with nutritional and metabolic status may be substantially influenced by inflammatory activity, complicating their interpretation in patients with advanced neurological diseases. This study aimed to characterize within-patient changes in inflammatory biomarkers and laboratory parameters related to nutritional and metabolic status in multimorbid patients receiving long-term enteral nutrition, with particular emphasis on differences between infectious and clinically non-infectious periods. A secondary objective was to propose a conceptual framework for interpreting these laboratory parameters in the context of inflammatory activity. Materials and Methods: A prospective longitudinal observational case series was conducted in an inpatient palliative care facility. Of 23 patients who entered prospective follow-up, five completed the predefined nine-month observation period and were included in the final longitudinal analysis. Twenty-three predefined assessment time points were evaluated for each participant. Inflammatory parameters, serum proteins, micronutrient-related measurements, and vitamin concentrations were analyzed descriptively in relation to the clinical course of each patient. Results: Patients with recurrent infectious episodes demonstrated pronounced increases in inflammatory biomarkers accompanied by decreases in serum proteins, particularly albumin, transferrin, and total protein, as well as changes in serum iron (Fe) and zinc (Zn). The greatest fluctuations were observed in patients with the highest inflammatory burden, whereas patients without recurrent infections showed comparatively stable laboratory profiles. Vitamin A, vitamin D, and folate concentrations showed relatively limited fluctuations. Importantly, all five patients received continued enteral nutritional support, with daily energy intake ranging from 1450 to 1800 kcal, and body weight increased by 1.0–3.0 kg during follow-up. Based on these case-based observations and current evidence, a conceptual framework for interpreting laboratory parameters in the context of inflammatory activity was proposed. Conclusions: In patients with advanced neurological diseases, laboratory parameters associated with nutritional and metabolic status should be interpreted in the context of inflammatory activity and should not be used independently to determine nutritional status. The proposed conceptual framework is intended as an interpretative aid for laboratory findings rather than as an alternative diagnostic system for malnutrition and requires validation in larger prospective studies. Full article

Background: Non-infectious uveitis (NIU) is an inflammatory ocular condition often associated with systemic immune-mediated disorders. The pathogenesis of NIU is complex and involves genes from the Human Leukocyte Antigen (HLA) class I system. Endoplasmic reticulum aminopeptidase 1 (ERAP1) cleaves antigenic peptides to an optimal length for loading and presentation on HLA I molecules. In this systematic review, we aimed to summarize the current knowledge regarding how susceptibility to developing NIU is linked to single-nucleotide polymorphisms (SNPs) in the ERAP1 gene, a locus well known for its connection to autoinflammatory disorders. Methods: We performed a PRISMA protocol-guided systematic review in various databases. Results: Out of the 311 screened studies, only 12 met the inclusion criteria. Five of them are Genome-Wide Association Studies. We found relevant research for three types of NIU: acute anterior uveitis, Behçet’s uveitis, and birdshot chorioretinopathy. A frequently observed association between ERAP1 SNPs and disease risk was noted when analyzing the HLA-carriers (B*27, B*51, A*29), which points toward an epistatic interaction between ERAP1 and the MHC class I complex. Conclusions: Our findings suggest that ERAP1 gene variation is understudied in relation to NIU. A significant amount of research has also concentrated on risk stratification, and reliable markers for patient selection are needed, especially in view of emerging anti-ERAP therapies. Full article

Atrial cardiomyopathy (ACM) is defined as any complex of structural, architectural, functional, electrophysiological, and molecular changes affecting the atria that may result in clinically significant health consequences. ACM can be caused by a variety of factors, including age-related changes, valvular or vascular disease, genetic diseases, congestive heart failure, metabolic diseases, cardiovascular disease (CVD) risk factors such as arterial hypertension (AH) or obesity, obstructive sleep apnea, and other infectious or noninfectious diseases predisposing to chronic inflammation. The diagnosis of ACM relies on several modalities, including electrocardiography, echocardiography, cardiac magnetic resonance imaging (MRI), computed tomography (CT), electroanatomical mapping (EAM), genetic studies, and biomarkers, which can detect and characterize structural, mechanical, and electrical atrial dysfunction. These changes often include structural atrial remodeling (fibrosis), abnormal structure of the atrial wall and its components, and contractile and electrical dysfunctions. When assessing aspects of ACM, structural changes in the atria such as left atrium (LA) size and fibrosis; LA architectural changes such as the expression of remodeling; changes in LA mechanics such as echocardiographic stress indices; changes in reservoir function and changes in contraction; biological factors determining changes in biomarkers; possible genetic predispositions and higher expression of certain genes encoding certain proteins; and arrhythmogenic factors associated with a higher risk of atrial fibrillation (AF) and stroke and a worse short- and long-term prognosis are very important. When considering the challenges of diagnosing ACM, it should be noted that without standardized diagnostics, most ACM diagnostic situations remain primarily research tools rather than practical clinical diagnostic methods. This review critically evaluates the evidence and translational gaps in the diagnosis of ACM, synthesizing the emerging role of advanced diagnostics and their clinical and prognostic implications as a key future tool for individual risk stratification. Full article

Distinguishing tuberculous spondylodiscitis (TS) from pyogenic spondylodiscitis (PS) on imaging remains clinically important because the two entities differ in antimicrobial strategy, surgical timing, and the urgency of microbiological workup, yet no single imaging sign is pathognomonic for either diagnosis. When tissue confirmation is delayed, imaging becomes the primary tool that shifts pre-test probability and guides biopsy strategy—a role that is particularly critical in culture-negative disease, in antibiotic-exposed patients, and in settings with limited access to rapid mycobacterial diagnostics. This narrative review maps the imaging evidence most useful for routine TS-versus-PS differentiation, synthesizes recent prediction models and quantitative tools, and evaluates the adjunctive role of ¹⁸F-FDG PET/CT in biopsy-oriented decision-making. PubMed/MEDLINE was searched from inception to 31 March 2026 using pre-specified search terms across three concept blocks. This was a narrative review without formal pooling of quantitative estimates or prospective protocol registration. A secondary Scopus search identified no additional eligible records. Priority was given to comparative TS-versus-PS imaging cohorts, meta-analyses of MRI and CT features, prediction-model studies, and the recent radiomics and PET/CT literature; radiomics and deep-learning studies are appraised as a distinct evidence tier given their retrospective single-center derivation and predominantly internal-only validation. In total, 14 comparative TS-versus-PS imaging cohorts, three meta-analyses, and 14 prediction-model, quantitative-scoring, radiomics, or deep-learning studies formed the core evidence base. MRI yields the greatest differentiating information and is the recommended first-line modality. Features favoring TS include thoracic predominance, multilevel or non-contiguous involvement, relative early disc preservation, subligamentous spread, intraosseous abscesses, thin-walled paravertebral collections, and severe vertebral collapse with kyphotic deformity. Features favoring PS include lumbar predominance, early disc-endplate destruction, homogeneous inflammatory enhancement, facet-joint arthritis, and epidural phlegmon. CT adds osseous detail—including sequestra, subligamentous bone erosion, and paravertebral calcification—and remains the primary guidance modality for biopsy. Recent prediction models combining imaging and laboratory variables have shown high discriminative performance in derivation cohorts; however, most remain internally validated and locally calibrated. Pending external validation, none should be considered ready for unmodified clinical adoption, and they should therefore be used as probability modifiers rather than stand-alone diagnostic rules. ¹⁸F-FDG PET/CT is complementary rather than primary and is most useful when MRI is contraindicated or equivocal, in hardware-associated infection, and for whole-body staging in suspected disseminated tuberculosis. Non-infectious mimics and alternative infectious diagnoses should be considered when the imaging pattern is internally inconsistent or when standard cultures remain negative. Imaging findings should refine etiological probability and guide biopsy strategy, but should not replace tissue confirmation when tissue is feasible. Non-imaging evidence complementing this review is addressed in the companion manuscript, Part I. Full article

Background: Although diagnosis of pulmonary nontuberculous mycobacterial (NTM) disease typically relies on bronchoscopic sampling, microbiologic testing, and histopathologic evaluation, definitive diagnosis is often difficult. This study investigated whether cytokine profiling of bronchoalveolar lavage (BAL) fluid can help differentiate NTM pulmonary disease (NTM-PD) from other diffuse lung disorders. Methods: From January 2023 to July 2025, we prospectively evaluated 50 patients presenting with undiagnosed micronodular or diffuse pulmonary opacities. BAL fluid was collected during bronchoscopy and analyzed for cytokines and related biomarkers. Eleven patients were clinically diagnosed with NTM-PD based on American Thoracic Society/Infectious Diseases Society of America criteria. Cytokine and cellular profiles were compared between patients with NTM-PD and those with other diffuse lung disorders. A secondary analysis compared infectious granulomatous disease (NTM-PD; n = 11) with noninfectious granulomatous disease (e.g., sarcoidosis; n = 8). Results: BAL fluid from patients with NTM-PD showed significantly higher levels of interleukin-8 (IL-8), interleukin-13 (IL-13), YKL-40, and neutrophils compared to those with other diffuse lung disorders. In subgroup analysis, IL-8, IL-13, and neutrophil proportions remained significantly elevated in infectious granulomatous NTM-PD compared with noninfectious granulomatous disorders. Conclusions: NTM-PD is associated with a distinct BAL cytokine profile characterized by elevated IL-8, IL-13, YKL-40, and neutrophil proportions. Full article

Background: Fever of unknown origin (FUO) remains a major diagnostic challenge due to its heterogeneous etiologies and nonspecific clinical manifestations. Although metagenomic next-generation sequencing (mNGS) represents a promising diagnostic tool, its clinical utility in adult patients with FUO remains incompletely characterized. Methods: In this study, we retrospectively analyzed adult FUO patients who underwent mNGS testing at Peking Union Medical College Hospital between March 2022 and April 2024. Clinically meaningful diagnostic contribution was determined according to the final clinical diagnosis following multidisciplinary adjudication. Diagnostic performance, pathogen spectrum, therapeutic impact, specimen type, and predictors of clinically meaningful mNGS results were evaluated. Results: A total of 127 FUO patients were included in the study. Infectious diseases accounted for 53.5% of final diagnoses, followed by noninfectious inflammatory diseases (11.0%), malignancies (10.2%), and undiagnosed conditions (19.7%). mNGS made a clinically meaningful diagnostic contribution in 31.5% (40/127) of patients, despite an overall positivity rate of 56.7% (72/127), and showed a higher sensitivity than conventional culture for infectious etiologies (69.1% vs. 16.9%), though with a lower specificity (57.6% vs. 96.0%). Diagnostic contribution varied significantly by specimen type, with drainage fluid/abscess samples showing the highest diagnostic yield (90.9%). Lower white blood cell count was independently associated with clinically meaningful mNGS results (OR 0.87, 95% CI 0.77–0.98). Conclusions: mNGS provides clinically meaningful diagnostic value in adult patients with FUO, particularly for identifying occult infectious etiologies. Lesion-directed sampling, whenever feasible, and careful interpretation of sequencing results in the clinical context are essential to maximize the diagnostic utility of this approach. A lower white blood cell count was independently associated with clinically meaningful mNGS results, although this finding requires validation in larger prospective studies. Full article

Objective: This study aimed to assess the relationship between excess epicardial fat and the patterns of perioperative myocardial remodeling in patients undergoing surgical correction of mitral valve (MV) disease. Methods: A total of 148 patients with acquired non-infectious MV disease scheduled for surgical correction under cardiopulmonary bypass were screened in this prospective observational non-randomized study. Preoperative computed tomography (CT) of the heart was performed to assess epicardial adipose tissue (EAT) volume. Transthoracic echocardiography (Echo), including evaluation of left ventricular (LV) global longitudinal strain (GLS), right ventricular (RV) free-wall longitudinal strain, and RV systolic function (3D Echo), was conducted preoperatively, as well as postoperatively during one year after surgery. Analysis of postoperative myocardial remodeling and complications within one year after surgery was performed. Patients were divided into groups before surgical correction of MV based on the (1) EAT volume, associated with atrial fibrillation (AF) presence (EAT volume less than or > 115.1 cm³ by CT), and (2) EAT volume, associated with the presence of at least three metabolic factors (EAT volume less than or ≥100.6 cm³). Results: Prior to MV correction, Echo showed that patients with EAT volume > 115.1 cm³ exhibited larger left and right atrial (LA/RA) volumes and more pronounced RV systolic dysfunction. An EAT volume of >115.1 cm³ was associated with a 4.6-fold increase in the odds of detecting a preoperative TAPSE value < 1.7 cm (OR: 4.6 [95% CI: 1.2543; 16.7481]; p = 0.02). In the early postoperative period, patients with EAT volume > 115.1 cm³ exhibited larger RA dimensions and higher RV end-systolic volumes, as well as impaired RV–pulmonary artery coupling. At the one-year follow-up, patients with EAT volume > 115.1 cm³ exhibited larger indexed atrial volumes and basal RV dimensions. By the one-year follow-up, the group with EAT volume ≤ 115.1 cm³ was characterized by dynamic improvements, including a 10.7% increase in LV GLS (p = 0.02), a 33.6% reduction in the indexed LA volume (p = 0.004), a 28% reduction in the LV mass index (p = 0.003), and a 10.3% reduction in the LV end-diastolic dimension (p = 0.01). Furthermore, this group exhibited a 15% increase in LV stroke volume (p = 0.009), a 17.6% increase in TAPSE (p = 0.02), and a 6.5% increase in RV ejection fraction (p = 0.04) (3D Echo), none of which were observed in the group with EAT volume > 115.1 cm³. Patients with EAT volume ≥100.6 cm³ had more pronounced impairment of LV GLS before and one month after surgery compared with those with EAT < 100.6 cm³ (p = 0.046; p = 0.045). One month after surgery, worsening of RV GLS was observed specifically in the group with EAT ≥ 100.6 cm³ (p = 0.031). By the one-year follow-up, significant improvement in RV systolic function was observed only in the group with EAT volume < 100.6 cm³. Conclusions: The presence of excess epicardial fat (verified by cardiac CT) in cardiac surgery patients with acquired MV disease is associated with less favorable preoperative remodeling of both the left and right cardiac chambers and impaired reverse myocardial remodeling within one year post-surgery. Further studies in larger, independent cohorts are needed to confirm the prognostic and clinical relevance of the EAT cut-off in patients with mitral valve disease. Full article

Diabetes mellitus (DM) may influence susceptibility to ocular infection and inflammatory phenotypes; however, disease-specific DM prevalence across uveitis etiologies has not been well characterized. We evaluated DM prevalence across major uveitis entities in a large single-center cohort and compared observed prevalence estimates with age-specific national estimates from Japan. A total of 3163 adult patients out of 4751 patients newly diagnosed with uveitis at the Tokyo Medical University Hospital between 2004 and 2023 were included in the final analysis after excluding patients without DM-related laboratory parameters, those who declined to participate through the institutional opt-out procedure, pediatric patients, and patients with diabetic iritis. DM was defined as an HbA1c level of 6.5% or higher, current treatment for DM, or a random blood glucose level of 200 mg/dL or higher. For reference comparisons with national data, we analyzed seven adult entities with sufficient sample size. For each entity, expected DM prevalence was based on the corresponding national age-stratum estimate according to the mean age category of that entity. Observed and expected DM prevalence estimates were compared using Fisher’s exact test. Multivariable modified Poisson regression with robust variance estimation was used to estimate adjusted prevalence ratios (aPRs) for DM prevalence, including age, sex, and individual uveitis entities as covariates. Overall, 166 of 3163 patients (5.3%) had DM. DM prevalence differed substantially by etiology, ranging from 18.0% in endophthalmitis to 1.8% in Behçet disease. Selected infectious uveitis showed a higher crude prevalence of DM than selected noninfectious uveitis (10.4% vs. 3.8%; p < 0.01). In comparisons with corresponding national age-stratum estimates, the observed prevalence estimates of DM in endophthalmitis and herpetic iritis did not differ significantly from the expected national prevalence estimates, whereas acute retinal necrosis, intraocular lymphoma, sarcoidosis, and Behçet disease showed a significantly lower prevalence than expected. In the multivariable analysis, older age (aPR per 10-year increase, 1.49; 95% CI, 1.33–1.67; p < 0.001) and male sex (aPR, 2.11; 95% CI, 1.40–3.19; p < 0.001) were independently associated with DM prevalence, whereas individual uveitis entities were not significantly associated after adjustment for covariates. DM prevalence in uveitis is heterogeneous and disease-specific. Although selected infectious uveitis showed a higher crude prevalence of DM, comparisons with corresponding national age-stratum estimates suggested that some of the differences reflected the underlying age structure. Older age and male sex were the strongest independent correlates of DM prevalence. Full article

Background/objectives: Catheter-related bloodstream infection (CRBSI) is a frequent and morbid complication in catheter-dependent maintenance hemodialysis, and rapid risk stratification is needed while awaiting cultures. This study aimed to evaluate admission complete blood count-derived indices—neutrophil-to-lymphocyte ratio (NLR), lymphocyte-to-monocyte ratio (LMR), platelet-to-lymphocyte ratio (PLR), systemic immune-inflammation index (SII), and pan-immune-inflammation value (PIV)—for identifying CRBSI. Methods: This single-center retrospective study (1 January 2011–31 October 2024) included adult catheter-dependent hemodialysis patients classified as CRBSI or controls. CRBSI required compatible clinical findings and concordant growth of the same microorganism(s) in paired simultaneous catheter and peripheral blood cultures. Controls were hospitalized for non-infectious reasons without infection during the index admission. Indices were calculated from admission blood counts. Discrimination was assessed using ROC analysis, and adjusted associations were evaluated using multivariable logistic regression. Results: Among 286 patients (147 CRBSI, 139 controls), CRBSI cases had higher NLR, SII, and PIV and lower LMR; PLR did not differ. NLR showed the numerically highest discriminatory performance among the evaluated indices (AUC 0.737; cut-off 5.96; sensitivity 68.7%, specificity 68.3%; p < 0.001). SII (cut-off 1189.21; AUC 0.693) and PIV (cut-off 821.62; AUC 0.686) had moderate discrimination, and LMR was modest (cut-off 1.65; AUC 0.642); PLR was not discriminatory (AUC 0.559; p = 0.086). In models adjusted for age, sex, hypertension, and cardiovascular disease, NLR remained associated with CRBSI (OR 1.159; p < 0.001), together with hypertension (OR 2.441; p = 0.017) and cardiovascular disease (OR 2.626; p < 0.001). Conclusions: Admission hematologic inflammation indices, particularly NLR, showed moderate ability to discriminate CRBSI from non-infectious admissions in catheter-dependent hemodialysis patients and may provide rapid adjunctive information while awaiting microbiological confirmation. Full article

Background: Healthcare-associated bloodstream infections (HABSIs) are among the main categories of nosocomial infections. This analysis aims to identify the clinical characteristics of patients in the emergency department (ED) who will develop a HABSI during their hospital stay. Methods: Main outcome measures were HABSI and the cumulative survival rate at 30 days. The features tested in a logistic model were age, sex, vitals by the National Early Warning Score (NEWS), priority levels, main complaints, comorbidities by the Charlson Comorbidity Index (CCI), trauma-related disease, main diagnosis and ED length of stay. Results: In 414 (2.3%) out of 18,304 patients, aged 75 (16) years, mean (SD), a diagnosis of HABSI was recorded. HABSIs occurred in subjects with main diagnosis of diseases of the respiratory system (N = 116; 28.0%), digestive system (N = 72; 17.4%), and circulatory system (N = 68; 16.4%). The main key clinical features selected by the logistic model were: NEWS > 6, diagnosis of neoplasms, CCI > 4, and diagnosis of diseases of the digestive system. The ROC curve for the HABSI risk score was 0.703 ± 0.027 in predicting the outcome, (sensitivity 79%, specificity 51%, at optimal cut-off score). The overall hazard mortality risk was twofold higher in patients with HABSIs (hazard ratio: 2.319; 95% confidence interval: 1.871–2.875; p-value: <0.001). The overall 30-day survival rate was lower among patients with HABSIs (33%) vs. non-HABSI patients (62%). Conclusions: A group of main clinical features in subjects without suspect of infectious disease in the ED are associated with HABSIs. These features negatively impact survival rate during hospital stays. Full article

The complement system is a key link between innate and adaptive immunity, contributing to pathogen elimination, immune regulation, and tissue homeostasis. Its activation is not only crucial in infections, such as COVID-19, but also plays a major role in the pathomechanism of several non-infectious respiratory diseases, such as asthma, COPD, sarcoidosis and lung cancer. Complement components can modulate the quality of the adaptive immune responses, including through the regulation of T2 immunity and eosinophilic inflammation, thereby linking natural defense to complex immune processes. In recent years, it has become increasingly clear that dysregulated complement activity contributes to inflammation, thrombosis and tissue damage in a wide range of respiratory diseases. The study of the various components of this cascade system may therefore be promising from both a diagnostic and therapeutic point of view. Some of its components may serve as biomarkers for distinguishing between different phenotypes of certain lung diseases, while their targeted inhibition or modulation may open the way towards new treatment options. A better understanding of the complement system’s integrative and regulatory role not only allows for a deeper insight into immunological interactions but may also bring us closer to phenotype-oriented, immunology-based pulmonology, which may have real clinical benefits in the future. Full article

Local immune cell activation and vascular remodelling are characteristic pathogenic features of pulmonary arterial hypertension (PAH). HIV and schistosome infections have been individually associated with PAH. However, whether co-infection with these pathogens has a distinct impact on the development of pulmonary vascular disease remains poorly understood, partly due to the lack of experimental animal models. In a novel non-infectious model of HIV and Schistosoma pulmonary co-exposure based on lung embolisation of S. mansoni eggs in HIV-transgenic (HIV) mice, we previously reported exacerbated endothelial remodelling and dysfunction, along with increased pulmonary arterial pressure; which were associated with a unique profile of pro-inflammatory cytokines in the lung. In the present study, we used flow cytometric analysis of isolated lung leukocytes and immunofluorescence staining to characterise the pulmonary immune cell landscape associated with individual or combined exposure to HIV and schistosome. Compared with mice exposed to HIV (untreated HIV mice) or schistosome (egg-treated wild-type mice), co-exposed (egg-treated HIV mice) animals showed significantly increased numbers of interstitial and alveolar macrophages, patrolling-type monocytes, NKT and γδ T cells, and reduced CD8⁺ αβ T cells. Other lung immune cells, including inflammatory-type monocytes, eosinophils/neutrophils, dendritic cells, CD4⁺ αβ T cells, NK cells and B cells were not significantly affected in the co-exposure condition. Taken together, these results show for the first time that combined pulmonary exposure to HIV and Schistosoma, as it may occur in co-infected individuals, alters the local immune cell landscape in a manner distinct from that of individual exposure. Furthermore, these findings may contribute to a better understanding of the complex inflammatory processes involved in the pathogenesis of PAH, thereby supporting the development of therapies targeting pathogenic immune cells in pulmonary vascular disease associated with HIV and Schistosoma co-morbidity. Full article

White dot syndromes (WDS), a purely descriptive term formulated in 1995, grouped inflammatory conditions primarily affecting the choroid that shared the clinical appearance of white dots on fundus examination, thus presenting similar white dots on fundus examination, which, however, had diverse pathophysiologies and different primary localizations of inflammation that did not warrant their association. Some of them, including birdshot retinochoroiditis (BRC) and sympathetic ophthalmia, produced predominant inflammation in the choroidal stroma, while others, including multiple evanescent white dot syndrome (MEWDS) and acute posterior multifocal placoid pigment epitheliopathy (APMPPE), caused inflammatory non-perfusion of the choriocapillaris. The aim of this review is to build upon a pathophysiology-based classification framework that groups these entities under the broader concept of non-infectious choroiditis and subdivides them according to their predominant inflammatory mechanisms and anatomical localization. This framework distinguishes diseases characterized by inflammatory occlusion of the choriocapillaris with consequent perfusion dysfunction, including MEWDS, APMPPE, multifocal choroiditis (MFC), and serpiginous choroiditis, from conditions primarily involving inflammatory infiltration of the choroidal stroma, such as birdshot retinochoroiditis (BRC), Vogt-Koyanagi-Harada disease (VKH), and sympathetic ophthalmia. Moreover, the review raises the question of how such an inadequate term, based on unscientific assumptions, could gain such quick access to ophthalmic practice and literature, be immediately adopted in major textbooks, and persist for so long being unrecognized as a misnomer. The slow and progressive reevaluation of this nomenclature over more than two decades is related and contextualized. Full article

Background/Objective: Leishmaniasis is a neglected tropical disease caused by species of the genus Leishmania, with a broad clinical spectrum that can overlap with other infectious and non-infectious conditions. Accurate species identification is critical for appropriate treatment and prognosis; however, parasitological methods are limited by suboptimal sensitivity, specificity, and inability to reliably differentiate species. This study aimed to develop and validate a real-time PCR assay based on melting-curve analysis (Leish-qPCR) for the detection of Leishmania spp. and the differentiation of Leishmania (Leishmania) amazonensis and Leishmania (Viannia) braziliensis. Methods and Results: Genus-specific primers were designed based on the kDNA (kinetoplast DNA) minicircle consensus sequences of Leishmania species, while species-specific primers targeted the internal transcribed spacer 2 (ITS2) consensus regions of the ribosomal RNA locus of L. (L.) amazonensis and L. (V.) braziliensis. Analytical performance was evaluated in silico and in vitro using a panel of protozoa, fungi, and bacteria, exhibiting 100% specificity with no cross-amplification. The limit of detection was one copy per reaction for all targets using positive controls. Clinical validation was performed using skin biopsy specimens from patients with granulomatous lesions. The optimized Leish-qPCR assay, performed in separate reaction tubes within the same run, demonstrated reliable analytical specificity and sensitivity, with distinct and reproducible melting temperature (Tm) peaks across plasmid controls, parasite DNA, and clinical samples. Comparative analysis with histopathological examination demonstrated moderate agreement between the methods, supporting the applicability of the assay for sensitive detection and species-level discrimination of Leishmania spp. in clinical samples. Conclusions: The Leish-qPCR assay presented high sensitivity, specificity, and diagnostic accuracy, representing a promising tool for routine diagnosis of leishmaniasis and for the differentiation of L. (L.) amazonensis and L. (V.) braziliensis in clinical samples. Full article