Serological diagnosis of visceral leishmaniasis by an enzyme immunoassay using protein A in naturally infected dogs

A rapid indirect enzyme-linked immunosorbent assay (ELISA) was developed for measuring antibodies against Leishmania chagasi using total antigen from lysed promastigotes. Fifty symptomatic mixed breed dogs from a region of high incidence of visceral leishmaniasis in Brazil were examined. The results showed that in the positive animals, diagnosed by cytological examination, the ELISA using protein A assay system (mean optical density ± SD / 2.078 ± 0.631) detected more antibodies than the anti-IgG assay (mean optical density ± SD / 1.008 ± 0.437), while in the negative animals, the results by both systems were similar. These results suggest that the ELISA assay using protein A peroxidase conjugated could be useful to detect early infected animals in endemic areas, and thus help to control the spread of the infection.

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Leishmaniasp. Amastigotes Identification in Canine Transmissible Venereal Tumor

Case Reports in Veterinary Medicine ◽

10.1155/2014/603852 ◽

2014 ◽

Vol 2014 ◽

pp. 1-4 ◽

Cited By ~ 1

Author(s):

Vinicius Soares Carreira ◽

Heitor Flávio Ferrari ◽

Ingeborg Maria Langohr ◽

Charles Mackenzie ◽

Luiz Carlos Montezzo ◽

...

Keyword(s):

Visceral Leishmaniasis ◽

Tumor Tissue ◽

Canine Visceral Leishmaniasis ◽

Leishmania Chagasi ◽

Neoplastic Tissue ◽

Mixed Breed ◽

Route Of Transmission ◽

Vector Borne ◽

Polymerase Chain ◽

Male Genital

Leishmaniasis is a vector-borne disease withLeishmania chagasibeing the etiological agent of canine visceral leishmaniasis in South America. Canine venereal tumor is a transplantable round cell tumor of histiocytic origin which is mostly observed in sexually active male and female intact dogs. It has been shown thatLeishmaniaamastigotes have higher tropism for the canine male genital tract tissues and venereal leishmaniasis transmission has been documented in dogs but, to date, a canine venereal tumor-dependent transmission route has not been fully demonstrated. In this report, a 10-year-old, mixed breed, intact female dog presented a vaginal venereal transmissible tumor but no other clinical abnormalities otherwise. Unexpectedly, tumor tissue imprint smears examination revealedLeishmaniasp. amastigotes within infiltrating macrophages. In addition to the cytological direct identification, the protozoan was confirmed within the neoplastic tissue by means of immunohistochemistry and polymerase chain reaction. This report illustrates an asymptomaticLeishmaniasp. infection that may have started on or from the canine venereal tumor tissue, the latter option further supporting previous evidence of such an alternative vector-independent route of transmission for canine visceral leishmaniasis in areas where these diseases coexist.

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Specific Serodiagnosis of Canine Visceral Leishmaniasis Using Leishmania Species Ribosomal Protein Extracts

Clinical and Vaccine Immunology ◽

10.1128/cvi.00295-09 ◽

2009 ◽

Vol 16 (12) ◽

pp. 1774-1780 ◽

Cited By ~ 25

Author(s):

Eduardo A. F. Coelho ◽

Laura Ramírez ◽

Mariana A. F. Costa ◽

Vinicio T. S. Coelho ◽

Vivian T. Martins ◽

...

Keyword(s):

Visceral Leishmaniasis ◽

Ribosomal Proteins ◽

High Sensitivity ◽

Leishmania Species ◽

Cross Reactivity ◽

Enzyme Linked Immunosorbent Assay ◽

Canine Visceral Leishmaniasis ◽

Leishmania Chagasi ◽

Serum Samples ◽

Potential Tool

ABSTRACT In the present work, we have analyzed the antigenicity of Leishmania species ribosomal proteins (LRPs). To accomplish this, Leishmania infantum ribosomes were biochemically purified from promastigote cytosolic extracts, and their reactivities were analyzed by using the sera from dogs naturally infected with L. infantum. Since antibodies reacting against different ribosomal proteins were observed in all the serum samples obtained from dogs with symptomatic visceral leishmaniasis tested, we have analyzed the potential usefulness of the LRP extracts in the development of an enzyme-linked immunosorbent assay (ELISA) for the serodiagnosis of canine visceral leishmaniasis (CVL) in an area of Brazil where visceral leishmaniasis is endemic due to infection by Leishmania chagasi. A comparative ELISA with crude soluble Leishmania chagasi antigen (SLA) and L. infantum LRPs was performed. LRP- and SLA-based ELISAs gave similar sensitivities for the diagnosis of symptomatic CVL, but the LRP extract provided a very high sensitivity for the detection of oligosymptomatic and asymptomatic dogs. In addition, an LRP-based ELISA showed a higher specificity when the sera from dogs harboring other infections were included in the analysis. The LRP antigen displayed no cross-reactivity with sera from dogs that had any of the other diseases tested, notably, Chagas' disease. Our findings suggest that LRPs are a potential tool for the diagnosis of CVL and will be particularly useful for the diagnosis of asymptomatic CVL.

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First description of autochthonous canine visceral leishmaniasis in the metropolitan region of Vitória, State of Espírito Santo, Brazil

Revista da Sociedade Brasileira de Medicina Tropical ◽

10.1590/s0037-86822012000600019 ◽

2012 ◽

Vol 45 (6) ◽

pp. 754-756 ◽

Cited By ~ 4

Author(s):

Marco André Loureiro Tonini ◽

Elenice Moreira Lemos ◽

Alexandre Barbosa Reis ◽

Wendel Coura Vital ◽

Edelberto Santos Dias ◽

...

Keyword(s):

Visceral Leishmaniasis ◽

Positive Culture ◽

Human Case ◽

Metropolitan Region ◽

Enzyme Linked Immunosorbent Assay ◽

Canine Visceral Leishmaniasis ◽

Leishmania Chagasi ◽

Molecular Tests ◽

Leishmania Spp ◽

Polymerase Chain

INTRODUCTION: We investigated autochthonous canine visceral leishmaniasis (CVL) in the metropolitan region of Vitória (MRV), an area in which a human case was previously reported. METHODS: Serological, parasitological, and molecular tests were performed in 201 dogs. RESULTS: Twenty-six (13%) and 12 (6%) dogs were identified as positive using in-house enzyme-linked immunosorbent assay (ELISA) and rK39 tests, respectively. Two dogs had a positive culture for Leishmania chagasi, and 4 were polymerase chain reaction (PCR)-positive for Leishmania spp. One positive dog belonged to the aforementioned patient. CONCLUSIONS: Although the responsible vector was not found, our results provide evidence of autochthonous CVL in the MRV, a non-endemic area for VL.

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An enzyme-linked immunosorbent assay (ELISA) for the detection of IgM antibodies against Leishmania chagasi in dogs

Pesquisa Veterinária Brasileira ◽

10.1590/s0100-736x2009000200006 ◽

2009 ◽

Vol 29 (2) ◽

pp. 120-124 ◽

Cited By ~ 1

Author(s):

Débora Carvalho ◽

Trícia M.F.S. Oliveira ◽

Cristiane D. Baldani ◽

Rosangela Z. Machado

Keyword(s):

Visceral Leishmaniasis ◽

Immunosorbent Assay ◽

Fluorescent Antibody ◽

Domestic Dog ◽

Enzyme Linked Immunosorbent Assay ◽

Leishmania Chagasi ◽

Serum Samples ◽

Igm Antibodies ◽

Specificity And Sensitivity ◽

Belo Horizonte

Visceral leishmaniasis is an emergent zoonosis with an increasing number of new cases in Brazil where the domestic dog is an important parasite reservoir in the infectious cycle of Leishmania chagasi. An enzyme-linked immunosorbent assay (ELISA), based upon the use of a total soluble antigenic preparation of L. chagasi, was adapted for the detection of IgM antibodies in the serum of infected dogs. Optimal dilutions of the antigen, using positive and negative reference sera, were determined by checkboard titrations. The specificity and sensitivity of the ELISA were 100 %. A total of 110 serum samples were taken from dogs in Belo Horizonte, Minas Gerais, Brazil, and examined for anti-L. chagasi IgM antibody by ELISA and indirect fluorescent antibody test (IFAT). About 25% (n=27) of all the dogs tested were found serologically positive for L. chagasi by IFAT, while 89.09% (n=98) were seropositive by ELISA. The results obtained by ELISA and IFAT were significantly different (P<0.01). The combined use of ELISA and IFAT is recommended in order to enable veterinary services to more efficiently detect canine visceral leishmaniasis.

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The increased presence of repetitive motifs in the KDDR-plus recombinant protein, a kinesin-derived antigen from Leishmania infantum, improves the diagnostic performance of serological tests for human and canine visceral leishmaniasis

PLoS Neglected Tropical Diseases ◽

10.1371/journal.pntd.0009759 ◽

2021 ◽

Vol 15 (9) ◽

pp. e0009759

Author(s):

Williane Fernanda Siqueira ◽

Agostinho Gonçalves Viana ◽

João Luís Reis Cunha ◽

Leticia Mansur Rosa ◽

Lilian Lacerda Bueno ◽

...

Keyword(s):

Amino Acid ◽

Visceral Leishmaniasis ◽

Diagnostic Performance ◽

Leishmania Donovani ◽

Protein A ◽

Enzyme Linked Immunosorbent Assay ◽

Serological Tests ◽

Appropriate Treatment ◽

Human Sera ◽

Fatal Form

Visceral leishmaniasis (VL) is caused by protozoa belonging to the Leishmania donovani complex and is considered the most serious and fatal form among the different types of leishmaniasis, if not early diagnosed and treated. Among the measures of disease control stand out the management of infected dogs and the early diagnosis and appropriate treatment of human cases. Several antigens have been characterized for use in the VL diagnosis, among them are the recombinant kinesin-derived antigens from L. infantum, as rK39 and rKDDR. The main difference between these antigens is the size of the non-repetitive kinesin region and the number of repetitions of the 39 amino acid degenerate motif (6.5 and 8.5 repeats in rK39 and rKDDR, respectively). This repetitive region has a high antigenicity score. To evaluate the effect of increasing the number of repeats on diagnostic performance, we designed the rKDDR-plus antigen, containing 15.3 repeats of the 39 amino acid degenerate motif, besides the absence of the non-repetitive portion from L. infantum kinesin. Its performance was evaluated by enzyme-linked immunosorbent assay (ELISA) and rapid immunochromatographic test (ICT), and compared with the kinesin-derived antigens (rKDDR and rK39). In ELISA with human sera, all recombinant antigens had a sensitivity of 98%, whereas the specificity for rKDDR-plus, rKDDR and rK39 was 100%, 96% and 71%, respectively. When evaluated canine sera, the ELISA sensitivity was 97% for all antigens, and the specificity for rKDDR-plus, rKDDR and rK39 was 98%, 91% and 83%, respectively. Evaluation of the ICT/rKDDR-plus, using human sera, showed greater diagnostic sensitivity (90%) and specificity (100%), when compared to the IT LEISH (79% and 98%, respectively), which is based on the rK39 antigen. These results suggest that the increased presence of repetitive motifs in the rKDDR-plus protein improves the diagnostic performance of serological tests by increasing the specificity and accuracy of the diagnosis.

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D-Dimer Determination to Exclude Pulmonary Embolism: a Two-Step Approach Using Latex Assay as a Screening Tool

Thrombosis and Haemostasis ◽

10.1055/s-0038-1648817 ◽

1994 ◽

Vol 72 (01) ◽

pp. 089-091 ◽

Cited By ~ 21

Author(s):

P de Moerloose ◽

Ph Minazio ◽

G Reber ◽

A Perrier ◽

H Bounameaux

Keyword(s):

Pulmonary Embolism ◽

Screening Tool ◽

Screening Tests ◽

Enzyme Linked Immunosorbent Assay ◽

Elisa Assay ◽

Elisa Technique ◽

Diagnostic Step ◽

D Dimer ◽

Rule Out ◽

Latex Test

SummaryD-dimer (DD), when measured by a quantitative enzyme-linked immunosorbent assay (ELISA), is a valuable test to exclude venous thromboembolism (VTE). However, DD ELISA technique is not appropriate for emergency use and the available agglutination latex assays are not sensitive enough to be used as an alternative to rule out the diagnosis of VTE. Latex assays could still be used as screening tests. We tested this hypothesis by comparing DD levels measured by ELISA and latex assays in 334 patients suspected of pulmonary embolism. All but one patient with a positive (DD ≥500 ng/ml) latex assay had DD levels higher than 500 ng/ml with the ELISA assay. Accordingly, ELISA technique could be restricted to patients with a negative result in latex assay. This two-step approach would have spared about 50% of ELISA in our cohort. In conclusion, our data indicate that a latex test can be used as a first diagnostic step to rule out pulmonary embolism provided a negative result is confirmed by ELISA and the performance of the latex assay used has been assessed properly.

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Cutaneous Leishmaniasis in Algeria; Highlight on the Focus of M’Sila

Microorganisms ◽

10.3390/microorganisms9050962 ◽

2021 ◽

Vol 9 (5) ◽

pp. 962

Author(s):

Razika Beniklef ◽

Karim Aoun ◽

Karim Boudrissa ◽

Meriem Ben Abid ◽

Kamel Cherif ◽

...

Keyword(s):

Risk Factors ◽

Visceral Leishmaniasis ◽

Cutaneous Leishmaniasis ◽

Leishmania Infantum ◽

Psammomys Obesus ◽

Phlebotomus Papatasi ◽

Wild Rodents ◽

High Incidence ◽

Medical Importance ◽

Underlying Risk

Algeria ranks second after Afghanistan for the incidence of cutaneous leishmaniasis (CL) worldwide. Here, we report a 34-years retrospective analysis of CL in Algeria and focused on the most affected region, the M’Sila province. All 66 cutaneous isolates corresponded to Leishmania (L.) major. Our study of the sandfly and rodent fauna further highlighted the high density of Phlebotomus papatasi and additional phlebotomine species of medical importance, not previously identified in M’Sila. Wild rodents belonging to nine species were trapped in M’Sila, and Psammomys obesus and Meriones shawi were found infected by L. major. In addition, Leishmania infantum was isolated from two visceral leishmaniasis cases, one dog and its proven vectors (P. perniciosus, P. longicuspis, and P. perfiliewi) inventoried during the survey. The high incidence of CL in the M’Sila province is likely a consequence of the increase in minimum temperatures recorded that constitutes suitable conditions for establishing a high endemicity and leads to an explosive rise in leishmaniases cases in this region. A thorough investigation of the underlying risk factors is urgently needed to detect new cases earlier. All these would improve the preparedness to fight the disease.

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Canine visceral leishmaniasis in riverside communities of the Cuiabá river watershed

Semina Ciências Agrárias ◽

10.5433/1679-0359.2019v40n6supl2p3313 ◽

2019 ◽

Vol 40 (6Supl2) ◽

pp. 3313

Author(s):

Valéria Régia Franco Sousa ◽

Álvaro Felipe de Lima Ruy Dias ◽

Juliana Yuki Rodrigues ◽

Mariana de Medeiros Torres ◽

Janaína Marcela Assunção Rosa Moreira ◽

...

Keyword(s):

Bone Marrow ◽

Visceral Leishmaniasis ◽

Urban Areas ◽

Axenic Culture ◽

Antibody Test ◽

Enzyme Linked Immunosorbent Assay ◽

Lutzomyia Longipalpis ◽

Cross Sectional ◽

Mato Grosso ◽

River Watershed

Visceral Leishmaniasis (VL) is a parasitic zoonosis expanding in Brazil. Several municipalities in the state of Mato Grosso including those on the river Cuiabá have reported the incidence of both human and canine cases and the identification of sandfly vector, Lutzomyia longipalpis and Lu. cruzi. Dogs are considered the main reservoir of Leishmania chagasi in the urban areas, hence, we devised a cross-sectional study aimed at assessing the prevalence of the infection in the dogs of riverside communities on Cuiabá River watershed by parasitological (parasitic isolation in culture), serological, and molecular methods. Of the 248 surveyed dogs, 24 were positive in enzyme linked immunosorbent assay (ELISA) or immunofluorescence antibody test (IFAT), with a prevalence of 9.7%. The riverside communities located in the town of Santo Antonio do Leverger displayed a higher prevalence of the disease than the cities of Cuiabá and Várzea Grande; however, the difference was not statistically significant (p > 0.05). Dogs born in the communities had a 3.24-fold higher risk of acquiring the infection. Promastigote were isolated in the axenic culture from the bone marrow samples and intact skin. Further, DNA of Leishmania sp. was detected in the bone marrow samples, lymph nodes, leukocyte cover, and skin of only one examined dog. These samples were sequenced and they showed 99% homology to L. infantum. To conclude, we observed a higher prevalence of infection in Riverside communities of Santo Antonio do Leverger and the confirmation of autochthony in these areas justifies the surveillance actions to minimise the risk of transmission within the riverine community itself, besides its dissemination to other areas by tourism.

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A comparison between enzyme immunoassay and HPLC for ochratoxin A detection in green, roasted and instant coffee

Brazilian Archives of Biology and Technology ◽

10.1590/s1516-89132007000200020 ◽

2007 ◽

Vol 50 (2) ◽

pp. 349-359 ◽

Cited By ~ 21

Author(s):

Simone Fujii ◽

Elisabete Yurie Sataque Ono ◽

Ricardo Marcelo Reche Ribeiro ◽

Fernanda Garcia Algarte Assunção ◽

Cássia Reika Takabayashi ◽

...

Keyword(s):

Monoclonal Antibody ◽

Ochratoxin A ◽

High Performance ◽

Screening Tool ◽

Correlation Coefficients ◽

Enzyme Linked Immunosorbent Assay ◽

Elisa Assay ◽

Quality And Safety ◽

Green Coffee ◽

Instant Coffee

An indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) for ochratoxin A (OTA) detection in green, roasted and instant coffees was developed using anti-OTA monoclonal antibody. Immunological reagents prepared were OTA-BSA (4.76 mg/mL), anti-OTA.7 MAb (2x10³-fold dilution) and HRP-anti IgG (10³-fold dilution). The detection limit was 3.73 ng OTA/g and correlation coefficients (r) between this immunoassay and high performance liquid chromatography were 0.98 for green coffee, 0.98 for roasted and 0.86 for instant. OTA levels detected by ic-ELISA were higher than by HPLC, with ELISA/HPLC ratio of 0.66 - 1.46 (green coffee), 0.96 - 1.11 (roasted) and 0.93 - 1.82 (instant). ELISA recoveries for OTA added to coffee (5 - 70 ng/g) were 81.53 % for green coffee, 46.73 % for roasted and 64.35 % for instant, while recoveries by HPLC were 80.54 %, 45.91 % and 55.15 %, respectively. Matrices interferences were minimized by samples dilution before carrying out the ELISA assay. The results indicate that MAb-based ic-ELISA could be a simple, sensitive and specific screening tool for OTA detection, contributing to quality and safety of coffee products.

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