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 Table of Contents  
ORIGINAL ARTICLE
Year : 2016  |  Volume : 9  |  Issue : 1  |  Page : 24-30

Detection of T. gondii infection in blood donors in Alexandria, Egypt, using serological and molecular strategies


1 Department of Applied and Molecular Parasitology, Parasitology, Medical Research Institute, Alexandria University, Alexandria, Egypt
2 Department of Hematology, Medical Research Institute, Alexandria University, Alexandria, Egypt
3 Department of Applied Medical Chemistry, Medical Research Institute, Alexandria University, Alexandria, Egypt
4 Department of Clinical Pathology, Military Medical Academy, Armed Forces Hospital, Alexandria, Egypt

Date of Submission26-Aug-2015
Date of Acceptance16-Mar-2016
Date of Web Publication25-Oct-2016

Correspondence Address:
Omnya A El-Geddawi
Department of Applied and Molecular Parasitology, Medical Research Institute, Alexandria University, Alexandria
Egypt
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/1687-7942.192992

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  Abstract 

Background
Toxoplasmosis is a widespread disease caused by the opportunistic parasite Toxoplasma gondii, with variable overall prevalence according to the different geographical areas. Blood donors pose as possible contributors for transfer of infection.
Objective
The aim of this study was to estimate the prevalence of exposure to Toxoplasma in blood donors using sensitive techniques in a cross-sectional study.
Materials and methods
An aggregate of 150 blood donors from the blood donation center of Alexandria University participated in this study. The blood samples were tested for the presence of T. gondii immunoglobulin (Ig) G antibody and target gene B1 using enzyme-linked immunosorbent assay and real-time PCR, respectively.
Results
Of 150 participants, 65.3% tested were positive for anti-Toxoplasma IgG, and 10% showed parasitemia as B1 gene was successfully amplified in nine seropositive samples and in six seronegative samples.
Conclusion
The recorded IgG seropositivity in this selected group of individuals may be considered an indication of the general prevalence of toxoplasmosis in Alexandria. Detected parasitemia using real-time PCR draws attention to the possibility of transmission through blood transfusion even from seronegative donors and emphasizes the importance of specialized Toxoplasma DNA screening before donation of blood.

Keywords: B1 gene, blood donors, enzyme-linked immunosorbent assay, real-time PCR, Toxoplasma gondii


How to cite this article:
El-Geddawi OA, El-Sayad MH, Sadek NA, Hussien NA, Ahmed MA. Detection of T. gondii infection in blood donors in Alexandria, Egypt, using serological and molecular strategies. Parasitol United J 2016;9:24-30

How to cite this URL:
El-Geddawi OA, El-Sayad MH, Sadek NA, Hussien NA, Ahmed MA. Detection of T. gondii infection in blood donors in Alexandria, Egypt, using serological and molecular strategies. Parasitol United J [serial online] 2016 [cited 2019 Dec 9];9:24-30. Available from: http://www.new.puj.eg.net/text.asp?2016/9/1/24/192992


  Introduction Top


Toxoplasma gondii is a parasitic protozoan of worldwide distribution in most warm-blooded creatures. Around 33% of the world’s human populace is reportedly infected with this parasite [1],[2],[3]. There is remarkable variation in the seroprevalence of toxoplasmosis between countries (10–80%) and even in a given country [4],[5]. Transmission of T. gondii to humans occurs by ingestion of cysts in improperly cooked infected meat, or feline mature oocysts contaminating food or water. Vertical transmission from mother’s placenta to fetus has drastic results. Organ transplantation and blood transfusion from infected donors are other important routes [6],[7],[8].

Toxoplasmosis can bring about severe manifestations and complications in immunocompromised individuals and babies; however, it is typically understated and self-restricting in immunocompetent individuals [1],[5]. Diagnosis of toxoplasmosis cannot rely solely on clinical manifestations; confirmatory parasitological and serological assessments are required [9]. Several serological tests recognizing specific immunoglobulins (IgG and IgM) against T. gondii in sera of patients have been used effectively [10]. Among these assays, ELISA demonstrates a high sensitivity and specificity [11]; however, anti-Toxoplasma IgG or IgM may not be distinguished during the acute phase of infection, due to delayed production of these antibodies until after a few weeks of parasitemia. This is especially important in certain immunocompromised patients in whom the titers of specific anti-Toxoplasma antibodies may not be detectable [12].

PCR assays are extremely important for providing results of high sensitivity, especially when real-time PCR is used as it is useful for accurate and early diagnosis of toxoplasmosis. In real-time PCR, the 5′-nuclease movement of Taq DNA polymerase is used to slice a non-extendible, fluorescence-marked hybridization probe during the extension period of PCR. The fluorescence of the intact probe is replaced by 6-carboxy-tetramethyl-rhodamine (TAMRA) fluorescent dye. During PCR, the hybridization probe discharges an extinguishing impact of quenching bringing about an increase in fluorescence, corresponding to the measure of PCR product that can be observed by a sequence indicator [13]. The regularly utilized focus for the location of T. gondii using PCR is the repetitive B1 gene [14], due to its presence in a repetitive manner as a 35-copy gene in the genome, and its conservation in each of the six T. gondii strains tested to date [15],[16],[17],[18].

The variable predominance of T. gondii antibodies reported in past investigations of blood donors [19],[20],[21],[22],[23],[24],[25],[26],[27] portrays clear geographic and worldwide contrasts in transmission patterns between nations. In Egypt, information on the prevalence of T. gondii in the general population varied between 30 and 60% [28], with few reports focused on blood donors. An early study in 1986 on prevalence in blood donors to Ain Shams University Hospitals in Cairo revealed an IgG seropositivity of 19.5% [29]. In another report from Mansoura governorate in 2009, 59.6% of cases were seropositive for Toxoplasma IgG antibodies using enzyme-linked immunosorbent assay (ELISA) [30]. This information inspired us to conduct this cross-sectional study to distinguish between exposure to infection and T. gondii disease among blood donors in Alexandria, Egypt, using ELISA and real-time PCR.


  Materials and methods Top


Study design

This cross-sectional study was performed during the period from March to July 2013 wherein 150 blood samples collected from donors attending the Blood Bank of Medical Research Institute, Alexandria University, were tested for anti-Toxoplasma IgG antibodies using ELISA and for circulating parasite DNA using quantitative real-time PCR. The study was approved by the ethics committee, Alexandria University. Donors were asked to participate in the study after clear explanation of the study objectives and informed verbal consent was obtained from all participants.

Blood sample collection

A volume of 10 ml of blood was collected from each donor and divided into two tubes: 4 ml was collected in a plain tube to allow coagulation of blood and separation of serum, and 6 ml was collected in a tube with EDTA for buffy coat separation. A total of 150 blood samples were enrolled in the study after excluding HIV, hepatitis B virus, hepatitis C virus, and Treponema pallidum infections.

Serological testing

Serum samples of blood donors were tested for anti-T. gondii IgG using commercially available ELISA (Biocheck Inc., Foster City, California, USA). The test was performed according to the manufacturer’s instructions; samples giving quantities of 32 or greater international units (IU/ml) were considered positive.

Molecular testing

Buffy coat preparation

Anticoagulated blood of 3 ml was mixed with an equal volume of PBS, pH 7.4, and slowly layered on 3 ml Ficoll-Paque, and then centrifuged at 1900 rpm for 30 min at 25°C. The leukocyte-rich supernatant was collected, washed in PBS, and centrifuged at 1400 rpm for 10 min at 25°C. The pellets were kept frozen at −80°C to extract and subsequently amplify DNA [31],[32].

Preparation of T. gondii tachyzoites

T. gondii tachyzoites, utilized as positive control and for standard curve preparation, were kindly provided by the Parasitology Department, Faculty of Medicine, Alexandria University. Tachyzoites were maintained in the laboratory by serial intraperitoneal inoculation of Swiss albino mice every 3 days. Parasites collected from mouse ascitic fluid were washed several times in PBS. The number of tachyzoites was determined using a hemocytometer (Neubauer chamber).

DNA extraction

Total DNA was extracted from buffy coat and parasites using the genomic preparation blood DNA isolation kit (GeneJET Whole Blood Genomic DNA Purification Kit; Thermo Fisher Scientific Inc., Waltham, Massachusetts, USA) according to the manufacturer’s directions. Successful DNA extraction from the 150 samples was verified by running the extraction products on 1% agarose gel for 15 min at 100 V and stained with ethidium bromide ([Figure 1]). The extracted DNAs were kept frozen at −80°C until further testing using real-time PCR.
Figure 1 Example of DNA extraction product (total DNA) from 24 stained samples visualized with ethidium bromide staining.

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Quantitative detection of T. gondii B1 gene using real-time PCR

Real-time PCR was performed as described by Mei-Hui et al. [33] using the forward primer (TOXO-F) (5 μmol/l, 5′-TCCCCTCTGCTGGCGAAAAGT-3′), reverse primer (TOXO-R) (5 μmol/l, 5′-AGCGTTCGTGGTCAACTATCGATTG-3′), and Taqman probe (2 μmol/l, 6FAM-TCTGTGCAACTTTGGTGTATTCGCAG-TAMRA) to amplify the 98 bp segment of the 35-repititive B1 gene. The primers and the TaqMan probe were purchased from Life technologies, Applied Biosystems, Carlsbad, California, USA. The amplification reactions were performed in final volumes of 25 μl containing 5 μl of template DNA, 12.5 μl of 2× Taqman universal master mix, 2.5 μl of the forward primer (TOXO-F), 2.5 μl of the reverse primer (TOXO-R), and 2.5 μl of Taqman probe using a StepOne Real-Time PCR System (Applied Biosystems, CA, USA). After initial activation of AmpliTaqGold DNA polymerase at 95°C for 10 min, 40 PCR cycles of 95°C for 15 s (denaturation step) and 60°C for 1 min (annealing and extension steps) were performed. Ribonuclease P (RNase P) housekeeping gene was utilized as internal control to check the potential PCR inhibitors in the analyzed specimens.

The standard curve for the real-time PCR quantification was set by amplification of serial dilutions of T. gondii DNA corresponding to 10 000, 1000, 100, 10, 2, 0.2, 0.02, and 0.002 tachyzoites. The CT values resolved were plotted against log amount of equivalent tachyzoites ([Table 1] and [Figure 2]).
Table 1 CT values corresponding to DNA-tachyzoite equivalents

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Figure 2 T. gondii DNA standard curve.

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Statistical analysis

The SPSS software statistical program (version 20; SPSS Inc., Chicago, Illinois, USA) was utilized for both data presentation and statistical analysis of results.


  Results Top


The age of the 150 participants ranged from 18 to 60 with a mean age of 32±8 years. Seropositivity of IgG T. gondii antibodies showed that, of the 150 blood donors, 98 (65.3%) were seropositive for toxoplasmosis ([Table 2]).
Table 2 Seroprevalence of T. gondii IgG antibodies in 150 blood samples of donors

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As demonstrated in [Table 3], the B1 gene was effectively amplified in only 15 blood tests (10%) ([Figure 3]), including nine samples with seropositive results and six samples with seronegative results. The measure of tachyzoites in every positive sample, as regards the CT values, is presented in [Table 4]. The values differed from 0.009 to 68.5 tachyzoite-equivalent.
Table 3 T. gondii B1 gene in 150 blood samples of donors

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Figure 3 Amplification plot of B1-gene in positive samples represented by the different colors.

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Table 4 Tachyzoite equivalent in real-time PCR positive samples (n=15) compared with ELISA

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  Discussion Top


Toxoplasmosis is prevalent everywhere throughout the world as an important protozoan parasite [2]. Being an opportunistic pathogen, it affects organ transplant recipients and other immunocompromised patients, causing critical or life-debilitating complications as a consequence of either acute infection or reactivation of a latent dormant infection [34].

As reported previously, the predominance of toxoplasmosis among blood donors in Egypt varies according to locality and time [28]. Accordingly, this study aimed to not only update the prevalence of toxoplasmosis in an important sector of the population in the second largest governorate in Egypt but also to evaluate the possibility of transmission of active infection through blood donation. To achieve this goal, assessment was carried out using serological (for prevalence) and molecular (for active infection) determinations. The outcome showed that the general prevalence of T. gondii IgG antibodies among the 150 blood donors was 65.3%. This rate varied, being slightly higher than that reported from Mansoura governorate, Egypt, in 2009, in which the prevalence of Toxoplasma -specific IgG in blood donors, as revealed using ELISA, was 59.6% [30]; both of these rates were higher than the 19.5% incidence reported much earlier in 1986 from Cairo [29].

When compared with different countries, our recorded seropositivity was much higher than that reported in blood donors in Mexico (7.4%) [27], Thailand (9%) [22], Chile (21.2%) [21], Malaysia (28.1%) [24], Czech Republic (33.1%) [20], Mali (41.2%) [23], and Saudi Arabia (52.1%) [19]. Moreover, seropositivity was lower than that reported in blood donors in Brazil (79.0%) [26] and in Cuba (73.43%) [25]. The variability in the prevalence levels of T. gondii infection among blood donor populaces may be attributed to dissimilarities in environmental pollutions as well as the individual habits and characteristics of the indigenous populations.

Diagnosis of toxoplasmosis is not conclusive without the monitoring of rise in antibody levels in an infected individual. Moreover, the serological diagnosis of active toxoplasmosis is restricted when reactivation of infection does not promote changes in the antibody levels, and when IgM antibodies persist during the chronic phase of infection, thus confusing interpretation of the serological results. Besides, serological techniques may not recognize below threshold levels of specific anti-Toxoplasma IgG or IgM produced during the active phase of disease in immunocompromised patients. In AIDS patients, these antibodies may be produced late following a few weeks of acquiring the contamination [35].

To improve diagnosis, the real-time PCR Taqman probe-based test was introduced to increase the sensitivity and specificity of the amplification reaction [36],[37]. Numerous studies utilized T. gondii B1 gene for toxoplasmosis detection using real-time PCR [37],[38]. Our results showed that the real-time PCR utilized for the identification of B1 gene proved to be sensitive, having recorded an equivalent of 0.002 tachyzoites, thus confirming its capacity to quantify the contamination load in a clinical example. It was successfully amplified in 15 (10%) blood donors’ samples (nine were IgG seropositive and six were IgG seronegative). This low detection rate cannot exclude recent infection and may be due to a small number of parasites circulating in the blood, a low amount of parasite DNA present in clinical samples compared with the total volume of blood in the human body, or short period of parasitemia duration [39],[40],[41]. The seronegative cases, whose B1 gene was decidedly identified as positive using real-time PCR, may be an indication of the presence of circulating Toxoplasma tachyzoites unassociated with antibodies. The proposed clarification is that it could be an extremely recent infection at the time of serological testing, prompting an inadequate distinguishable generation of antibodies by serology [42].

About 54% of blood donors were in the age group 26–35 years and 22.7% were in age group 36–45 years; those in the age group 18–25 years constituted 16% and those older than 45 years constituted 7.3%. Our study showed that the percentage of IgG seropositivity was nearly equal in two of the age groups, from 36–45 years and those over 45 years (73.5 and 72.7%, respectively); it was lower in the younger age groups. This finding is in agreement with many other studies reporting that the increased age-related rate of Toxoplasma infection is due to the greater likelihood of exposure to infection with increase in age [19],[43],[44],[45].

As with other techniques, PCR may experience poor outcome due to variable execution. The absence of a consistent standard technique implies that sensitivity and specificity of molecular tests cannot generally be compared with those of other classical systems (e.g. serological testing, and cell cultures and animal inoculation). Proper correlation between protocols confirmed the high regard for molecular procedures, especially due to the shorter time needed to obtain results [46],[47],[48]. Accordingly, updating the diagnosis of toxoplasmosis using real-time PCR techniques will benefit the quick shut tube framework, and the reproducible quantitative results without the danger of pollution. This technique has great potential for use as a standardized technique applied at the clinical level [42]. Furthermore, this strategy may be suitable for routine screening of T. gondii infection in the clinical research facility in conjunction with other diagnostic methods.


  Conclusion Top


The revealed data represents the first report on the prevalence of toxoplasmosis in blood donors from Alexandria governorate, Egypt. The IgG seropositivity rate in this group of individuals may be considered as an indication of the general prevalence of exposure to infection in this governorate. The unexpected detection of circulating B1 gene in a number of seropositive as well as seronegative donors indicates the possibility of a high risk for transmission of infection to blood recipients. Thus, the development of a general screening system for toxoplasmosis in blood contributors in Alexandria is recommended, especially as transmission of the disease from seronegative individuals cannot be excluded.

Authors contribution

All authors participated actively in the study and are in agreement with its contents which have not been published or considered for publication elsewhere. MH Sayad designed the study and was in charge of performing ELISA technique; NA Sadek provided the blood donors’ cases. NA Hussein, MA Ahmed, OA Geddawi were in charge of performing the molecular analysis (Real-Time PCR). OA Geddawi was in charge of collecting the samples, performing the ELISA and extracting the DNA for molecular analysis.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.

 
  References Top

1.
Montoya JG, Liesenfeld O. Toxoplasmosis. Lancet 2004; 363:1965–1976.  Back to cited text no. 1
[PUBMED]    
2.
Hill DE, Chirukandoth S, Dubey JP. Biology and epidemiology of Toxoplasma gondii in man and animals. Anim Health Res Rev 2005; 6:41–61.  Back to cited text no. 2
    
3.
Pappas G, Roussos N, Falagas ME. Toxoplasmosis snapshots: global status of Toxoplasma gondii seroprevalence and implications for pregnancy and congenital toxoplasmosis. Int J Parasitol 2009; 39:1385–1394.  Back to cited text no. 3
    
4.
Sukthana Y. Toxoplasmosis: beyond animals to humans. Trends Parasitol 2006; 22:137–142.  Back to cited text no. 4
    
5.
Robert-Gangneux F, Dardé ML. Epidemiology of and diagnostic strategies for toxoplasmosis. Clin Microbiol Rev 2012; 25:264–296.  Back to cited text no. 5
    
6.
Siegel SE, Lunde MN, Gelderman AH, Halterman RH, Brown JA, Levine AS et al. Transmission of toxoplasmosis by leukocyte transfusion. Blood 1971; 37:388–394.  Back to cited text no. 6
    
7.
Derouin F, Pelloux H. Prevention of toxoplasmosis in transplant patients. Clin Microbiol Infect 2008; 14:1089–1101.  Back to cited text no. 7
    
8.
Remington JS, Klein JO. Infectious diseases of the fetus and newborn infant. Philadelphia, PA: Saunders 2010. 918–1041.  Back to cited text no. 8
    
9.
Petersen E, Borobio MV, Guy E, Liesenfeld O, Meroni V, Naessens A et al. European multicenter study of the LIAISON automated diagnostic system for determination of Toxoplasma gondii-specific immunoglobulin G (IgG) and IgM and the IgG avidity index. J Clin Microbiol 2005; 43:1570–1574.  Back to cited text no. 9
    
10.
Gharavi MJ, Jalali S, Khademvatan S, Heydari S. Detection of IgM and IgG anti-Toxoplasma antibodies in renal transplant recipients using ELFA, ELISA and ISAGA methods: comparison of pre- and post-transplantation status. Ann Trop Med Parasitol 2011; 105:367–371.  Back to cited text no. 10
    
11.
Fuccillo DA, Madden DL, Tzan N, Sever JL. Difficulties associated with serological diagnosis of Toxoplasma gondii infections. Diagn Clin Immunol 1987; 5:8–13.  Back to cited text no. 11
    
12.
Porter SB, Sande MA. Toxoplasmosis of the central nervous system in the acquired immunodeficiency syndrome. N Engl J Med 1992; 327:1643–1648.  Back to cited text no. 12
    
13.
Holland PM, Abramson RD, Watson R, Gelfand DH. Detection of specific polymerase chain reaction product by utilizing the 5′-3′ exonuclease activity of Thermus aquaticus DNA polymerase. Proc Natl Acad Sci USA 1991; 88:7276–7280.  Back to cited text no. 13
    
14.
Burg JL, Grover CM, Pouletty P, Boothroyd JC. Direct and sensitive detection of a pathogenic protozoan, Toxoplasma gondii, by polymerase chain reaction. J Clin Microbiol 1989; 27:1787–1792.  Back to cited text no. 14
    
15.
Colin D, Narciss O, Peter A, Sharro T, Susan L. Comparison of PCR detection methods for B1, P30 and 18SrDNA genes of Toxoplasma gondii in aqueous humor. Invest Opthalmol Vis Sci 2000; 41:634–644.  Back to cited text no. 15
    
16.
Contini C, Cultrera R, Seraceni S, Segala D, Romani R, Fainardi E et al. The role of stage-specific oligonucleotide primers in providing effective laboratory support for the molecular diagnosis of reactivated Toxoplasma gondii encephalitis in patients with AIDS. J Med Microbiol 2002; 51:879–890.  Back to cited text no. 16
    
17.
Joseph P, Calderón MM, Gilman RH, Quispe ML, Cok J, Ticona E et al. Optimization and evaluation of a PCR assay for detecting toxoplasmic encephalitis in patients with AIDS. J Clin Microbiol 2002; 40:4499–4503.  Back to cited text no. 17
    
18.
Khan A, Su C, German M, Storch GA, Clifford DB, Sibley LD. Genotyping of Toxoplasma gondii strains from immunocompromised patients reveals high prevalence of type I strains. J Clin Microbiol 2005; 43:5881–5887.  Back to cited text no. 18
    
19.
Al-Amari OM. Prevalence of antibodies to Toxoplasma gondii among blood donors in Abha, Asir Region, south-western Saudi Arabia. J Egypt Public Health Assoc 1994; 69:77–88.  Back to cited text no. 19
    
20.
Svobodová V, Literák I. Prevalence of IgM and IgG antibodies to Toxoplasma gondii in blood donors in the Czech Republic. Eur J Epidemiol 1998; 14:803–805.  Back to cited text no. 20
    
21.
Zamorano CG, Contreras MC, Villalobos S, Sandoval L, Salinas P. Seroepidemiological survey of human toxoplasmosis in Osorno, Region X, Chile, 1998. Bol Chil Parasitol 1999; 54:33–36.  Back to cited text no. 21
    
22.
Pinlaor S, Ieamviteevanich K, Pinlaor P, Maleewong W, Pipitgool V. Seroprevalence of specific total immunoglobulin (Ig), IgG and IgM antibodies to Toxoplasma gondii in blood donors from Loei Province, Northeast Thailand. Southeast Asian J Trop Med Public Health 2000; 31:123–127.  Back to cited text no. 22
    
23.
Maïga I, Kiemtoré P, Tounkara A. Prevalence of anti-Toxoplasma antibodies in patients with acquired immunodeficiency syndrome and blood donors in Bamako. Bull Soc Pathol Exot 2001; 94:268–270.  Back to cited text no. 23
    
24.
Nissapatorn V, Kamarulzaman A, Init I, Tan LH, Rohela M, Norliza A et al. Seroepidemiology of toxoplasmosis among HIV-infected patients and healthy blood donors. Med J Malaysia 2002; 57:304–310.  Back to cited text no. 24
    
25.
Martin-Hernlndez I, Garcya-Izquierdo SM. Prevalence of IgG antibodies to Toxoplasma gondii in Cuban blood donors. Rev Biomed 2003; 14:247–251.  Back to cited text no. 25
    
26.
Coêlho RA, Kobayashi M, Carvalho LB Jr. Prevalence of IgG antibodies specific to Toxoplasma gondii among blood donors in Recife, Northeast Brazil. Rev Inst Med Trop Sao Paulo 2003; 45:229–231.  Back to cited text no. 26
    
27.
Alvarado-Esquivel C, Mercado-Suarez MF, Rodríguez-Briones A, Fallad-Torres L, Ayala-Ayala JO, Nevarez-Piedra LJ et al. Seroepidemiology of infection with Toxoplasma gondii in healthy blood donors of Durango, Mexico. BMC Infect Dis 2007; 7:75.  Back to cited text no. 27
    
28.
El Nassef N, Mohamed M, El Nahas N, Hassanain MD, Shams ElDin S, Ammar A. Seroprevalence and genotyping of Toxoplasma gondii in Menoufia governorate. Menoufia Med J 2015; 28:617–626.  Back to cited text no. 28
    
29.
Azab ME, Safar EH, el-Shennawt SF, Hassan FA. Serological evidence of infection with Plasmodium and Toxoplasma in blood donors to Ain-Shams University Hospital. J Egypt Soc Parasitol 1986; 16:163–170  Back to cited text no. 29
    
30.
Elsheikha HM, Azab MS, Abousamra NK, Rahbar MH, Elghannam DM, Raafat D. Seroprevalence of and risk factors for Toxoplasma gondii antibodies among asymptomatic blood donors in Egypt. Parasitol Res 2009; 104:1471–1476.  Back to cited text no. 30
    
31.
Böyum A. Isolation of mononuclear cells and granulocytes from human blood. Isolation of monuclear cells by one centrifugation, and of granulocytes by combining centrifugation and sedimentation at 1 g. Scand J Clin Lab Invest Suppl 1968; 97:77–89.  Back to cited text no. 31
    
32.
Bøyum A. Isolation of lymphocytes, granulocytes and macrophages. Scand J Immunol 1976; 5(Suppl 5):9–15.  Back to cited text no. 32
    
33.
Mei-Hui L, Tse-Ching C, Tseng-tong K, Ching-Chung T. Real-Time PCR. for quantitative detection of Toxoplasma gondii. J Clin Microbiol 2000; 38:4121.  Back to cited text no. 33
    
34.
Aspinall TV, Guy EC, Roberts KE, Joynson DH, Hyde JE, Sims PF. Molecular evidence for multiple Toxoplasma gondii infections in individual patients in England and Wales: public health implications. Int J Parasitol 2003; 33:97–103.  Back to cited text no. 34
    
35.
O’Driscoll JC, Holliman RE. Toxoplasmosis and bone marrow transplantation. Rev Med Microbiol 1991; 2:215–222.  Back to cited text no. 35
    
36.
Calderaro A, Piccolo G, Gorrini C, Peruzzi S, Zerbini L, Bommezzadri S et al. Comparison between two real-time PCR assays and a nested-PCR for the detection of Toxoplasma gondii. Acta Biomed 2006; 77:75–80.  Back to cited text no. 36
    
37.
Su C, Shwab EK, Zhou P, Zhu XQ, Dubey JP. Moving towards an integrated approach to molecular detection and identification of Toxoplasma gondii. Parasitology 2010; 137:1–11.  Back to cited text no. 37
    
38.
Filisetti D, Gorcii M, Pernot-Marino E, Villard O, Candolfi E. Diagnosis of congenital toxoplasmosis: comparison of targets for detection of Toxoplasma gondii by PCR. J Clin Microbiol 2003; 41:4826–4828.  Back to cited text no. 38
    
39.
Fuentes I, Rubio JM, Ramírez C, Alvar J. Genotypic characterization of Toxoplasma gondii strains associated with human toxoplasmosis in Spain: direct analysis from clinical samples. J Clin Microbiol 2001; 39:1566–1570.  Back to cited text no. 39
    
40.
Gallego C, Saavedra-Matiz C, Gómez-Marín JE. Direct genotyping of animal and human isolates of Toxoplasma gondii from Colombia (South America). Acta Trop 2006; 97:161–167.  Back to cited text no. 40
    
41.
Ferreira IM, Vidal JE, Costa-Silva TA, Meira CS, Hiramoto RM, Penalva de Oliveira AC et al.. Toxoplasma gondii: genotyping of strains from Brazilian AIDS patients with cerebral toxoplasmosis by multilocus PCR-RFLP markers. Exp Parasitol 2008; 118:221–227.  Back to cited text no. 41
    
42.
Kompalic-Cristo A, Frotta C, Suarez M, Fernandes O, Britto C. Evolution of a real-time PCR assay based on the repetitive B1 gene for the detection of Toxoplasma gondii in human peripheral blood. Parasitol Res 2007; 101:619–625.  Back to cited text no. 42
    
43.
Bobic B, Jevremovic I, Marinkovic J, Sibalic D, Olgica D. Risk factors for Toxoplasma infection in reproductive age in female population in the area of Belgrade, Yogoslavia. J Epidem 1998; 14:605–610.  Back to cited text no. 43
    
44.
Jumaian NF. Seroprevalence and risk factors for Toxoplasma infection in pregnant women in Jordan. East Mediterr Health J 2005; 11:45–51.  Back to cited text no. 44
    
45.
Chacin-Bonilla L, Sanchez-Chavez Y, Monsalve F, Estevez J. Seroepidemiology of toxoplasmosis in amerindians from western Venezuela. Am J Trop Med Hyg 2001; 65:131–135.  Back to cited text no. 45
    
46.
Reischl U, Bretagne S, Krüger D, Ernault P, Costa JM. Comparison of two DNA targets for the diagnosis of Toxoplasmosis by real-time PCR using fluorescence resonance energy transfer hybridization probes. BMC Infect Dis 2003; 3:7.  Back to cited text no. 46
    
47.
Simon A, Labalette P, Ordinaire I, Fréalle E, Dei-Cas E, Camus D et al. Use of fluorescence resonance energy transfer hybridization probes to evaluate quantitative real-time PCR for diagnosis of ocular toxoplasmosis. J Clin Microbiol 2004; 42:3681–3685.  Back to cited text no. 47
    
48.
Cassaing S, Bessières MH, Berry A, Berrebi A, Fabre R, Magnaval JF. Comparison between two amplification sets for molecular diagnosis of toxoplasmosis by real-time PCR. J Clin Microbiol 2006; 44:720–724.  Back to cited text no. 48
    


    Figures

  [Figure 1], [Figure 2], [Figure 3]
 
 
    Tables

  [Table 1], [Table 2], [Table 3], [Table 4]


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4 Toxoplasma gondii: Are There any Implications for Routine Blood Screening?
Masoud Foroutan,Hamidreza Majidiani
International Journal of Infection. 2017; In Press(In Press)
[Pubmed] | [DOI]



 

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