Frequency of anti- Toxoplasma gondii IgA, IgM, and IgG antibodies in high-risk pregnancies, in Brazil

Abstract:

INTRODUCTION

Toxoplasmosis during pregnancy can be severe; thus, it is essential to diagnose the disease via serological tests.

METHODS

An enzyme-linked immunosorbent assay (ELISA) was used to investigate anti-Toxoplasma gondii immunoglobulin A (IgA), M (IgM) and G (IgG) antibodies in 62 high-risk pregnant women.

RESULTS

Forty-three (69.4%) women were positive for IgA, 31 (50%) for IgG, and 57 (91.9%) for IgM; 4 (6,5%) were positive for IgA but negative for IgM; 10 (16.1%) were negative for IgA and IgM but positive for IgG.

CONCLUSIONS

Testing for these antibodies can help diagnose infection in pregnant women, thereby contributing to clinical management.

Keywords:
Toxoplasma gondii; Acute infection; High-risk pregnancy

Congenital toxoplasmosis is a serious disease that occurs when the Toxoplasma gondii parasite crosses the placental barrier and affects the fetus during pregnancy. The severity of the disease may be associated with the type of strain acquired, the immune status of the mother, and the gestational period in which maternal infection and fetal transmission occurs^{1) (2}.

Among the different types of antibodies, the anti-Toxoplasma gondii immunoglobulin G (IgG) and immunoglobulin M (IgM) are the most commonly reported among pregnant women worldwide^{1) (3}, and the presence of anti-T. gondii antibodies is an important factor in determining the time of infection. IgA and IgM class antibodies are associated with recent infections (acute infection) while IgG class antibodies are associated with chronic infection^{3) (4) (5) (6}. However, the individual analysis of these antibodies does not differentiate the time of infection as IgA and IgM antibodies may be detectable for several months or even be undetectable in acute infections, and thus searching for more than one type of antibody is required to confirm the acute infection. In addition, the IgM antibody test is widely used to diagnose acute infections, as these antibodies usually appear 1 week after infection and may disappear within 6 months^{7) (8}. However, the IgA antibody test is not commonly used, and most studies only use this test to detect the disease in cases of congenital infection. Thus, the IgA antibody test is an important marker for T. gondii infection in newborn babies. On the other hand, IgM antibodies during infection can arise at the beginning of pregnancy but be undetectable in the newborn serum at birth^{6) (9) (10}. Furthermore, although the detection of IgA antibodies is similar to IgM antibodies, the peak titers of IgA antibodies occurs later^{8) (10}. However, few studies, especially in Brazil, have described the behavior and use of IgA in the diagnosis of infection in pregnant women. Thus, considering the severity of toxoplasmosis during pregnancy and the contribution of serological diagnosis to identifying the disease, we aimed to investigate the frequency of anti-T. gondii IgM, IgA, and IgG antibodies to identify the incidence of infection during high-risk pregnancies among women attending a Brazilian National Health Service tertiary teaching hospital.

Peripheral blood samples were collected in dry tubes to obtain serum and were stored at -20°C until used to test for IgM, IgA, and IgG antibodies against T. gondii. The reactions were performed using an enzyme-linked immunosorbent assay (ELISA) with the ETI-TOXOK-A (for IgA), ETI-TOXOK-G (for IgG), and ETI-TOXOK-M (for IgM) commercial assays (DiaSorin, Italy) according to the manufacturer's instructions.

The mean age of the pregnant women was 24.9 ± 6.9 (range: 14-43). Table 1 shows the results of the anti-T. gondii antibody analysis, and Table 2 shows the serological profile of the T. gondii antibodies. IgM antibodies were more common than IgA antibodies (91.9% and 69.4%, respectively). However, IgA antibodies were detected without IgM antibodies in 4 samples (6.5%). This supports the results found by Fricker-Hidalgo et al., who found that 5 samples were positive for IgA but negative for IgM in pregnant women, as well as Roc et al., who identified 9 samples from neonates that were positive for IgA, only three of which were also positive for IgM^{11) (12}.

Table 1
Anti-Toxoplasma gondii IgA, IgM, and IgG antibodies (ELISA) in high-risk pregnant women (n=62) in São José do Rio Preto, São Paulo, Brazil.

• IgA: immunoglobulin A; IgM: immunoglobulin M; IgG: immunoglobulin G; ELISA: enzyme-linked immunosorbent assay.

Table 2
Anti-Toxoplasma gondii IgA, IgM, and IgG antibody profiles (ELISA) in high-risk pregnant women (n=62) in São José do Rio Preto, São Paulo, Brazil.

• IgA: immunoglobulin A; IgM: immunoglobulin M; IgG: immunoglobulin G; ELISA: enzyme-linked immunosorbent assay.

We also found that 10 (16.1%) samples were negative for IgA and IgM antibodies but were positive for IgG antibodies. These results may suggest reactivation of a past infection, but clinical and serological follow-up of these pregnant women as well as an IgG avidity test, which was not conducted in the current study, would be necessary to verify this hypothesis. Moreover, treatment of pregnant women can modify the results of serological tests, and this may have contributed to the negative results for the IgA and IgM classes of antibodies, as previously described^{13) (14}.

In addition, diagnosing toxoplasmosis in Brazil is difficult because there is no specific countrywide policy and no standard commercial assay to identify the type of infection hindering management of the different forms of the disease^{1) (13) (15) (16}.

In France there are flowcharts for interpretation of serological results in pregnant women to prevent congenital toxoplasmosis. Villard et al. proposed other flowcharts for other forms of the disease, such as congenital toxoplasmosis, immunosuppressed situations, and ocular toxoplasmosis, highlighting the importance of the serological test chosen in identifying infection and the subsequent prevention of the disease. The authors also described the importance of the interpretation of the results and the association of IgA with IgM class antibodies characterizing acute infection⁸. However, in Brazil, most public health services do not perform more than one serological assay during antenatal care, and most of them do not offer an avidity assay to estimate the date of maternal infection. Nonetheless, the use of these laboratory assays performed during pregnancy can help guide clinical research and confirm the presence or absence of acute maternal infection^{6) (14) (16}.

In conclusion, our study showed that the identification of anti-T. gondii IgA antibodies as well as routine testing for IgM and IgG antibodies can help diagnose toxoplasmosis during pregnancy thereby contributing to clinical management.

Ethicalconsiderations

This study was approved by the Research Ethics Committee of the São José do Rio Preto Medical School (FAMERP-CAAE 32259714.8.0000.5415). Serum samples from 62 high-risk pregnant women suspected of having toxoplasmosis or who had anti-T. gondii IgM antibodies at any time during pregnancy were analyzed. The pregnant women attended and were clinically evaluated in the High-Risk Antenatal Care and Fetal Medicine Outpatient Clinic of the Hospital de Base, Fundação Faculdade Regional de Medicina, São José do Rio Preto, São Paulo, Brazil.

Acknowledgments

We offer our deepest thanks to the Auckland University of Technology in New Zealand, particularly Professor Stephen Henry who provided library access.

• ¹
  Dubey JP, Lago EG, Gennari SM, Su C, Jones JL. Toxoplasmosis in humans and animals in Brazil: high prevalence, high burden of disease, and epidemiology. Parasitology 2012; 139:1375-1424.
• ²
  Bichara CNC, Canto GAC, Tostes CL, Freitas JJS, Carmo EL, Póvoa MM, et al. Incidence of congenital toxoplasmosis in the city of Belém, state of Pará, northern Brazil, determined by a neonatal screening program: preliminary results. Rev Soc Bras Med Trop 2012; 45:122-124.
• ³
  Reis MM, Tessaro MM, D'Azevedo PA. Toxoplasma-IgM and IgG-avidity in single samples from areas with a high infection rate can determine the risk of mother-to-child transmission. Rev Inst Med Trop Sao Paulo 2006; 48:93-98.
• ⁴
  Dhakal R, Gajurel K, Pomares C, Talucod J, Press CJ, Montoya JG. Significance of a positive Toxoplasma Immunoglobulin M test result in the United States. J Clin Microbiol 2015; 53:3601-3605.
• ⁵
  Tanyuksel M, Guney C, Araz E, Saracli MA, Doganci L. Performance of the immunoglobulin G avidity and enzyme immunoassay IgG/IgM screening tests for differentiation of the clinical spectrum of toxoplasmosis. J Microbiol 2004; 42:211-215.
• ⁶
  Dard C, Fricker-Hidalgo H, Brenier-Pinchart MP, Pelloux H. Relevance of and New Developments in Serology for Toxoplasmosis. Trends Parasitol 2016; 32:492-506.
• ⁷
  Paquet C, Yudin MH, Society of Obstetricians Gynaecologists of Canada. Toxoplasmosis in pregnancy: prevention, screening, and treatment. J Obstet Gynaecol Can 2013; 35:78-81.
• ⁸
  Villard O, Cimon B, L'Ollivier C, Fricker-Hidalgo H, Godineau N, Houze S, et al. Serological diagnosis of Toxoplasma gondii infection. Diagn Microbiol Infect Dis 2016; 84:22-33.
• ⁹
  Gontijo da Silva M, Clare Vinaud M, de Castro AM. Prevalence of toxoplasmosis in pregnant women and vertical transmission of Toxoplasma gondii in patients from basic units of health from Gurupi, Tocantins, Brazil, from 2012 to 2014. PLoS One 2015; 10:e0141700.
• ¹⁰
  Bessières MH, Berrebi A, Rolland M, Bloom MC, Roques C, Cassaing S, et al. Neonatal screening for congenital toxoplasmosis in a cohort of 165 women infected during pregnancy and influence of in utero treatment on the results of neonatal tests. Eur J Obstet Gynecol Reprod Biol 2001; 94:37-45.
• ¹¹
  Fricker-Hidalgo H, Cimon B, Chemla C, Darde ML, Delhaes L, L'Ollivier C, et al. Toxoplasma seroconversion with negative or transient immunoglobulin M in pregnant women: myth or reality? A French multicentre retrospective study. J Clin Microbiol 2013; 51:2103-2111.
• ¹²
  Roc ML, Palacián MP, Lomba E, Monforte ML, Rebaje V, Revillo Pinilla MJ. Serologic diagnosis of congenital toxoplasmosis. Enferm Infecc Microbiol Clin 2010; 28:517-519.
• ¹³
  Rodrigues IM, Costa TL, Avelar JB, Amaral WN, Castro AM, Avelino MM. Assessment of laboratory methods used in the diagnosis of congenital toxoplasmosis after maternal treatment with spiramycin in pregnancy. BMC Infect Dis 2014; 14:349.
• ¹⁴
  Pomares C, Montoya JG. Laboratory diagnosis of congenital toxoplasmosis. J Clin Microbiol 2016; pii: JCM.00487-16.
• ¹⁵
  Fochi MML, Baring S, Spegiorin LCJF, Vaz-Oliani DCM, Galão EA, Oliani AH, et al. Prematurity and low birth weight did not correlate with anti-Toxoplasma gondii maternal serum profiles - a Brazilian report. PLoS One 2015; 10:e0132719.
• ¹⁶
  Augustine SAJ. Towards universal screening for toxoplasmosis: rapid, cost-effective and simultaneous detection of Toxoplasma anti-IgG, IgM and IgA antibodies using very small serum volumes. J Clin Microbiol 2016; 54:1684-1685.

• This study was supported by research grants from Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) #2012/07716-9 to LCM; #2013/15879-8 to FHAM; #2014/01706-7 to MNF; and #2014/09496-1 to VLPC), the Brazilian Ministry of Science, Technology and Innovation - Conselho Nacional de Desenvolvimento Científico e Tecnológico (PIBIC-CNPq #135144/2014-0 to NSC and #135229/2014-6 to GSS), Fundação de Apoio à Pesquisa e Extensão de São José do Rio Preto (FAPERP) #175/2015 to FHAM and #129/2015 to MNF), and the institutional research grant Bolsa Auxílio à Pesquisa (BAP-FAMERP to CCBM). The opinions, assumptions, and conclusions or recommendations expressed in this material are the responsibility of the authors and do not necessarily reflect the views of FAPESP.

Publication Dates

• Publication in this collection
  Jul-Aug 2016

History

• Received
  29 Mar 2016
• Accepted
  09 June 2016