Key words : Graft dysfunction, Mortality, Opportunistic infections

Introduction

Potent immunosuppressive regimen have reduced the risk of graft loss due to acute rejection; however, patients still experience increased risk of infectious complications.¹ Infections are the second leading cause of death in renal allograft recipients after cardiovascular complications.² Urinary tract infection (UTI) is considered to be the most common infection after renal transplant, with recurrent bacterial UTIs contributing to reduced graft survival.³ BK polyoma virus nephropathy is considered an important viral infection causing allograft failure, with prevalence of definite BK virus nephropathy ranging from 3% to 20%.^4-6 Although previous studies have shown prevalence of individual infections affecting the renal allograft, literature is scarce on the spectrum of renal opportunistic infections encountered in the renal allograft in patients undergoing renal transplant and their effects on graft survival. In this study, we aimed to determine the prevalence, clinicopathologic features, and outcomes of bacterial, fungal, and viral infections diagnosed on indication graft biopsies and graft nephrectomy specimens from patients with renal graft dysfunction.

Materials and Methods

We conducted a single-center retrospective study that analyzed 2019 renal transplants performed from December 2011 to December 2021. We retrospectively analyzed all indication renal graft biopsies and nephrectomy specimens from renal transplants performed during this period for presence of bacterial, viral, and fungal opportunistic infections. Analysis of allograft pathology was conducted according to Banff classification.⁷ We retrieved epidemiological records, clinical details, and laboratory workup, including microbiological investigations, from electronic records. Follow-up of the study patients was recorded in terms of graft failure (<15 mL/min/1.73 m2), death, or stable graft function.

Statistical analyses
We presented categorical variables as percentages and continuous variables as means ± SD using SPSS software version 17.0.0.

Results

Between December 2011 and December 2021, our center performed 2019 renal transplant procedures. During this period, 47 episodes of renal oppor-tunistic infections were diagnosed in either allograft biopsies or graft nephrectomy specimens from 47 renal allograft recipients. The mean age of patients was 44.59 ± 13.72 years. Among the study group, there were 39 males and 8 females.

The mean serum creatinine level at presentation was 1.85 ± 0.30 mg/dL. The mean estimated glomerular filtration rate (eGFR) at time of biopsy was 47.06 ± 10.48 mL/min/1.73 m². Donors were living related donors and included mother, father, sister, brother, and wife (emotionally related donor). Overall follow-up of study patients posttransplant ranged from 1 year to 10 years. The duration from transplant to detection of infection in renal allograft biopsy or graft nephrectomy ranged from 14 days to 39 months. Table 1 shows the spectrum of infections encountered, with corresponding duration from time of transplant to time of detection and follow-up. Table 2 shows detailed epidemiological and clinical profiles of patients, including baseline serum creatinine, serum creatinine at time of biopsy, duration between transplant and detection of opportunistic infection on biopsy, organism detected on biopsy, serological tests, and culture results wherever applicable.

Viral infections
Twenty-four patients showed viral opportunistic infections on indication renal allograft biopsy (51% of all opportunistic infections diagnosed in allograft). Of 24 patients with viral infections, 22 had BK polyoma virus nephropathy. The clinical presentation in all patients was a progressive rise in serum creatinine. In the BK virus nephropathy group, there were 20 adult and 2 pediatric renal allograft recipients. Duration from transplant to detection of BK virus nephropathy ranged from 4 months to 39 months. Prior acute rejection episodes were shown in 2 patients (1 acute cellular at month 4 posttransplant, 1 acute antibody-mediated rejection at month 1 posttransplant). One of the patients also had a history of cytomegalovirus (CMV) gastritis 1 month after diagnosis of BK polyoma virus nephropathy. Blood serology was performed in 18 of 22 patients at the time of biopsy, and polymerase chain reaction (PCR) was positive in all of these patients (Artus BKV RG PCR kit; Qiagen). Serologic details are listed in Table 2.

Histology showed BK polyoma viral inclusions in all cases with polyoma virus load, with polyoma virus load score ranging from 1 to 3, ci scores ranging from 0 to 2, and overall class ranging from class 1 in 23% to class 2 in 77% (based on Banff working classification of definitive BK polyoma virus nephropathy6). There were no cases of class 3. All cases showed positive nuclear staining on immuno-histochemistry against SV40 large T-cell antigen. These patients were treated by stopping antipro-liferative agents (mycophenolate mofetil, azathioprine) and reducing baseline immunosuppression (tacrolimus, cyclosporine). Patients had reduction of dose of antiproliferative agents by 50% and then reduction of calcineurin inhibitors (tacrolimus, cyclosporine) by 25% to 50%. Steroid dose was reduced by 25% in all patients. Target trough level was 4 to 6 pg/L for tacrolimus and 75 to 150 ng/mL for cyclosporine. Five patients received leflunomide instead of mycophenolate mofetil for variable periods (6 months to 2 years). On follow-up, 45% patients had stable graft function, 23% developed graft failure, and 32% patients died.

Cytomegalovirus nephritis was detected at month 4 and month 5 posttransplant in 2 patients. Presentation was fever and leukopenia in both patients. Both patients presented with graft dysfunction. Evidence of acute antibody-mediated rejection was shown at month 1 posttransplant in 1 of the patients. Serology for CMV (CMV quantitative real-time PCR) showed count of 11?789 IU/mL in 1 patient and 13?392 IU/mL in the other patient. Both patients showed viral inclusions in endothelial cells of peritubular capillaries and tubular epithelial cells with positive immunohistochemistry accompanied by varying amounts of mixed inflammation and white blood cell casts in tubules.

Cytomegalovirus disease and infection were managed by reduction of immunosuppression, simi-lar to management of BK virus-associated nephritis. Intravenous ganciclovir (5 g/kg every 12 h) was given for severe disease or high viral load patients (dose adjusted to eGFR), until disease severity or viral loads had reduced. Oral valganciclovir was given in mild cases at a dose of 900 mg every 12 hours (dose adjusted to eGFR) for 3 months. Cytomegalovirus replication was monitored with serum PCR methods with weekly testing until a negative viral load was shown. After a negative report was shown, antimetabolites were reintroduced in patients. One patient developed graft failure 4 months after diagnosis, whereas the other patient had stable graft function on follow up.

Bacterial graft pyelonephritis
There were 12 cases of bacterial graft pyelonephritis (25% of all renal allograft infections). Time from transplant to diagnosis ranged from as early as 14 days to 30 months following transplant. Clinical presentation was fever in 12 of 12 patients (100%), graft tenderness in 10 of 12 patients (83%), pyuria in 12 of 12 patients (100%), leukocytosis in 8 of 12 patients (66%), and graft dysfunction in 11 of 12 patients (92%) of patients. Escherichia coli was isolated in urine culture of 10 patients, and Klebsiella species was identified in 2 patients (Table 2). These patients underwent computed tomography scan of their allograft to confirm the diagnosis and to look for obstruction or abscess. Renal abscess was seen in 5 cases, which were managed with intervention and pus aspiration. All patients with bacterial graft pyelonephritis were managed by antibiotics (oral or intravenous) as per the sensitivity pattern. Two patients experienced acute cellular rejection (at 1 month and 10 months posttransplant), and 1 patient had prior acute antibody-mediated rejection at 1 month posttransplant. All patients had neutrophilic casts in renal tubules with mild to moderate interstitial inflammation and edema. Two patients had concomitant acute cellular rejection. C4d staining was negative in peritubular capillaries in these cases. On follow-up, 6 patients developed graft failure, 4 patients had stable graft, and 2 patients died from cardiovascular complications.

Tubercular and granulomatous interstitial nephritis
There were 2 cases of tubercular granulomatous inflammation (18 and 20 months posttransplant) and 1 case of granulomatous interstitial nephritis following E coli urinary tract infection, as confirmed on urine culture at 5 months posttransplant (Table 2). Patients presented with weight loss and persistent pyuria. Both cases of tubercular disease in graft biopsy had prior history of pulmonary tuberculosis and showed granuloma along with small foci of caseous necrosis and Langhan giant cells. Both patients with tubercular disease were given antitubercular therapy (isoniazid, pyrazinamide, ethambutol, fluoroquinolone) without rifampicin for at least 1 year. Intensive therapy with 4 drugs was prescribed for 2 months, and then 2 drugs (isoniazid, fluoroquinolone) were continued until 12 months. The patients with E coli infection showed ill-formed epithelioid cell granulomas without caseous necrosis and accompanying interstitial edema and mixed inflammation along with neutrophilic casts in tubules. The patient received antibiotics as per sensitivity pattern. One patient with tubercular granuloma had stable graft function, whereas the other patient and the patient with granulomatous interstitial nephritis (secondary to E coli UTI) developed graft failure on follow-up.

Fungal infections
Eight patients had fungal etiology, accounting for 17% of cases of allograft infection. Clinical presen-tation was variable in all patients, with fever in 6 of 8 (75%), leukocytosis in 4 of 8 (50%), pyuria in 8 of 8 (100%), and graft tenderness in 7 of 8 patients (87%).

Three patients had mucormycosis (2 detected on graft nephrectomy and 1 on open biopsy), with all 3 cases detected during the first 45 days posttransplant. One patient had prior history of COVID-19 infection and had also experienced acute antibody-mediated rejection 1 week after renal transplant. This patient also had simultaneous detection of rhino-orbital mucormycosis. Urine culture test for fungus and serum test for galactomannan and beta glucan were also done. Urine culture showed 2 cases of Mucor species and 1 of Rhizopus (Table 2). Radiological imaging in the form of ultrasonography was performed for these cases. Renal histology showed large areas of ischemic necrosis with marked interstitial inflammation, neutrophilic microabscess, and foreign body type of giant cells. Angioinvasion by ribbon-like broad nonseptate hyphae was seen in 2 cases. All 3 patients with mucormycosis received intravenous liposomal amphotericin B at 0.1 to 0.5 mg/kg/day and/or posaconazole at 300 mg/day (2 patients) or isavuconazole at 300 mg/day (1 patient). Treatment was continued until remission of fungal infection. On follow up, 2 patients died from complications of mucormycosis; the other patient became dialysis dependent.

There were 2 cases of invasive aspergillus infection detected at 3 months and 30 months posttransplant. Cases were diagnosed on renal allograft biopsy and graft nephrectomy, respectively. One patient also had history of pulmonary aspergillosis. Urine culture showed Aspergillus flavus species in both cases. Both cases showed presence of slender septate hyphae with acute angle branching within the interstitium accompanied by moderate mixed inflammation. Acute tubular injury was seen in only 1 case. Both patients with invasive aspergillus infection had experienced prior acute rejection episodes (1 acute cellular rejection, 1 antibody-mediated rejection) in the first 3 months post-transplant. Both patients received a combination of liposomal amphotericin B (0.5 mg/kg/day for 3 weeks) and voriconazole (400 mg/day). Both patients died at follow-up as a complication of aspergillus infection.

There were 2 cases of renal candidiasis detected at month 2 and month 3 posttransplant on graft biopsies. Patients showed presence of budding yeast forms and pseudohyphae accompanied by neutrophilic interstitial infiltrate and edema. Fungal culture confirmed the species as Candida albicans in both cases. Patients received fluconazole and liposomal amphotericin B. One patient developed graft failure, and the other patient had stable graft function on follow-up.

There was only 1 case of Cryptococcus infection detected at 3 months posttransplant. Analysis showed neutrophilic casts in tubules along with presence of budding yeast forms with prominent cell walls on periodic acid-Schiff and silver methenamine stains accompanied by neutrophilic interstitial infiltrate and edema. Fungal culture confirmed the species as Cryptococcus neoformans. The patient was treated with a combination of liposomal amphotericin B (1.0 mg/kg) and isavuconazole (300 mg/day). This patient died on follow-up from complications of infection. There was no history of any systemic involvement in this patient.

Discussion

Infection can involve as high as 70% of renal allograft recipients during the first 3 years after transplant and can be the second leading cause of mortality after cardiovascular causes.1,2,8 BK virus nephropathy is considered as the most common infection affecting the renal allograft, with reported incidence of about 6%. The incidence is higher in ABO incompatible grafts (18%) and in highly sensitized recipients (20%) following a desensitization protocol with considerable variations among different centers.5,6,9 BK virus nephropathy was the most common opportunistic infection encountered in renal allografts in our center. Patients can present with slowly deteriorating graft function, ureteric stenosis, or hemorrhagic cystitis. Histology shows tubulo-interstitial nephritis with viral inclusions in tubular epithelial cells, both within the cortex and medulla and occasionally within parietal epithelial cells of the Bowman capsule.

Treatment includes optimizing immunosup-pressive medication in the form of reducing target concentration of calcineurin inhibitors, replacement of tacrolimus by cyclosporine, replacement of calcineurin inhibitors by mechanistic target of rapamycin inhibitors like sirolimus, and reduction in the dose of mycophenolate. The risk of graft loss in affected patients can be as high as 50%.^10,11 The prevalence of other viral infections like CMV and adenovirus is low. Although latent CMV infections are common in the renal allograft, tissue-invasive disease is not common, with prevalence of CMV nephritis in renal allograft of <1%; a recent study reported a prevalence of as low as 0.2%.¹² Low prevalence is due to regular use of prophylaxis given to renal transplant recipients.^12,13 The gastrointestinal tract, liver, lung, retina, and gall bladder are the more common sites of invasive of CMV infection.

Most patients with CMV nephritis experience graft failure on follow-up, and a substantial number of patients can have increased episodes of acute rejection because of the immune modulator effects of CMV.¹² Patients with tissue invasive CMV disease have at least 4 weeks of therapy with ganciclovir or valganciclovir until symptoms or CMV viremia are resolved, as well as reduction in immunosuppression and immunomodulation.^12-14

Patients with bacterial graft pyelonephritis receive antibiotics for 14 to 21 days for the first infection episode and 28 days for recurrent episodes. Treatment is aimed to prevent decline in graft function, to reduce the episodes of recurrent UTIs, and to prevent potential progression to sepsis.^15,16 Patients with early acute graft pyelonephritis have increased patient mortality compared with patients who do not experience episodes. Subsequent recurrent graft pyelonephritis episodes can also result in poorer patient survival. In addition, late presentation of graft pyelonephritis at 6 months posttransplant resulted in 2 times increased risk of patient mortality.¹⁷

The 3 most common fungal infections involving the renal allograft include Mucor, Aspergillus, and Candida. Renal mucormycosis can be seen as a part of disseminated disease or as isolated renal involvement, with the former being more common.¹⁸ Patients with renal mucormycosis are treated with nephrectomy along with antifungal agents, including amphotericin B and posaconazole.¹⁹ A high rate of graft failure (95%) has been shown in patients with bilateral involvement,¹⁸ and invasive renal mucormycosis also carries a high mortality rate (44%).¹⁹ Invasive aspergillosis is an uncommon opportunistic infection of the renal allograft and primarily affects the lung, followed by the gastro-intestinal tract, brain, and liver. Primary pulmonary infection is common, whereas disseminated disease is rare.²⁰ In an autopsy study of posttransplant patients with invasive aspergillosis, 94% of patients had pulmonary aspergillosis followed by invol-vement in the gastrointestinal tract (21%) and brain (13%). Renal involvement and liver involvement was seen in 12% each.²¹

In our study, invasive aspergillus infection with renal involvement was seen as multiple focal abscesses, aspergillus cast in renal pelvis, and ascending urothelial infection. On histology, apart from abscesses, infarction and papillary necrosis have been described.¹⁸ Patients with invasive aspergillosis receive appropriate antifungal drugs together with surgical therapy and reduction in immunosuppressive medication.²⁰ Monotherapy with voriconazole or, for severe cases, a combination therapy of voriconazole and echinocandin has been used.²² In patients who cannot tolerate voriconazole or in those who develop side effects, posaconazole or isavuconazole can be used.^23,24 Graft nephrectomy may be required in some cases.²⁵ Renal candidiasis can present as microabscess, renal papillary necrosis, and emphysematous pyelonephritis with clinical manifestation ranging from fever, dysuria, flank pain, pyuria, or hematuria.18 Patients are initially given echinocandin and then can be switched to oral antifungal therapy with fluconazole, according to the organism isolated and the response to treatment.²⁶ No definite role of surgical therapy has been documented, although a combination of medical therapy and graft nephrectomy in the management of renal candidiasis has been reported.²⁶

Limitations
Our study describing the spectrum and outcome of opportunistic renal allograft infections diagnosed on renal allograft biopsies and graft nephrectomy specimen had limitations. The study did not include cases diagnosed on clinical grounds or based on just microbiological examination without tissue diagnosis.

Conclusions

Our study provided insight into the spectrum of opportunistic infections involving the renal allograft in renal transplant recipients and their outcomes. Although BK virus nephropathy was the most common infection of the renal allograft, fungal infections were associated with the highest mortality and bacterial graft pyelonephritis infection resulted in the highest rate of graft failure. Renal allograft opportunistic infections are important but under-recognized causes of graft dysfunction. Early and correct identification along with prompt treatment for these infections may help in prolonging graft and patient survival.

References:

1. Dharnidharka VR, Agodoa LY, Abbott KC. Risk factors for hospitalization for bacterial or viral infection in renal transplant recipients--an analysis of USRDS data. Am J Transplant. 2007;7(3):653-661. doi:10.1111/j.1600-6143.2006.01674.x
   CrossRef - PubMed
   
2. Snyder JJ, Israni AK, Peng Y, Zhang L, Simon TA, Kasiske BL. Rates of first infection following kidney transplant in the United States. Kidney Int. 2009;75(3):317-326. doi:10.1038/ki.2008.580
   CrossRef - PubMed
   
3. Santithanmakorn C, Vanichanan J, Townamchai N, et al. Bacterial urinary tract infection and early asymptomatic bacteriuria in kidney transplantation still negatively affect kidney transplant outcomes in the era of modern immunosuppression and cotrimoxazole prophylaxis. Biomedicines. 2022;10(11). doi:10.3390/biomedicines10112984
   CrossRef - PubMed
   
4. Myint TM, Chong CHY, Wyld M, Nankivell B, Kable K, Wong G. Polyoma BK virus in kidney transplant recipients: screening, monitoring, and management. Transplantation. 2022;106(1):e76-e89. doi:10.1097/TP.0000000000003801
   CrossRef - PubMed
   
5. Sharma R, Zachariah M. BK virus nephropathy: prevalence, impact and management strategies. Int J Nephrol Renovasc Dis. 2020;13:187-192. doi:10.2147/IJNRD.S236556
   CrossRef - PubMed
   
6. Nickeleit V, Singh HK, Randhawa P, et al. The Banff Working Group classification of definitive polyomavirus nephropathy: morphologic definitions and clinical correlations. J Am Soc Nephrol. 2018;29(2):680-693. doi:10.1681/ASN.2017050477
   CrossRef - PubMed
   
7. Loupy A, Haas M, Roufosse C, et al. The Banff 2019 Kidney Meeting Report (I): updates on and clarification of criteria for T cell- and antibody-mediated rejection. Am J Transplant. 2020;20(9):2318-2331. Doi:10.1111/ajt.15898
   CrossRef - PubMed
   
8. Nambiar P, Silibovsky R, Belden KA. Infection in kidney transplantation. Contemp Kidney Transplant. 2018:307-327 doi:10.1007/978-3-319-19617-6_22
   CrossRef - PubMed
   
9. Nickeleit V, Singh HK, Dadhania D, et al. The 2018 Banff Working Group classification of definitive polyomavirus nephropathy: a multicenter validation study in the modern era. Am J Transplant. 2021;21(2):669-680. doi:10.1111/ajt.16189
   CrossRef - PubMed
   
10. Gupta P, Gupta A, Bhalla AK, et al. BK Virus nephropathy in living donor renal allograft recipients: An observational study from a large transplant center in India. Saudi J Kidney Dis Transpl. 2018;29(6):1366-1370. doi:10.4103/1319-2442.248313
    CrossRef - PubMed
    
11. Patel R. Infections in recipients of kidney transplants. Infect Dis Clin North Am. 2001;15(3):901-952, xi. doi:10.1016/s0891-5520(05)70178-1
    CrossRef - PubMed
    
12. Swanson KJ, Djamali A, Jorgenson MR, et al. Cytomegalovirus nephritis in kidney transplant recipients: epidemiology and outcomes of an uncommon diagnosis. Transpl Infect Dis. 2021;23(5):e13702. doi:10.1111/tid.13702
    CrossRef - PubMed
    
13. Morgantetti GF, Balancin ML, de Medeiros GA, Dantas M, Silva GEB. Cytomegalovirus infection in kidney allografts: a review of literature. Transl Androl Urol. 2019;8(Suppl 2):S192-S197. doi:10.21037/tau.2018.10.14
    CrossRef - PubMed
    
14. Kotton CN, Kumar D, Caliendo AM, et al. The Third International Consensus Guidelines on the management of cytomegalovirus in solid-organ transplantation. Transplantation. 2018;102(6):900-931. doi:10.1097/TP.0000000000002191
    CrossRef - PubMed
    
15. Kumar R, Pereira M, Taimur S, True K, Detwiler R, van Duin D. Duration of antibiotic treatment for acute graft pyelonephritis: what’s the standard of care? Transpl Infect Dis. 2023;25(1):e13996. doi:10.1111/tid.13996
    CrossRef - PubMed
    
16. Goldman JD, Julian K. Urinary tract infections in solid organ transplant recipients: Guidelines from the American Society of Transplantation Infectious Diseases Community of Practice. Clin Transplant. 2019;33(9):e13507. doi:10.1111/ctr.13507
    CrossRef - PubMed
    
17. Pacaud M, Colas L, Kerleau C, et al. Impact of late and recurrent acute graft pyelonephritis on long-term kidney graft outcomes. Front Immunol. 2022;13:824425. doi:10.3389/fimmu.2022.824425
    CrossRef - PubMed
    
18. Gupta K. Fungal infections and the kidney. Indian J Nephrol. 2001;11(4):147-154.
    CrossRef - PubMed
    
19. Didehdar M, Chegini Z, Khoshbayan A, Moradabadi A, Shariati A. Clinical presentations, diagnosis, management, and outcomes of renal mucormycosis: an overview of case reports. Front Med (Lausanne). 2022;9:983612. doi:10.3389/fmed.2022.983612
    CrossRef - PubMed
    
20. Sigera LS, Denning DW. Invasive aspergillosis after renal transplantation. J Fungi (Basel). 2023;9(2).255 doi:10.3390/jof9020255
    CrossRef - PubMed
    
21. Marr KA, Schlamm HT, Herbrecht R, et al. Combination antifungal therapy for invasive aspergillosis: a randomized trial. Ann Intern Med. 2015;162(2):81-89. doi:10.7326/M13-2508
    CrossRef - PubMed
    
22. Maertens JA, Rahav G, Lee DG, et al. Posaconazole versus voriconazole for primary treatment of invasive aspergillosis: a phase 3, randomised, controlled, non-inferiority trial. Lancet. 2021;397(10273):499-509. doi:10.1016/S0140-6736(21)00219-1
    CrossRef - PubMed
    
23. Maertens JA, Raad, II, Marr KA, et al. Isavuconazole versus voriconazole for primary treatment of invasive mould disease caused by Aspergillus and other filamentous fungi (SECURE): a phase 3, randomised-controlled, non-inferiority trial. Lancet. 2016;387(10020):760-769. doi:10.1016/S0140-6736(15)01159-9
    CrossRef - PubMed
    
24. Trnacevic S, Mujkanovic A, Nislic E, et al. Invasive aspergillosis after kidney transplant-treatment approach. Med Arch. 2018;72(6):456-458. doi:10.5455/medarh.2018.72.456-458
    CrossRef - PubMed
    
25. Tan J, Wild A, Reid G, Shantier M. Management of early graft candidiasis in a kidney transplant recipient. BMJ Case Rep. 2022;15(11): e250890.doi:10.1136/bcr-2022-250890
    CrossRef - PubMed