Diagnosis Of Leishmaniasis Research Articles

Evaluation of Loopamp Leishmania detection kit for the diagnosis of cutaneous leishmaniasis in Ethiopia

BackgroundCutaneous leishmaniasis (CL) in Ethiopia and some parts of Kenya is predominantly caused by Leishmania aethiopica. While skin-slit (SS) microscopy is routinely used for CL diagnosis, more sensitive molecular tests are available. The Loopamp™ Leishmania detection kit (Loopamp) is a robust loop-mediated isothermal amplification (LAMP) assay with the potential for implementation in primary healthcare facilities. In this study, we comparatively assessed the diagnostic accuracy of four methods currently used to diagnose CL: Loopamp, kinetoplast DNA (kDNA) PCR, spliced leader RNA (SL-RNA) PCR and SS microscopy.MethodsA study on 122 stored tape disc samples of suspected CL patients was conducted in Gondar, northwestern Ethiopia. Routine SS microscopy results were obtained from all patients. Total nucleic acids were extracted from the tapes and subjected to PCR testing targeting kDNA and SL-RNA, and Loopamp. Diagnostic accuracy was calculated with SS microscopy as a reference test. The limit of detection (LoD) of Loopamp and kDNA PCR were determined for cultured L. aethiopica and Leishmania donovani.ResultsOf the 122 patients, 64 (52.5%) were identified as CL cases based on SS microscopy. Although the PCR tests showed a sensitivity of 95.3% (95% confidence interval [CI] 91.6–99.1), Loopamp only had 48.4% (95% CI 39.6–57.3) sensitivity and 87.9% (95% CI 82.1–93.7) specificity. The LoD of Loopamp for L. donovani was 100-fold lower (20 fg/µl) than that for L. aethiopica (2 pg/µl).ConclusionsThe Loopamp™ Leishmania detection kit is not suitable for the diagnosis of CL in Ethiopia, presumably due to a primer mismatch with the L. aethiopica 18S rRNA target. Further research is needed to develop a simple and sensitive point-of-care test that allows the decentralization of CL diagnosis in Ethiopia.Graphical

Otimização do conjugado Anti-IGG canino no kit diagnóstico da Leishmaniose Visceral Canina (LVC)

Introduction: The diagnosis of canine visceral leishmaniasis (CVL) is of great importance in the control of visceral leishmaniasis, a disease that is neglected worldwide and represents a significant public health problem. Enzyme-linked immunosorbent assay (ELISA) test is widely used because of the possibility of large-scale diagnosis and low cost. Objective: The objective of this study was to optimize the stoichiometry of the conjugate used in the EIE/CVL kit; for this purpose, some conjugation and monitoring parameters were modified through molecular exclusion chromatography to improve the final product. Method: The conjugate produced by Bio-Manguinhos was optimized by purification using SEC gel filtration chromatography and evaluated using the EIE-leishmaniasis-visceral-canine-Bio-Manguinhos kit (EIE-LVC). The conjugates were produced with different stoichiometries (IgG X peroxidase): current conjugate, optimized conjugate and optimized and purified conjugate. Results: The results obtained were validated using the manual Canine Visceral Leishmaniasis test. The homogeneity of the conjugate means in the ELISA test was evaluated using the chi-square test, thereby demonstrating the normality and reliability of the data. Based on statistical data, positive samples obtained by the ELISA test showed homogeneity at a significance level of 0.05, i.e. a data reliability level of 95%. The optimization of the production of the canine anti-IgG conjugate, an extremely important component used in the composition of the Bio-Manguinhos EIE LVC KIT, contributed to ensuring the quality of the kit and improving the diagnosis of canine visceral leishmaniasis disease, making it more sensitive and inexpensive for public health. Conclusions: This work demonstrates that it is possible to optimize the anti-IgG canine peroxidase (HRP) conjugate used in the EIE CVL kit to make it more specific and ultimately more economical.

Rapid antigen detection test for diagnosis of post kala-azar dermal leishmaniasis: application of CL Detect™ rapid test for active case detection in the endemic area.

Post-kala-azar dermal leishmaniasis (PKDL) is a skin condition that occurs in a small percentage of people who have been cured of visceral leishmaniasis (VL), and contributes to transmission of VL. The rK39 rapid test cannot decisively diagnose PKDL due to presence of antileishmanial antibodies from past VL episodes. CL Detect™ Rapid Test, an in-vitro diagnostic test that detects Leishmania antigen peroxidoxin, was assessed for diagnosing PKDL. The CL Detect RDT had 73.3% sensitivity and 100% specificity in the study. The test can be used as a primary screening tool to monitor PKDL in endemic regions and identify active Leishmania infection.

Antigenic reactivity of Leishmania (Viannia) lainsoni axenic amastigote proved to be a suitable alternative for optimizing Montenegro skin test

BackgroundLaboratory diagnosis of American cutaneous leishmaniasis (ACL) requires a tool amenable to the epidemiological status of ACL in Brazil. Montenegro skin test (MST), an efficient immunological tool used for laboratory diagnosis of ACL, induces delayed-type hypersensitivity (DTH) response to the promastigote antigens of Leishmania; however, human immune responses against infection are modulated by the amastigote of the parasite. Leishmania (V.) lainsoni induces strong cellular immunity in humans; therefore, the antigenic reactivity of its axenic amastigote (AMA antigen) to MST was evaluated for the laboratory diagnosis of ACL.MethodsAmong 70 individuals examined, 60 had a laboratory-confirmed diagnosis of ACL; 53 had localized cutaneous leishmaniasis (LCL), and 7 had mucosal leishmaniasis (ML). Patients were treated at the Evandro Chagas Institute’s leishmaniasis clinic, Pará State, Brazil. Ten healthy individuals with no history of ACL (control group) were also examined. Leishmania (V.) braziliensis promastigote antigen (PRO) was used to compare the reactivity with that of AMA antigen. Paired Student’s t-test, kappa agreement, and Spearman test were used to evaluate the reactivity of AMA and PRO.ResultsThe mean reactivity of AMA in ACL patients was 19.4 mm ± 13.3, which was higher (P < 0.001) than that of PRO: 12.1 mm ± 8.1. MST reactivity according to the clinical forms revealed that AMA reactivity in LCL and ML, 18.8 mm ± 13.3 and 24.3 mm ± 13.7, was higher (P < 0.001) than that of PRO, 11.8 mm ± 8.2 and 14.6 mm ± 8.4, respectively.ConclusionAMA reactivity was higher than that of PRO, indicating that AMA is a promising alternative for optimizing MST in the laboratory diagnosis of ACL.Graphical abstract

The Epidemiology of PCR-Confirmed Cutaneous Leishmaniasis in Israel: A Nationwide Study.

Leishmaniasis, mainly cutaneous leishmaniasis (CL), is endemic in Israel. In recent years, the diagnosis of leishmaniasis has transitioned to a molecular diagnosis. To summarize all cases of leishmaniasis and the identified species seen in Israel based on molecular diagnosis. A retrospective study was performed of patients diagnosed with leishmaniasis between January 2017 and December 2022. All five medical centers in Israel in which Leishmania diagnosis is performed were included: Soroka, HaEmek, Hadassah, Rambam, and Sheba, all utilized molecular diagnostic methods. Data on the annual number of cases, species, age, and gender were retrieved. During the years 2017-2022, a total of 4168 cases of leishmaniasis were diagnosed, which corresponds with ~7/100,000 inhabitants. L. (L.) major and L. (L.) tropica accounted for 84% and 14%, respectively. During the years 2020-2021, L. (L.) infantum emerged as a new form of cutaneous disease [2.7% of cases during this period]. Visceral L. (L.) infantum was found in five cases. Imported New World leishmaniasis accounted for 1% of the cases. L. (L.) major affected more males (67%) while L. (L.) tropica commonly affected more children and caused more facial lesions. The mean annual number of cases during these years is ~700. The dominant species is L. (L.) major. Since 2020, cutaneous L. (L.) infantum is an emerging infection in Israel.

Comparison of urine and serum IgG detection ELISA for tegumentary leishmaniasis diagnosis and prognosis

Laboratorial diagnosis of tegumentary leishmaniasis (TL) is hampered by variable sensitivity and/or specificity of the tests, which are still hampered by blood́ invasive collection. In this context, in the present study, we develop a serum- and urine-based ELISA to TL diagnoses. A recombinant protein (rLiHyA), which was previously showed to be antigenic for the disease, as well as a B-cell epitope produced as synthetic peptide and a Leishmania antigenic extract (SLA), were used as antigens. A total of paired 205 urine and serum samples were used, which were comprised by samples from cutaneous (n = 30) and mucosal (n = 30) leishmaniasis patients, as well as from healthy individuals living in endemic region of disease (n = 45), of patients with Chagas disease (n = 30), leprosy (n = 35), malaria (n = 15) or HIV-infected (n = 20). Results showed that serum-based ELISA presented sensitivity of 24.0 %, 100 % and 41.0 %, when SLA, rLiHyA and synthetic peptide were used as antigens, and specificity of 98.4 %, 98.4 % and 98.4 %, respectively. The area under the curve (AUC) was calculated and results were 0.74, 1.0, and 0.71, respectively, when SLA, rLiHyA and synthetic peptide were used as antigens. Performing an urine-based ELISA, sensitivity was 28.0 %, 100 % and 75.0 %, respectively, when SLA, rLiHyA, and synthetic peptide were used, while specificity values were of 98.4 %, 98.4 % and 98.4 %, respectively. In addition, the AUC values were 0.82, 1.0, and 0.94, respectively. A significant drop in specific antibodies levels in both patientś serum and urine samples was found six months after treatment, suggesting a prognostic role of rLiHyA for TL. In conclusion, preliminary data suggest the potential of use patient urine to TL diagnoses.

Oral Leishmaniasis in HIV-Positive and HIV-Negative Patients: A Comparative Analysis with Two New Case Reports.

This case report examines the clinical presentation, diagnosis, treatment, and outcomes of mucocutaneous leishmaniasis with primary oral involvement in HIV-positive and HIV-negative patients diagnosed in Brazil. We discuss the clinical manifestations, diagnostic methods, and therapeutic strategies, highlighting the clinical and histopathologic diagnostic features and distinct progression patterns based on HIV status. Our findings are compared with patterns observed in other countries, emphasizing the differences between the Americas and Europe, Asia, and Africa. In the Americas, particularly in Brazil, mucocutaneous leishmaniasis often presents with localized oral lesions, even in the presence of systemic immunosuppression, whereas in the Europe, Asia, and Africa, oral involvement is typically associated with visceral leishmaniasis in immunocompromised patients. These differences were due to variations in the parasite species involved. This comparison underscores the importance of regional and immunological factors in diagnosing and managing this neglected infectious disease.

Clinical manifestations and diagnostic approaches in cases of canine leishmaniasis in Bulgaria.

Leishmaniasis, a parasitic disease found in many parts of southern Europe, is transmitted in both humans and canines through the bite of phlebotomine sandflies, and can present in a variety of ways, such as cutaneous, mucocutaneous, diffuse, and visceral. In Bulgaria there are endemic areas of canine leishmaniasis, with sporadic cases in humans. However, no detailed studies of the animal population and vectors have been performed. Here we describe a few clinical cases of canine visceral leishmaniasis in two districts in western Bulgaria: one endemic and one without previously detected cases in humans or dogs. Diagnosis was confirmed serologically and molecularly using both real time and conventional PCR. Specific anti-leishmanial antibodies were confirmed in three of the cases via ELISA, with 50% of them returning extremely high values. In the majority of the cases DNA fragments were detected in the skin or lymph node aspirate but not in the blood. This paper highlights the need for further studies updating the current knowledge on the epidemiology, diagnosis, and control of visceral leishmaniasis in the reservoir host population.

A Diagnostic Algorithm for Detection of Leishmania spp. in Human Fresh and Fixed Tissue Samples.

Leishmaniasis is an important travel-related parasitic infection in the United States. Treatment regimens vary by Leishmania species and require an accurate diagnosis. The sensitivity and specificity of diagnostic methods depend on the type and condition of specimen analyzed. To identify the best algorithm for detection of parasites in fresh and fixed tissue samples, we evaluated parasite cultures, two PCR methods, and Leishmania immunohistochemistry (IHC) in samples received by the CDC from 2012 through 2019. The sensitivity and specificity of IHC assays were evaluated in fresh specimens tested. Diagnostic accuracy for formalin-fixed tissue was evaluated by using PCR-based methods and IHC. Of 100 suspected cases with fresh tissue available, Leishmania spp. infection was identified by PCR in 56% (56/100) of specimens; from these, 80% (45/56) were positive by parasite culture and 59% (33/56) by IHC. Of 420 possible cases where only fixed specimens were available, 58% (244/420) were positive by IHC and/or PCR. Of these, 96% (235/420) were positive by IHC and 84% (204/420) by PCR-based methods. Overall parasite detection using all methodologies was similar for fresh and formalin-fixed tissue specimens (56% versus 58%, respectively). Although PCR-based methods were superior for diagnosis of leishmaniasis and species identification in fresh samples, IHC in combination with PCR increased the accuracy for Leishmania spp. detection in fixed samples. In conclusion, PCR is the most effective method for detecting Leishmania infection in fresh tissue samples, whereas for formalin-fixed samples, IHC and PCR-based methods should be used in combination.

Non-invasive urine-based ELISA using a recombinant Leishmania protein to diagnose tegumentary leishmaniasis

The diagnosis of tegumentary leishmaniasis (TL) is hampered by variable sensitivity and/or specificity of the tests. Serological assays are suitable to diagnose visceral leishmaniasis (VL); however, they present low performance for the detection of TL cases. Additionally, blood collection to obtain patient serum represents a challenge, as it is an invasive and uncomfortable procedure, requiring laboratorial infrastructure and trained professionals. In this context, the present study proposed to evaluate patient urine to detect TL, given that this analyte has proven to be effective in ELISA experiments for the detection of VL cases. For this, a Leishmania protein called LiHyV, two specific B-cell epitopes derived from protein amino acid sequence, and a Leishmania antigenic extract (SLA) were used as antigens. A total of 215 paired urine and serum samples were evaluated, and results showed that, when serum was employed as an analyte, rLiHyV, Peptide1, Peptide2, and SLA presented a sensitivity of 85 %, 29 %, 58 %, and 31 %, respectively, and a specificity of 97.5 %, 98 %, 100 %, and 97.5 %, respectively, in the diagnosis of TL. When urine was used, rLiHyV, Peptide1, Peptide2, and SLA presented a sensitivity of 95 %, 74 %, 67 %, and 52 %, respectively, and a specificity of 100 %, 99 %, 98 %, and 86 %, respectively. In conclusion, preliminary data suggest that urine could be considered as an alternative biological sample for the detection of TL cases.

Molecular Diagnosis of Human Cutaneous Leishmaniasis and Identification of the Causative Leishmania Species in Iran: A Narrative Review

Cutaneous leishmaniasis (CL) is a common form of leishmaniasis in underdeveloped countries. Although CL tends to be self-limiting, it can cause significant scars and may progress to more severe manifestations. Additionally, Leishmania species vary in susceptibility to the available treatments. The selection of treatment and clinical outcome of CL depend on the accurate determination of the Leishmania species. This mini-review aims to provide an overview of the molecular diagnosis techniques such as PCR-based assays, nucleic acid sequence-based amplification, and loop-mediated isothermal amplification utilized in the identification of Leishmania species in Iran.

Histopathological findings and diagnosis of cutaneous leishmaniasis, confirmed by PCR, in an endemic region of Brazil.

Diagnosis of cutaneous leishmaniasis (CL) is difficult, and the correct use of histopathological criteria can be useful in clinical practice. The present study evaluates the association between histopathological findings and the results of polymerase chain reaction (PCR) in clinically suspected cases of CL. Skin samples were received in a laboratory from an endemic region of Brazil for over nine years. Associations were analyzed by means of the Chi square test with a 5% level of significance. Of the 222 examined samples, 190 (85.6%) tested positive by PCR. All 25 cases identified by microscopic examination also tested positive by PCR. Except for the more intense inflammatory infiltrate, all other evaluated histological variables (ulceration, epidermal hyperplasia, hyperkeratosis, presence of granuloma, neutrophils, histiocytes, lymphocytes, plasmocytes, and necrosis) were not significantly associated with PCR positivity. The intensity of the inflammatory infiltrate is a good indicator of the occurrence of CL. Histopathological aspects are useful to increase the predictive values of CL diagnoses, but PCR is still necessary to confirm or exclude the disease.

ELISA with recombinant antigen Lb6H validated for the diagnosis of American tegumentary leishmaniasis.

American tegumentary leishmaniasis (ATL) diagnosis is an open question, and the search for a solution is urgent. The available tests that detect the etiological agent of the infection are specific for ATL diagnosis. However, they present disadvantages, such as low sensitivity and the need for invasive procedures to obtain the samples. Immunological methods (leishmanin skin test and search for anti-Leishmania antibodies) are good alternatives to the etiological diagnosis of ATL. Presently, we face problems with disease confirmation due to the discontinuity in the production of leishmanin skin test antigen, particularly in resource-poor settings. Aiming to diagnose ATL, we validated rLb6H-ELISA for IgG antibodies using 1,091 samples from leishmaniasis patients and healthy controls, divided into four panels, living in 19 Brazilian endemic and non-endemic states. The rLb6H-ELISA showed a sensitivity of 98.6% and a specificity of 100.0%, with the reference panel comprising 70 ATL patient samples and 70 healthy controls. The reproducibility evaluation showed a coefficient of variation of positive samples ≤ 8.20% for repeatability, ≤ 17,97% for reproducibility, and ≤ 8.12% for homogeneity. The plates sensitized with rLb6H were stable at 4°C and -20°C for 180 days and 37°C for seven days, indicating 12 months of validity. In samples of ATL patients from five research and healthcare centers in endemic and non-endemic areas, rLb6H-ELISA showed a sensitivity of 84.0%; no significant statistical difference was observed among the five centers (chi-square test, p = 0.13). In samples of healthy controls from four areas with different endemicity, a specificity of 92.4% was obtained; lower specificity was obtained in a visceral leishmaniasis high endemicity locality (chi-square test, p<0.001). Cross-reactivity was assessed in 166 other disease samples with a positivity of 13.9%. Based on the good diagnostic performance and the reproducibility and stability of the antigen, we suggest using ELISA-rLb6H to diagnose ATL.

Comparison of SensPERT® Leishmania rapid test with two other immunochromatographic tests for the diagnosis of canine visceral leishmaniasis in Brazil.

In urban environments, dogs serve as the primary reservoir for visceral leishmaniasis (VL). Rapidly diagnosing canine VL through tests enables early treatment and a favorable prognosis. This study aimed to assess the diagnostic performance of the SensPERT® Leishmania test kit (Dechra®), Alere® Leishmaniasis Ac test kit, and the rapid test dual path platform (TR-DPP®) Bio-Manguinhos in detecting VL. 30 serum samples from reactive VL dogs and 30 serum samples from healthy dogs were employed for assessing the sensitivity and specificity variation between SensPERT® Leishmania test kit, Alere® Leishmaniasis Ac test kit, and rapid test dual platform - TR-DPP®. The SensPERT® Leishmania test outperformed Alere® and TR-DPP® in terms of sensitivity, specificity, positive and negative predictive values and demonstrated near-perfect concordance with Alere® and substantial concurrence with TR-DPP®. The SensPERT® Leishmania rapid test proved to be a promising test in the detection of VL in dogs.

Feline Leishmaniasis: Evidence-based Treatments and Challenges to be Solved.

Leishmaniasis is a neglected tropical disease caused by protozoa parasites from the Leishmania genus. Vertebrate hosts acquire the infection through the bite of a female sandfly, initiating a complex parasite development cycle. Contrary to previous beliefs regarding cats' resistance, these animals have recently been identified as potential reservoirs for leishmaniasis. Clinical symptoms in cats can manifest in diverse forms, including cutaneous, mucocutaneous, and visceral manifestations. The diagnosis of feline leishmaniasis is complicated by nonspecific symptoms and the relatively lower specificity of serological tests. The recommended treatment for feline leishmaniasis involves the administration of medications; however, success varies in each cat. This review aims to present cases of feline leishmaniasis, highlighting clinical symptoms, diagnostic methods, therapy schedules, and outcomes. Among the 24 cases documented in the available literature, 12 achieved successful treatment without relapses, resulting in a reduced parasite load and improved symptoms. Three cases responded well but presented persistent sequelae. Two feline leishmaniasis cases initially had treatment success but later experienced recurrences. Finally, no response was observed in seven cases, leading to the euthanasia of cats due to ineffectiveness or irregularities along the therapy. Conventional treatments, despite potential hepatotoxicity and nephrotoxicity, exhibit a high efficacy in reducing parasitic load, thereby improving clinical symptoms and increasing the life expectancy of affected cats. Nevertheless, consistent adherence is crucial, as interruptions may render the therapy ineffective and contribute to parasite resistance. Therefore, addressing the challenges associated with feline leishmaniasis treatment necessitates the development of new strategies to ensure a more effective and sustained approach.

Synthetic peptides-based SPR biosensor evaluation towards canine Visceral leishmaniasis diagnosis: A simple and effective approach

A vital component of surface plasmon resonance (SPR) biosensors is the recognition element. It generates the sensor selectivity towards the target biomarker and plays an important role in defining the sensor’s analytical sensitivity. In serological diagnostics, recombinant proteins (purified antigens) have presented better selectivity compared to soluble crude antigens in SPR immunosensors. However, recombinant DNA technology to obtain proteins is only explored by specialized laboratories, and commercial recombinant proteins still have a high cost. Thus, with the aim of developing a much simpler and lower-cost method, we investigated the prospect of using synthetic peptides as recognition elements for constructing an SPR biosensor. Two synthetic peptides, named PEP13 and PEP16, were tested for the serological diagnosis of Canine Visceral Leishmaniasis (CVL), an endemic neglected disease that affects humans and dogs worldwide. Between the two peptides tested, PEP13 was more sensitive when evaluating its responses against purified antibodies in buffer solution (LOD = 1.05 nmol L-1), and it also discriminated the response better when applied in diluted serum samples of infected dogs compared to diluted healthy dogs’ samples. For this reason, the PEP13 immunosensor was used to analyze canine serum samples, precisely identifying all the positive (n = 7) and negative (n = 7) CVL cases (p = 0.00136) in less than 12 min. In brief, this study explored promising biostructures through a simple and fast methodology for serological diagnosis, addressing the suitability of synthetic peptides for use in biosensors in the urgent field of neglected diseases.

Simplified molecular diagnosis of visceral leishmaniasis: Laboratory evaluation of miniature direct-on-blood PCR nucleic acid lateral flow immunoassay.

Diagnosis of visceral leishmaniasis (VL) in resource-limited endemic regions is currently based on serological testing with rK39 immunochromatographic tests (ICTs). However, rK39 ICT frequently has suboptimal diagnostic accuracy. Furthermore, treatment monitoring and detection of VL relapses is reliant on insensitive and highly invasive tissue aspirate microscopy. Miniature direct-on-blood PCR nucleic acid lateral flow immunoassay (mini-dbPCR-NALFIA) is an innovative and user-friendly molecular tool which does not require DNA extraction and uses a lateral flow strip for result read-out. This assay could be an interesting candidate for more reliable VL diagnosis and safer test of cure at the point of care. The performance of mini-dbPCR-NALFIA for diagnosis of VL in blood was assessed in a laboratory evaluation and compared with the accuracy of rK39 ICTs Kalazar Detect in Spain and IT LEISH in East Africa. Limit of detection of mini-dbPCR-NALFIA was 650 and 500 parasites per mL of blood for Leishmania donovani and Leishmania infantum, respectively. In 146 blood samples from VL-suspected patients from Spain, mini-dbPCR-NALFIA had a sensitivity of 95.8% and specificity 97.2%, while Kalazar Detect had a sensitivity of 71.2% and specificity of 94.5%, compared to a nested PCR reference. For a sample set from 58 VL patients, 10 malaria patients and 68 healthy controls from Ethiopia and Kenya, mini-dbPCR-NALFIA had a pooled sensitivity of 87.9% and pooled specificity of 100% using quantitative PCR as reference standard. IT LEISH sensitivity and specificity in the East African samples were 87.9% and 97.4%, respectively. Mini-dbPCR-NALFIA is a promising tool for simplified molecular diagnosis of VL and follow-up of treated patients in blood samples. Future studies should evaluate its use in endemic, resource-limited settings, where mini-dbPCR-NALFIA may provide an accurate and versatile alternative to rK39 ICTs and aspirate microscopy.

Artificial Intelligence in Cutaneous Leishmaniasis Diagnosis: Current Developments and Future Perspectives.

Cutaneous Leishmaniasis (CL) is a major global health problem requiring appropriate diagnosis methods. Its diagnosis is challenging, particularly in resource-limited settings. The integration of Artificial Intelligence (AI) into medical diagnostics has shown promising results in various fields, including dermatology. In this systematic review, we aim to highlight the value of using AI for CL diagnosis and the AI-based algorithms that are employed in this process, and to identify gaps that need to be addressed. Our work highlights that only a limited number of studies are related to using AI algorithms for CL diagnosis. Among these studies, seven gaps were identified for future research. Addressing these considerations will pave the way for the development of robust AI systems and encourage more research in CL detection by AI. This could contribute to improving CL diagnosis and, ultimately, healthcare outcomes in CL-endemic regions.

Synthetic Peptides Selected by Immunoinformatics as Potential Tools for the Specific Diagnosis of Canine Visceral Leishmaniasis.

Diagnosing canine visceral leishmaniasis (CVL) in Brazil faces challenges due to the limitations regarding the sensitivity and specificity of the current diagnostic protocol. Therefore, it is urgent to map new antigens or enhance the existing ones for future diagnostic techniques. Immunoinformatic tools are promising in the identification of new potential epitopes or antigen candidates. In this study, we evaluated peptides selected by epitope prediction for CVL serodiagnosis in ELISA assays. Ten B-cell epitopes were immunogenic in silico, but two peptides (peptides No. 45 and No. 48) showed the best performance in vitro. The selected peptides, both individually and in combination, were highly diagnostically accurate, with sensitivities ranging from 86.4% to 100% and with a specificity of approximately 90%. We observed that the combination of peptides showed better performance when compared to peptide alone, by detecting all asymptomatic dogs, showing lower cross-reactivity in sera from dogs with other canine infections, and did not detect vaccinated animals. Moreover, our data indicate the potential use of immunoinformatic tools associated with ELISA assays for the selection and evaluation of potential new targets, such as peptides, applied to the diagnosis of CVL.

Refinement of the rKLi8.3-Based Serodiagnostic ELISA Allows Detection of Canine Leishmaniosis in Dogs with Low Antibody Titers.

The diagnosis of canine leishmaniasis (CanL) still represents a challenge due to the variable clinical manifestations and the large number of asymptomatic dogs. Serological tests are most commonly used to detect infected animals, revealing anti-Leishmania antibodies, mainly of the IgG isotype. Recently, a new diagnostic antigen, rKLi8.3, containing 8.3 kinesin tandem repeats (TR) from a Leishmania infantum strain from Sudan, has been shown to provide excellent specificity and sensitivity for the detection of Leishmania-infected humans and dogs. However, asymptomatic animals with very low antibody titers are often difficult to detect by serodiagnosis. Thus, we wondered whether the addition of an anti-IgG-enhancing step in the protein A/G-based rKLi8.3-ELISA will improve the diagnostic performance without decreasing the specificity. For this, parasitologically confirmed CanL cases with low or high clinical scores, uninfected healthy controls and dogs with other infections were tested by rKLi8.3-ELISA as well as two different immunochromatographic rapid tests, rKLi8.3-lateral flow test (LFT) and Dual Path Platform (DPP®) based on the rK28 antigen. Our results show that the diagnostic accuracies of the rKLi8.3-ELISA and LFT were similar to that of DPP, missing several asymptomatic animals. However, the addition of a secondary, amplifying anti-dog IgG antibody in the protein A/G-based rKLi8.3-ELISA enabled the detection of nearly all asymptomatic dogs without compromising its specificity.