|Year : 2012 | Volume
| Issue : 3 | Page : 178-181
Parvovirus B19, rubella, Epstein–Barr, and cytomegalovirus and idiopathic thrombocytopenia in Egyptian children, single-center study
Maysaa El Sayed Zaki, Hanaa Morcous
Department of Clinical Pathology, Mansoura Faculty of Medicine, Mansoura University, Mansoura, Egypt
|Date of Submission||21-Feb-2012|
|Date of Acceptance||24-Mar-2012|
|Date of Web Publication||21-Jun-2014|
Maysaa El Sayed Zaki
MD, Department of Clinical Pathology, Mansoura Faculty of Medicine, Mansoura University, 35516 Mansoura
Source of Support: None, Conflict of Interest: None
Idiopathic thrombocytopenic purpura (ITP) in children is usually a self-limiting disorder. It may follow a viral infection or immunization and is caused by an inappropriate response of the immune system. Many viruses, such as parvovirus B19, cytomegalovirus (CMV), Epstein–Barr virus (EBV), and rubella, are implicated in the occurrence of ITP.
The aim of the study was to investigate the occurrence of viral-associated ITP in Egyptian children.
Materials and methods
Viral studies included specific immunoglobulin M for rubella, EBV, and CMV. In addition, molecular detection for parvovirus B19 was carried out.
Positive viral markers either by positive serology immunoglobulin M for rubella, CMV, or EBV or by a molecular study for parvovirus B19 were detected in 19 patients (38.8%). The most common viral infection was parvovirus B19 (30.6%), followed by EBV (16.3%), rubella (12.12%), and CMV (10.2%).
We conclude that in a large proportion of children with ITP in our region, an association with markers of acute viral infections similar to those of rubella, EBV, and CMV is present. Moreover, a significant proportion of the children had occult parvovirus B19 viremia. A study of occult viral infections is recommended in children with ITP.
Keywords: cytomegalovirus, Epstein– Barr virus, idiopathic thrombocytopenic purpura, parvovirus B19, rubella
|How to cite this article:|
Zaki ME, Morcous H. Parvovirus B19, rubella, Epstein–Barr, and cytomegalovirus and idiopathic thrombocytopenia in Egyptian children, single-center study. Egypt J Haematol 2012;37:178-81
|How to cite this URL:|
Zaki ME, Morcous H. Parvovirus B19, rubella, Epstein–Barr, and cytomegalovirus and idiopathic thrombocytopenia in Egyptian children, single-center study. Egypt J Haematol [serial online] 2012 [cited 2018 Sep 19];37:178-81. Available from: http://www.ehj.eg.net/text.asp?2012/37/3/178/135056
| Introduction|| |
Immune thrombocytopenic purpura (ITP), a common hematological disorder in children, is usually self-limiting and has low morbidity and mortality 1,2. The causes of ITP are unclear in most pediatric patients. Preceding or concomitant infection is often implicated, particularly if the onset is acute 3. In pediatric acute ITP cases, it is usually possible to determine an underlying pathology; among them, viral infections are the leading causative agents 4. The possibility of an immune mechanism has been suggested by the finding of increased amounts of platelet-associated immunoglobulin G (IgG) in viral-associated ITP 5. Many viruses, such as the HIV, cytomegalovirus (CMV), Epstein–Barr virus (EBV), varicella, rubeola, mumps, and parvovirus, have been implicated in childhood ITP 6. The varicella virus and EBV were the most frequent pathogens among the identifiable viruses, but previously published small-sample studies have found them responsible for less than 10% of cases of ITP, with nonspecific viral infections predominating 7.
In Egypt, the viral association with ITP has not been studied clearly. In the present study, EBV, rubella, and CMV serology was used in order to investigate the occurrence of viral-associated ITP in children at Mansoura University Children Hospital. Moreover, detection of parvovirus B19 (PB19) was attempted using a PCR. The present study aims to show that in a significant proportion of cases of ITP in children, there is an association with viral infections.
| Material and methods|| |
The study was carried out on 49 children who presented with ITP to Mansoura University Children Hospital during the period from February 2010 to December 2010. The study was approved by the ethical committee of Mansoura Faculty of Medicine and all parents of the children signed approval written consents.
The diagnosis of ITP was established by an assessment of a history of spontaneous bruising, petechiae, epistaxis, or other mucous membrane bleeding, together with a platelet count less than 50×109/l, the absence of clinical and laboratory findings of any coexisting disease (negative direct Coombs’ test, antinuclear antibody, and lupus test), and no evidence of exposure to drugs and toxins that are known to cause thrombocytopenia. The family histories of these patients were negative for collagen vascular disease, thrombocytopenia, and other hematological disorders. Bone marrow aspirations were performed only in patients with abnormal hematological parameters other than isolated thrombocytopenia or a failure to achieve remission within 2–3 weeks.
In viral serological studies, the full serological panels’ studies for IgM for EBV, CMV, and rubella IgM antibodies were investigated for all patients at diagnosis. A molecular study for PB19 was carried out for all patients’ sera.
Polymerase chain reaction for parvovirus B19
Extracted DNA from serum samples using the commercially available kit (Qiagen GmbH, Hilden, Germany) was used for PCR. Primers were designed to bracket a well-conserved region in PB19. The primers were pair A (5′-TGT GGT AAG AAA AAT AC-3′), (5′-TCA TTAAAT GGA TTT-3′) and primer B (5′-GGA ACAGACTTAGAGCTTATTC-3′), (5′-ACC CATCCTCTC TGT TTG ACT TAG TTG CTC GTAT-3′). Amplification was carried out in a 50 ml reaction volume of 67 mmol/l Tris (pH 8.8), 16.6 mmol/l (NH4)2SO4, 6.7 mmol/l MgCl2, 10 mmol/l mercaptoethanol, 6.7 mmol/l EDTA, 170 mg of BSA/ml, 50 pmol of each primer, and 1 U Taq polymerase overlaid with mineral oil. Positive control DNA to establish the specificity of the reaction was a cloned PB19 virus. An internal control for the β-globin gene was included as an internal control for each sample to exclude false-negative results. The reaction was carried on a programmable heating block. The temperature and the time regime used were as follows: 5 min at 95°C, 30 s at 50°C, and 35 cycles of 91°C for 1 min, 50°C for 1 min, and 67°C for 3 min. Gel electrophoresis was performed and a positive band was considered at (218 bp) and that for β-globin was found at 268 bp 8.
| Results|| |
The study was carried out on 49 children complaining of ITP. They were 27 males and 22 females. Their mean age was 5.4±3.9 years.
Positive viral markers either by positive serology IgM for rubella, CMV, or EBV or by a molecular study for PB19 were detected in 19 patients (38.8%). The most common viral infection was PB19 (30.6%), followed by EBV (16.3%), rubella (12.12%), and CMV (10.2%) [Table 1].
The clinical features did not vary between infected and noninfected patients in rash or lymphadenopathy (data not shown), except for the association of fever with PB19 (P<0.001). Although the hematological parameters were lower for infected children than for noninfected children, this decrease was not statistically significant [Table 2].
|Table 2: Comparative study between patients with positive viral markers and those with negative viral markers|
Click here to view
| Discussion|| |
The categorization of ITP implies that it is a single clinical-pathologic entity. However, variability in natural history and response to therapy suggests that ITP comprises heterogeneous disorders that eventuate in the production of platelet autoantibodies. ITP is characterized by increased destruction of antibody-coated platelets in the reticuloendothelial system. ITP of childhood is a common hematological disorder. In the majority of children with acute ITP, thrombocytopenia occurs within 1–3 weeks after an infectious disease. ITP may also occur after rubella, rubeola, chickenpox, or live virus vaccination 9.
Certain cases of otherwise ‘typical ITP’ are secondary to persistent, often in apparent infections (e.g. Helicobacter pylori or hepatitis C), or are accompanied by coexisting antibodies that might influence outcome (e.g. antiphospholipid antibodies). Insights from secondary forms (e.g. coexisting immune deficiency and molecular mimicry after infection) suggest that platelet-reactive antibodies arise through diverse mechanisms. In addition, environmental and genetic factors may impact platelet turnover, propensity to bleed, and response to ITP-directed therapy.
In children – unlike in adults – most cases of ITP are of acute onset and are often preceded by a viral-like infection 10–12. In a few cases, a specific viral infection (EBV, rubella, and varicella) is diagnosed 12. In the present study, the aim was to evaluate the presence of viral markers indicating an acute stage of infections associated with ITP in children.
The proportion of children with evidence of infections in our study was 38.8%. This represents a large proportion compared with other studies. In one study, the proportion of children whose ITP was associated with documented acute viral infection was 13.3% 6. In the present study, clinical manifestations and laboratory data of virus-associated or not associated groups are similar, except for the association of PB19 with fever (P<0.001). This finding may indicate the presence of a viral infection associated with ITP; however, its absence does not exclude that association. Therefore, the association of acute viral infections with ITP should be confirmed by laboratory investigations.
It was interesting to note that the highest frequency was found for PB19 detection by PCR (30.6%). This indicates that B19 infection is implicated in the development of cytopenia in some of our patients, in whom treatment with intravenous immunoglobulin might be beneficial. Several studies have shown that B19 infection may cause neutropenia and thrombocytopenia 13–16. Lee et al. 17 reported that four of 40 patients with ITP (10%) had active or recent infection with B19, as evidenced by the presence of the B19 IgM antibody and viral DNA in serum and/or the bone marrow in Taiwanese children with ITP. The significant association between B19 infection and pediatric ITP indicates tissue tropism of B19 beyond the erythroid progenitor cell 16. Previous studies have also shown that NS-1 protein, produced by PB19, could inhibit megakaryocyte colony formation 18,19. Whether B19 infection is implicated in the pathogenesis of ITP in adult patients remains controversial 20,21. The number of ITP patients in our study is small. Therefore, further study is required to examine the etiological role of B19 in pediatric patients with ITP.
Destructive thrombocytopenia of peripheral origin may result from immunologic-mediated antiplatelet antibody production with subsequent excessive platelet clearance in the reticuloendothelial system 15,22.
A number of viruses, including varicella, CMV, and EBV as well as vaccinations, are known to cause thrombocytopenia 10,12.
In the present study on Egyptian children, positive IgM for EBV was present in 16.3% of children with ITP. In areas where EBV is endemic, as in Taiwan, in a survey of 1350 healthy children younger than 15 years of age in Taipei city 23, the prevalence of a positive VCA antibody was 29.2% in children younger than 1 year of age and increased to 65 and 87.5% at the age of 2 and 3 years, respectively. Thus, the authors of that study concluded that most EBV infections occur in Taiwan during the first 1–4 years of life. The findings of other study by Hsiao 24 reported that the seropositivity rate of the ITP patients by age 4 was 83%. It has been speculated that this high prevalence may be because of the intimate relationships in Chinese family culture, such as prechewing food for children. In our locality, the prevalence of EBV may be related to overcrowding and intimate contacts between family members, leading to the distribution of droplet infection. Although there is a close association of EBV infection with ITP, the role of EBV in the pathogenesis of ITP in our locality requires further investigation, because this does not establish a causal relationship. Indeed, the lack of atypical lymphocytes casts some doubt on the significance of the EBV serological data. It was claimed that autoantibodies to platelet glycoproteins GPIIb/IIIa and GPIb/IX on platelets and in plasma may contribute to the occurrence of immune thrombocytopenia associated with EBV-related infectious mononucleosis. Moreover, there was effective therapeutic modification of ITP with the use of intravenous immunoglobulin 25. Recently, it has been speculated that EBV pathogenesis is likely because of improper immune modulation in response to the infection 26. Dysfunctional cellular immunity is considered to be essential to the pathophysiology of EBV–ITP 27.
Rubella IgM was present in 12.12% of our patients, although those patients were vaccinated according to the schedule of vaccination in Egypt at the age of 1 year. Second, antibody levels are declining, albeit less so for rubella 28.
Thrombocytopenia occurs occasionally after a naturally occurring infection with CMV, rubella, EBV, varicella zoster virus, the severe acute respiratory syndrome coronavirus, and many others 29–31. Thrombocytopenia may be immune, because of infection of megakaryocytes or progenitors, or may result from peripheral consumption, for example, purpura fulminans because of varicella zoster virus.
CMV commonly causes severe congenital thrombocytopenia and delayed platelet recovery after bone marrow transplantation. CMV can infect megakaryocytes, progenitor cells, and supporting stroma. An acute ITP-like syndrome occurs in immunocompetent and immunosuppressed individuals. In our patients, CMV IgM was detected in 10.2%. In another series of patients, three of 28 children and three of 80 adults with typical ITP had CMV in their urine, but there was no correlation between urinary viral clearance and the resolution of thrombocytopenia 32. However, in another study, platelet counts improved in four refractory patients after treatment for CMV 33. Eradication of CMV with antiviral therapy improved the ITP in these cases 34.
| Conclusion|| |
We conclude that in a large proportion of children with ITP, in our region, there is an association with acute viral markers similar to those of rubella, EBV, and CMV. Moreover, a significant proportion of them had occult PB19 viremia. A study of occult viral infections is recommended in children with ITP.
| References|| |
|1.||Schulman I. Idiopathic (immune) thrombocytopenic purpura in children: pathogenesis and treatment. Pediatr Rev. 1983;5:173–178 |
|2.||Robb LG, Tiedeman K. Idiopathic thrombocytopenic purpura: predictors of chronic disease. Arch Dis Child. 1990;65:502–506 |
|3.||Walker RW, Walker W. Idiopathic thrombocytopenic purpura: initial illness and long term follow up. Arch Dis Child. 1984;59:316–322 |
|4.||Tavil B, Unal S, Aytaç-Elmas S, Yetgin S. Weekly long-term intravenous immunoglobulin for refractory parvovirus B19 and Epstein–Barr virus-induced immune thrombocytopenic purpura. Turk J Pediatr. 2008;50:74–77 |
|5.||Winiarski J. Antibodies to platelet membrane glycoprotein antigensin three cases of infectious mononucleosis-induced thrombocytopenic purpura. Eur J Haematol. 1989;43:29–34 |
|6.||Yenicesu I, Yetgin S, Ozyürek E, Aslan D. Virus-associated immune thrombocytopenic purpura in childhood. Pediatr Hematol Oncol. 2002;19:433–437 |
|7.||Buchanan GR. Overview of ITP treatment modalities in children. Blut. 1989;59:96–104 |
|8.||Johnson G, Nelson S, Petric M, Tellier R. Comprehensive PCR-based assay for detection and species identification of human herpes viruses. J Clin Microbiol. 2000;38:3274–3279 |
|9.||Akbayram S, Akgun C, Dogan M, Caksen H, Oner AF. Acute ITP due to insect bite: report of 2 cases. Clin Appl Thromb Hemost. 2011;17:408–409 |
|10.||Kaplan C, Morinet F, Cartron J. Virus-induced autoimmune thrombocytopenia and neutropenia. Semin Hematol. 1992;29:34–44 |
|11.||Baranski B, Young N. Hematologic consequences of viral infections. Hematol Oncol Clin North Am. 1987;1:167–183 |
|12.||Wright JF, Blanchette VS, Wang H, Arya N, Petric M, Semple JW, et al. Characterization of platelet-reactive antibodies in children with varicella-associated acute immune thrombocytopenic purpura (ITP). Br J Haematol. 1996;95:145–152 |
|13.||Saunders PW, Ried MM, Cohen BJ. Human parvovirus induced cytopenias: a report of five cases. Br J Haematol. 1986;63:407–410 |
|14.||Doran HM, Teall AJ. Neutropenia accompanying erythroid aplasia in human parvovirus infection. Br J Haematol. 1988;69:287–288 |
|15.||Inoue S, Kinra NK, Mukkamala SR, Gordon R. Parvovirus B19infection: aplastic crisis, erythema infectiosum and idiopathic: aplastic crisis, erythema infectiosum and idiopathic thrombocytopenic purpura. Pediatr Infect Dis J. 1991;10:251–253 |
|16.||Murray JC, Kelley PK, Hogrefe WR, McClain KL. Childhood idiopathic thrombocytopenic purpura: association with human parvovirus B19 infection. AmJ Pediatr Hematol Oncol. 1994;16:314–319 |
|17.||Lee YM, Tsai WH, You JY, Ing-Tiau Kuo B, Liao PT, Ho CK, Hsu HC. Parvovirus B19 infection in Taiwanese patients with hematological disorders. J Med Virol. 2003;71:605–609 |
|18.||Ozawa K, Ayub J, Kajigaya S, Shimada T, Young N. The gene encoding the nonstructural protein of B19 (human) parvovirus may parvovirus B19 infection. Am J Pediatr Hematol Oncol. 1988;16:314–319 |
|19.||Srivastava A, Bruno E, Briddell R, Cooper R, Srivastava C, Van Besien K, Hoffman R. Parvovirus B19-induced perturbation of human megakaryocytosis in vitro. Blood. 1990;76:1997–2004 |
|20.||Lefrere JJ, Courouce AM, Kaplan C. Parvovirus and idiopathic thrombocytopenic purpura. Lancet. 1989;1:279 |
|21.||Van Elsacker-Niele AMW, Weiland HT, Kroes ACM, Kappers-Klunne MC. Parvovirus B19 infection and idiopathic thrombocyto-penic purpura. Ann Hematol. 1996;72:141–144 |
|22.||Oeda E, Shinohara K, Inoue H, Nomiyama J. Parvovirus B19 infection causing severe peripheral blood thrombocytopenia and persistent viremia. Am J Hematol. 1994;45:274–275 |
|23.||Tsai WS, Chang MH, Chen JY, Lee CY, Liu YG. Seroepidemiological study of Epstein–Barr virus infection in children in Taipei. Chung Hua Min Kuo Hsiao Erh Ko I Hseuh Hui Tsa Chih. 1989;30:81–86 |
|24.||Hsiao C-C. Epstein–Barr virus associated with immune thrombocytopenic purpura in childhood: a retrospective study. J Paediatr Child Health. 2000;36:445–448 |
|25.||Tanaka M, Kamijo T, Koike K, Ueno I, Nakazawa Y, Kurokawa Y, et al. Specific autoantibodies to platelet glycoproteins in Epstein–Barr virus-associated immune thrombocytopenia. Int J Hematol. 2003;78:168–170 |
|26.||Barnard D, Li JKK. Commentary on immune system associated diseases caused by viruses: the role of EBV. Front Microbiol. 2011;2:28 |
|27.||Jin CQ, Liu F, Dong HX, Zhang J, Zhou JW, Song L, et al. Type 2 polarized immune response holds a major position in Epstein–Barr virus-related idiopathic thrombocytopenic purpura (EBV-ITP). Int J Lab Hematol. 2011;34:164–171 |
|28.||Peltola H, Jokinen S, Paunio M, Hovi T, Davidkin I. Measles, mumps, and rubella in Finland: 25 years of a nationwide elimination programme. Lancet Infect Dis. 2008;8:796–803 |
|29.||Chou AL, Huang WW, Tsao SM, Li CT, Su CC. Human herpesvirus type 8 in patients with cirrhosis: correlation with sex, alcoholism, hepatitis B virus, disease severity, and thrombocytopenia. Am J Clin Pathol. 2008;130:231–237 |
|30.||Hui DS. Review of clinical symptoms and spectrum in humans with influenza A/H5N1 infection. Respirology. 2008;13:S10–S13 |
|31.||Rafailidis PI, Mourtzoukou EG, Varbobitis IC, Falagas ME. Severe cytomegalovirus infection in apparently immunocompetent patients: a systematic review. Virol J.. 2008;5:47 |
|32.||Wright JG. Severe thrombocytopenia secondary to asymptomatic cytomegalovirus infection in an immunocompetent host. J Clin Pathol. 1992;45:1037–1038 |
|33.||Psaila B, Bussel JB. Refractory immune thrombocytopenic purpura: current strategies for investigation and management. Br J Haematol. 2008;143:16–26 |
|34.||DiMaggio D, Anderson A, Bussel JB. Cytomegalovirus can make immune thrombocytopenic purpura refractory. Br J Haematol. 2009;146:104–112 |
[Table 1], [Table 2]