© 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd Transplant Infectious Disease, ISSN 1398-2273
Cryptosporidium infection after renal transplantation in an endemic area D. Bhadauria, A. Goel, A. Kaul, R.K. Sharma, A. Gupta, V. Ruhela, A. Gupta, H. Vardhan, N. Prasad. Cryptosporidium infection after renal transplantation in an endemic area. Transpl Infect Dis 2015: 17: 48–55. All rights reserved Abstract: Background. Cryptosporidium is one of the common causes of infective diarrhea in post-transplant patients in endemic areas. However, data are limited on Cryptosporidium infection in recipients of solid organ transplantation. The aim of this study was to determine the incidence, disease manifestation, management, and outcome of Cryptosporidium infection in living-donor renal transplant recipients (RTR). Methods. We performed a detailed retrospective review of the data on all RTR who had diarrheal illness requiring evaluation and hospitalization, and Cryptosporidium infection. Results. During the study period, 119/1235 (8.98%) RTR developed diarrhea, and Cryptosporidium was found in 34/119 (28.5%). Nine of 680 (1.3%) patients were on a cyclosporine (CSA)-based regimen, and 25/643 (3.8%) patients were on a tacrolimus (Tac)-based regimen. The relative risk of developing Cryptosporidium infection was lower on the CSA-based regimen, compared with the Tac-based regimen (odds ratio [OR]: 0.35, 95% confidence interval [CI]: 0.17– 0.72, P = 0.003). Twelve of the 34 patients had acute graft dysfunction, mainly caused by combined Tac toxicity and dehydration. Mean serum creatinine and trough Tac level were 2.04 0.65 mg/dL and 8.24 1.19 ng/dL, respectively. Nitazoxanide alone was used in 13 patients, and nitazoxanide in combination with fluoroquinolone in 21 patients, with duration of treatment ranging from 16 to 60 days. Tac was changed to CSA in 8/ 11 patients. The clearance of cysts and response to nitazoxanide alone were significantly lower, compared with combination therapy (61.53% vs. 95.23%, P = 0.01, 38.46 vs. 85.71%, P = 0.004, respectively). The OR for cyst clearance and response was also significantly lower with nitazoxanide alone, in comparison with combination therapy (OR: 0.65, 95% CI: 0.34–0.92, P = 0.01, OR: 0.45, 95% CI: 0.21–0.81, respectively). Four (16%) of 24 patients with response had relapse. Conclusion. Patients with Tac and mycophenolate mofetil combination therapy had a significantly high risk of Cryptosporidium infection. Cryptosporidial infection may require prolonged nitazoxanide therapy, either alone or in combination, with or without reduction in immunosuppression.
Diarrhea is one of the most common complications observed in solid organ transplant (SOT) recipients (1). SOT recipients are at increased risk for infectious diarrhea. Cryptosporidium has been increasingly
48
D. Bhadauria1, A. Goel2, A. Kaul1, R.K. Sharma1, A. Gupta1, V. Ruhela1, A. Gupta1, H. Vardhan1, N. Prasad1 1
Nephrology, Sanjay Gandhi Post Graduate Institute of Medical Sciences, Lucknow, Uttar Pradesh, India, 2 Gastroenterology, Sanjay Gandhi Post Graduate Institute of Medical Sciences, Lucknow, Uttar Pradesh, India
Key words: Cryptosporidium; renal transplant; diarrhea; immunosuppression Correspondence to: Narayan Prasad, Nephrology, Sanjay Gandhi Post Graduate Institute of Medical Sciences, Type-4/11, SGPGIMS campus Rae Bareli Road Lucknow, Lucknow, Uttar Pradesh 226014 India Tel: 91-522-2495188 Fax: 91-522-2668017 E-mail: [email protected]
Received 22 April 2014, revised 21 August 2014, 26 October 2014, accepted for publication 26 October 2014 DOI: 10.1111/tid.12336 Transpl Infect Dis 2015: 17: 48–55
recognized as one of the common causes of infectious diarrhea in the post-transplant period, particularly in endemic areas like India (2, 3). Cryptosporidium is an intracellular protozoan parasite that infects the
Bhadauria et al: Cryptosporidium in renal transplant patients
epithelial cells of the digestive tract. The illness is typically self-limited in immunocompetent hosts, but may be severe and prolonged in patients with deficient cellular and humoral immunity, as seen in patients with human immunodeficiency virus (HIV) infection, organ transplantation, cancer treatment with immunosuppression, or immunoglobulin-A deficiency or hypogammaglobulinemia (4–7). Cryptosporidium are ubiquitous in natural water sources throughout the world and are acquired through contaminated food or water, usually leading to illness (4–6). The largest outbreak, following fecal contamination of drinking water, took place in Milwaukee, Wisconsin, USA, in the early 1990s (8). Cryptosporidial infection in immunocompromised hosts has been reported mainly in patients with acquired immunodeficiency syndrome (4, 5). The data are mainly limited to either case series (7–9) or case reports (5, 9–11), which are insufficient to determine incidence, risk factors, and the optimal antimicrobial regimen in renal transplant recipients (RTR) or other SOT patients. In the United States and Europe, cryptosporidiosis in SOT recipients is an uncommonly recognized illness; most reports are confined to single cases or small series (12–15). A few case series and epidemiological studies of stool carriage of Cryptosporidium in SOT recipients have been reported from endemic areas of the Middle East, India, and South America (2, 3, 6, 16–22). Children, individuals who reside in or travel to endemic areas, and intestinal graft recipients are particularly at increased risk of cryptosporidial infection (2, 10, 17, 23, 24). In a recently published report of 10 SOT recipients with cryptosporidial gastroenteritis, elevated levels of tacrolimus (Tac) were observed (10). The aim of this study was to determine the incidence, disease manifestation, management, and outcome of cryptosporidial infection in living-donor RTR.
Protocols of treatment of diarrheal illness and cryptosporidial infection The protocol of evaluation and treatment of diarrheal illness is summarized in Figure 1. We treat diarrheal illness according to the etiology of the diarrhea (Table 1). A thorough workup for diarrhea was performed, including clinical examination, assessment of hematological and biochemical investigations, examination for cytomegalovirus (CMV) viremia, standard stool culture, and a parasitological examination (Cryptosporidium, Isospora, Microsporidium, Entamoeba, and Giardia), plus Clostridium difficile was searched for in the stool (culture and toxin). In cases where weight loss and/or biological malabsorption symptoms occurred, an upper endoscopy with duodenal biopsies, with or without colonoscopy, was performed. Cryptosporidium in stool is diagnosed by modified acidfast bacilli (AFB) staining. For 3 consecutive days, freshly collected stool samples were transported to the laboratory within 60 min and were examined by modified AFB staining technique. This technique is used to detect oocysts of coccidian species (Cryptosporidium, Isospora, and Cyclospora). Unlike the Ziehl-Neelsen stain, this AFB stain does not require the heating of reagents for staining. Under the microscope, Cryptosporidium oocysts appear pink-red against a green background. Cryptosporidial infection is treated initially with nitazoxanide for 5–7 days. We included the patients with persistent, symptomatic, and clinically significant diarrheal illness persisting for ≥7 consecutive days that necessitated evaluation and hospitalization (Table 2). Response to therapy was defined as improvement in clinical symptoms with stool clearance for cysts.
Patients and methods In this retrospective observational study, we reviewed the data of all living-related RTR who had diarrheal illness requiring evaluation and hospitalization from January 2002 to June 2013. This study was carried out in a major transplant center in the northern part of India. The demographic profiles, immunosuppression regimen, details of diarrheal illness, hospitalization, management, and outcomes of the patients were retrieved from the electronic Hospital Information System of the institute.
Fig. 1. Protocol for diarrheal diseases. MMF, mycophenolate mofetil; CMV, cytomegalovirus; Cl., Clostridium; D2, second part of duodenum.
Transplant Infectious Disease 2015: 17: 48–55
49
Bhadauria et al: Cryptosporidium in renal transplant patients
Etiology of diarrhea and other opportunistic infections (N = 119) Etiologies of diarrhea
n (%)
Cryptosporidium
34/119 (28.5%)
Non-cryptosporidial infection CMV
48/119 (48.3%) 18
Candida albicans
8
Giardia
7
Clostridium difficile
4
Escherichia coli
3
Isospora belli
5
Microsporidium
3
Drug-induced
35 (29.4%)
PTLD
2 (1.6%)
Other opportunistic infections
17 (50%) patients
CMV (other than enteritis)
7 (20.5%) patients
Systemic fungal
8 (23.5%) patients
Cutaneous fungal
4 (11.7) patients
BKVIN
2 (5.8%) patients
CMV, cytomegalovirus; PTLD, post-transplant lymphoproliferative disorder; BKVIN, BK virus interstitial nephropathy.
Table 1 Clinical symptoms and signs of cryptosporidial infection (N = 34) Symptoms and signs
Number of patients (%)
Fever
11 (32.35)
Malaise
25 (73.25)
Profuse watery diarrhea
12 (35.29)
Crampy abdominal pain
17 (50)
Weight loss
9 (26.47)
Vomiting
18 (52.94)
Dehydration
15 (44.11)
Hypotension
8 (23.52)
Table 2
Relapse of disease was defined as re-appearance of symptoms with stool positive for cysts, after clinical response to treatment and normal stool examination, 2 weeks after stopping treatment.
Immunosuppression regimen All patients received a triple-immunosuppression regimen including steroid, calcineurin inhibitors (either cyclosporine [CSA] or Tac), and mycophenolate mofetil
50
Transplant Infectious Disease 2015: 17: 48–55
(MMF). CSA was used until December 2006, and Tac was used from January 2007 onward. The CSA dose was adjusted to maintain a trough level between 300 and 400 lg/L, and then gradually tapered to 100– 200 lg/L at 12 weeks. The dose of Tac was adjusted to maintain a trough level between 8 and 12 ng/mL for the first 6 months, and thereafter between 4 and 6 ng/ mL. All patients received methylprednisolone 500 mg intravenously during the intraoperative period, subsequently 20 mg of prednisolone daily until 8 weeks, and thereafter the dose of prednisolone was tapered every week by 2.5 mg to reach the nadir dose of 10 mg daily by 12 weeks. Interleukin-2 receptor blocker was used as induction in low immunological risk patients who opted for this treatment. Anti-thymocyte globulin (ATG) was used for induction in high-risk patients (those with re-transplantation, previous cross-match positivity, or high panel-reactive antibodies), and for treatment of acute rejection, as per indication. All rejection episodes were treated with 3–5 doses of 500 mg intravenous methylprednisolone initially, followed by oral prednisolone to reach the nadir in the next 4 weeks. Steroid-resistant rejection was treated with ATG, and antibody-mediated rejection was treated with intravenous immunoglobulin and plasmapheresis, as per our institute protocols. Cotrimoxazole was used universally as chemoprophylaxis for at least 6 months after transplant. AntiCMV prophylaxis was added to patients who received ATG.
Treatment regimen for Cryptosporidium All patients with Cryptosporidium infection were treated with either nitazoxanide alone (at a dose of 500 mg 3 times a day), or as a combination of nitazoxanide and fluoroquinolone (dosed according to glomerular filtration rate). The patient was first evaluated for response after 5–7 days of initial therapy. Clinical improvement and stool examination for clearance of Cryptosporidium cysts was performed weekly. The treatment was stopped if both stool clearance and clinical improvement were observed. In cases of partial response to initial nitazoxanide (improvement in symptoms, but stool still positive for cysts), the duration of treatment was extended up to 2 months. In case of non-response to initial nitazoxanide, fluoroquinolone was added and the duration of treatment was extended up to 2 months. Patients were observed for recurrence during follow-up. The patients who relapsed were treated with triple therapy with nitazoxanide, fluoroquinolone, cotrimoxazole
Bhadauria et al: Cryptosporidium in renal transplant patients
(double-strength tablet twice daily), and azithromycin (500 mg once a day), along with reduction in immunosuppression.
Modification of immunosuppression Immunosuppression was modified in case of nonresponse or partial response to antimicrobials, relapse, or in the presence of other opportunistic infections. MMF was switched to azathioprine (Aza), except in patients with concomitant Tac nephrotoxicity, in whom Tac was switched to CSA (Fig. 2).
Statistical analysis All data are presented in form of mean standard deviation. The categorical variables were compared using Fisher exact test and chi-square test. A P-value of
Cryptosporidium infection after renal transplantation in an endemic area D. Bhadauria, A. Goel, A. Kaul, R.K. Sharma, A. Gupta, V. Ruhela, A. Gupta, H. Vardhan, N. Prasad. Cryptosporidium infection after renal transplantation in an endemic area. Transpl Infect Dis 2015: 17: 48–55. All rights reserved Abstract: Background. Cryptosporidium is one of the common causes of infective diarrhea in post-transplant patients in endemic areas. However, data are limited on Cryptosporidium infection in recipients of solid organ transplantation. The aim of this study was to determine the incidence, disease manifestation, management, and outcome of Cryptosporidium infection in living-donor renal transplant recipients (RTR). Methods. We performed a detailed retrospective review of the data on all RTR who had diarrheal illness requiring evaluation and hospitalization, and Cryptosporidium infection. Results. During the study period, 119/1235 (8.98%) RTR developed diarrhea, and Cryptosporidium was found in 34/119 (28.5%). Nine of 680 (1.3%) patients were on a cyclosporine (CSA)-based regimen, and 25/643 (3.8%) patients were on a tacrolimus (Tac)-based regimen. The relative risk of developing Cryptosporidium infection was lower on the CSA-based regimen, compared with the Tac-based regimen (odds ratio [OR]: 0.35, 95% confidence interval [CI]: 0.17– 0.72, P = 0.003). Twelve of the 34 patients had acute graft dysfunction, mainly caused by combined Tac toxicity and dehydration. Mean serum creatinine and trough Tac level were 2.04 0.65 mg/dL and 8.24 1.19 ng/dL, respectively. Nitazoxanide alone was used in 13 patients, and nitazoxanide in combination with fluoroquinolone in 21 patients, with duration of treatment ranging from 16 to 60 days. Tac was changed to CSA in 8/ 11 patients. The clearance of cysts and response to nitazoxanide alone were significantly lower, compared with combination therapy (61.53% vs. 95.23%, P = 0.01, 38.46 vs. 85.71%, P = 0.004, respectively). The OR for cyst clearance and response was also significantly lower with nitazoxanide alone, in comparison with combination therapy (OR: 0.65, 95% CI: 0.34–0.92, P = 0.01, OR: 0.45, 95% CI: 0.21–0.81, respectively). Four (16%) of 24 patients with response had relapse. Conclusion. Patients with Tac and mycophenolate mofetil combination therapy had a significantly high risk of Cryptosporidium infection. Cryptosporidial infection may require prolonged nitazoxanide therapy, either alone or in combination, with or without reduction in immunosuppression.
Diarrhea is one of the most common complications observed in solid organ transplant (SOT) recipients (1). SOT recipients are at increased risk for infectious diarrhea. Cryptosporidium has been increasingly
48
D. Bhadauria1, A. Goel2, A. Kaul1, R.K. Sharma1, A. Gupta1, V. Ruhela1, A. Gupta1, H. Vardhan1, N. Prasad1 1
Nephrology, Sanjay Gandhi Post Graduate Institute of Medical Sciences, Lucknow, Uttar Pradesh, India, 2 Gastroenterology, Sanjay Gandhi Post Graduate Institute of Medical Sciences, Lucknow, Uttar Pradesh, India
Key words: Cryptosporidium; renal transplant; diarrhea; immunosuppression Correspondence to: Narayan Prasad, Nephrology, Sanjay Gandhi Post Graduate Institute of Medical Sciences, Type-4/11, SGPGIMS campus Rae Bareli Road Lucknow, Lucknow, Uttar Pradesh 226014 India Tel: 91-522-2495188 Fax: 91-522-2668017 E-mail: [email protected]
Received 22 April 2014, revised 21 August 2014, 26 October 2014, accepted for publication 26 October 2014 DOI: 10.1111/tid.12336 Transpl Infect Dis 2015: 17: 48–55
recognized as one of the common causes of infectious diarrhea in the post-transplant period, particularly in endemic areas like India (2, 3). Cryptosporidium is an intracellular protozoan parasite that infects the
Bhadauria et al: Cryptosporidium in renal transplant patients
epithelial cells of the digestive tract. The illness is typically self-limited in immunocompetent hosts, but may be severe and prolonged in patients with deficient cellular and humoral immunity, as seen in patients with human immunodeficiency virus (HIV) infection, organ transplantation, cancer treatment with immunosuppression, or immunoglobulin-A deficiency or hypogammaglobulinemia (4–7). Cryptosporidium are ubiquitous in natural water sources throughout the world and are acquired through contaminated food or water, usually leading to illness (4–6). The largest outbreak, following fecal contamination of drinking water, took place in Milwaukee, Wisconsin, USA, in the early 1990s (8). Cryptosporidial infection in immunocompromised hosts has been reported mainly in patients with acquired immunodeficiency syndrome (4, 5). The data are mainly limited to either case series (7–9) or case reports (5, 9–11), which are insufficient to determine incidence, risk factors, and the optimal antimicrobial regimen in renal transplant recipients (RTR) or other SOT patients. In the United States and Europe, cryptosporidiosis in SOT recipients is an uncommonly recognized illness; most reports are confined to single cases or small series (12–15). A few case series and epidemiological studies of stool carriage of Cryptosporidium in SOT recipients have been reported from endemic areas of the Middle East, India, and South America (2, 3, 6, 16–22). Children, individuals who reside in or travel to endemic areas, and intestinal graft recipients are particularly at increased risk of cryptosporidial infection (2, 10, 17, 23, 24). In a recently published report of 10 SOT recipients with cryptosporidial gastroenteritis, elevated levels of tacrolimus (Tac) were observed (10). The aim of this study was to determine the incidence, disease manifestation, management, and outcome of cryptosporidial infection in living-donor RTR.
Protocols of treatment of diarrheal illness and cryptosporidial infection The protocol of evaluation and treatment of diarrheal illness is summarized in Figure 1. We treat diarrheal illness according to the etiology of the diarrhea (Table 1). A thorough workup for diarrhea was performed, including clinical examination, assessment of hematological and biochemical investigations, examination for cytomegalovirus (CMV) viremia, standard stool culture, and a parasitological examination (Cryptosporidium, Isospora, Microsporidium, Entamoeba, and Giardia), plus Clostridium difficile was searched for in the stool (culture and toxin). In cases where weight loss and/or biological malabsorption symptoms occurred, an upper endoscopy with duodenal biopsies, with or without colonoscopy, was performed. Cryptosporidium in stool is diagnosed by modified acidfast bacilli (AFB) staining. For 3 consecutive days, freshly collected stool samples were transported to the laboratory within 60 min and were examined by modified AFB staining technique. This technique is used to detect oocysts of coccidian species (Cryptosporidium, Isospora, and Cyclospora). Unlike the Ziehl-Neelsen stain, this AFB stain does not require the heating of reagents for staining. Under the microscope, Cryptosporidium oocysts appear pink-red against a green background. Cryptosporidial infection is treated initially with nitazoxanide for 5–7 days. We included the patients with persistent, symptomatic, and clinically significant diarrheal illness persisting for ≥7 consecutive days that necessitated evaluation and hospitalization (Table 2). Response to therapy was defined as improvement in clinical symptoms with stool clearance for cysts.
Patients and methods In this retrospective observational study, we reviewed the data of all living-related RTR who had diarrheal illness requiring evaluation and hospitalization from January 2002 to June 2013. This study was carried out in a major transplant center in the northern part of India. The demographic profiles, immunosuppression regimen, details of diarrheal illness, hospitalization, management, and outcomes of the patients were retrieved from the electronic Hospital Information System of the institute.
Fig. 1. Protocol for diarrheal diseases. MMF, mycophenolate mofetil; CMV, cytomegalovirus; Cl., Clostridium; D2, second part of duodenum.
Transplant Infectious Disease 2015: 17: 48–55
49
Bhadauria et al: Cryptosporidium in renal transplant patients
Etiology of diarrhea and other opportunistic infections (N = 119) Etiologies of diarrhea
n (%)
Cryptosporidium
34/119 (28.5%)
Non-cryptosporidial infection CMV
48/119 (48.3%) 18
Candida albicans
8
Giardia
7
Clostridium difficile
4
Escherichia coli
3
Isospora belli
5
Microsporidium
3
Drug-induced
35 (29.4%)
PTLD
2 (1.6%)
Other opportunistic infections
17 (50%) patients
CMV (other than enteritis)
7 (20.5%) patients
Systemic fungal
8 (23.5%) patients
Cutaneous fungal
4 (11.7) patients
BKVIN
2 (5.8%) patients
CMV, cytomegalovirus; PTLD, post-transplant lymphoproliferative disorder; BKVIN, BK virus interstitial nephropathy.
Table 1 Clinical symptoms and signs of cryptosporidial infection (N = 34) Symptoms and signs
Number of patients (%)
Fever
11 (32.35)
Malaise
25 (73.25)
Profuse watery diarrhea
12 (35.29)
Crampy abdominal pain
17 (50)
Weight loss
9 (26.47)
Vomiting
18 (52.94)
Dehydration
15 (44.11)
Hypotension
8 (23.52)
Table 2
Relapse of disease was defined as re-appearance of symptoms with stool positive for cysts, after clinical response to treatment and normal stool examination, 2 weeks after stopping treatment.
Immunosuppression regimen All patients received a triple-immunosuppression regimen including steroid, calcineurin inhibitors (either cyclosporine [CSA] or Tac), and mycophenolate mofetil
50
Transplant Infectious Disease 2015: 17: 48–55
(MMF). CSA was used until December 2006, and Tac was used from January 2007 onward. The CSA dose was adjusted to maintain a trough level between 300 and 400 lg/L, and then gradually tapered to 100– 200 lg/L at 12 weeks. The dose of Tac was adjusted to maintain a trough level between 8 and 12 ng/mL for the first 6 months, and thereafter between 4 and 6 ng/ mL. All patients received methylprednisolone 500 mg intravenously during the intraoperative period, subsequently 20 mg of prednisolone daily until 8 weeks, and thereafter the dose of prednisolone was tapered every week by 2.5 mg to reach the nadir dose of 10 mg daily by 12 weeks. Interleukin-2 receptor blocker was used as induction in low immunological risk patients who opted for this treatment. Anti-thymocyte globulin (ATG) was used for induction in high-risk patients (those with re-transplantation, previous cross-match positivity, or high panel-reactive antibodies), and for treatment of acute rejection, as per indication. All rejection episodes were treated with 3–5 doses of 500 mg intravenous methylprednisolone initially, followed by oral prednisolone to reach the nadir in the next 4 weeks. Steroid-resistant rejection was treated with ATG, and antibody-mediated rejection was treated with intravenous immunoglobulin and plasmapheresis, as per our institute protocols. Cotrimoxazole was used universally as chemoprophylaxis for at least 6 months after transplant. AntiCMV prophylaxis was added to patients who received ATG.
Treatment regimen for Cryptosporidium All patients with Cryptosporidium infection were treated with either nitazoxanide alone (at a dose of 500 mg 3 times a day), or as a combination of nitazoxanide and fluoroquinolone (dosed according to glomerular filtration rate). The patient was first evaluated for response after 5–7 days of initial therapy. Clinical improvement and stool examination for clearance of Cryptosporidium cysts was performed weekly. The treatment was stopped if both stool clearance and clinical improvement were observed. In cases of partial response to initial nitazoxanide (improvement in symptoms, but stool still positive for cysts), the duration of treatment was extended up to 2 months. In case of non-response to initial nitazoxanide, fluoroquinolone was added and the duration of treatment was extended up to 2 months. Patients were observed for recurrence during follow-up. The patients who relapsed were treated with triple therapy with nitazoxanide, fluoroquinolone, cotrimoxazole
Bhadauria et al: Cryptosporidium in renal transplant patients
(double-strength tablet twice daily), and azithromycin (500 mg once a day), along with reduction in immunosuppression.
Modification of immunosuppression Immunosuppression was modified in case of nonresponse or partial response to antimicrobials, relapse, or in the presence of other opportunistic infections. MMF was switched to azathioprine (Aza), except in patients with concomitant Tac nephrotoxicity, in whom Tac was switched to CSA (Fig. 2).
Statistical analysis All data are presented in form of mean standard deviation. The categorical variables were compared using Fisher exact test and chi-square test. A P-value of
