Laura Iniesta

Leishmania infantum-specific IgG, IgG1 and IgG2 antibody responses in healthy and ill dogs from endemic areas: Evolution in the course of infection and after treatment

The expression of IgG, IgG1 and IgG2 specific antibodies for Leishmania infantum was studied in five groups of dogs in Catalonia (Spain): I, 99 asymptomatic dogs (infected and uninfected) from a highly endemic area for leishmaniosis; II, 139 untreated dogs with clinically patent leishmaniosis; III, 11 naturally infected asymptomatic dogs monitored for up to 5 years since they were found seropositive to Leishmania antigen and without treatment; IV, 25 naturally infected dogs with clinically patent leishmaniosis and treated with either meglumine antimoniate and allopurinol or allopurinol alone and V, six experimentally infected dogs, treated with meglumine antimoniate and controlled for 5 years. The levels (ELISA units) of IgG, IgG1 and IgG2 in asymptomatic dogs (group I) were very variable (24+/-33, 32+/-31 and 26+/-31, respectively), and, as expected, lower than in ill dogs (group II) (168+/-34, 84+/-71 and 172+/-31, respectively). In both groups, the correlation between IgG and IgG2 levels (r=0.95, P<0.001 in group I and r=0.63, P<0.001 in group II) was higher than between IgG and IgG1 levels (r=0.01, P>0.05 in group I and r=0.31, P<0.001 in group II). In group III, IgG and IgG2 expression increased during infection, while IgG1 expression remained the same. In dogs of group IV, IgG levels after 1 year of treatment decreased more in responsive (mean values, 163+/-42 before treatment (b.t.) and 100+/-36 after treatment (a.t.)) than in unresponsive dogs (158+/-29 b.t. and 124+/-51 a.t.), especially for IgG1 (94+/-89 b.t. and 20+/-21 a.t. in responsive dogs and 35+/-25 b.t. and 22+/-13 a.t. in unresponsive dogs) rather than for IgG2 (156+/-16 b.t. and 114+/-45 a.t. in responsive and 151+/-11 b.t. and 125+/-36 a.t. in unresponsive dogs). Similar results were observed in the evolution of experimentally infected animals after consecutive and specific treatments. Overall results show the great variation in Leishmania-specific IgG1 expression in asymptomatic and symptomatic dogs, their lack of correlation with that of IgG2 and chemotherapy is more effective in dogs with initially high expression of IgG1.

Diagnostic Techniques To Detect Cryptic Leishmaniasis in Dogs

ABSTRACT This study of several techniques for detecting cryptic leishmaniasis in dogs from areas in Spain where Leishmania infantum is highly endemic concludes that immunological techniques (enzyme-linked immunosorbent assay, immunofluorescence antibody test, Western blotting, delayed-type hypersensitivity reaction, and in vitro lymphocyte proliferation assay) do not clearly differentiate between noninfected and infected asymptomatic dogs and that culture and PCR are more reliable diagnostic tools.

Detection of Anti-Leishmania Immunoglobulin G Antibodies in Urine Specimens of Dogs with Leishmaniasis

ABSTRACT For years, anti-Leishmania immunoglobulin G (IgG) antibodies have been detected in the sera of dogs living in areas of leishmaniasis endemicity. They have also been found in the aqueous humor and cerebrospinal fluid. In contrast, a review of the literature failed to identify the detection of anti-Leishmania antibodies in urine samples from dogs with leishmaniasis. Ninety-five dog urine samples were examined for the presence of anti-Leishmania antibodies by using a protein A enzyme-linked immunosorbent assay (ELISA). Twenty additional urine samples were collected from healthy dogs as controls. An IgG2 ELISA was performed on 26 urine samples found positive by the protein A ELISA. Twenty-three urine samples found positive to anti-Leishmania antibodies were tested for the local production of anti-Leishmania antibodies in the urinary tract by means of the urine antibody coefficient. Ten urine samples (and the corresponding serum samples) were compared by Western blot (WB) analysis. Thirty-five out of the 95 urine samples were found positive, 57 were found negative, and 3 were found inconclusive for antibody detection by the protein A ELISA. A high correlation between protein A and IgG2 levels was found in positive urine samples. Anti-Leishmania antibodies were present in the urine of dogs that had leishmaniasis, urinary protein/creatinine (U P/C) ratios of greater than one, and normal urinary sediment. A statistically significant correlation was observed between the U P/C ratios and the levels of anti-Leishmania antibodies in positive urine samples. In general, WB analysis and the urine antibody coefficient suggested that the presence of anti-Leishmania antibodies in urine was the consequence of an impairment of filtration of the glomerular barrier. However, in some dogs, WB analysis could be interpreted as suggesting that the presence of anti-Leishmania antibodies was caused, to a lesser extent, by local antibody production in the urinary tract. Antibody detection in urine could be a noninvasive method for leishmaniasis diagnosis and prognosis in dogs with glomerulonephropathies.

Transfusion-transmitted leishmaniasis: a practical review

In the Balearic Islands, as in other areas in southern Europe, there are a significant proportion of asymptomatic Leishmania infantum–infected blood donors. Theoretically, these donors may represent an important challenge for blood transfusion safety. However, despite an active search of multiply transfused patients, there have been, so far, no cases of transfusion‐transmitted leishmaniasis (TTL) in our region. On the other hand, there is scarce evidence of the TTL in the literature. A review of asymptomatic Leishmania‐infected blood donors’ studies in endemic areas and TTL reports published in the English literature were performed, to ascertain the factors that determine the real risk of transfusion transmission of Leishmania.

Pathogen inactivation technology applied to a blood component collected from an asymptomatic carrier of Leishmania infantum: a case report

Asymptomatic Leishmania infections have been the main cause of transfusion transmission in endemic areas. Polymerase chain reaction has been used to detect L. infantum DNA in the peripheral blood of asymptomatic Leishmania carriers. In our region, the prevalence of asymptomatic L. infantum infection in donors is markedly high (5·9% of donors studied). We investigated the ability of pathogen inactivation technology, using amotosalen and UVA illumination, to eliminate L. infantum in a blood component collected from an asymptomatic L. infantum infected donor. This is the first report of the INTERCEPT system being used to eliminate a parasite from a component collected from a donor.

The challenge of discordant serology in Chagas disease: The role of two confirmatory techniques in inconclusive cases

Serodiscordance in Chagas disease (CD) remains a challenge since individuals with inconclusive results are clinically complicated to manage. This work, conducted outside the endemic area, aims to compare two different confirmatory techniques for the diagnosis of CD in individuals without a definitive diagnosis, to analyze the performance of the screening techniques in this group of patients, and to describe the serological follow-up of these subjects over time. Sera from 48 individuals with repeatedly discordant results by one recombinant enzyme immunoassay (r-ELISA) and one native ELISA (n-ELISA), were included in the study. Confirmatory procedures were performed through TESA-blot, using trypomastigote antigens of Trypanosoma cruzi, and in-house WB (IH-WB) using a lysate from T. cruzi epimastigotes. Of the 48 sera, TESA-blot confirmed 22 (45.8%) cases and IH-WB 17 (35.4%). Both techniques showed a substantial agreement (k = 0.604). Confirmation defined as the positivity of one of the ELISA and at least one of the confirmatory tests was reached in 24/48 (50%) cases. We found a great dispersion of r-ELISA index values, especially among individuals with confirmatory negative results, ranging from 0.03-6.2. Additionally, n-ELISA yielded a better performance than r-ELISA in this cohort of patients, showing a significantly greater agreement with the confirmatory methods. Our results indicate that either confirmatory test could be an efficient tool to solve inconclusive cases regardless of which form of the parasites life cycle they use. Also, most individuals remain with discordant serology throughout the short-term follow-up period time of study. Finally, we consider that it is necessary to establish a reference test feasible and commercialized in all areas to solve the problem of inconclusive results.