Evaluation of the performance of three serological tests for diagnosis of Leishmania infantum infection in dogs using latent class analysis

Abstract Canine leishmaniasis (CanL) is a disease caused by Leishmania infantum. Serological methods are the most common diagnostic techniques used for the diagnosis of the CanL. The objective of our study was to estimate the sensitivity and specificity of one in-house ELISA kit (ELISA UNIZAR) and three commercially available serological tests (MEGACOR Diagnostik GmbH) including an immunochromatographic rapid test (FASTest LEISH®), an immunofluorescent antibody test (MegaFLUO LEISH®) and an enzyme-linked immunosorbent assay (MegaELISA LEISH®), using latent class models in a Bayesian analysis. Two hundred fifteen serum samples were included. The highest sensitivity was achieved for FASTest LEISH® (99.38%), ELISA UNIZAR (99.37%), MegaFLUO LEISH® (99.36%) followed by MegaELISA LEISH® (98.49%). The best specificity was obtained by FASTest LEISH® (98.43%), followed by ELISA UNIZAR (97.50%), whilst MegaFLUO LEISH® and MegaELISA LEISH® obtained the lower specificity (91.94% and 91.93%, respectively). The results of present study indicate that the immunochromatographic rapid test evaluated FASTest LEISH® show similar levels of sensitivity and specificity to the quantitative commercial tests. Among quantitative serological tests, sensitivity and specificity were similar considering ELISA or IFAT techniques.

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Diagnostic performance of a commercial ELISA used as a complementary test for bovine tuberculosis in two bovine herds with different disease status

Arquivo Brasileiro de Medicina Veterinária e Zootecnia ◽

10.1590/1678-4162-11071 ◽

2020 ◽

Vol 72 (1) ◽

pp. 1-8

Author(s):

P.M. Soares Filho ◽

A.K. Ramalho ◽

A.M. Silva ◽

M.A. Issa ◽

P.M.P.C. Mota ◽

...

Keyword(s):

Bovine Tuberculosis ◽

Diagnostic Performance ◽

Latent Class ◽

Serological Test ◽

Antibody Test ◽

Latent Class Models ◽

Lower Sensitivity ◽

Kappa Index ◽

Serological Tests ◽

Test Kit

ABSTRACT Bovine tuberculosis is a worldwide spread zoonotic disease. Intradermal tuberculinizations are the most used diagnostic tests in the world. Serological tests can be an ancillary diagnosis for bovine tuberculosis. The objective of this study was to evaluate the diagnostic performance of the ELISA Mycobacterium Bovis Antibody Test Kit IDEXX ™ in infected herds, which were in different disease control stages. One hundred and twenty animals from two dairy herds of Minas Gerais state, Brazil, were subjected to the ELISA serological test and the comparative cervical tuberculin test (CCT). Diagnostic test parameters were estimated using Bayesian latent class models and concordance between tests estimated by the frequentist approach. The ELISA test presented lower sensitivity than CCT in both herds. Its sensitivity was higher in the herd in sanitation process. Specificity estimates were above 95% in both herds. Kappa index indicated low concordance or even disagreement between tests. According to the results, the ELISA IDEXX should not be used as substitution for CCT. The tests must not be associated in series. Parallel association increased diagnostic sensitivity in the herd which was in the process of sanitation.

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Toward Diagnosing Leishmania infantum Infection in Asymptomatic Dogs in an Area Where Leishmaniasis Is Endemic

Clinical and Vaccine Immunology ◽

10.1128/cvi.00268-08 ◽

2009 ◽

Vol 16 (3) ◽

pp. 337-343 ◽

Cited By ~ 45

Author(s):

D. Otranto ◽

P. Paradies ◽

D. de Caprariis ◽

D. Stanneck ◽

G. Testini ◽

...

Keyword(s):

Leishmania Infantum ◽

Clinical Signs ◽

Antibody Test ◽

Diagnostic Methods ◽

Canine Leishmaniasis ◽

Enzyme Linked Immunosorbent Assay ◽

Canine Monocytic Ehrlichiosis ◽

Immunofluorescence Antibody Test ◽

Immunofluorescence Antibody

ABSTRACT The most frequently used diagnostic methods were compared in a longitudinal survey with Leishmania infantum-infected asymptomatic dogs from an area of Italy where leishmaniasis is endemic. In February and March 2005, 845 asymptomatic dogs were tested by an immunofluorescence antibody test (IFAT), a dipstick assay (DS), and an enzyme-linked immunosorbent assay (ELISA) for L. infantum and by IFAT for Ehrlichia canis. Dogs seronegative for L. infantum were further parasitologically evaluated by microscopic examination of lymph node tissues and PCR of skin samples. A total of 204 animals both serologically and parasitologically negative for L. infantum at the first sampling were enrolled in the trial and were further examined for canine leishmaniasis (CanL) and canine monocytic ehrlichiosis in November 2005 (i.e., the end of the first sandfly season) and March 2006 and 2007 (1- and 2-year follow-ups, respectively). At the initial screening, the overall rates of L. infantum seroprevalence were 9.5% by IFAT, 17.1% by ELISA, and 9.8% by DS and the overall rate of E. canis seroprevalence was 15%. The rates of concordance between the results of IFAT and DS were almost equal, whereas the rate of concordance between the results of IFAT and DS and those of the ELISA was lower. The results of the annual incidence of Leishmania infection were variable, depending on the test employed, with the highest values registered for PCR (i.e., 5.7% and 11.4% at the 1- and 2-year follow-ups, respectively), followed by ELISA, IFAT, and DS. Over the 2 years of observation, 55 animals (i.e., 26.9%) became positive for L. infantum by one or more diagnostic tests at different follow-up times, with 12.7% showing clinical signs related to CanL, while the remaining 87.3% were asymptomatic. A diagnostic scheme for assessment of the L. infantum infection status in asymptomatic dogs is suggested.

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Serological Diagnosis of Brucella Infections in Odontocetes

Clinical and Vaccine Immunology ◽

10.1128/cvi.00413-08 ◽

2009 ◽

Vol 16 (6) ◽

pp. 906-915 ◽

Cited By ~ 11

Author(s):

Gabriela Hernández-Mora ◽

Charles A. Manire ◽

Rocío González-Barrientos ◽

Elías Barquero-Calvo ◽

Caterina Guzmán-Verri ◽

...

Keyword(s):

Sensitivity And Specificity ◽

Brucella Melitensis ◽

Enzyme Linked Immunosorbent Assay ◽

Dot Blot ◽

Serum Samples ◽

Igg Antibody ◽

Serological Tests ◽

Diagnostic Potential ◽

Control Serum ◽

History Of

ABSTRACT Brucella ceti causes disease in Odontoceti. The absence of control serum collections and the diversity of cetaceans have hampered the standardization of serological tests for the diagnosis of cetacean brucellosis. Without a “gold” standard for sensitivity and specificity determination, an alternative approach was followed. We designed an indirect enzyme-linked immunosorbent assay (iELISA) that recognizes immunoglobulins G (IgGs) from 17 odontocete species as a single group. For the standardization, we used Brucella melitensis and Brucella abortus lipopolysaccharides, serum samples from seven resident odontocetes with no history of infectious disease displaying negative rose bengal test (RBT) reactions, and serum samples from seven dolphins infected with B. ceti. We compared the performance of the iELISA with those of the protein G ELISA (gELISA), the competitive ELISA (cELISA), and the immunofluorescence (IF) and dot blot (DB) tests, using 179 odontocete serum samples and RBT as the reference. The diagnostic potential based on sensitivity and specificity of the iELISA was superior to that of gELISA and cELISA. The correlation and agreement between the iELISA and the gELISA were relatively good (R i/g 2 = 0.65 and κi/g = 0.66, respectively), while the correlation and agreement of these two ELISAs with cELISA were low (R i/c 2 = 0.46, R g/c 2 = 0.37 and κi/c = 0.62, κg/c = 0.42). In spite of using the same anti-odontocete IgG antibody, the iELISA was more specific than were the IF and DB tests. An association between high antibody titers and the presence of neurological symptoms in dolphins was observed. The prediction is that iELISA based on broadly cross-reacting anti-dolphin IgG antibody would be a reliable test for the diagnosis of brucellosis in odontocetes, including families not covered in this study.

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rSodC is a potential antigen to diagnose Corynebacterium pseudotuberculosis by enzyme-linked immunoassay

AMB Express ◽

10.1186/s13568-020-01125-0 ◽

2020 ◽

Vol 10 (1) ◽

Author(s):

Antonio Pedro Fróes de Farias ◽

José Tadeu Raynal Rocha Filho ◽

Silvana Beutinger Marchioro ◽

Luan Santana Moreira ◽

Andressa Souza Marques ◽

...

Keyword(s):

Sensitivity And Specificity ◽

Recombinant Proteins ◽

Corynebacterium Pseudotuberculosis ◽

Semiarid Region ◽

Enzyme Linked Immunosorbent Assay ◽

Serum Samples ◽

Serological Tests ◽

Control Programs ◽

Epidemiological Surveys ◽

Purified Antigen

Abstract Caseous lymphadenitis (CL) is a chronic infectious disease that affects sheep and goats. Many serological tests have been developed to detect the disease; one of the most widely used is the enzyme-linked immunosorbent assay (ELISA), due to its advantages, which include acceptable cost-effectiveness, applicability, sensitivity and specificity. ELISA formulations using recombinant proteins can exhibit significant sensitivity and specificity when using a single purified antigen. DTxR, Trx, TrxR, LexA, SodC, SpaC, NanH, and PknG recombinant proteins can be considered target proteins for ELISA development due to its extracellular or on the cell surface location, which allows a better recognition by the immune system. Therefore, the objectives of this study were to evaluate the antigenic reactivity of Corynebacterium pseudotuberculosis recombinant proteins in goat and sheep serum. Of eight proteins evaluated, rSodC was selected for validation assays with small ruminant serum samples from the semiarid region of the state of Bahia, Brazil. Validation assays with goat serum samples showed that ELISA-rSodC presented sensitivity and specificity of 96% and 94%, respectively. Validation assays with sheep serum showed that ELISA-rSodC exhibited sensitivity and specificity of 95% and 98%, respectively. Analysis of 756 field serum samples showed that rSodC identified 95 positive samples (23%) in goats and 75 positive samples (21%) in sheep. The ELISA with recombinant SodC protein developed in this study discriminated positive and negative serum samples with high levels of sensitivity and specificity. This formulation is promising for epidemiological surveys and CL control programs. Trial registration AEC No 4958051018. 12/18/2018, retrospectively registered

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rSodC is a potential antigen to diagnose Corynebacterium pseudotuberculosis by enzyme-linked immunoassay

10.21203/rs.3.rs-54005/v2 ◽

2020 ◽

Author(s):

Antonio Pedro Fróes de Farias ◽

José Tadeu Raynal Rocha Filho ◽

Silvana Beutinger Marchioro ◽

Luan Santana Moreira ◽

Andressa Souza Marques ◽

...

Keyword(s):

Sensitivity And Specificity ◽

Recombinant Proteins ◽

Corynebacterium Pseudotuberculosis ◽

Semiarid Region ◽

Enzyme Linked Immunosorbent Assay ◽

Serum Samples ◽

Serological Tests ◽

Control Programs ◽

Epidemiological Surveys ◽

Purified Antigen

Abstract Caseous lymphadenitis (CL) is a chronic infectious disease that affects sheep and goats. Many serological tests have been developed to detect the disease; one of the most widely used is the enzyme-linked immunosorbent assay (ELISA), due to its advantages. ELISA formulations using recombinant proteins can exhibit significant sensitivity and specificity when using a single purified antigen. DTxR, Trx, TrxR, LexA, SodC, SpaC, NanH, and PknG recombinant proteins can be considered target proteins for ELISA development. Therefore, the objectives of this study were to evaluate the antigenic reactivity of Corynebacterium pseudotuberculosis recombinant proteins in goat and sheep serum. Of eight proteins evaluated, rSodC was selected for validation assays with small ruminant serum samples from the semiarid region of the state of Bahia, Brazil. Validation assays with goat serum samples showed that rSodC presented sensitivity and specificity of 96% and 94%, respectively. Validation assays with sheep serum showed that recombinant SodC exhibited sensitivity and specificity of 95% and 98%, respectively. Analysis of 756 field serum samples showed that rSodC identified 95 positive samples (23%) in goats and 75 positive samples (21%) in sheep. The ELISA with recombinant SodC protein developed in this study discriminated positive and negative serum samples with high levels of sensitivity and specificity. This formulation is promising for epidemiological surveys and CL control programs.

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Serodiagnosis of Visceral Leishmaniasis in Northeastern Italy: Evaluation of Seven Serological Tests

Microorganisms ◽

10.3390/microorganisms8121847 ◽

2020 ◽

Vol 8 (12) ◽

pp. 1847

Author(s):

Margherita Ortalli ◽

Daniele Lorrai ◽

Paolo Gaibani ◽

Giada Rossini ◽

Caterina Vocale ◽

...

Keyword(s):

Visceral Leishmaniasis ◽

Western Blot ◽

Rapid Test ◽

Antibody Test ◽

Screening Tests ◽

Serum Samples ◽

Serological Tests ◽

Northeastern Italy ◽

Indirect Immunofluorescent ◽

Immunofluorescent Antibody Test

This study compares the performance of seven assays, including two ELISA (Leishmania ELISA IgG + IgM, Vircell Microbiologists; Leishmania infantum IgG ELISA, NovaTec), three rK39-based immunochromatographic tests (rK39-ICTs) (Leishmania Dipstick Rapydtest, Apacor; On Site Leishmania IgG/IgM Combo Rapid Test, CTK Biotech; LEISHMANIA Strip quick Test, Cypress Diagnostic), one indirect immunofluorescent antibody test (IFAT) (Leishmania-Spot IF, BioMérieux), and one western blot (WB) (Leishmania WESTERN BLOT IgG, LDBio Diagnostics) for serodiagnosis of visceral leishmaniasis (VL). Serum samples from 27 VL patients living in northeastern Italy were analyzed, as well as the serum samples from 50 individuals in whom VL diagnosis was excluded. The WB and the IFAT had 96% sensitivity, followed by the ELISA (63% and 74%, respectively). The rK39-ICT exhibited the worst performance among the serological tests, with sensitivities ranging from 52% to 70%. By combining selected ELISA/ICT, the sensitivity of VL detection reached 89%. IFAT and WB outperformed ELISA and rK39-ICT by possessing optimal sensitivity, but their high cost and complexity of execution would not allow their employment as screening tests. In conclusion, the combination of easy-to-perform tests, such as ICT and ELISA, could improve sensitivity in the serodiagnosis of Mediterranean VL.

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COMPARISON OF A RAPID ENZYME-LINKED IMMUNOSORBANT ASSAY TEST WITH AN INDIRECT IMMUNOFLUORESCENT ANTIBODY TEST IN DIAGNOSING EHRLICHIA AND ANAPLASMA INFECTIONS IN DOGS

Trakia Journal of Sciences ◽

10.15547//tjs.2019.04.009 ◽

2019 ◽

Vol 17 (4) ◽

pp. 346-352

Author(s):

Kr. Gospodinova ◽

G. Zhelev ◽

V. Petrov

Keyword(s):

Serum Antibody ◽

Rapid Test ◽

Antibody Test ◽

Diagnostic Value ◽

Enzyme Linked Immunosorbent Assay ◽

Serum Samples ◽

Immunofluorescent Antibody ◽

Antibody Titers ◽

Indirect Immunofluorescent ◽

Immunofluorescent Antibody Test

PURPOSE: The objective of this study is to compare the diagnostic value of commercial enzyme-linked immunosorbent assay (ELISA) with indirect immunofluorescent antibody test (IFA) in detecting immunoglobulin-G (IgG) antibodies to Ehrlichia canis and Anaplasma phagocytophilum. METHODS: Seventy-four serum samples, obtained from dogs believed to be naturally infected with E. canis or A. phagocytophilum, were analyzed. RESULTS: By ELISA, 48 (64.9%) samples were found positive for IgG to E. canis, 10 (13.5%) to A. phagocytophilum, 12 (16.2%) to both E. canis and A. phagocytophilum, and in 4 (5.4%) samples no presence of antibodies was detected. The number of serologically positive dogs for IgG was 44 (59.5%) to E. canis, 10 (13.5%) to A. phagocytophilum, 16 (21.6%) to both E. canis and A. phagocytophilum, and 4 (5.4%) were determined negative by means of IFA. In most samples the antibody titer did not exceed 1:80 but in 5 it reached a level of 1:320, and in other 4 of even above 1:640. CONCLUSIONS: This study shows that IFA assay is more sensitive than commercial ELISA rapid test when serum antibody titers are low.

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Leishmania tarentolae and Leishmania infantum in humans, dogs and cats in the Pelagie archipelago, southern Italy

PLoS Neglected Tropical Diseases ◽

10.1371/journal.pntd.0009817 ◽

2021 ◽

Vol 15 (9) ◽

pp. e0009817

Author(s):

Roberta Iatta ◽

Jairo Alfonso Mendoza-Roldan ◽

Maria Stefania Latrofa ◽

Antonio Cascio ◽

Emanuele Brianti ◽

...

Keyword(s):

Leishmania Infantum ◽

Molecular Diagnostic ◽

Enzyme Linked Immunosorbent Assay ◽

Molecular Testing ◽

Molecular Diagnostic Techniques ◽

Serum Samples ◽

Serological Tests ◽

Leishmania Tarentolae ◽

Human Samples ◽

Animal Populations

Visceral leishmaniasis (VL) caused by Leishmania infantum is endemic in the Mediterranean basin with most of the infected human patients remaining asymptomatic. Recently, the saurian-associated Leishmania tarentolae was detected in human blood donors and in sheltered dogs. The circulation of L. infantum and L. tarentolae was investigated in humans, dogs and cats living in the Pelagie islands (Sicily, Italy) by multiple serological and molecular testing. Human serum samples (n = 346) were tested to assess the exposure to L. infantum by immunofluorescence antibody test (IFAT), enzyme-linked immunosorbent assay (ELISA) and Western blot (WB) and to L. tarentolae by IFAT. Meanwhile, sera from dogs (n = 149) and cats (n = 32) were tested for both Leishmania species by IFAT and all blood samples by specific sets of real time-PCR for L. infantum and L. tarentolae. The agreement between serological tests performed for human samples, and between serological and molecular diagnostic techniques for both human and animal samples were also assessed. Overall, 41 human samples (11.8%, 95% CI: 8.9–15.7) were positive to L. infantum (5.2%, 95% CI: 3.3–8.1), L. tarentolae (5.2%, 95% CI: 3.3–8.1) and to both species (1.4%, 95% CI: 0.6–3.3) by serology and/or molecular tests. A good agreement among the serological tests was determined. Both Leishmania spp. were serologically and/or molecularly detected in 39.6% dogs and 43.7% cats. In addition to L. infantum, also L. tarentolae circulates in human and animal populations, raising relevant public health implications. Further studies should investigate the potential beneficial effects of L. tarentolae in the protection against L. infantum infection.

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Evaluation of the NovaLisa™ Leishmania Infantum IgG ELISA in A Reference Diagnostic Laboratory in A Non-Endemic Country

Antibodies ◽

10.3390/antib8010020 ◽

2019 ◽

Vol 8 (1) ◽

pp. 20 ◽

Cited By ~ 1

Author(s):

Christen Stensvold ◽

Amalie Høst ◽

Salem Belkessa ◽

Henrik Nielsen

Keyword(s):

Western Blot ◽

Leishmania Infantum ◽

Antibody Test ◽

Cost Effective ◽

Enzyme Linked Immunosorbent Assay ◽

Serum Samples ◽

Diagnostic Laboratory ◽

Cost Effective Alternative ◽

Immunofluorescence Antibody Test ◽

Immunofluorescence Antibody

Anti-Leishmania antibodies may be detectable in patients with leishmaniasis. Here, we compared a commercial enzyme-linked immunosorbent assay (ELISA) for the detection of anti-Leishmania antibodies, with an immunofluorescence antibody test (IFAT) that is no longer commercially available. Eighty-six serum samples from 73 patients were tested. The results obtained by the NovaLisa™ Leishmania infantum IgG ELISA, interpreted according to the instructions of the manufacturer, but with a modified cut-off for borderline positive values, were compared with the IFAT results that were already available. Moreover, Leishmania Western blot IgG results were available for 43 of the samples. The overall concordance of ELISA and IFAT was 67%. The ELISA and IFAT tests scored as 24% and 15% of the samples being positive, respectively, while 13% and 33% scored as borderline-positive, respectively. Using a Western blot (WB) as the reference, the sensitivities and specificities for the positive plus borderline-positive samples combined was 95.5% (95% confidence interval (CI), 77.2–99.9%) and 81.0% (95% CI, 58.1–94.6%) for ELISA, and 95.5% (95% CI, 77.2–99.9%) and 42.9% (95% CI, 21.8–66.0%) for IFAT, respectively. Overall, the ELISA proved to be a cost-effective alternative to the IFAT, due to its higher accuracy and specificity, and with a consequently lower number of confirmatory WB tests being required. Lastly, we also present data on the associations between seroconversion and the type of leishmaniasis.

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Development of Recombinant Chimeric Antigen Expressing Immunodominant B Epitopes of Leishmania infantum for Serodiagnosis of Visceral Leishmaniasis

Clinical and Diagnostic Laboratory Immunology ◽

10.1128/cdli.12.5.647-653.2005 ◽

2005 ◽

Vol 12 (5) ◽

pp. 647-653 ◽

Cited By ~ 45

Author(s):

A. Boarino ◽

A. Scalone ◽

L. Gradoni ◽

E. Ferroglio ◽

F. Vitale ◽

...

Keyword(s):

Visceral Leishmaniasis ◽

Confidence Interval ◽

Leishmania Infantum ◽

Serological Test ◽

Antibody Test ◽

Point Of View ◽

Canine Leishmaniasis ◽

Enzyme Linked Immunosorbent Assay ◽

Gene Fragment ◽

Antigenic Properties

ABSTRACT Wild canids and domestic dogs are the main reservoir of zoonotic visceral leishmaniasis (VL) caused by Leishmania infantum (syn.: Leishmania chagasi). Serological diagnosis of VL is therefore important in both human and dog leishmaniasis from a clinical and epidemiological point of view. Routine diagnosis of VL is traditionally carried out by immunofluorescent antibody test (IFAT), which is laborious and difficult to standardize and to interpret. In the last decade, however, several specific antigens of Leishmania infantum have been characterized, allowing the development of a recombinant-based immunoassay. Among them, the whole open reading frame encoding K9 antigen, the gene fragment encoding the repetitive sequence of K26, and the 3′-terminal gene fragment of the kinesin-related protein (K39sub) were previously evaluated as diagnostic markers for canine leishmaniasis and proved to be independent in their antibody reactivity. Since sensitivity of serological test is usually higher in multiple-epitope format, in this study the relevant epitopes of K9, K26, and K39 antigens were joined by PCR strategy to produce the chimeric recombinant protein. The resulting mosaic antigen was found highly expressed in Escherichia coli and efficiently purified by affinity chromatography. Antigenic properties of this recombinant antigen were evaluated by indirect enzyme-linked immunosorbent assay (ELISA) using a panel of human and dog sera previously characterized by parasitological and/or serological techniques. Chimeric ELISA showed 99% specificity in both human (n = 180) and canine (n = 343) control groups, while sensitivity was higher in canine VL (96%, n = 213) than in human VL (82%, n = 185). Accordingly, concordance between IFAT and canine chimeric ELISA (k = 0.95, 95% confidence interval = 0.93 to 0.98) was higher than between IFAT and human chimeric ELISA (k = 0.81, 95% confidence interval = 0.76 to 0.87). Results suggest the potential use of this new antigen for routine serodiagnosis of VL in both human and canine hosts.

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