| Abstract|| |
Reactivation of infection with the cytomegalovirus (CMV) in renal transplant recipients may cause significant morbidity and mortality. To evaluate factors associated with activation of CMV replication, we followed prospectively a group of 68 renal transplant recipients for 12 months. The control group consisted of healthy blood donors (n = 37). Sera were collected periodically from these patients and analyzed for the presence of specific anti-CMV antibodies. Enzyme-linked immunoassay based on recombinant CMV proteins was used to detect the following antibody specificities: IgG and IgM. During the first year after transplantation, reactivation of CMV occurred in 48 recipients (70.6%). Detailed analysis did not show any association of reactivation with the type of basic immunosuppressive therapy, prophylactic or therapeutic use of anti-lymphocyte antibodies, as well as occurrence of acute rejection episodes. There was a borderline association (P=0.068) between the presence of CMV infection and EBV reactivation. In conclusion, results of our study suggest that CMV infection may represent a factor activating EBV replication
Keywords: Cytomegalovirus, Reactivation, Acute rejection, Epstein-Barr virus, Immunosuppressive therapy
|How to cite this article:|
Khameneh ZR. Occurrence of Cytomegalovirus Infection and Factors Causing Reactivation of the Infection among Renal Transplant Recipients: A Single Center Study. Saudi J Kidney Dis Transpl 2008;19:41-5
|How to cite this URL:|
Khameneh ZR. Occurrence of Cytomegalovirus Infection and Factors Causing Reactivation of the Infection among Renal Transplant Recipients: A Single Center Study. Saudi J Kidney Dis Transpl [serial online] 2008 [cited 2020 Nov 26];19:41-5. Available from: https://www.sjkdt.org/text.asp?2008/19/1/41/37431
| Introduction|| |
Viral infections constitute the single greatest cause of infectious-disease related morbidity and mortality in organ transplant recipients. 
Cytomegalovirus (CMV), which frequently causes latent asymptomatic infection in healthy adults, may evade immune surveillance in immune compromised patients and start to replicate. Reactivation of chronic Epstein Barr Virus (EBV) infection sometimes results in clinical symptoms (fever, leukopenia, pharyngitis, hepatitis, lymphadenopathy) and eventually may lead to uncontrolled proliferation of B cells terminating in post-transplant lymphoproliferative disease (PTLD). ,,,, The risk of PTLD is mostly deter-mined by the prevalence of anti-EBV sero-positivity in transplanted population. The probability of PTLD is further increased by concomitant use of antilymphocytic anti-bodies (ALG) and infection with the CMV.
It has been assumed that similar factors may affect the rates of reactivation of EBV during the post-transplant course. We therefore undertook this prospective study in a group of kidney allograft recipients to investigate the effect of type of immunosuppressive regimen, incidence of CMV infection and the number of rejection episodes, on EBV reactivation as assessed by serological markers.
| Patients and Methods|| |
We studied 68 renal allograft recipients (38 male and 30 female) who were transplanted at The Transplantation Institute, Medical University of Urmia. The most frequent causes of renal insufficiency of the native kidneys in the studied patients were chronic glomerulonephritis, pyelonephritis, and polycystic kidney disease. Immunosuppressive regimen consisted of cyclosporine (CsA), azathioprine (Aza), prednisone (Pred) and mycophenolate mofetil (MMF). Thirty-five patients received triple drug immunosuppression (Aza, CsA and Pred), eight patients received (Pred, CsA and MMF), while 25 patients were administered double drug regimen (Pred and CsA). A total of 19 episodes of acute rejection were diagnosed; all were treated with 0.5 to 1.0 gm of methylprednisolone, given for three days. In eight cases of resistant rejection, antibody therapy in the form of ALG was used in addition. None of the 68 study recipients had clinically evident active EBV infection. There were no cases of PTLD, or infectious mononucleosis among the patients studied.
A total of 35 healthy blood donors (n = 35; mean age 34.31 ± 11.46 years; range: 19-35 19-35 years) constituted the control group.
| Diagnostic Assays of Cytomegalovirus Infection|| |
Serum samples were collected prospectively from all patients before transplantation and at 3, 6, 9 months and 1 year after transplantation. Sera were stored at -70 o Celsius. The samples were tested for IgG and IgM anti-CMV antibodies. The levels of antibodies were determined using commercially available sensitive enzyme-linked immunosorbent assay (ELISA) method.
| Statistical analysis|| |
Data were analyzed using Student's t-test or Mann-Whitney test where appropriate. Rates were compared by means of chi-square test.
| Results|| |
Pre-transplant CMV serology
Both renal allograft recipients and subjects from the control group had high rate of seropositivity against CMV. At the time of transplantation, 94.1% of the recipients had serological evidence of previous infection in comparison to the 88.6% positivity among healthy blood donors. Additionally, serological evidence of reactivation (appearance of anti-IgM in previously seropositive patients) was detected in 5.9% of the recipients, which was lower than among the control group (8.6%): these frequencies, however, were not significantly different.
Serological evidence of reactivation of CMV infection in the post-transplant period
In the sera collected before transplantation, 8.8% of the samples among the study subjects tested positive for CMV-IgM as against 13.5% among the control group. However, 12 months after transplantation, the CMV-IgM positivity rate increased from 8.8% to 72.1%. Although this increase in seropositivity did not reach statistical significance (X 2 =2.53, P=0.112), it shows that in some recipients, reactivation of CMV infection occurred during the posttransplant period.
The effect of immunosuppressive therapy on CMV serology
A total of 40 recipients (58.8%) had serological evidence of reactivation. In this group, 20 subjects were treated with Pred + Aza + CsA, 17 patients received Pred + CsA, while three recipients were treated with Pred + MMF + CsA. There was no statistically significant difference in the frequency of reactivation in the patient-groups receiving different immunosuppressive regimens.
The effect of acute rejection on CMV infection
Among the 68 patients studied, acute rejection was diagnosed in 19. The acute rejection episodes occurred after a mean period of 3.78 ± 0.12 months following transplantation. All these patients received therapy with pulse methylprednisolone, while eight recipients with steroid-resistant rejection were treated with ALG in addition.
Ten recipients had serological markers of previous CMV infection in sera collected prior to the occurrence of the rejection episode. In serum samples obtained after a rejection episode and therapy, only three patients had serological markers of reactivation of CMV infection. Rejection episodes as well as anti-rejection therapy did not affect the serological status of CMV infection. Also, there was no effect of ALG administration on serological markers of CMV infection
The effect of EBV infection on CMV activation
Reactivation of, or occurrence of primary EBV infection is a common complication during the post-transplant course. Before transplantation, 85.3% of the recipients were latently infected with EBV while only 14.7% of the recipients had serological evidence of EBV reactivation just prior to transplantation. During the first 12 months after transplantation, reactivation of EBV infection occurred in 40 recipients (58.8%). No primary infection with EBV was diagnosed during this period.
Since EBV infection causes a transient immunosuppression, which may exacerbate CMV replication, analysis of relationship between CMV and EBV infection was carried out. Among the 40 patients who had reactivation of EBV infection, serological markers of CMV reactivation and replication developed in 23 recipients. Although this prevalence is not statistically significant (X 2 = 3.22, p=0.068), it indicates that EBV infection may play a role in reactivation of CMV infection.
| Discussion|| |
In this study, we studied CMV serology during the post-transplant period in sera consecutively collected over a period of one year. At the time of transplantation, four subjects (5.9%) had serological evidence of reactivation of latent infection. During the first post-transplant year the number of recipients who developed reactivation doubled up to 48 patients (70.6%). The diagnosis of reactivation was based on seroconversion of anti IgM. Using the same parameter, Hornef detected reactivation in 24.4% recipients observed during 10-59 weeks after transplantation. In another group of renal allograft recipients, reactivation occurred in 29.5% of patients during 649 days of observation. ,, For comparison, in a retrospective study conducted on 67 heart-lung and 295 heart transplant recipients, Gray et al.  found reactivation of past infection in 17.4% of the recipients. This is a very interesting finding since these recipients were routinely treated with prophylactic ALG despite which the rate of reactivation was at the same level as in patients in our study, most of whom did not receive ALG or OKT3. There is a common notion that intensification of immunosuppression increases the risk of reactivation of CMV. , There was no significant difference noted in the rate of reactivation among our patients who received different basic immunosuppressive protocols. Unfortunately, the number of recipients in our study is too small to draw any conclusions but our data provide some suggestions regarding the effect of basic immunosuppression with MMF on the rate of CMV reactivation. It seems that the use of MMF increases the risk of CMV reactivation.
There are only few studies in the literature estimating the rates of seroconversion with the use of different immunosuppressive protocols. Hornef et al.  did not find any differrence in seroconversion rate (IgM) between recipients treated or not treated with ALG induction therapy. However, when they analyzed their data including the rise in antiIgG OD value as well as a two-fold rise in anti-IgA value, an increase in the reactivation rate was noted in the group on quadruple immunosuppression.
Rejection can be associated with activation of some infections. This may be due to injury induced by alloantigen-driven inflammation or intensification of immunosuppressive regimen as part of anti-rejection therapy. Also, viral infections may facilitate the development of rejection. In our study, rejection episodes occurred in 19 recipients, all of whom received therapy with high doses of methylprednisolone while eight patients with steroid-resistant rejection were treated with anti-lymphocyte antibodies, in addition. No change was detected in viral serology in the sera collected before and after rejection. These data suggest no significant association between rejection episodes and EBV reactivation. On the contrary, Hornef et al. found 43.3% reactivation rate in recipients with acute rejection. They found that serological evidence of reactivation preceded the occurrence of acute rejection in some patients (23.1%) while in others (76.9%), the rejection episodes were followed by serological signs of CMV reactivation. The authors also described a significant correlation between groups of patients who had experienced at least one rejection episode versus patients who did not have any, and the incidence of a greater than four-fold rise of the anti-IgG OD value. In their data, patients suffering from graft rejection showed constant high amounts of anti- EA-IgG.
These apparent discrepancies between our data and of Hornef et al. may be explained by some differences in the patient population studied. In the group of patients studied by Hornef et al,  there was a much higher proportion of recipients who lost their grafts and who had received much more intensified immunosuppressive regimen. Also, Hornef et al  detected serological evidence of CMV reactivation in sera collected even after 205 days after the rejection episode. In the majority of their patients, reactivation occurred after rejection, suggesting causal role of anti-rejection therapy. Also, the majority of rejection episodes were detected during the first post-transplant month while in our study the mean time of diagnosis of acute rejection was 3.78 months. Therefore, it is possible that at least in some recipients in our study, serological symptoms of reactivation could have been detected if their sera had been collected after longer duration than the one year as in our study.
In a small group of pediatric renal allograft recipients, Acott et al  found CMV reactivation in only one of the eight patients who had acute rejection, which matches our data on very low rate of CMV reactivation during acute rejection. In this short communication, the authors did not provide any data on the changes in CMV serological status during longitudinal observation.
Nine patients (13.8%) in our study had active CMV infection. Unfortunately, there is no data on donor's CMV serology, and thus it cannot be established if the transplanted kidney was a source of the virus in these cases. Six patients with active CMV infection developed serological evidence of EBV reactivation in the post-transplant period; of the remaining recipients only 10 had EBV reactivation. This difference reaches the border of statistical significance (X 2 =3.22, p=0.088), and might indicate association between CMV infection and EBV reactivation.
This relationship is also suggested by other authors. Hornef et al, found a striking coincidence of active, and particularly symptomatic CMV infection and serological evidence of EBV reactivation. According to their data, serological reactivation of EBV infection generally occurred within a short period of CMV infection (less than 3 weeks). O'Neill and Shirodaria  have reported on higher anti-VCA IgG titers in renal allograft recipients without CMV infection than in patients with CMV infection. In the majority of their patients, they found a significant increase in anti-VCA IgG and anti-EA IgG as well as appearance of anti-VCA IgM. They also concluded that these findings might represent reactivation of these viruses during CMV infection.
Similar common occurrence (around 40%) of EBV reactivation and CMV disease was reported in a study by Merlino et al.  Also, the role of CMV infection in PTLD has been implicated. Results from liver and heart transplant recipients show that the risk of this complication is significantly increased when primary EBV infection is accompanied by CMV disease. 
It must be emphasized that immuno competent individuals with acute primary CMV infection also show the serological parameters of EBV reactivation in 40% of cases.  Only two recipients in our study had CMV disease with EBV reactivation, associated with prior anti-lymphocyte therapy. From these data, it does not seem that ALG therapy predisposes to concomitant activation of both viral infections.
In conclusion, results of our study suggest that CMV infection may represent a factor activating EBV replication. We did not make any effort to evaluate the potential mechanisms of this interaction. Even in the available literature, there are no data, which may explain this phenomenon and further studies are required in this regard.
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Zakieh Rostamzadeh Khameneh
Assistant Professor of Virology, Urmia University of Medical Sciences, Urmia