The Essentials of Cell-Mediated Immunity Assays for the Management of CMV Infection after Solid Organ Transplantation

Alexis Guenette ^1,* , Valentina Stosor ^1,2

1. Division of Infectious Diseases, Department of Medicine, Northwestern University Feinberg School of Medicine, Chicago, Illinois, USA

2. Division of Organ Transplantation, Department of Surgery, Northwestern University Feinberg School of Medicine, Chicago, Illinois, USA

* Correspondence: Alexis Guenette 

Academic Editor: Priyamvada Singh

Special Issue: Cytomegalovirus Management in Solid Organ Transplant Recipients

Received: March 12, 2025 | Accepted: September 01, 2025 | Published: September 04, 2025

OBM Transplantation 2025, Volume 9, Issue 3, doi:10.21926/obm.transplant.2503256

Recommended citation: Guenette A, Stosor V. The Essentials of Cell-Mediated Immunity Assays for the Management of CMV Infection after Solid Organ Transplantation. OBM Transplantation 2025; 9(3): 256; doi:10.21926/obm.transplant.2503256.

© 2025 by the authors. This is an open access article distributed under the conditions of the Creative Commons by Attribution License, which permits unrestricted use, distribution, and reproduction in any medium or format, provided the original work is correctly cited.

1. Introduction

Cytomegalovirus (CMV) is one of the most common opportunistic pathogens encountered in addition to being an important cause of mortality and morbidity after solid organ transplantation (SOT). Clinical presentation can present as primary infection or reactivation of latent virus with disease manifestation ranging from fever with malaise, leukopenia, thrombocytopenia and mild hepatic transaminitis to dissemination to various organs, including the allograft [1]. Pretransplant CMV-IgG serologic testing of the donor and recipient remains the standard method for risk stratification regarding post-transplant CMV disease; however there has been some concern that serologic results do not correlate with immunity. Pre-transplant serologic assessment in conjunction with the immunosuppressive regimen and organ type transplanted guide the post-transplant prevention of CMV disease.

The development of CMV and its severity depends on the organ type transplanted and donor/recipient CMV serologies. This along with induction and maintenance immunosuppression are factors that can affect CMV CMI. Regarding induction immunosuppression, T-cell depleting agents such as alemtuzumab and antithymocyte globulin (ATG) cause the depletion of T lymphocytes which will significantly impair CMV CMI compared to basiliximab, an IL-2 receptor antagonist, which inhibits T-cell activation and proliferation and is associated with less risk of post-transplant CMV [2,3]. Regarding maintenance immunosuppression, calcineurin inhibitors (CNIs) directly suppress CMV-specific T-cell activity and in combination with other immunosuppressants like corticosteroids can lead to a weaker CMV CMI [4,5]. Triple drug maintenance immunosuppression including combinations of CNIs, corticosteroids and another agent such as mycophenolate mofetil or an mTOR inhibitor can significantly impact CMV-specific T-cell functionality [5]. Increasing usage of belatacept, a fusion protein combining cytotoxic T-lymphocyte antigen-4 and Fc region of human IgG1, as part of a CNI-sparing regimen has demonstrated a progressive loss of CMV CMI theoretically driven by T-cell exhaustion [6]. Interestingly mTOR inhibitors may improve T-cell function and memory along with inhibition of select pathways involved in CMV replication [7] along with a decrease in CNIs is an alternative option to help limit CMV post-transplant depending on the serologic status, organ transplanted, and risk of rejection [8].

T cells are vital in controlling CMV replication but fail to eliminate the virus. Latency is established in myeloid progenitor cells after an initial lytic gene expression is silenced through epigenetic repression [9]. Patients that undergo SOT are maintained on life-long pharmacologic immunosuppression to prevent allograft rejection; these therapies affect T lymphocytes by obstructing the function of these cells and consequently increasing the risk of opportunistic, including CMV reactivation and de novo infection [1]. Currently, there are two approaches for management of post-transplant CMV infection, antiviral prophylaxis and preemptive therapy (PET). Despite these two approaches, each with their respective advantages and disadvantages, there are still overall limitations which has prompted interest in immune monitoring to measure nonspecific and CMV specific T-cell quantity and/or function to further inform risk stratification and management of CMV in transplant recipients.

2. CMV Cell-Mediated Immunity Assays

Primarily, the objective of CMV cell-mediated immunity (CMI) laboratory assays is the measurement of cytokines, including interferon gamma (IFN-γ), produced by CD4⁺ and/or CD8⁺ T lymphocytes in response to the stimulation of peripheral blood mononuclear cells (PBMCs) or whole blood with CMV antigens, lysate, or overlapping peptides. “High” CMV CMI, indicating adequate CMV-specific CD4⁺ and/or CD8⁺ T cell immunity, predicts protection against clinically significant CMV [3]. Current methods to measure CMV-specific T-cell function include flow cytometry, enzyme-linked immunosorbent spot assay, enzyme-linked immunosorbent assay, cytokine profiling, tetramer staining, T cell receptor spectra typing, and expansion protocols [10]. Table 1 provides currently available, research or commercially, CMV CMI assays.

Table 1 Available CMV T cell-mediated immunity assays. Reprinted [or adapted] from “Utility of Cytomegalovirus Cell-Mediated Immunity Assays in Solid Organ Transplantation” by V. Hall, 2022, Journal of Clinical Microbiology, 60 [10]. Reprinted [or adapted] with permission.

3. Potential Clinical Applications of CMV CMI

3.1 CMV CMI in Pretransplant Setting

CMV-IgG serology is the standard pre-transplant assessment to determine recipient and donor (latent) infection; however, emerging data has suggested that CMV-IgG does not always correlate with an effective CMV-CMI. Abate et colleagues demonstrated that there was 18% disagreement between serology and CMV-CMI amongst 100 CMV-IgG positive individuals. In a study of 44 CMV-IgG positive lung and kidney transplant candidates, 68% has detectable CMV-CMI [11]. Lastly, a study with 583 kidney transplant recipients demonstrated 8% of CMV-IgG had undetectable or low-level CMV-CMI prior to transplant [3].

Therefore, there are concerns that the absence of CMV CMI in CMV-seropositive transplant candidates may correlate with higher risk of post-transplant CMV infection based on several small studies. In a series of 44 CMV-seropositive lung and kidney recipients, Cantisan and colleagues demonstrated that the rate of post-transplant CMV reactivation was significantly higher among patients with non-reactive versus reactive pre-transplant CMV-CMI, 7 of 14 [50%] vs. 4 of 30 [13.3%] [11]. Additionally, Bhugra et all, studied thirty living donor liver transplant recipients of which thirteen patients with positive pre-transplant CMV-CMI demonstrated lower risk of CMV disease (15.4% vs. 58.8%), prompt elimination of virus (7 days vs. 21 days), and truncated duration of antiviral treatment (13 days vs. 28 days) when compared to CMV-seropositive liver recipients with negative pre-transplant CMV-CMI [12]. Finally, in a prospective interventional trial of 160 CMV-seropositive kidney recipients, those with undetectable or low pre-transplant CMV-CMI had significantly higher rates of post-transplant CMV infection [2].

Some groups advocate for CMV CMI measurement in transplant candidates, along with pretransplant serology, to further inform post-transplant risk stratification (high vs. low risk), however it is uncertain whether this combination is a better predictor of posttransplant viral replication than serology alone, and as such, the role of CMV CMI for this indication after transplantation remains under clinical investigation [10,13].

3.2 CMV CMI in Posttransplant Setting: Prophylaxis

3.2.1 D+R-

Assessing CMV CMI prior to or after completion of prophylaxis is a potential method of mitigating delayed-onset CMV disease; however, in this group there is a concern regarding the lack of CMV CMI [14] therefore CMV CMI has a limited role in this CMV D/R serostatus group. When utilizing prophylaxis in this risk group, the antiviral is suppressing reactivation of donor-origin virus, thus preventing antigenic exposure and immune priming [1]. Extended duration of antiviral prophylaxis is an alternative prevention strategy, although CMV viremia will occur in up to 25% of transplant recipients after discontinuation of therapy [14]. An alternative approach is a CMV surveillance program that utilizes routine CMV viral load monitoring for up to 12 months after discontinuation of antiviral prophylaxis, with the thought that development of CMV viremia will elicit CMV CMI. In this setting concomitant usage of CMV CMI testing may inform the need for treatment and guide the duration of viral surveillance [14].

With the advent of PET, it has been hypothesized that concurrent surveillance of CMV CMI can assess the need for antiviral treatment of CMV viremia, given that viral load thresholds for initiation of treatment have been poorly defined. Abate and colleagues demonstrated that allowing DNAemia to occur at low level may contribute to the development of CMV CMI. In a prospective study of 37 solid organ transplant recipients with low-level DNAemia, (<1,000 copies/mL), QFN-CMV was measured at the first detection of CMV with an institutional threshold of 15,000 copies/mL for starting treatment. It was determined that a positive CMV CMI was associated with spontaneous clearance of DNAemia suggesting that low-level DNAemia does not require antiviral intervention [15].

3.2.2 R+

Assessing CMV CMI prior to or after completion of antiviral prophylaxis offers the potential for mitigating delayed-onset CMV along shortened duration of the prophylaxis irrespective of induction immunosuppression. Recently, Solera and colleagues, demonstrated earlier discontinuation of prophylaxis using a positive QTF- CMV, ≥0.2 IU/mL, with only 12% of patients developing CMV viremia ≥1000 IU/mL in SOT patients, including kidney, kidney-pancreas, liver, and heart recipients and no evidence of CMV disease, which is consistent with more recent studies [16,17,18]. Earlier discontinuation leads to less neutropenia which is a common reason for alternative antiviral prophylaxis when using valganciclovir along with potential cost-savings.

3.3 CMV CMI in Posttransplant Setting: Treatment

Assessing CMV CMI upon or prior to completion of clinically significant CMV viremia may inform need for secondary prophylaxis in conjunction with surveillance CMV PCRs. Dioverti et al., demonstrated that none of the patients who had reactive QFT-CMV at the end of treatment had a relapse or recurrence of CMV infection [14]. They also hypothesized that patients with negative CMV CMI at the end of treatment are over-immunosuppressed and therefore are high risk of CMV relapse despite secondary prophylaxis unless immunosuppression is minimized.

3.4 CMV CMI in Posttransplant Setting: Rejection

As with prophylaxis, the same strategy can be applied during the treatment of rejection, especially with T-cell depleting agents. CMV CMI is obtained prior to or upon completion of the treatment of rejection, thereby assessing if there is a continued need for CMV prophylaxis [10].

4. Limitations

Primarily, CMV CMI assays are for patients with CMV immunity therefore this is not applicable to the D-/R- group although if they develop a primary infection, CMV CMI can be applied to determine secondary prophylaxis. Overall, a key shortcoming is the lack of widespread availability, clinical validation, and standardization of CMV-CMI. Available assays measure only CD4⁺ and/or CD8⁺ T cell function via interferon gamma release after ex vivo CMV antigenic stimulation however current assays do not consider other characteristics of pathogen-specific immune responses thereby implying a need for a more comprehensive measure of CMV-CMI. Lastly, there is a lack of prospective, controlled, and interventional studies to support the role of CMV-CMI [1].

CMV CMI is an additional cost which varies based on the platform, however there is room to investigate the overall cost-effectiveness in whichever cohort it is applied. When implemented in a cohort of CMV D+R- or CMV R+ with ATG in kidney and liver transplant recipients, the duration of prophylaxis was shorter in the CMV CMI-monitored group thereby resulting in a significant reduction of antiviral prophylaxis [8,17]. This study along with a trial from Paez-Vega which looked at CMV R+ kidney transplant recipients that received ATG induction suggests that CMV CMI resulted in a reduction in the duration of antiviral prophylaxis without increasing post-transplant CMV disease but does not have similar outcomes that include CMV DNAemia [17,18]. A Brazilian study did demonstrate cost-effectiveness in preemptive CMV therapy of kidney transplant recipients when CMV CMI monitoring was applied [8,19].

5. Conclusion

Currently there are several assays available with several potential clinical scenarios, pretransplant and posttransplant, that CMV CMI can be utilized however which assay and in what scenario still needs to be determined.

Acknowledgments

A.G. receives grant support from Viracor Eurofins. V.S. has no relevant disclosures.

Author Contributions

A.G and V.S drafted the manuscript and revised the paper.

Competing Interests

The authors have declared that no competing interests exist.