A multicentre, patient- and assessor-blinded, non-inferiority, randomised and controlled phase II trial to compare standard and torque teno virus-guided immunosuppression in kidney transplant recipients in the first year after transplantation: TTVguideIT.

BACKGROUND: Immunosuppression after kidney transplantation is mainly guided via plasma tacrolimus trough level, which cannot sufficiently predict allograft rejection and infection. The plasma load of the non-pathogenic and highly prevalent torque teno virus (TTV) is associated with the immunosuppression of its host. Non-interventional studies suggest the use of TTV load to predict allograft rejection and infection. The primary objective of the current trial is to demonstrate the safety, tolerability and preliminary efficacy of TTV-guided immunosuppression. METHODS: For this purpose, a randomised, controlled, interventional, two-arm, non-inferiority, patient- and assessor-blinded, investigator-driven phase II trial was designed. A total of 260 stable, low-immunological-risk adult recipients of a kidney graft with tacrolimus-based immunosuppression and TTV infection after month 3 post-transplantation will be recruited in 13 academic centres in six European countries. Subjects will be randomised in a 1:1 ratio (allocation concealment) to receive tacrolimus either guided by TTV load or according to the local centre standard for 9 months. The primary composite endpoint includes the occurrence of infections, biopsy-proven allograft rejection, graft loss, or death. The main secondary endpoints include estimated glomerular filtration rate, graft rejection detected by protocol biopsy at month 12 post-transplantation (including molecular microscopy), development of de novo donor-specific antibodies, health-related quality of life, and drug adherence. In parallel, a comprehensive biobank will be established including plasma, serum, urine and whole blood. The date of the first enrolment was August 2022 and the planned end is April 2025. DISCUSSION: The assessment of individual kidney transplant recipient immune function might enable clinicians to personalise immunosuppression, thereby reducing infection and rejection. Moreover, the trial might act as a proof of principle for TTV-guided immunosuppression and thus pave the way for broader clinical applications, including as guidance for immune modulators or disease-modifying agents. TRIAL REGISTRATION: EU CT-Number: 2022-500024-30-00.

Prediction of humoral and cellular immune response to COVID-19 mRNA vaccination by TTV load in kidney transplant recipients and hemodialysis patients.

BACKGROUND: Immunosuppressed individuals such as kidney transplant recipients (KTR) and hemodialysis patients (DP) show impaired immune responses to COVID-19 vaccination. Plasma Torque Teno Virus (TTV) DNA load is used as surrogate for the individual degree of immunosuppression. We now assessed the association of TTV load at time of COVID-19 vaccination with humoral and cellular immune response rates to vaccination in KTR, DP, and healthy medical personnel (MP). METHODS: A total of 100 KTR, 115 DP and 54 MP were included. All were SARS-CoV-2 seronegative at the time of vaccination with either BNT162b2 or mRNA-1273. Plasma TTV loads were assessed at the time of first vaccination. After two-dose vaccination, seroconversion (de novo detection of SARS-CoV-2 S1-IgA and/or IgG) was determined. In addition, cellular responses as assessed by interferon γ release and neutralizing antibodies were assessed in a subset of participants. ROC analyses were performed to define TTV load cut-offs predicting specific immune responses to vaccination. RESULTS: Plasma TTV loads at the time of first vaccination were negatively associated with seroconversion after two-dose vaccination in KTR (OR 0.87, 95% CI 0.76-0.99). TTV loads were significantly lower in KTR who developed humoral and cellular immune responses to vaccination compared to non-responders (p = 0.0411 and 0.0030, respectively). Of patients with TTV loads above 106 copies/ml, none developed cellular immune responses against SARS-CoV-2, and only 2 of 17 (12%) seroconverted in response to vaccination. CONCLUSION: Plasma TTV loads at the time of first vaccination in immunosuppressed individuals may be useful to predict individual vaccine-specific immune responses.

Obesity is associated with a higher Torque Teno viral load compared to leanness.

Introduction: Obesity affects a rising proportion of the population and is an important risk factor for unfavorable outcomes in viral disease including severe acute respiratory syndrome coronavirus 2- associated diseases. Torque Teno virus (TTV) is a ubiquitous and apathogenic virus which reflects the immune function of its host. The aim of this study was to investigate the association between obesity and TTV load - an indirect marker of compromised viral immune response. Methods: TTV was quantified by TTV R-GENE® PCR in a total of 89 participants of which 30 were lean (BMI <25 kg/m2) and 59 were obese (BMI >30 kg/m2). For 38 subjects, follow-up was available after bariatric surgery. Results: TTV load was higher in individuals with obesity (median 2.39, IQR: 1.69-3.33 vs. 1.88, IQR 1.08-2.43 log10 copies/mL; p = 0.027). Multivariable linear modeling revealed an independent association between TTV load and obesity. TTV was positively correlated with waist-to-hip ratio and inversely with 25OH vitamin D levels. Interleukin 6 and fasting insulin resistance were confounders of the association between TTV and obesity, while age was an effect modifier. TTV load increased by 87% (95% CI 2-243%) in the year following bariatric surgery. Discussion: A higher TTV load in obese individuals may reflect compromised immune function and thus might serve for risk stratification of unfavorable outcomes during infectious disease, including coronavirus disease 2019, in this population. Our data warrant further analysis of TTV-based risk assessment in obese individuals in the context of infectious disease-associated outcomes.

Detection of Torquetenovirus and Redondovirus DNA in Saliva Samples from SARS-CoV-2-Positive and -Negative Subjects.

OBJECTIVES: Torquetenovirus (TTV) and Redondovirus (ReDoV) are the most prevalent viruses found in the human respiratory virome in viral metagenomics studies. A large-scale epidemiological study was performed to investigate their prevalence and loads in saliva samples according to SARS-CoV-2 status. METHODS: Saliva samples from 448 individuals (73% SARS-CoV-2 negative and 27% SARS-CoV-2 positive) aged 23-88 years were tested. SARS-CoV-2 and TTV were determined in saliva by specific qualitative and quantitative real-time PCRs, respectively. A sub-cohort of 377 subjects was additionally tested for the presence and load of ReDoV in saliva, and a different sub-cohort of 120 subjects for which paired saliva and plasma samples were available was tested for TTV and ReDoV viremia at the same timepoints as saliva. RESULTS: TTV in saliva was 72% prevalent in the entire cohort, at a mean DNA load of 4.6 log copies/mL, with no difference regardless of SARS-CoV-2 status. ReDoV was found in saliva from 61% of the entire cohort and was more prevalent in the SARS-CoV-2-negative subgroup (65% vs. 52%, respectively). In saliva, the total mean load of ReDoV was very similar to the one of TTV, with a value of 4.4 log copies/mL. The mean viral loads in subjects infected with a single virus, namely, those infected with TTV or ReDoV alone, was lower than in dually infected samples, and Tukey's multiple-comparison test showed that ReDoV single-infected samples resulted in the only true outlier (p = 0.004). Differently from TTV, ReDoV was not detected in any blood samples. CONCLUSIONS: This study establishes the prevalence and mean value of TTV and ReDoV in saliva samples and demonstrates the existence of differences between these two components of the human virome.

Metagenomic Next-Generation Sequencing Reveals the Profile of Viral Infections in Kidney Transplant Recipients During the Covid-19 Pandemic

Purpose: To study the clinical application of metagenomic next-generation sequencing (mNGS) in the detection of viral infections in kidney transplant recipients (KTRs) during the COVID-19 pandemic. Method(s): Using mNGS techniques, 50 human fluid samples of KTRs were detected in Henan Province People's Hospital between May 2020 to May 2021, including 20 bronchoalveolar lavage fluid (BALF) samples, 21 urine samples and 9 blood samples. The detected nucleic acid sequences were compared and analyzed with the existing viral nucleic acid sequences in the database, and the virus infection spectrum of KTRs was drawn. Result(s): The viral nucleic acids of 15 types of viruses were detected in 96.00% (48/50) of the samples, of which 11 types of viruses were in BALF (95.00%, 19/20), and the dominant viruses were torque teno virus (TTV) (65.00%;13/20), cytomegalovirus (CMV) (45.00%;9/20) and human alphaherpesvirus 1 (25.00%;5/20). 12 viruses (95.24%, 20/21) were detected in the urine, and the dominant viruses were TTV (52.38%;11/21), JC polyomavirus (52.38%;11/21), BK polyomavirus (42.86%;9/21), CMV (33.33%;7/21) and human betaherpesvirus 6B (28.57%;6/21). 7 viruses were detected in the blood (100.00%, 9/9), and the dominant virus was TTV (100.00%;9/9). Four rare viruses were detected in BALF and urine, including WU polyomavirus, primate bocaparvovirus 1, simian virus 12, and volepox virus. Further analysis showed that TTV infection with high reads indicated a higher risk of acute rejection (P<0.05). Conclusion(s): mNGS detection reveals the rich virus spectrum of infected persons after kidney transplantation, and improves the detection rate of rare viruses. TTV may be a new biomarker for predicting rejection. (Figure Presented).

Can torque teno virus be a predictor of SARS-CoV-2 disease progression in cancer patients?

INTRODUCTION: Cancer patients with SARS-CoV-2 infection can experience a broad range of clinical manifestations and outcomes. Previous studies have demonstrated an association between torque teno virus (TTV) load and deficiencies of the immune system. The impact of SARS-CoV-2 and TTV viral loads in cancer patients is unknown. METHODS: In this retrospective study, 157 cancer patients and 191 noncancer controls were analysed for SARS-CoV-2 RNA and TTV DNA presence. RESULTS: SARS-CoV-2 RNA was detected in 66.2% of cancer patients and in 68.6% of noncancer control subjects. In SARS-CoV-2-positive patients, TTV was detectable in 79.8% of cancer patients, while in controls, TTV was detected in 71.7% of subjects. No statistically significant correlation was found between TTV and SARS-CoV-2 loads in cancer patients. However, the 100-day survival rate in cancer patients who died from COVID-19 was significantly lower in the TTV-positive group than in the TTV-negative group (P = 0.0475). In the cancer TTV-positive group, those who died also had a higher load of TTV than those who did not die (P = 0.0097). CONCLUSIONS: Our findings indicated that the presence of TTV in nasopharyngeal swabs from cancer patients was related to a higher number of deaths from COVID-19 and to a higher TTV DNA load.

Metagenomic Next-Generation Sequencing Reveals the Profile of Viral Infections in Kidney Transplant Recipients During the COVID-19 Pandemic.

Background: To study the clinical application of metagenomic next-generation sequencing (mNGS) in the detection of viral infections in kidney transplant recipients (KTRs) during the COVID-19 pandemic. Methods: Using mNGS technology, 50 human fluid samples of KTRs were detected, including 20 bronchoalveolar lavage fluid (BALF) samples, 21 urine samples and 9 blood samples. The detected nucleic acid sequences were compared and analyzed with the existing viral nucleic acid sequences in the database, and the virus infection spectrum of KTRs was drawn. Results: The viral nucleic acids of 15 types of viruses were detected in 96.00% (48/50) of the samples, of which 11 types of viruses were in BALF (95.00%, 19/20), and the dominant viruses were torque teno virus (TTV) (65.00%; 13/20), cytomegalovirus (CMV) (45.00%; 9/20) and human alphaherpesvirus 1 (25.00%; 5/20). 12 viruses (95.24%, 20/21) were detected in the urine, and the dominant viruses were TTV (52.38%; 11/21), JC polyomavirus (52.38%; 11/21), BK polyomavirus (42.86%; 9/21), CMV (33.33%; 7/21) and human betaherpesvirus 6B (28.57%; 6/21). 7 viruses were detected in the blood (100.00%, 9/9), and the dominant virus was TTV (100.00%; 9/9). Four rare viruses were detected in BALF and urine, including WU polyomavirus, primate bocaparvovirus 1, simian virus 12, and volepox virus. Further analysis showed that TTV infection with high reads indicated a higher risk of acute rejection (P < 0.05). Conclusions: mNGS detection reveals the rich virus spectrum of infected KTRs, and improves the detection rate of rare viruses. TTV may be a new biomarker for predicting rejection.

Torque teno virus DNA load as a predictive marker of antibody response to a three-dose regimen of COVID-19 mRNA-based vaccine in lung transplant recipients.

BACKGROUND: Previous studies have reported that lung transplant recipients (LTR) develop a poor response to two doses of COVID-19 vaccine, but data regarding the third dose are lacking. We investigated the antibody response after three doses of mRNA vaccine in LTR and its predictive factors. METHODS: A total of 136 LTR, including 10 LTR previously infected and 126 COVID-19-naive LTR, were followed during and after three doses of mRNA vaccine. We retrospectively measured anti-receptor-binding domain (RBD) IgG response and neutralizing antibodies. In a posthoc analysis, we used a multivariate logistic regression model to assess the association between vaccine response and patient characteristics, including viral DNA load (VL) of the ubiquitous Torque teno virus (TTV) (optimal cut-off set by ROC curve analysis), which reflects the overall immunosuppression. RESULTS: After 3 doses, 47/126 (37.3%) COVID-19-naive LTR had positive anti-RBD IgG (responders) and 14/126 (11.1%) had antibody titers above 264 Binding Antibody Units/mL. None neutralized the omicron variant versus 7 of the 10 previously infected LTR. Nonresponse was associated with TTV VL ≥6.2 log10 copies/mL before vaccination (Odds Ratio (OR) = 17.87, 95% confidence interval (CI95) = 3.02-105.72), mycophenolate treatment (OR = 4.73, CI95 = 1.46-15.34) and BNT162b2 (n = 34; vs mRNA-1273, n = 101) vaccine (OR = 6.72, CI95 = 1.75-25.92). In second dose non-responders, TTV VL ≥6.2 or <3.2 log10 copies/mL before the third dose was associated with low (0/19) and high (9/10) rates of seroconversion. CONCLUSION: COVID-19-naive LTR respond poorly to three doses of mRNA vaccine, especially those with high TTV VL. Future studies could further evaluate this biomarker as a guide for vaccine strategies.

Monitoring of Torque Teno virus DNAemia in critically ill COVID-19 patients: May it help to predict clinical outcomes?

BACKGROUND: Torque teno virus (TTV) DNA load in plasma directly associates with the net state of immunosuppression and inflammation in different clinical settings, including transplantation and chronic inflammatory diseases. OBJECTIVES: We investigated whether plasma TTV DNA load may predict the occurrence of certain infectious events and overall mortality in critically ill COVID-19 patients. PATIENTS AND METHODS: 50 patients (median age, 65.5 years) were recruited. TTV DNA load was quantitated in serial plasma specimens by real-time PCR. Serum levels of interleukin-6, C-reactive protein, ferritin, lactate dehydrogenase, Gamma-Glutamyl Transferase (GGT), alanine transaminase (ALT) and aspartate transaminase (AST) and absolute lymphocyte counts (ALC) in paired specimens were available. Nosocomial bloodstream infections and ventilator-associated pneumonia and overall mortality were the clinical outcomes. RESULTS: TTV DNA was detected in 38 patients (76%). A weak inverse correlation (Rho=-0.28; P = 0.004) was observed between TTV DNA loads and ALC. No direct correlation was found between TTV DNA load and serum levels of any of the above biomarkers. Patients with detectable TTV DNA had an increased risk of subsequently developing infectious events (HR 9.28; 95% CI, 1.29-69.5; P = 0.03). A trend (P = 0.05) towards higher TTV DNA area under a curve between days 7 and 17 after ICU admission (AUC7-17) was observed in patients who died, as compared to survivors. CONCLUSION: Our findings suggested that plasma TTV DNA load monitoring may be helpful for predicting the occurrence of severe nosocomial infections and mortality in critically ill COVID-19 patients.