NEPHROLOGY 2002; 7, 125-129
Original Article
Cytomegalovirus disease among renal transplant recipients in India VINAY SAKHUJA,'VIVEKANAND JHA,'KUSUMJOSHI,' RITAMBRANADAI2KAMALSUD,' HARBIRS KOHLI,' KRISHANL GUPTA' AND SHOBHA SEHGAL3 Departments of 'Nephrology , 2Histopathologyand 31mmunopathology,Postgraduate Institute of Medical Education
and Research, Chandigarh, lndia SUMMARY: There is a paucity of data on cytomegalovirus (CMV) disease in renal transplant recipients from India. We have encountered this problem with increasing frequency at our centre, and document our experience in this paper. Case records of 514 recipients of renal transplants performed between 1980 and 1997, including 96 autopsies, were reviewed. Diagnosis was made by demonstrationof classic cytomegaly and intranuclear inclusions on histology, and/or detection of CMV pp-65 antigen in the peripheral blood leucocytes. Cytomegalovirus disease was documented in 21 cases (11 at autopsy, six by antigen detection, and four by a combination of histological demonstration and antigenemia detection). The incidence of this infection rose from 0.6% in 1980-1991 (precyclosporin era) to 5.5% (4.4% lethal) during 1992-1 997 (postcyclosporin era). At autopsy, the lung was the most frequently involved organ followed by the gastrointestinal tract. Ninety-five per cent of patients had co-infection with other organisms, and fungi were isolated in 80% of all patients with coinfection. Graft dysfunction was present in 17 cases (81%). We conclude that there has been a fourfold increase in the incidence of lethal CMV disease in renal transplant recipients after the introduction of cyclosporin immunosuppression. There is a high incidence of co-infection with other organisms, with fungi being the commonest offenders in tropical environment. This, along with allograft dysfunction, could account for the high mortality seen with CMV disease. KEY WORDS: cyclosporine, cytomegalovirus, infection, kidney transplantation.
INTRODUCTION
CMV antigenemia assay. The present report describes our experience with CMV disease occurring in renal
Both the intensity of immunosuppression and the type of epidemiological exposure determine the nature of infections in renal transplant recipients.' Common lifethreatening infections reported from tropical countries like India include tuberculosis and systemic fungal infections.'" Cytomegalovirus (CMV) infection has largely gone unrecognised in India, both because of a lack of awareness and non-availability of cheap and reliable confirmatory tests. At our institute, we encountered CMV infection at autopsy in renal transplant recipients, with increasing frequency in the last few years. More recently, clinical diagnosis of this infection could be confirmed by using a
transplant recipients from an area where it was previously unreported.
Correspondence: Professor Vinay Sakhuja, Head, Department of Nephrology, Postgraduate lnstitute of Medical Education and Research, Chandigarh 160 012, India. Email:
[email protected] Accepted for publication 7 September 2001.
METHODS Material for this retrospective study was obtained from two sources: (i) Case records of 514 recipients of renal transplants performed between 1980 and 1997 were analysed. Of these, 452 living related and four cadaver donor transplants were performed at our institute (Post Graduate Institute of Medical Education and Research) and 58 (49 living unrelated and nine living related) were operated elsewhere; ( i i ) autopsy records of 96 renal transplant recipients who died in the hospital from 1980 to 1997 were also reviewed. Details of history and physical findings were reviewed. Special note was made of the organ systems that showed clinical or laboratory evidence of involvement and the presence, type and extent of coexisting infections. A t autopsy, a note was made of the organs showing evidence of CMV infection. Criteria for diagnosis included demonstration of: (i) cells that were two- to fourfold larger than surrounding cells and showed
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intranuclear inclusions surrounded by a clear halo on histopathological examination of tissue obtained at autopsy or biopsy; and/or (ii) CMV antigenemia by indirect immunofluorescence in peripheral blood polymorphonuclear leucocytes using monoclonal antibodies to human CMV pp-65 antigen in patients with a clinical picture consistent with CMV disease. Patients with unexplained fever with leucopenia and/or transaminitis in the absence of other causes were considered to have a clinical picture consistent with CMV disease. The pretransplant CMV antibody status of kidney donors and recipients was not known. The immunosuppressive regimen in 154 patients who received a transplant until 1991 consisted of azathioprine 2mg/kg per day along with prednisolone 100mg/day for 1 week, reduced by 5 mg every other day to a maintenance dose of 1015 mg/day. After 1991, 272 patients received a triple drug regimen that consisted of cyclosporin A (CsA) 8 mg/kg per day for the first 14 days, reduced by 1 mg/kg per month to a maintenance dose of 4-5 mg/kg per day, prednisolone 30mg/day for the first 14 days and tapered gradually to 10-15 mg/day over 3 months and azathioprine 1-1.5 m&g per day. in the remaining 88 patients, CsA was administered at 10m&g per day for 1 month, then reduced by 1m&g per month to a maintenance dose of 5mg/kg per day. Prednisolone was given at 30mg/day for 2 months and tapered to 15mg/day over the next 2 months. Cyclosporine was replaced by azathioprine Zmgjkg per day after an overlap period of 6-8 weeks 1 year after transplantation in the latter two groups. Four cadaver transplant recipients received quadruple therapy including OKT3 for a period of 2 weeks followed by a tripledrug regimen. Acute rejection was diagnosed by using standard ciinical criteria and treated with either IV methylprednisolone 5001000mg/day or oral prednisolone 300-500 mg/day for 3-5 days. Twelve patients were treated with OKT3 at a dose of 2.5 mglkg per day for 10-14 days for steroid resistant rejection. Cytomegalovirus disease diagnosed ante-rnortem was treated with intravenous ganciclovir in a dose of 5 mgFg per twice daily (adjusted for renal function) for 2-3 weeks.
RESULTS Cytomegalovirus disease was documented in 21 (4.1%) renal transplant recipients. Diagnosis was established by using histopathological examination of tissues obtained at autopsy in 11 cases. Demonstration of pp-65 antigen in the circulating polymorphonuclear cells was the sole basis of diagnosis in six cases with clinical and laboratory features consistent with CMV disease. In the remaining four cases, antigen positivity was corroborated by histological evidence of infection, on colonic and esophageal biopsies in one case each, and at autopsy in two cases. All these 10 cases were diagnosed after 1996. The demographic characteristics of these patients are given in Table 1. The median interval between transplantation and development of CMV disease was 4 months. Over 66% of patients presented within 6 months of transplantation. The clinical features and laboratory findings of these patients are presented in Table2. Fever was the commonest presenting symptom, followed by cough and dyspnea. The commonest laboratory abnormalities were anaemia and leukopenia. Post-transplant diabetes mellitus was noted in 38% of cases. Seventeen patients (80.9%) were on cyclosporin at the time of development
V Sakhuja et nl.
Table 1 Demographic data of patients with cytomegalovirus disease
21
No. cases Age (years) Mean Range Sex ratio (Male : Female) Duration of symptoms before presentation (days1 Mode of diagnosis Autopsy only Serology only Serology biopsy Serology + autopsy
+
Time of onset 1-6 months 6-12 months > 12 months
44.165 15.28 16-41 17:4 14.6 f8.11
11 6 -, L
2
14 4 3
Serum creatinine (mg/dL) At onset Peak
3.9f1.34 6.8k2.19
Table 2 Clinical and Laboratory manifestations of patients with cytomegalovirus disease Parameter Clinical features Fever Cough Dyspnea CNS manifestations Jaundice Dysphagia Lower GI bleed Laboratory abnormalities Anemia Leukopenia Transaminasemia
PTDM Leukocytosis Thrombocytopenia
No.
Per cent
21 17 15 10 8 3 2
100 80.9 71.4 7.6 38.1 14.2 9.5
11 11 9 8
52.4 52.4 42.9 38.1 23.8 19
5 4
PTDM, post-transplant diabetes mellitus; GI, gastrointestinal; CNS, central nervous system.
of this infection. Eleven cases (53.4%) had received antirejection treatment in the 3-month period preceding the diagnosis (methylprednisolone in 10 cases and methylprednisolone followed by OKT3 in one case). Out of 27 autopsies of renal transplant recipients carried out between 1980 and 1991 (the period when azathioprine and prednisolone immunosuppression was used), only one patient (4%) showed evidence of CMV infection. In contrast, CMV was seen in 12 out of 69 (17%) transplant autopsies carried out between 1992 and 1997 (P=0.036), during which cyclosporin was added to the immunosuppressive regimen. Distribution of organ
CMV disease in renal transplantation
Lung Oesophagus Colon Adrenal Native kidney Allograft Stomach Pancreas
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=
involvement is shown in Fig. 1. The lungs were the most commonly involved organ followed by the oesophagus and the colon. Of the four cases who showed esophageal involvement at autopsy, two patients had complained of dysphagia. In another case, upper gastrointestinal endoscopy showed ulcers and biopsy revealed cytomegalic cells containing characteristic inclusion bodies. Two patients, who presented with lower gastrointestinal bleeding, showed colonic ulcers (one each at colonoscopy and at autopsy). Cells containing CMV inclusions were demonstrated in the mucosa on histopathological examination. Along with the CMV disease, simultaneous infection with other organisms was documented in 20 (95.2%) patients (Table 3). Fungal infections were the commonest and were seen in 16 patients (80%) with a superinfection. Candidal infection of the orophanynx, oesophagus, colon or urinary tract was documented in 10 cases. The other common organisms isolated during life were pyogenic bacteria and Mycobacterium tuberculosis. Acute allograft dysfunction was noted in a total of 17 patients (81%), and in six of the 10 cases in whom the diagnosis was made during life. Eleven patients required dialysis. Of the three dialysis-dependent patients who received ganciclovir therapy, renal function improved in two. A graft biopsy was done in one case and did not show any abnormality. A t autopsy, evidence of acute allograft rejection was seen in three cases. Two of these also showed CMV inclusions alongside the interstitial inflammation. One patient exhibited glomerular involvement with CMV. In the others, the graft dysfunction was probably caused by acute tubular necrosis, secondary to sepsis or nephrotoxic drugs, Five patients (23.8%) received intravenous ganciclovir for 2-3 weeks, four of whom showed good recovery. The diagnosis was made on the basis of positive antigenemia in all these cases. Two patients relapsed 3 and 6 weeks after the completion of therapy; one of these responded to a second 3-week course of ganciclovir. After explaining the risks to the patients and their families, immunosuppression was withdrawn in two, in one after the first episode, and in the other after she
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Table 3 Associated infections in patients with cytomegalovirus
disease *
Mode of Diagnosis Clinical Autopsy Clinical + Autopsy Fungal Candidiasis Aspergillosis Pneumocystis carinii Mucormycosis Pyogenic Tuberculosis Nocardiosis
6 3
7
Total
2
1
21 (51.2) 11 (26.8) 4 (9.8)
1
2
3 (7.3)
0 3
3 2 1 0
3 (7.3) 13 (31.7) 6 (19.6) 1 (2.4)
2 0
1
Figures in parentheses are percentages. *Twenty patients had 41 infections, 16 patients had infection with more than one organism.
relapsed. Withdrawal of immunosuppression was the only measure used in three other cases (as ganciclovir was not available in India at that time), all of whom died. In total, there were 17 deaths.
DISCUSSION Cytomegalovirus is the most important viral pathogen encountered in renal transplant recipients in Western nations with a temperate ~lirnate.“”~Approximately two-thirds of transplant recipients show evidence of infection, and 30% of these develop clinical d i ~ e a s e . ~ A direct correlation has been shown between the prevalence of anti-CMV antibody and the socioeconomic condition of the population.8 Hot and humid climate, overcrowding, poor hygiene and large families in the developing countries of the tropical region provide ideal conditions for its spread. Among Indian adults, the antibody prevalence is 85-100%,”’* and hence CMV disease seen in the post transplant period is likely to be almost entirely caused by reactivation or superinfection. This is the first study from India to systematically document the pattern of CMV disease in renal transplant recipients. The incidence of 4% is an underestimate of the problem in the present era of cyclosporin and monoclonal/ polyclonal antibody use. Several factors are likely to be responsible for this low figure. The pp-65 CMV antigenemia assay was introduced at this Institute only in 1996, and patients tested with the relatively insensitive CMV antibody testing used before this, very likely escaped detection. It is also possible that mild reactivations involving transient pyrexias and hepatitis or leukopenia were attributed to other diagnoses, and thus, the diagnostic tests were applied only to those with severe disease. Also, a significant proportion of patients went back to their primary physicians for follow up (where tests for CMV are not available), and hence, were
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not seen at our centre. The immunosuppression policies also mitigated against this infection; monoclonal and polyclonal antibodies started to be used at this Institute in 1995 and even then, only a small proportion of patients who needed these agents for various indications could actually afford them. The present study has also confirmed an increase in the incidence of CMV disease in the postcyclosporin era. Over 95% of the cases were seen after 1991, when cyclosporin was introduced in the immunosuppressive regime. Although a low index of suspicion of this infection could account for the low incidence in the precyclosporin era, CMV was documented fourfold more frequently, even at autopsy during 1992-1997 compared with 1980-1991, suggesting that the increase was indeed real. The autopsy rates as percentages of total transplants were similar in both time periods, but we were unable to precisely determine the autopsy/death ratios in the two eras because the migrant nature of our patient population prevented us from establishing the exact number of deaths. The histological diagnosis was based on light microscopy in the present study, and it is possible that the use of more sensitive techniques like immunohisto. chemistry might have detected more cases. Cytomegalovirus disease among renal transplant recipients presents with a non-specific febrile illness, malaise, anorexia and increasing fatigue. Other symptoms depend upon the organ involved. In the present study, the lung was the most frequently involved organ, followed by the gastrointestinal (GI) tract. The GI tract is involved in approximately half of all patients with disseminated CMV disease, almost all of whom are AIDS patients or organ transplant recipients.12 Gastrointestinal bleeding and perforation secondary to direct mucosal involvement or CMV vasculitis are common presenting features.13In the present series too, involvement of the colonic mucosa leading to massive lower gastrointestinal bleeding was the mode of presentation in two cases. Endoscopic biopsy from the involved part of the GI tract is useful for confirming invasive CMV disease, even in the absence of CMV antigenemia.14 Post-transplant diabetes mellitus (PTDM) was seen in 38% of our patients, a figure that is much higher than that reported in the 1iterat~re.l~ In an earlier study from our centre, PTDM was noted in 9.8% of all renal transplant recipients on cyclosporin.16 Hjelmesaeth et al. documented PTDM and impaired glucose tolerance in 42 and 32% patients treated for CMV infection, respecti~e1y.l~ In morphological studies on the pancreas of infants with generalised CMV infection, islet cell changes were found in approximately half of the Fox and Murphy showed development of diabetes mellitus in 10 out of 24 Octodon deguses infected with CMV.” All these studies suggest the possibility of a direct role of CMV in the causation of PTDM. In the present study, however, CMV was demonstrated in the pancreas in only one out of 11 autopsied cases.
V Sakhuja et al.
Graft function improved in two of the three patients who received ganciclovir therapy. Also, CMV inclusions were seen in the midst of interstitial inflammation at autopsy in two patients. These findings suggest a direct etilogical role for CMV in the tubulo-interstitial nephritis and the graft dysfunction. The exact mechanism of this phenomenon is not certain, and the postulations include the upregulation of human leucocyte antigen (HLA) antigens on the graft by cytokines that are elaborated in response to CMV, and immunological damage caused because of homology between CMV and histocompatibility antigens.21 Opportunistic infections with fungi and M. tuberculosis were noted in a majority of our patients in close temporal relationship with the CMV disease, and contributed to the mortality. A high index of suspicion is necessary for diagnosing and treating multiple lifeethreatening infections in those with CMV disease. This entails use of invasive diagnostic techniques like bronchoscopy, aspiration cytology and tissue biopsies. Non-availability of ganciclovir was a major hindrance in treatment until 1998. Even after it became available, high cost continues to keep it beyond the reach of many patients. The approximate cost of a 3-week course is US$800. Added to the cost of cyclosporin-based immunosuppression (approximately US$300400/month), the financial burden on the patients becomes prohibitive in a country with an annual per capita gross national product (GNP) of US$400 and no national or private health insurance schemes. The prophylactic and ‘pre-emptive’ treatment strategies for CMV advocated in the Western are not suitable for developing countries because of these reasons. We therefore restricted the use of ganciclovir to those viremic patients who showed clinical evidence of CMV disease. This strategy is similar to the ‘deferred therapy’ described by Brennan et a1.,12and seems suitable for countries having patients with limited tinancia1 resources. Sensitive and specific diagnostic tests are, however, necessary for optimal use of this strategy. The CMV antigenemia test is now offered free of charge at our hospital, which is funded by the government. To the best of our knowledge, the pp-65 antigenemia assay is not done elsewhere in the country. The approximate cost of CMV antibody testing and PCR in private laboratories is US$15-20 and 100-125, respectively. In conclusion, CMV disease has become increasingly common among renal transplant recipients on cyclosporin in India. Lungs and the gastrointestinal tract are the most frequently involved sites. A high proportion of patients have coexistent post-transplant diabetes mellitus. A variety of fungal, bacterial and mycobacterial superinfections contribute to the high mortality. Detection of pp-65 antigen in the circulating polymorphs allows early and accurate diagnosis. A high index of suspicion, rapid and accurate diagnosis and early treatment of both the CMV disease and associated infections is necessary to reduce the mortality and morbidity.
CMV disease in renal transplantation
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