Kinetics of foot-and-mouth disease vaccine-induced antibody responses in buffaloes (Bubalus bubalis): avidity ELISA as an alternative to the virus neutralization test.

The role of water buffaloes in foot-and-mouth disease (FMD) epidemiology as one of the major hosts of the virus that can develop persistent asymptomatic infection highlights the importance of sustaining surveillance on the antibody response elicited by vaccination in these animals. There is gap in the knowledge on how serological assays that measure antibodies against capsid proteins perform with buffalo samples and which would be the most reliable test to substitute the virus neutralization test (VNT) a cumbersome and low-throughput tool for field surveillance. Alternatively, the liquid-phase blocking sandwich ELISA (LPBE) is commonly used. Previous data from our laboratory demonstrated that the vaccine-induced antibodies assessed by the LPBE yielded low specificity with buffaloes' samples. In contrast, a single-dilution avidity ELISA (AE) aimed to detect high-avidity antibodies against exposed epitopes, combined with an indirect ELISA (IE) to assess IgG levels, produced more reliable results. Here we analyzed for the first time the kinetics of the antibodies induced by vaccination in two different buffalo herds (n = 91) over 120 days using AE, IE, LPBE, and the VNT. Kinetics were similar in the different assays, with an increase of antibodies between 0- and 14-days post-vaccination (dpv) which were maintained thereafter. VNT and AE results were concordant (Kappa value = 0.76), and both assays revealed a decay in the antibody response in calves with maternal antibodies at 90 and 120 dpv, which was not evidenced by the LPBE. These results show that kinetics of antibody responses to FMD vaccination are similar in buffalo and cattle, and support the use of indirect ELISA assays, in particular Avidity ELISA, as alternatives to the VNT for vaccine-immunity monitoring irrespectively of the animal's passive or active immune status.

Modified ELISA for antibody avidity evaluation: The need for standardization.

Antibody avidity is an important parameter to evaluate immune response, being useful to evaluate vaccine responses and helping to distinguish acute and latent infection. The antibody avidity can be measured by different methods, yet the most common is a modified ELISA. The utilization of commercial kits or in-house methods to evaluate antibody avidity have been adopted more and more, although the lack of standardization between different assays may generate a lot of variation in the process, making it hard to compare the results generated.

Effect of chaotropes in Chagas disease and leishmaniasis cross-reacting serology assays for epidemiological surveys

Abstract INTRODUCTION: Serological cross-reactivity between leishmaniasis and Chagas disease, especially at low titers, leads to difficulties of the seroepidemiological interpretation. METHODS: We have studied the ability of urea as a chaotrope to select high-avidity antibodies in IgG ELISA, thus reducing low-avidity IgG cross-reactivity in serologically positive samples in both assays. RESULTS: Using 0.5M urea for diluting the sample efficiently defined leishmaniasis or double infections in high-avidity IgG ELISA and eliminated false-positive results. CONCLUSIONS: The use of a chaotropic diluting agent is useful for improving the specificity of Chagas disease and leishmaniasis immunoassays.

Effect of chaotropes in Chagas disease and leishmaniasis cross-reacting serology assays for epidemiological surveys

INTRODUCTION: Serological cross-reactivity between leishmaniasis and Chagas disease, especially at low titers, leads to difficulties of the seroepidemiological interpretation. METHODS: We have studied the ability of urea as a chaotrope to select high-avidity antibodies in IgG ELISA, thus reducing low-avidity IgG cross-reactivity in serologically positive samples in both assays. RESULTS: Using 0.5M urea for diluting the sample efficiently defined leishmaniasis or double infections in high-avidity IgG ELISA and eliminated false-positive results. CONCLUSIONS: The use of a chaotropic diluting agent is useful for improving the specificity of Chagas disease and leishmaniasis immunoassays.

Passive acquisition of anti-Staphylococcus aureus antibodies by newborns via transplacental transfer and breastfeeding, regardless of maternal colonization

OBJECTIVE: To investigate the transmission of anti-Staphylococcus aureus (Sa) IgG, IgG1 and IgG2 via placental transfer and the transfer of IgA via the colostrum according to maternal Sa carrier status at delivery. METHODS: We evaluated anti-Sa IgG, IgG1 and IgG2 in maternal and cord sera and IgA in colostrum from a case (n=49, Sa+) and a control group (n=98, Sa-). RESULTS: Of the 250 parturients analyzed for this study, 49 were nasally colonized with S. aureus (prevalence of 19.6%). Ninety-eight non-colonized subjects were selected for the control group. The anti-Sa IgG, IgG1 and IgG2 levels and the IgG avidity indexes in the maternal and cord sera did not differ between the groups, with a low transfer ratio of anti-Sa IgG to the newborns in both groups. The anti-Sa IgG2 titers were significantly higher than the IgG1 titers in the maternal and cord sera. Inversely, the transfer ratios were higher for anti-Sa IgG1 compared with IgG2; however, no differences between the groups were detected. The Sa-specific IgA levels and avidity indexes in the colostrum were equivalent between groups. CONCLUSIONS: Maternal Sa nasal colonization at delivery is not associated with higher antibody levels in the mother or newborns. The high titers of anti-Sa IgG2 found in the cord serum indicate a greater reactivity with non-protein antigens, which may further contribute to the susceptibility to staphylococcal infections at birth. The presence of IgA in the colostrum with avidity to S. aureus reinforces the importance of breastfeeding shortly after birth.

Kinetics and avidity of anti-Toxocara antibodies (IgG) in rabbits experimentally infected with Toxocara canis / Cinética e avidez de anticorpos (IgG) anti-Toxocara em coelhos experimentalmente infectados com Toxocara canis

An evaluation was made of the kinetics and avidity of anti-Toxocara antibodies (IgG) in rabbits experimentally infected with embryonated Toxocara canis eggs. Seventeen four month old New Zealand White rabbits were distributed into two groups. In the experimental group, twelve rabbits were infected orally with 1,000 embryonated T. canis eggs. A second group (n = 5), uninfected, was used as a control. Serum samples were collected for analysis on days 7, 14, 21, 28 and 60 post-infection (DPI). An indirect ELISA test was performed to evaluate the reactivity index (RI) of IgG anti-T. canis antibodies and to calculate the avidity index (AI). The animals showed seroconversion from the 14th DPI, with high AI (over 50%) except for one animal, which presented an intermediate AI. At 60 DPI, all the animals were seropositive and maintained a high AI. The data indicated that specific IgG antibodies formed early (14 DPI) in rabbits infected with T. canis, with a high avidity index that persisted throughout the course of the infection.(AU)

O objetivo deste estudo foi o de avaliar a cinética e a avidez de anticorpos anti-Toxocara canis, em coelhas infectadas experimentalmente com ovos embrionados de Toxocara canis. Foram utilizados 17 coelhos New Zealand de linhagem branca, com quatro meses de idade, distribuídos em dois grupos. No grupo experimental, doze coelhas foram infectadas, oralmente, com 1.000 ovos larvados de T. canis. Um segundo grupo (n=5), não infectado, foi utilizado como controle. Nos dias 7, 14, 21, 28 e 60 pós-infecção (DPI), foram coletadas amostras de soro para análise. O teste de ELISA indireto foi realizado para avaliar o índice de reatividade (IR) de anticorpos IgG anti-T. canis e para cálculo do índice de avidez (IA). A soroconversão nos animais ocorreu a partir do140 DPI, com verificação de alto IA (superior a 50%), com exceção de um animal, que apresentou médio IA. Aos 60 DPI, todos os animais foram soropositivos e mantiveram alto IA. Os dados mostram que em coelhos infectados por T. canis, anticorpos IgG específicos formam-se precocemente (14 DPI), apresentando alto índice de avidez e que se mantém durante o curso da infecção.(AU)

Kinetics and avidity of anti-Toxocara antibodies (IgG) in rabbits experimentally infected with Toxocara canis / Cinética e avidez de anticorpos (IgG) anti-Toxocara em coelhos experimentalmente infectados com Toxocara canis

Abstract An evaluation was made of the kinetics and avidity of anti-Toxocara antibodies (IgG) in rabbits experimentally infected with embryonated Toxocara canis eggs. Seventeen four month old New Zealand White rabbits were distributed into two groups. In the experimental group, twelve rabbits were infected orally with 1,000 embryonated T. canis eggs. A second group (n = 5), uninfected, was used as a control. Serum samples were collected for analysis on days 7, 14, 21, 28 and 60 post-infection (DPI). An indirect ELISA test was performed to evaluate the reactivity index (RI) of IgG anti-T. canis antibodies and to calculate the avidity index (AI). The animals showed seroconversion from the 14th DPI, with high AI (over 50%) except for one animal, which presented an intermediate AI. At 60 DPI, all the animals were seropositive and maintained a high AI. The data indicated that specific IgG antibodies formed early (14 DPI) in rabbits infected with T. canis, with a high avidity index that persisted throughout the course of the infection.

Resumo O objetivo deste estudo foi o de avaliar a cinética e a avidez de anticorpos anti-Toxocara canis, em coelhas infectadas experimentalmente com ovos embrionados de Toxocara canis. Foram utilizados 17 coelhos New Zealand de linhagem branca, com quatro meses de idade, distribuídos em dois grupos. No grupo experimental, doze coelhas foram infectadas, oralmente, com 1.000 ovos larvados de T. canis. Um segundo grupo (n=5), não infectado, foi utilizado como controle. Nos dias 7, 14, 21, 28 e 60 pós-infecção (DPI), foram coletadas amostras de soro para análise. O teste de ELISA indireto foi realizado para avaliar o índice de reatividade (IR) de anticorpos IgG anti-T. canis e para cálculo do índice de avidez (IA). A soroconversão nos animais ocorreu a partir do140 DPI, com verificação de alto IA (superior a 50%), com exceção de um animal, que apresentou médio IA. Aos 60 DPI, todos os animais foram soropositivos e mantiveram alto IA. Os dados mostram que em coelhos infectados por T. canis, anticorpos IgG específicos formam-se precocemente (14 DPI), apresentando alto índice de avidez e que se mantém durante o curso da infecção.

Kinetics and avidity of anti-Toxocara antibodies (IgG) in rabbits experimentally infected with Toxocara canis

Abstract An evaluation was made of the kinetics and avidity of anti-Toxocara antibodies (IgG) in rabbits experimentally infected with embryonated Toxocara canis eggs. Seventeen four month old New Zealand White rabbits were distributed into two groups. In the experimental group, twelve rabbits were infected orally with 1,000 embryonated T. canis eggs. A second group (n = 5), uninfected, was used as a control. Serum samples were collected for analysis on days 7, 14, 21, 28 and 60 post-infection (DPI). An indirect ELISA test was performed to evaluate the reactivity index (RI) of IgG anti-T. canis antibodies and to calculate the avidity index (AI). The animals showed seroconversion from the 14th DPI, with high AI (over 50%) except for one animal, which presented an intermediate AI. At 60 DPI, all the animals were seropositive and maintained a high AI. The data indicated that specific IgG antibodies formed early (14 DPI) in rabbits infected with T. canis, with a high avidity index that persisted throughout the course of the infection.

Resumo O objetivo deste estudo foi o de avaliar a cinética e a avidez de anticorpos anti-Toxocara canis, em coelhas infectadas experimentalmente com ovos embrionados de Toxocara canis. Foram utilizados 17 coelhos New Zealand de linhagem branca, com quatro meses de idade, distribuídos em dois grupos. No grupo experimental, doze coelhas foram infectadas, oralmente, com 1.000 ovos larvados de T. canis. Um segundo grupo (n=5), não infectado, foi utilizado como controle. Nos dias 7, 14, 21, 28 e 60 pós-infecção (DPI), foram coletadas amostras de soro para análise. O teste de ELISA indireto foi realizado para avaliar o índice de reatividade (IR) de anticorpos IgG anti-T. canis e para cálculo do índice de avidez (IA). A soroconversão nos animais ocorreu a partir do140 DPI, com verificação de alto IA (superior a 50%), com exceção de um animal, que apresentou médio IA. Aos 60 DPI, todos os animais foram soropositivos e mantiveram alto IA. Os dados mostram que em coelhos infectados por T. canis, anticorpos IgG específicos formam-se precocemente (14 DPI), apresentando alto índice de avidez e que se mantém durante o curso da infecção.

Antibody avidity following secondary dengue virus type 2 infection across a range of disease severity.

BACKGROUND: The four dengue virus serotypes (DENV1-4) are responsible for the most prevalent mosquito-borne viral illness in humans. DENV causes a spectrum of disease from self-limiting dengue fever (DF) to severe, life-threatening dengue hemorrhagic fever/dengue shock syndrome (DHF/DSS). Antibodies from one infection can contribute to either protection or increased disease severity in a subsequent infection with a distinct DENV serotype. The effectiveness of the antibody response is modulated by both the affinity and avidity of the antibody/antigen interaction. OBJECTIVES: We investigated how antibody avidity developed over time following secondary DENV2 infection across different disease severities. STUDY DESIGN: We analyzed sera from 42 secondary DENV2-infected subjects (DF, n=15; DHF, n=16; DSS, n=11) from a pediatric hospital-based dengue study in Nicaragua. IgG avidity against DENV2 virions was measured in samples collected during acute and convalescent phases as well as 3, 6, and 18 months post-illness using a urea enzyme-linked immunosorbent assay. RESULTS: The data show a significant increase in avidity from acute to convalescent phase followed by a decrease from convalescent phase to 3 months post-symptom onset, then a plateau. Linear regression analysis comparing antibody avidity between disease severity groups over time indicate that individuals with more severe disease (DHF/DSS) experienced greater decay in antibody avidity over time compared to less severe disease (DF), and ROC curve analysis showed that at 18 months post-illness, lower avidity was associated with previously having experienced more severe disease. CONCLUSIONS: These data suggest that increased dengue disease severity is associated with lower antibody avidity at later time-points post-illness.

Avaliação comparativa entre o desempenho dos reagentes paratoxoplasmose AxSYM, VIDAS e VIDIA em um país latinoamericano com alta soroprevalência / Comparative evaluation of AxSYM, VIDAS and VIDIA toxoplasmosis reagent performance in a high seroprevalence Latin American country

Aims: The purpose of this study was to compare the performance of three automated immunoassays for the detection of IgM and IgG Toxoplasma gondii antibodies using sera of pregnant women living in Colombia, a Latin American country with a high seroprevalence. Methods: A total of 905 sera were tested for IgM antibodies and 914 for IgG antibodies with AxSYM, VIDAS and VIDIA immunoassays. Discrepancies were resolved by using the dye test for IgG antibodies, and the ISAGA test for IgM. Results: The overall agreement between AxSYM, VIDAS and VIDIA assays was excellent for detection of IgG and IgM antibodies, and discrepancies were relatively rare (3.6% and 5.5% of sera for IgG and IgM antibodies, respectively). The performance of the three immunoassays was similar for the detection of IgG antibodies with high sensitivity (100.00% for VIDIA, 99.59% for VIDAS, 99.38% for AxSYM) and specificity (99.04% for VIDIA, 98.82% for AxSYM, 98.57% for VIDAS). The specificity for IgM antibodies was excellent for the three immunassays (99.88% for VIDIA, 99.76% for AxSYM and VIDAS). The sensitivity of the detection of IgM antibodies was higher with VIDIA (95.12%) than with VIDAS (76.74%) and AxSYM (61.90%) assays. The correlation between IgG titers was limited between AxSYM and VIDAS assays and between AxSYM and VIDIA assays, but was excellent between VIDIA and VIDAS assays. Conclusions: Our study performed with Latin American sera confirmed the excellent specificity of AxSYM, VIDAS and VIDIA assays for the detection of IgG and IgM antibodies already reported in other countries. The sensitivity of the detection of IgG antibodies was slightly higher with VIDIA than with VIDAS and AxSYM assays. The sensitivity of the detection of IgM antibodies was higher with VIDIA than with VIDAS and AxSYM assays.

Toxoplasmose e gestação: resultados perinatais e associação do teste de avidez de IgG com infecção congênita em gestantes com IgM anti-Toxoplasma gondii reagente / Toxoplasmosis and pregnancy: perinatal results and association of the IgG avidity test with congenital infection in pregnant women with positive anti-Toxoplasma gondii IgM

Objetivos: verificar os desfechos perinatais em gestantes com toxoplasmose aguda e se houve associação entre os resultados dos testes de avidez para anticorpos IgG anti-Toxoplasma gondii e a presença ou ausência de infecção fetal/neonatal. Métodos: um estudo transversal incluiu gestantes com diagnóstico sorológico de toxoplasmose apresentando IgM específica reagente, atendidas no Ambulatório de Gestação de Alto Risco da Faculdade de Medicina da Universidade Federal de Mato Grosso do Sul, no período de novembro de 2002 a novembro de 2007. Resultados do teste de avidez de IgG demonstrando índices superiores a 30% foram considerados alta avidez, enquanto valores inferiores a 30% foram considerados baixa avidez. Definiram-se como sendo de infecção fetal e/ou neonatal os casos com resultado positivo para a reação em cadeia da polimerase no líquido amniótico ou com IgM específica para toxoplasmose reagente no sangue do recém-nascido. Resultados: considerando-se todas as gestantes referidas para o ambulatório de gestação de alto risco no período estudado, a frequência de gestantes com IgM anti-Toxoplasma gondii reagente foi de 10,8% (176/1.634). A taxa de infecção congênita nessas pacientes foi de 4% (7/176). O teste de avidez de IgG foi realizado em 162 gestantes (92%), encontrando-se avidez alta em 144 (88,9%). Houve associação (p=0,003) entre avidez alta e ausência de toxoplasmose fetal/neonatal na amostra estudada, com razão de prevalência de 13,4 (intervalo de confiança [IC] 95% 2,2-86,6). O Valor preditivo positivo do teste de avidez (probabilidade de infecção congênita com avidez baixa) foi de 22% (IC 95% 6%-47%), enquanto o valor preditivo negativo (probabilidade de ausência da infecção congênita com avidez alta) foi de 98% (IC 95% 94%-99%).

Aims: To verify the perinatal outcomes in pregnant women with acute toxoplasmosis, and to determine if there was association between the results of Toxoplasma gondii-specific IgG avidity test and the presence or absence of fetal/neonatal infection. Methods: A cross-sectional study included pregnant women with serological diagnosis of acute toxoplasmosis (presenting a positive Toxoplasma gondii-specific IgM test) attended at the outpatient unit for high-risk pregnancy of the Faculty of Medicine, Federal University of Mato Grosso do Sul, Brazil, in the period from November 2002 to November 2007. Test results demonstrating IgG avidity index above 30% were considered high avidity, while values below 30% were considered low avidity. Fetal and/or neonatal infection was defined by positive result for the polymerase chain reaction in amniotic fluid, or by a positive Toxoplasma gondii-specific IgM test in the newborn?s serum. Results: Considering all pregnant women referred to the outpatient unit for high-risk pregnancy in the period of study, frequency of pregnant women with positive Toxoplasma gondii-specific IgM was 10.8% (176/1.634). The rate of congenital infection in these patients was 4% (7/176). The IgG avidity test was performed in 162 patients (92% of the 176 pregnant women with positive IgM), and the avidity was high in 144 (88.9%). There was an association (p=0.003) between high avidity and no fetal/neonatal toxoplasmosis in our sample, with a prevalence ratio of 13.4 (confidence interval [CI] 95% 2.2-86.6). The positive predictive value of the avidity test (probability of congenital infection with a low avidity) was 22% (95% 6%-47%), while the negative predictive value (probability of absence of congenital infection with a high avidity) was 98% (95% CI 94% -99%).

Pattern of functional antibody activity against Haemophilus influenzae type b (Hib) in infants immunized with diphtheria-tetanus-pertussis/Hib Brazilian combination vaccine

We evaluated the functional activity of Haemophilus influenzae B (Hib) antibodies elicited in a group of infants immunized with the diphtheria-tetanus-pertussis vaccine combined with an Hib vaccine produced totally in Brazil after technological transfer of Hib vaccine production from Glaxo SmithKline, Belgium. Blood samples from immunized infants (N = 985) were collected for the determination of Hib antibodies. Total Ig and IgM and IgG subclasses of antibodies against polyribosyl ribitol phosphate (PRP) were analyzed by ELISA. Almost all vaccinees (97.56 percent, 961/985) developed a strong anti-PRP IgG antibody response (¡Ý1.0 ¦Ìg/mL), while an anti-PRP IgM response was observed in 64.24 percent (634/985) of them (¡Ý0.15 ¦Ìg/mL). Only 18.88 percent (186/985) of the infants in the group with high PRP antibody IgG concentrations (¡Ý1.0 ¦Ìg/mL) developed a high IgM antibody response. Anti-PRP IgG antibody levels were significantly higher than anti-PRP IgM. These results demonstrate the predominance of IgG antibodies over IgM antibodies in response to PRP, with a ratio of 17:1. IgG antibodies were predominantly of the IgG1 subclass. An increase in IgG avidity was also observed during the course of immunization.