Visceral Leishmaniasis In Dogs Research Articles

Seroepidemiological survey for visceral leishmaniasis in dogs from an unaffected municipality of RS in the period 2021-2024

Canine visceral leishmaniasis (CVL) is a vector-borne parasitic disease that is expanding geographically, with autochthonous cases recorded in Rio Grande do Sul (RS) since 2008. Indigenous areas such as the municipalities of Pelotas, Rio Grande and Bagé are not part of active epidemiological surveillance for CVL, but the region has predisposing factors for the spread of the disease. The aim of this study was therefore to carry out a seroepidemiological survey of serum samples from dogs treated at a veterinary hospital in a disease-free area. The survey took place over a three-year period, following the current Ministry of Health (MoH) protocol. A total of 2,365 samples from asymptomatic animals were tested, of which 0.8% were seroreactive in the Dual Path Platform (DPP®) screening test. Of the samples that reacted to the DPP®, 37% were seroreactive to the ELISA (Bio-Manguinhos®), one was inconclusive (5.2%), 42% were negative and in 21% no confirmatory test was carried out. It can be concluded that Pelotas, Rio Grande and Bagé, the places of residence of seroreagent animals and CVL-free municipalities, have seroreagent dogs circulating according to the current MH protocol and require attention from epidemiological surveillance.

Visceral leishmaniasis seroprevalence and associated sociocultural factors in an endemic area of Sergipe, northeastern Brazil

The present study aimed to investigate the presence of anti-Leishmania antibodies in samples of domiciled dogs and associate them with epidemiological indicators in the city of Lagarto, Sergipe. The diagnosis of Canine Visceral Leishmaniasis (CVL) was carried out in 755 dogs, where the rapid two-way platform immunochromatographic test (TR-DPP) (Fiocruz/Biomanguinhos) and the serological test of Enzyme-Linked Immunosorbent Assay (ELISA) were applied. The results showed that all homes with seroreactive dogs were close to abandoned land, squares and/or green areas. There was a significant association between seroactive animals and type of shelter (p=0.008); 54.8% had an indoor shelter and 45.2% of the dogs had a shelter outside their homes. The disease is geographically distributed in 14 (73.68%) of the 19 sample areas in the municipality. Of these, five (26.31%) had cases of Visceral Leishmaniasis in dogs and humans. Eight cases of Human Visceral Leishmaniasis (HVL) were reported during the years 2017 to 2020 in the municipality. Dogs that are located in areas of greatest risk impact the maintenance of natural foci of Leishmaniasis transmission to human and animal hosts in areas endemic for Visceral Leishmaniasis. Such results are essential to assist epidemiological surveillance in the municipality of Lagarto-SE with the implementation of control measures aimed at the insect vector of the disease, the sand fly, to prevent human and canine cases.

Canine visceral leishmaniasis diagnosis by UV spectroscopy of blood serum and machine learning algorithms

Visceral leishmaniasis (VL) is a zoonotic disease caused by the protozoan Leishmania infantum, and dogs are considered the main urban hosts for future disease transmission. The first and most effective control against the spread of disease relies on identifying infected animals, followed by their treatment or sacrifice, to reduce the protozoan reservoirs. Despite the availability of various diagnostic tests for VL in dogs the development of a quick and accurate diagnosis is essential from a public health and ethical point of view. Here we analyze the use of UV–Vis spectroscopy as an alternative diagnostic method for VL diagnosis by using the antigen-antibody interaction in canine blood serum and machine learning algorithms. The main UV spectra in the 220 to 280 nm range exhibit nine electronic absorption bands, but no significative difference could be identified between the positive and negative group spectra. Finally, UV pre-proceed spectra by SNV (standard normal variate) were submitted to principal component analysis followed by Linear SVM algorithm, the prediction model was tested in a leave-one-out cross-validation and external validation test reaching an overall accuracy of 75%.

RMELEISH: A Novel Recombinant Multiepitope-Based Protein Applied to the Serodiagnosis of Both Canine and Human Visceral Leishmaniasis.

visceral leishmaniasis (VL) is a critical public health problem in over ninety countries. The control measures adopted in Brazil have been insufficient when it comes to preventing the spread of this overlooked disease. In this context, a precise diagnosis of VL in dogs and humans could help to reduce the number of cases of this disease. Distinct studies for the diagnosis of VL have used single recombinant proteins in serological assays; however, the results have been variable, mainly in relation to the sensitivity of the antigens. In this context, the development of multiepitope-based proteins could be relevant to solving such problem. a chimeric protein (rMELEISH) was constructed based on amino acid sequences from kinesin 39 (k39), alpha-tubulin, and heat-shock proteins HSP70 and HSP 83.1, and tested in enzyme-linked immunosorbent (ELISA) for the detection of L. infantum infection using canine (n = 140) and human (n = 145) sera samples. in the trials, rMELEISH was able to discriminate between VL cases and cross-reactive diseases and healthy samples, with sensitivity and specificity values of 100%, as compared to the use of a soluble Leishmania antigenic extract (SLA). the preliminary data suggest that rMELEISH has the potential to be tested in future studies against a larger serological panel and in field conditions for the diagnosis of canine and human VL.

In silico Discovery of Potential Natural Inhibitors against Trypanothione Synthetase in Canine leishmaniasis

Background: Leishmania infantum, one important intracellular parasite causes most potentially lethal diseases such as leishmaniasis i.e. Visceral leishmaniasis in dogs. Although certain chemical drugs such as pentostam, amphotericin B, miltefosine have been trailed against this disease, but all these drugs induced antibiotic resistance and toxicity in the host. Further, the trypanothione synthetase, a key enzyme of this parasite which catalyzes a reaction, maintaining thiol redox within the cell Methods: The binding study was carried out with selected natural/synthetic phytocompounds/drugs against the modeled trypanothione synthetase though molecular docking. Result: The generated protein model with lowest discrete potential energy (DOPE) -19960.97 was found good in quality with z score of -5.19 and quality factor of 61.83% and it was found that the natural inhibitors glycyrrhetic acid (GRA) and Theaflavin (TFN) showed highest binding energies of -7.34 and -6.95 Kcal/mol. This study may be concluded that the natural inhibitors glycyrrhetic acid (GRA) and Theaflavin (TFN) would be potential regimen in treatment of canine leishmaniasis.

Visceral leishmaniasis in a recent transmission region: 27.4% infectivity rate among seronegative dogs.

American visceral leishmaniasis (VL) is a parasitic disease whose main domestic reservoir in the urban environment is dog and is considered one of the most important zoonoses in the context of public health. Serological tests are typically used for the diagnostic screening of the disease. This study aimed to analyse the performance of different methodologies used in the diagnosis of VL in dogs sampled from a recent transmission area. The sample consisted of 52 dogs separated into groups based on the absence and presence of clinical signs of VL. The following serological techniques were carried out: the DPP® rapid test (RT), the ALERE® RT and an RT and immunoenzymatic assay with a recently developed protein (rKDDR-plus). In addition, molecular techniques were carried out with conjunctival swabs, and bone marrow aspirate samples and parasitological samples were obtained directly from bone marrow aspirates. It was concluded that 27.4% of seronegative dogs were infected, but the serological tests, used as screening tests, showed unsatisfactory sensitivity results (average: 51.2%) for dogs without clinical signs. It was suggested that polymerase chain reaction with conjunctival swabbing be used as a screening test for dogs without clinical signs, as this is a non-invasive collection technique with high-sensitivity values.

Molecular and serological evaluation of zoonotic visceral leishmaniasis in dogs in a rural area of Fars province, southern Iran, as a source of Leishmania infantum infection.

Canine visceral leishmaniasis (CVL) is endemic in the southern parts of Iran. The current study aimed at molecular and serological evaluation of zoonotic visceral leishmaniasis in dogs in Fars province, southern Iran. Blood samples were collected from 60 dogs in the three villages in Fars Province. Serum samples were tested for antibodies against L. infantum by direct agglutination test (DAT). DNA was extracted from each dog's buffy coat and tested by PCR, targeting the Leishmania ITS‐2 region. From a total of 60 studied dogs, 25 (41.7%) were female, and 35 (58.3%) were male. Dogs' age ranged from 1 to 7 years, with a mean age of 2.97 (±1.4) years. Anti‐Leishmania antibodies were detected in sera samples of 28 (46.7%) dogs, (titre ≥ 1:320). Out of 28 seropositive cases, 13 (46.4%) were female, and 15 (53.6%) were male. Association between seropositivity and dogs’ clinical signs was statistically significant (p < .05). Leishmania DNA was detected in the buffy coat of 3 of 60 studied dogs which were all seropositive by DAT. The PCR products were sequenced and molecular analysis showed that two of the isolates were Leishmania infantum, and one was L. tropica. The high proportion of seropositive dogs indicates the important role of these animals in the epidemiology of the disease in the region. Infected dogs with or without signs, especially those that are molecularly positive, can act as an active reservoir of the disease in the area.

Investigation of canine visceral leishmaniasis in a non-endemic area in Brazil and the comparison of serological and molecular diagnostic tests.

INTRODUCTION: Visceral leishmaniasis (VL) is an important zoonosis in Brazil. Previous identification of parasitized dogs can also help prevent the disease in humans, even in non-endemic areas of the country. The Brazilian Ministry of Health recommends diagnosis in dogs using a DPP® (rapid test) as a screening test and an immunoenzymatic assay (ELISA) as a confirmatory test (DPP®+ELISA), and culling infected dogs as a legal control measure. However, the accuracy of these serological tests has been questioned. METHODS: VL in dogs was investigated in a non-endemic area of the São Paulo state for three consecutive years, and the performances of different diagnostic tests were compared. RESULTS: A total of 331 dog samples were collected in 2015, 373 in 2016, and 347 in 2017. The seroprevalence by DPP®+ELISA was 3.3, 3.2, and 0.3%, respectively, and by indirect immunofluorescence assay (IFA), it was 3.0, 5.6, and 5.5%, respectively. ELISA confirmed 18.4% of DPP® positive samples. The concordance between the IFA and DPP® was 83.9%. The concordance between IFA and DPP®+ELISA was 92.9%. A molecular diagnostic test (PCR) was performed in 63.2% of the seropositive samples, all of which were negative. CONCLUSIONS: In non-endemic areas, diagnostic tests in dogs should be carefully evaluated to avoid false results.

Molecular detection of visceral leishmaniasis in dogs from Barão de Melgaço, Pantanal region of Mato Grosso, Brazil

ABSTRACT: The increasing expansion of visceral leishmaniasis (VL) in the Brazilian territory evidences the need for studies focused on the main reservoir of this parasite: the dog. This study aimed to conduct an epidemiological survey in the municipality of Barão de Melgaço, Pantanal region of the state of Mato Grosso (MT), Brazil. Conventional polymerase chain reaction (PCR) and qualitative SYBR®Green real-time PCR (qPCR) were used to diagnose canine VL (CVL) and characterize the factors associated with this infection. Of the 402 dogs that had blood samples collected, 31 presented the parasite DNA, representing a prevalence of 7.71% in the population studied. Positivity indices for PCR and qPCR were 3.48 (14/402) and 7.21% (29/402), respectively. Comparison of the results obtained by both techniques showed moderate agreement (Kappa = 0.5364). Of the independent variables analyzed, presence of clinical signs (p≤0.05) was the only one associated with CVL. Based on this study, we conclude that VL is a circulating disease, with relatively low prevalence, in dogs of Barão de Melgaço/MT, and that the presence of clinical signs is the only variable associated with canine infection.

Visceral Leishmaniasis in Dogs

Background: Canine visceral leishmaniasis (CVL) is a zoonosis of variable clinical presentation, either in systemic or cutaneous form. Clinical signs include anorexia, ophthalmopathies, and chronic kidney disease. In the state of Santa Catarina, the foci are concentrated in the capital and its adjacencies, in the east side of the state. The objective of this study is to outline the first three reported cases of CVL in the municipality of Curitibanos, since there are no reports to date in the region of the mountainous plateau, in the middle west of Santa Catarina.Cases: All dogs were treated at the Veterinary Clinic School of the Federal University of Santa Catarina, Campus Curitibanos. The animals, two males and one female, belonged to the same tutor, resided in Curitibanos, and were attended between 2016 and 2020. In the first case, attended in 2016, the complaint was of eye and skin changes about three months ago. The animal lived in an urban environment and came from Uruguaiana, Rio Grande do Sul. On physical examination, the animal presented skin peeling, wet and crusty lesions, bloody discharge in the ears and nasal hyperkeratosis, as well as signs suggestive of uveitis. In this case, euthanasia was carried out. The second case had complaints of respiratory, eye changes, hyporexia, and polydipsia. The dog was prostrate, dehydrated, with lymphadenomegaly and respiratory disorders, compatible with bacterial pneumonia. Bilateral corneal edema confirmed uveitis. Upon return, the animal remained dehydrated with enlarged lymph nodes. In the aspiration of the lymph nodes, suggestive forms of Leishmania sp. were observed. The recommended confirmatory tests were performed, leading to a definitive diagnosis of CVL. The patient was treated with miltefosine, but later died. The third case was attended for general evaluation after a positive diagnosis for CVL during an epidemiological survey of the second case. The animal was alert, tachycardic, and tachypneic with hyperemic mucous membranes. Miltefosine-based treatment and permanent use of deltamethrin-impregnated collar were prescribed and ovariohysterectomy was carried out. The patient is clinically well. In the search for vectors, in no place or moment of search, referring to the cases, vectors for CVL were found.Discussion: It is suggested that patients 2 and 3 are autochthonous cases, whose transmission form has not been fully elucidated, and vector transmission cannot be ruled out, as well as other less common forms of transmission. For case 1, it is suggested that it is an allochthone case, probably imported from the Uruguaiana region, where the disease is endemic. The conduct of a positive result for CVL is euthanasia, with the exception of dogs that have guardians who wish to perform treatment, but not always possible due to the high cost of miltefosine, the only drug approved for dog therapy in Brazil. In the first case, the owner opted for euthanasia, since in 2016 there was no possibility of treatment. For the last two cases, a treatment cycle was performed as recommended by the guidelines. Regarding the municipality of Curitibanos, in the first case diagnosed in 2016, the animal came from Uruguaiana. In conclusion, this study aimed to report the clinical and epidemiological characteristics related to the first three patients with CVL reported in Curitibanos.

Potential of Artesunate in the treatment of visceral leishmaniasis in dogs naturally infected by Leishmania infantum: Efficacy evidence from a randomized field trial.

Leishmaniasis is among the world’s most neglected diseases. Dogs are the main reservoirs/hosts of Leishmania infantum, causative agent of both canine and human visceral leishmaniosis. Canine leishmaniasis (CanL) represents a public health problem as one of the most prevalent zoonotic diseases worldwide. Current therapeutics present drawbacks; thus, there is a need for more effective, safer, and cheaper drugs. The aim of this study was to evaluate and to compare the efficacy of oral administration of artesunate or meglumine antimoniate/allopurinol in dogs with clinical leishmaniasis. Forty-two dogs with naturally occurring clinical leishmaniasis were included in this open-label, simple randomized positive-control clinical field trial with 6 months of follow-up. Dogs received meglumine antimoniate 100 mg/kg/day and allopurinol 30 mg/kg/day for 28 days (control group, n = 26) or artesunate 25 mg/kg/day for 6 days (test group, n = 16). The animals were evaluated for their clinical evolution, parasite load (by qPCR) and humoral response at different time points: 0, 30, 90, and 180 days after treatment. Data analyses showed a significant improvement in both groups in clinical scores, parasitemia and antibody titers after treatment. Compared to the control group, the artesunate group showed significantly lower clinical score (P = 0.0001), lower parasitemia (P = 0.0001) and antibody titers after 6 months of follow-up. Compared to baseline values, a rapid, significant reduction (P < 0.012) in antibody levels, 2.28- versus 3.04-fold for the control versus artesunate groups, respectively, was observed 30 days after treatment. Antibody levels continued to decrease further in the artesunate group, where 58% of cases became seronegative at the 6-month follow-up. All qPCR-positive dogs were negative after treatment with artesunate, while 14.3% remained positive with the appearance of two new cases in the control group. Artesunate was well tolerated, and no side effects were recorded. Treatment failures were similar in both groups with 27.27% (6/22), including 18.18% (4/22) mortality in the control group, versus 26.66% (4/15), including 13.33% (2/15) mortality in the artesunate group. This is the first report showing the potential of artesunate in the treatment of dogs with clinical leishmaniasis. Artesunate showed higher efficacy than the current first-line treatment for CanL without any adverse effects. It could be a good alternative chemotherapy for CanL, and may be considered for further studies in human leishmaniases. Further clinical trials are needed to confirm these findings, to determine if there are relapses after treatment and if dogs remain infective to sandflies, to define the ideal therapeutic dosage and duration of treatment with artesunate.

Fluralaner (Bravecto\xae) induces long-term mortality of Lutzomyia longipalpis after a blood meal in treated dogs

BackgroundLeishmania infantum is the etiological agent of visceral leishmaniasis (VL) in the New World, where the sand fly Lutzomyia longipalpis and domestic dogs are considered the main vector and host reservoirs, respectively. Systemic insecticides have been studied as an alternative to control vector-borne diseases, including VL. Fluralaner, an isoxazoline class compound, is a systemic insecticide used in dogs, with proven efficiency against different species of phlebotomine sand flies. However, to date no studies have demonstrated the efficacy of fluralaner on Lu. longipalpis. The aim of this study was to evaluate the insecticidal effect of fluralaner (Bravecto®) on the sand fly Lu. longipalpis after blood meal in treated dogs.MethodsHealthy mongrel dogs (n = 8) were recruited from the Zoonoses Control Center in the city of Natal, Rio Grande do Norte, Brazil, and randomized into two groups: fluralaner treated (n = 4) and non-treated control (n = 4). Colony-reared female specimens of Lu. longipalpis (n = 20) were allowed to feed on all dogs for 40 min before treatment (for fluralaner-treated dogs), at day 1 after treatment and then monthly until 1 year post-treatment.ResultsIn the treatment group, there was 100% mortality of Lu. longipalpis for up to 5 months after treatment initiation, decreasing to 72.5% at 6 months post-treatment initiation. The efficacy of fluralaner ranged from 100% at day 1 (P = 0.0002) to 68% ( P = 0.0015) at 6 months, decreasing to 1.4% at 1 year post-treatment. Sand fly mortality carried out blood meal in non-treated control dogs remained constant at ≤ 15%.ConclusionsTaken together, our results suggest that fluralaner may be used as a control strategy for VL in dogs in VL endemic areas.Graphical

Environmental Variables Associated with Distribution of Canine Visceral Leishmaniasis in Dogs in Ardabil Province, Northwestern Iran: A Systematic Review

Background: Visceral leishmaniasis (VL) is a zoonotic disease that currently occurs in some parts of Europe, Asia, Africa, and America. This study aimed to determine the distribution of the canine visceral leishmaniasis in dogs (Canis lupus familiaris) in northwestern Iran.&#x0D; Methods: The data were collected from 1994 to 2018 in Ardabil Province from electronic databases. An extensive literature search was conducted in different international and national databases, including Cochrane, MEDLINE/PubMed, Scopus, Web of Science, and Iran Medex to find articles with the words “visceral leishmaniasis in Ardabil” in their titles, and “canine visceral leishmaniasis in Ardabil” or “accidental reservoir hosts of visceral leishmaniasis in Ardabil” in their subtitles, irrespective of the type and duration of study. The GIS software and MaxEnt model were used to determine the ecologically suitable niches for the disease.&#x0D; Results: In total, 9088 dogs were examined, and the overall prevalence rate of CVL in dogs was estimated to be 14.56%. The most ecologically suitable areas of CVL occurrence were identified in four hotspots in Meshkinshahr, Germi, and two spots in Parsabad counties. The results of jackknife test showed that the environmental and climate variables with the highest gain, when used in isolation, were Isothermality, Bio3, Bio13, and Bio 4.&#x0D; Conclusion: A widely epidemic CVL has emerged among dogs, making a lot of risks on inhabitants of this area and increasing the probability of an outbreak of VL in humans.

Analysis and simulations with a multi-scale model of canine visceral leishmaniasis

Visceral leishmaniasis in dogs is believed to have an impact on the prevalence of the disease in human populations. Here, we continue the analysis of the nested immuno-epidemiological model of visceral leishmaniasis in dogs, including a proof of well-posedness using functional analytical methods. Once well-posedness is established, we continue stability analysis of the endemic equilibria and provide necessary and sufficient conditions for the presence of backward bifurcation, and prove the instability of the lower endemic equilibrium in the presence of backward bifurcation. Lastly, we provide a number of simulations of the model using a number of control strategies. Control measures currently in use attempt to reduce the parasite load in the host, reduce the vector population, reduce the vector biting rate, and remove infected hosts. We examine various combinations of these strategies and conclude that a strategy combining culling infected dogs and removing vectors from the population by means such as insecticide will be the most effective.

High seroprevalence and associated factors for visceral leishmaniasis in dogs in a transmission area of Paraíba state, Northeastern Brazil.

Dogs have been implicated as main reservoirs for visceral leishmaniasis (VL) in urban areas. Therefore, studies on this species provide important data for public health. Thus, the objective of the present study was to ascertain the seroprevalence of canine VL (CVL) and the associated factors in the Brejo Paraibano microregion, northeastern Brazil. A total of 409 dogs were sampled from the eight municipalities of the microregion: Alagoa Grande, Alagoa Nova, Areia, Bananeiras, Borborema, Matinhas, Pilões and Serraria. The diagnosis of CVL was made using an enzyme-linked immunosorbent assay (ELISA S7®), through which the prevalence observed was 29.3%. By robust Poisson regression analysis two factors were associated with seroprevalence: illiterate/incomplete elementary owner's education level (prevalence ratio = 1.57; 95% CI = 1.05-2.34; p-value = 0.027) and tick infestation (prevalence ratio = 1.82; 95% CI = 1.27-2.61; p-value = 0.001). It is concluded that the seroprevalence of CVL in the Brejo Paraibano microregion is high. The factors associated with seroprevalence indicated the importance to develop socioeducational actions on the population, and the finding that tick infestation was associated with seroprevalence shows that there is a need for investigation regarding the role of ticks in the epidemiology of CVL.

Recombinant Cysteine Proteinase B from Leishmania braziliensis and Its Domains: Promising Antigens for Serodiagnosis of Cutaneous and Visceral Leishmaniasis in Dogs.

Leishmaniasis represents a group of parasitic diseases caused by a protozoan of the genus Leishmania and is widely distributed in tropical and subtropical regions. Leishmaniasis is one of the major tropical neglected diseases, with 1.5 to 2 million new cases occurring annually. Diagnosis remains a challenge despite advances in parasitological, serological, and molecular methods. Dogs are an important host for the parasite and develop both visceral and cutaneous lesions. Our goal was to contribute to the diagnosis of canine cutaneous leishmaniasis (CL) and visceral leishmaniasis (VL) using the recombinant cysteine proteinase B (F-CPB) from Leishmania braziliensis and its N- and C-terminal domains (N-CPB and C-CPB) as antigens in an enzyme-linked immunosorbent assay (ELISA). Sera from dogs from Northwest Argentina diagnosed with CL were tested by ELISA against a supernatant of L. braziliensis lysate, the F-CPB protein, and its domains. We found values of sensitivity (Se) of 90.7%, 94.4%, and 94.3% and specificity (Sp) of 95.5%, 90.9%, and 91.3% for F-CPB and its N- and C-terminal domains, respectively. In sera from dogs diagnosed with VL from Northeast Argentina, we found Se of 93.3%, 73.3%, and 66.7% and Sp of 92.3%, 76.9%, and 88.5% for F-CPB and its N- and C-terminal domains, respectively. These results support CPB as a relevant antigen for canine leishmaniasis diagnosis in its different clinical presentations. More interestingly, the amino acid sequence of CPB showed high percentages of identity in several Leishmania species, suggesting that the CPB from L. braziliensis qualifies as a good antigen for the diagnosis of leishmaniasis caused by different species.

Recombinant Leishmania eukaryotic elongation factor-1 beta protein: A potential diagnostic antigen to detect tegumentary and visceral leishmaniasis in dogs and humans

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.

Performance of different serological tests in the diagnosis of natural infection by Leishmania infantum in dogs

Visceral leishmaniasis (VL) is a zoonosis caused by the parasite Leishmania infantum and the dog is its main reservoir in rural and urban areas. The diagnosis of infection is mainly based on the presence of anti-Leishmania IgG antibodies in the serum of infected dogs. In this study, the sensitivity and specificity of qualitative rapid tests (RTs) dual path platform (DPP) Bio-Manguinhos, rapid enzyme-linked immunosorbent assay (ELISA) IDEXX, Kalazar Detect and ALERE, as well as quantitative ELISA Bio-Manguinhos and in-house indirect immunofluorescence assay (IFA) tests were analyzed in sera from infected and uninfected dogs. Serial dilutions of the in-house IFA were compared with RTs and ELISA Bio-Manguinhos. The results showed that none of the tests reached 100% sensitivity and specificity. There was no statistical difference between the analyzed RTs. The most sensitive test was the DPP Bio-Manguinhos (97.9%), while the rapid ELISA IDEXX showed higher specificity (100%). In the treatment setting of infected and/or diseased animals, quantitative tests for monitoring the evolution of antibody titers are required, which indicates the maintenance of in-house IFA in animal handling. Furthermore, we demonstrate that the RTs present higher sensitivity in serum samples with superior antibody titers obtained in the in-house IFA. However, the RTs exhibited false negatives in samples with low titers of antibodies. Among the RTs, only the DPP Bio-Manguinhos presented better performance in this situation. Therefore, the use of RTs for the diagnosis of VL in dogs with low titers of antibodies, such as asymptomatic, should be carefully evaluated.

Leishmania infantum recombinant kinesin degenerated derived repeat (rKDDR): A novel potential antigen for serodiagnosis of visceral leishmaniasis.

Visceral leishmaniasis (VL) or kala-azar, the most severe form of leishmaniasis, can lead to death if not properly diagnosed and treated. Correct identification of infected patients and reservoirs is vital for controlling the spread of leishmaniasis. Current diagnostic kits for leishmaniasis show high sensitivity and specificity, but can also result in false negatives and cross reactions with related parasitic infections. New diagnostic methods with greater accuracy are urgently needed for diagnosis of leishmaniasis. In this study, we aimed to uncover a new highly effective antigen for the diagnosis of visceral leishmaniasis in dogs and humans, aiming to improve the accuracy compared with those of current methods of diagnosis. Initially, in-silico epitope prediction analyses identified several potential B-cell epitopes in the repetitive region of Leishmania infantum kinesin, which co-localized with predicted structural disordered regions, suggesting high potential for antigenicity. Based on this analysis, 8.5 genomic motifs, which encode the repetitive sequence of 39 degenerate amino acids, were selected for recombinant expression. BLASTn analysis of this repetitive region indicated that it is absent in the T. cruzi parasite, which is closely related to Leishmania, indicating the specificity of this region. This potentially antigenic protein, named recombinant kinesin degenerated derived repeat (rKDDR), was recombinantly expressed in Escherichia coli BL21-Star using the pET28a-TEV expression vector. We then evaluated the performance of rKDDR in correctly diagnosing Leishmania infection and compared this new assay with currently used diagnostic tests for leishmaniasis. rKDDR showed greater sensitivity and specificity in correctly diagnosing leishmaniasis both in human (sensitivity 92.86% and specificity 100%) and canine (sensitivity 88.54% and specificity 97.30%) sera compared with those of rK39 (human: sensitivity 90.48% and specificity 97.92%; canine: sensitivity 78.13% and specificity 90.09%). In addition, the rKDDR-ELISA outperformed the EIE-LVC kit, which is the serologic kit recommended by the Brazilian Ministry of Health for the diagnosis of canine visceral leishmaniasis. These results indicate that rKDDR is a highly promising candidate for diagnosis of visceral leishmaniasis, and is more accurate than the currently used gold-standard antigens.

A novel multi-scale immuno-epidemiological model of visceral leishmaniasis in dogs

Leishmaniasis is a neglected and emerging disease prevalent in Mediterranean and tropical climates. As such, the study and development of new models are of increasing importance. We introduce a new immuno-epidemiological model of visceral leishmaniasis in dogs. The within-host system is based on previouslyВ collectedВ and published data, showing the movement and proliferation of the parasite in the skin and the bone-marrow, as well as the IgG response. The between-host system structures the infected individuals inВ time-since-infection and is of vector-host type. The within-host system has a parasite-free equilibrium and at least one endemic equilibrium, consistent with the fact that infected dogs do not recover without treatment. We compute the basic reproduction number R0 of the immuno-epidemiological modelВ and provide the existence and stability results of the population-levelВ disease-free equilibrium. Additionally, we prove existence of an uniqueВ endemic equilibrium when R0 &gt; 1, and evidence of backward bifurcation and existence of multiple endemic equilibria when R0 &lt; 1.