Association between Toxoplasma gondii infection and history of blood transfusion: a case-control seroprevalence study.

Objectives This study was performed to determine the association between seropositivity to Toxoplasma gondii and a history of blood transfusion. Methods Patients who had undergone blood transfusion (n = 410) and age- and sex-matched controls who had not undergone blood transfusion (n = 1230) were examined for anti- T. gondii IgG and IgM antibodies using enzyme-linked immunoassays. Results Anti- T. gondii IgG antibodies were detected in 57 (13.9%) patients and in 129 (10.5%) controls with a borderline difference [odds ratio (OR) = 1.37, 95% confidence interval (CI) = 0.98-1.92]. High anti- T. gondii IgG antibody levels (>150 IU/mL) were found in 27 (47.4%) of the 57 anti- T. gondii IgG-positive patients and in 37 (28.7%) of the 129 anti- T. gondii IgG positive controls with a significant difference (OR = 2.23, 95% CI = 1.17-4.26). Anti- T. gondii IgM antibodies were found in 13 (22.8%) of the 57 seropositive patients and in 37 (28.7%) of the 129 seropositive controls, but the difference was not significant (OR = 0.73, 95% CI = 0.35-1.52). Seroprevalence was significantly higher in patients aged >50 years than in controls of the same age and in female patients than in female controls. Conclusions These findings indicate that a history of blood transfusion is a risk factor for T. gondii infection.

Introduction

Infection with the parasite Toxoplasma gondii is prevalent in humans and animals worldwide.[1,2] Most infections with T. gondii are asymptomatic. However, individuals who develop toxoplasmosis, the disease caused by T. gondii, may present several well-recognized clinical features: lymphadenopathy, chorioretinitis, and meningoencephalitis.[4,5] In addition, infection with T. gondii has been associated with heart disease,[6,7] schizophrenia,[8,9] depression,[10,11] and mixed anxiety and depression. Immunocompromised individuals infected with T. gondii may develop disseminated and life-threatening disease. Furthermore, a primary infection with T. gondii during pregnancy is a risk factor for infection of the fetus and consequent congenital toxoplasmosis.

Infections with T. gondii are usually acquired by ingestion of raw or undercooked meat containing tissue cysts or by ingestion of food or water contaminated with oocysts shed by cats. Transmission of T. gondii can also occur by organ transplantation. In addition, blood transfusion has been associated with T. gondii infection.[17,18] After infection, T. gondii disseminates from the blood to the host tissues.[19-21] In fact, T. gondii parasitemia has been diagnosed by examination of blood smears and by polymerase chain reaction. Infection with T. gondii has been detected in blood donors.[24-26] However, the extent of the association between blood transfusion and infection with T. gondii remains unclear. To the best of our knowledge, no case-control study has been performed to investigate this issue. Therefore, we sought to determine the association between infection with T. gondii and blood transfusion in a sample of people in the northern Mexican city of Durango.

Materials and methods

Study design and study population

This case-control seroprevalence study included patients with a history of blood transfusion (cases) and age- and sex-matched subjects without a history of blood transfusion (controls). The cases were enrolled in a public family health care center (Institute of Security and Social Services of State Workers) in Durango City, Mexico. All participants were examined for the presence of anti-T. gondii IgG and IgM antibodies. This study was performed from August 2015 to May 2016 in Durango City, Mexico. The inclusion criteria for the cases were a history of blood transfusion, age of ≥16 years, and agreement to participate in the study.

Sample size calculation

We calculated the sample size using a 95% confidence level, a power of 80%, a 1:3 proportion of cases and controls, a reference seroprevalence of 6.1% as the expected frequency of exposure in the controls, and an odds ratio (OR) of 2. The result of this calculation was 295 cases and 883 controls.

Clinical data of cases

The clinical characteristics of the subjects with a history of blood transfusion were recorded with the aid of a questionnaire. The clinical items were the presence of any illness, frequent headache or dizziness, history of surgery, lymphadenopathy, transplantation, or impairments in memory, reflexes, hearing, or vision.

Detection of T. gondii IgG and IgM antibodies

A sample of blood was obtained from each participant. Sera were obtained by centrifugation of whole blood and stored at −20°C until analyzed. The sera of participants were analyzed for anti-T. gondii IgG antibodies. For this purpose, we used the commercially available enzyme immunoassay kit “Toxoplasma IgG” (Diagnostic Automation Inc., Woodland Hills, CA, USA). Anti-T. gondii IgG antibody levels were expressed as international units (IU)/mL, and a cut-off of ≥8 IU/mL was used for seropositivity. Sera with anti-T. gondii IgG antibodies were further tested for anti-T. gondii IgM antibodies. For this purpose, we used the commercially available enzyme immunoassay “Toxoplasma IgM” kit (Diagnostic Automation Inc.). All tests were performed in accordance with the manufacturer’s instructions. We analyzed the positive and negative controls included in the kits in each run.

Statistical analysis

We statistically analyzed the data using the software SPSS version 15.0 (SPSS Inc. Chicago, IL, USA) and Epi Info version 7 (Centers for Disease Control and Prevention: http://wwwn.cdc.gov/epiinfo/). We compared the age of the cases and controls using Student’s t test. The frequency of blood transfusion among seropositive and seronegative individuals was assessed with Student t test and the Mann–Whitney U test. Pearson’s chi-square test was used to determine the association of T. gondii seropositivity with transfusional data. The ORs and 95% confidence intervals (CIs) were calculated, and statistical significance was set at a P value of  < 0.05.

Ethics statement

This project was approved by the Institutional Ethics Committee of the General Hospital of the Secretary of Health in Durango City, Mexico. Before enrollment, all participants were informed about the purpose and procedures of this study, and written informed consent was obtained from all subjects.

Results

This study included 410 cases and 1230 age- and sex-matched controls. The cases comprised 57 male and 353 female patients. This study included a higher number of female than male participants because a history of blood transfusion was more frequently observed in women than in men among the attendees of the family health care center surveyed. The cases were 16 to 80 years old (mean, 41.90 ± 11.58 years). The controls were randomly selected from the general population in the same city and matched with the cases by sex and age. Thus, the controls included 171 male and 1059 female participants aged 16 to 80 years (mean, 41.92 ± 11.55 years). The age of the controls was not significantly different from that of the cases.

Anti-T. gondii IgG antibodies were detected in 57 (13.9%) of the 410 cases and in 129 (10.5%) of the 1230 controls (OR = 1.37, 95% CI = 0.98–1.92, P = 0.05). Of the 57 anti-T. gondii IgG-positive cases, 27 (47.4%) had anti-T. gondii IgG antibody levels of >150 IU/mL and 30 (52.6%) had lower antibody levels. Of the 129 anti-T. gondii IgG-positive controls, 37 (28.7%) had anti-T. gondii IgG antibody levels of >150 IU/mL and 92 (71.3%) had lower antibody levels. Significantly more subjects had high IgG antibody levels among the cases than controls (OR = 2.23, 95% CI = 1.17–4.2, P = 0.01). Of the 57 cases seropositive for anti-T. gondii IgG antibodies, 13 (22.8%) were also positive for anti-T. gondii IgM antibodies compared with 37 (28.7%) of the 129 controls seropositive to anti-T. gondii IgG antibodies (OR = 0.73, 95% CI = 0.35–1.52).

Stratification by age and sex showed that the seroprevalence of T. gondii infection was significantly higher in cases aged ≥51 years than in controls of the same age and in female cases than in female controls (P < 0.05 for both) (Table 1).

Table 1.

Comparison of seroprevalence of Toxoplasma gondii infection between cases and controls according to age and sex.

	Cases			Controls
	Subjects tested	Seroprevalence of T. gondii infection		Subjects tested	Seroprevalence of T. gondii infection
Characteristics	n	n	%	n	n	%	P value
Age, years
≤30	72	9	12.5	210	22	10.5	0.63
31–50	236	28	11.9	716	79	11.0	0.72
>50	102	20	19.6	304	28	9.0	0.005
Sex
Male	57	7	12.3	171	29	17.0	0.40
Female	353	50	14.2	1059	100	9.4	0.01

With respect to the number of blood transfusions in the cases, seropositive subjects underwent 1 to 7 transfusions (mean, 1.87 ± 1.43; median, 1), whereas seronegative cases underwent 1 to 24 transfusions (mean, 1.80 ± 2.09; median, 1). No statistically significant difference in the number of blood transfusions was found between cases and controls (Mann–Whitney U test and Student’s t test).

With respect to the clinical characteristics, the frequency of T. gondii seropositivity was similar in cases regardless of the presence of any illness, frequent headache, history of surgery, lymphadenopathy, transplantation, or impairments in memory, reflexes, hearing, or vision. Cases with dizziness had a higher seroprevalence of T. gondii infection than cases without this clinical characteristic [30/168 (17.9%) vs 27/242 (11.2%), respectively; borderline significance: P = 0.05].

Discussion

After infection, T. gondii enters the bloodstream and disseminates into the infected host. Experiments in mice infected with T. gondii have shown that intracellular tachyzoites in the blood circulation mainly disseminate throughout the body, while extracellular tachyzoites hardly contribute to parasite dissemination. Therefore, the presence of T. gondii in blood can be a source of infection for patients receiving a blood transfusion and for laboratory workers, physicians, and nurses handling blood. However, this risk factor for T. gondii infection has been neglected. In fact, screening for T. gondii infection is not currently performed in blood banks. Epidemiological studies have shown considerable numbers of blood donors infected with T. gondii. For instance, a 7.4% to 13.5% seroprevalence of T. gondii infection in blood donors has been reported in Mexico[25,26] and Taiwan ; a 19.5% to 28.8% seroprevalence has been reported in Turkey, India, and Iran ; and a 59.6% and 79.0% seroprevalence has been reported in Egypt and Brazil, respectively. Although the current epidemiological knowledge suggests that T. gondii can be transmitted by blood transfusion, the magnitude of the association between T. gondii infection and blood transfusion has been poorly studied. To the best of our knowledge, no age- and sex-matched case-control study on the association of blood transfusion and infection with T. gondii has been performed. Therefore, in the present study, we aimed to determine this association in subjects from the northern Mexican city of Durango. In general, we found a slightly higher seroprevalence of T. gondii infection in subjects with than without a history of blood transfusion. The association between infection and blood transfusion was of borderline statistical significance (P = 0.05); however, stratification by age and sex showed a significant association between T. gondii infection and blood transfusion in subjects aged >50 years and in females. In addition, the frequency of high IgG antibody levels was significantly higher in subjects with than without a history of transfusion. Therefore, these results suggest that infection with T. gondii is associated with blood transfusion. Why older and female cases had a higher seroprevalence than their matched controls remains unclear. Older and female cases likely had a higher rate of exposure to T. gondii than their controls. Why cases had a higher frequency of high IgG levels than their matched controls also remains unclear. High levels of specific anti-T. gondii IgG antibodies can be present in patients with recent infections. In addition, high IgG levels may persist due to frequent contact with the parasite. We arbitrarily considered high levels of anti-T. gondii IgG antibodies when an IgG titer of >150 IU/mL was found. This category was selected because it is higher than the highest calibrator (150 IU/mL) of IgG included in the diagnostic kit used. The results of the present study, including the higher seroprevalence of T. gondii infection and higher frequency of high anti-T. gondii IgG antibody levels in cases than controls, suggest that T. gondii infection can be transmitted by blood transfusion in Durango, Mexico.

We selectively tested only anti-T. gondii IgG antibody-positive samples for anti-T. gondii IgM antibodies because anti-T. gondii IgG seropositivity appears early after infection. This fact supports the use of this marker in epidemiological studies; a single IgM result without an IgG result has limited value for the diagnosis of T. gondii infection because IgM enzyme immunoassays to detect anti-T. gondii IgM antibodies yield a high frequency of false-positive results.[38,39]

The present study has some limitations. First, we studied the association of T. gondii infection with a history of blood transfusion, but we did not determine the association between acute T. gondii infection and recent blood transfusion. Second, the median number of blood transfusions per individual was low; therefore, the association between T. gondii infection and a high median number of transfusions remains unclear. Third, we did not control for confounding factors. Finally, we used controls from the general population rather than controls from the same family health care center surveyed.

Conclusions

We conclude that individuals with a history of blood transfusion have a higher risk of infection with T. gondii than those without a history of blood transfusion. To the best of our knowledge, no previous age- and sex-matched case-control study of the association of infection with T. gondii and blood transfusion has been performed. Further studies to determine the magnitude of this epidemiological association should be conducted.