Safety and Feasibility of Third-Party Cytotoxic T Lymphocytes for High-Risk Patients with Covid-19.

Cytotoxic T lymphocytes (CTLs) destroy virally infected cells and are critical for the elimination of viral infections such as those caused by the SARS-CoV-2 virus. Delayed and dysfunctional adaptive immune responses to SARS-CoV-2 are associated with poor outcomes. Treatment with allogeneic SARS-CoV-2-specific CTLs may enhance cellular immunity in high-risk patients and provide a safe, direct mechanism of treatment. Thirty high-risk ambulatory patients with COVID-19 were enrolled on a phase I trial to assess the safety of 3rd party, SARS-CoV-2-specific CTLs. Twelve Interventional patients, 6 of whom were immunocompromised, matched the human leukocyte antigen (HLA)-A*02:01 restriction of the CTLs and received a single infusion of one of four escalating doses of a product containing 68.5% SARS-CoV-2-specific CD8+ CTLs/total cells. Symptom improvement and resolution in these patients was compared to an Observational group of eighteen patients lacking HLA-A*02:01 who could receive standard of care. No dose-limiting toxicities were observed at any dosing level. Nasal swab PCR data showed ≥ 88% and >99% viral elimination from baseline in all patients at 4- and 14-days post-infusion. The CTLs did not interfere with the development of endogenous anti-SARS-CoV-2 humoral or cellular responses. T-cell receptor beta (TCR) analysis comparing SARS-CoV-2-specific T-cell responses derived from the CTL donor versus recipients showed persistence of donor-derived CTLs through the end of the 6-month follow-up period. Interventional patients consistently reported symptomatic improvement 2-3 days after infusion, whereas improvement was more variable in Observational patients. This study shows that SARS-CoV-2-specific CTLs are a potentially feasible cellular therapy for COVID-19 illness. (Clinicaltrials.gov #NCT04765449).

PICC-associated infection with Escherichia hermannii: A case report and review of the literature.

Since its identification as a unique species in 1982, Escherichia hermannii has been implicated as a pathogenic organism in very few cases of human disease. Our report discusses a case of bacteremia with Escherichia hermannii identified by Matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) and RapID™ ONE analysis in a patient getting TPN through a peripherally-inserted CVC (PICC). The PICC was removed. The bloodstream infection was successfully treated with empiric piperacillin-tazobactam, which was then narrowed to trimethoprim-sulfamethoxazole based on sensitivity data for a 14 day course of antimicrobial therapy. E. hermannii's association with bloodstream infection in patients with central venous catheters supports data implicating biofilm formation as a key pathogenic feature of E. hermannii. Of the 9 previous cases of E. hermannii infection reviewed in the literature, 4 cases occurred in immunocompromised hosts, 2 were associated with trauma or injection, 2 were associated with central lines, and only one case had no identifiable risk factor. E. hermannii appears to act as an opportunistic pathogen, causing disease in an immunocompromised host or through a central access catheter, injection, or trauma. E. hermannii likely causes catheter-related bloodstream infections in these hosts through biofilm formation, demonstrating the importance of catheter removal in addition to antimicrobial therapy in the treatment of these infections.

Disseminated Nocardia cyriacigeorgia causing pancreatitis in a haploidentical stem cell transplant recipient.

We report the first published case of acute pancreatitis secondary to disseminated nocardiosis in a hematopoietic stem cell transplant (HSCT) recipient on chronic immunosuppression for graft-versus-host disease (GVHD). Nocardiosis in the HSCT population is relatively rare, and has not yet been described in haploidentical HSCT recipients. Our patient is a 28-year-old male with a history of haploidentical HSCT and GVHD of the skin and lung who was admitted to the hospital with acute pancreatitis. The workup for the etiology of his pancreatitis was initially unrevealing. He subsequently developed worsening sepsis and respiratory failure despite broad spectrum antimicrobials. After multiple bronchoscopies and pancreatic fluid sampling, he was found to have disseminated nocardiosis with Nocardia cyriacigeorgia.

A two-step approach to myeloablative haploidentical transplantation: low nonrelapse mortality and high survival confirmed in patients with earlier stage disease.

Haploidentical hematopoietic stem cell transplantation (HSCT) is an attractive alternative donor option based on the rapid availability of an acceptable donor for most patients and decreased cost compared with costs of other alternative donor strategies. The safety of haploidentical HSCT has increased in recent years, making it ethically feasible to offer to patients with earlier stage disease. We developed a 2-step approach to haploidentical HSCT that separates the lymphoid and myeloid portions of the graft, allowing fixed T cell dosing to improve consistency in outcome comparisons. In the initial 2-step trial, the subset of patients without morphologic disease at HSCT had high rates of disease-free survival. To confirm these results, 28 additional patients without evidence of their disease were treated and are now 15 to 45 (median, 31) months past HSCT. To date, the 2-year cumulative incidence of nonrelapse mortality is 3.6%, with only 1 patient dying of nonrelapse causes, confirming the safety of this approach. Based on low regimen toxicity, the probabilities of disease-free and overall survival at 2 years are 74% and 77%, respectively, consistent with the findings in the initial trial and supporting the use of this approach in earlier stage patients lacking a matched related donor.

Clostridium difficile colonization and disease in patients undergoing hematopoietic stem cell transplantation.

There was an increase in the Clostridium difficile infection (CDI) rate in our bone marrow transplantation unit. To evaluate the role of unit-based transmission, C. difficile screening was performed on adult patients admitted for hematopoietic stem cell transplantation (HSCT) over a 2-year period, and C. difficile isolates were typed. C. difficile testing was performed using a 2-step C. difficile glutamate dehydrogenase antigen plus toxin A/B enzyme immunoassay (EIA) and cytotoxin assay (or molecular toxin assay). Multilocus sequence typing (MLST) was performed on toxin-positive whole stool samples. A retrospective chart review was performed on all patients with a positive toxin assay. Sixteen of 150 patients (10.7%) had toxigenic C. difficile colonization (CDC) on admission. The overall incidence of CDI within 100 days after HSCT was 24.7% (37 of 150). The median time to diagnosis of CDI was 3.5 days after HSCT. In an adjusted logistic regression model, CDC on admission was a significant risk factor for CDI (odds ratio, 68.5; 95% confidence interval, 11.4 to 416.2). MLST on 22 unit patient toxin-positive stool specimens revealed 15 distinct strain types. Further analysis identified at least 1 potential cross-transmission event; some events may have been missed because of incomplete typing from other specimens. Despite aggressive infection control interventions, there was no decline in the number of CDI cases during the study period. These data suggest that prior CDC plays a major role in CDI rates in this high-risk patient population. It remains unclear if CDI was cross-transmitted in the unit.

Risk factors for surgical site infections following spinal fusion procedures: a case-control study.

BACKGROUND: Spinal fusion procedures are associated with a significant rate of surgical site infection (SSI) (1%-12%). The goal of this study was to identify modifiable risk factors for spinal fusion SSIs at a large tertiary care center. METHODS: A retrospective, case-control (1:3 ratio) analysis of SSIs following posterior spine fusion procedures was performed over a 1-year period. Clinical and surgical data were collected through electronic database and chart review. Variables were evaluated by univariate analysis and multivariable logistic regression. RESULTS: In total, 57 deep SSIs were identified out of 1587 procedures (3.6%). Infections were diagnosed a mean of 13.5 ± 8 days postprocedure. Staphylococcus aureus was the predominant pathogen (63%); 1/3 of these isolates were methicillin resistant. Significant patient risk factors for infection by univariate analysis included ASA score >2 and male gender. Among surgical variables, infected cases had significantly higher proportions of staged procedures and thoracic level surgeries and had a greater number of vertebrae fused. Notably, infected fusion procedures had a longer duration of closed suction drains than controls (5.1 ± 2 days vs 3.4 ± 1 day, respectively; P < .001). Drain duration (unit odds ratio [OR], 1.6 per day drain present; 95% confidence interval [CI], 1.3-1.9), body mass index (OR, 1.1; 95% CI, 1.0-1.1), and male gender (OR, 2.7; 95% CI, 1.4-5.6) were significant risk factors in the multivariate analysis. CONCLUSIONS: Prolonged duration of closed suction drains is a strong independent risk factor for SSI following instrumented spinal fusion procedures. Therefore, removing drains as early as possible may lower infection rates.

A 2-step approach to myeloablative haploidentical stem cell transplantation: a phase 1/2 trial performed with optimized T-cell dosing.

Studies of haploidentical hematopoietic stem cell transplantation (HSCT) have identified threshold doses of T cells below which severe GVHD is usually absent. However, little is known regarding optimal T-cell dosing as it relates to engraftment, immune reconstitution, and relapse. To begin to address this question, we developed a 2-step myeloablative approach to haploidentical HSCT in which 27 patients conditioned with total body irradiation (TBI) were given a fixed dose of donor T cells (HSCT step 1), followed by cyclophosphamide (CY) for T-cell tolerization. A CD34-selected HSC product (HSCT step 2) was infused after CY. A dose of 2 × 10(8)/kg of T cells resulted in consistent engraftment, immune reconstitution, and acceptable rates of GVHD. Cumulative incidences of grade III-IV GVHD, nonrelapse mortality (NRM), and relapse-related mortality were 7.4%, 22.2%, and 29.6%, respectively. With a follow-up of 28-56 months, the 3-year probability of overall survival for the whole cohort is 48% and 75% in patients without disease at HSCT. In the context of CY tolerization, a high, fixed dose of haploidentical T cells was associated with encouraging outcomes, especially in good-risk patients, and can serve as the basis for further exploration and optimization of this 2-step approach. This study is registered at www.clinicaltrials.gov as NCT00429143.

Comparison of human memory CD8 T cell responses to adenoviral early and late proteins in peripheral blood and lymphoid tissue.

Treatment of invasive adenovirus (Ad) disease in hematopoietic stem cell transplant (SCT) recipients with capsid protein hexon-specific donor T cells is under investigation. We propose that cytotoxic T cells (CTLs) targeted to the late protein hexon may be inefficient in vivo because the early Ad protein E3-19K downregulates HLA class I antigens in infected cells. In this study, CD8+ T cells targeted to highly conserved HLA A2-restricted epitopes from the early regulatory protein DNA polymerase (P-977) and late protein hexon (H-892) were compared in peripheral blood (PB) and tonsils of naturally infected adults. In tonsils, epitope-specific pentamers detected a significantly higher frequency of P-977+CD8+ T cells compared to H-892+CD8+ T cells; this trend was reversed in PB. Tonsil epitope-specific CD8+ T cells expressed IFN-γ and IL-2 but not perforin or TNF-α, whereas PB T cells were positive for IFN-γ, TNF-α, and perforin. Tonsil epitope-specific T cells expressed lymphoid homing marker CCR7 and exhibited lower levels of the activation marker CD25 but higher proliferative potential than PB T cells. Finally, in parallel with the kinetics of mRNA expression, P-977-specific CTLs lysed targets as early as 8 hrs post infection. In contrast, H-892-specific CTLs did not kill unless infected fibroblasts were pretreated with IFN-γ to up regulate HLA class I antigens, and cytotoxicity was delayed until 16-24 hours. These data show that, in contrast to hexon CTLs, central memory type DNA polymerase CTLs dominate the lymphoid compartment and kill fibroblasts earlier after infection without requiring exogenous IFN-γ. Thus, use of CTLs targeted to both early and late Ad proteins may improve the efficacy of immunotherapy for life-threatening Ad disease in SCT recipients.

Impact of a standardized protocol and antibiotic-impregnated catheters on ventriculostomy infection rates in cerebrovascular patients.

BACKGROUND: Ventriculostomy infections create significant morbidity. To reduce infection rates, a standardized evidence-based catheter insertion protocol was implemented. A prospective observational study analyzed the effects of this protocol alone and with antibiotic-impregnated ventriculostomy catheters. OBJECTIVE: To compare infection rates after implementing a standardized protocol for ventriculostomy catheter insertion with and without the use of antibiotic-impregnated catheters. METHODS: Between 2003 and 2008, 1961 ventriculostomies and infections were documented. A ventriculostomy infection was defined as 2 positive CSF cultures from ventriculostomy catheters with a concurrent increase in cerebrospinal fluid white blood cell count. A baseline (preprotocol) infection rate was established (period 1). Infection rates were monitored after adoption of the standardized protocol (period 2), institution of antibiotic-impregnated catheter A (period 3), discontinuation of antibiotic-impregnated catheter A (period 4), and institution of antibiotic-impregnated catheter B (period 5). RESULTS: The baseline infection rate (period 1) was 6.7% (22/327 devices). Standardized protocol (period 2) implementation did not change the infection rate (8.2%; 23/281 devices). Introduction of catheter A (period 3) reduced infections to 1.0% (2/195 devices, P=.0005). Because of technical difficulties, this catheter was discontinued (period 4), resulting in an increase in infection rate (7.6%; 12/157 devices). Catheter B (period 5) significantly decreased infections to 0.9% (9 of 1001 devices, P=.0001). The Staphylococcus infection rate for periods 1, 2, and 4 was 6.1% (47/765) compared with 0.2% (1/577) during use of antibiotic-impregnated catheters (periods 3 and 5). CONCLUSION: The use of antibiotic-impregnated catheters resulted in a significant reduction of ventriculostomy infections and is recommended in the adult neurosurgical population.

Characterization of a single-cycle rabies virus-based vaccine vector.

Recombinant rabies virus (RV)-based vectors have demonstrated their efficacy in generating long-term, antigen-specific immune responses in murine and monkey models. However, replication-competent viral vectors pose significant safety concerns due to vector pathogenicity. RV pathogenicity is largely attributed to its glycoprotein (RV-G), which facilitates the attachment and entry of RV into host cells. We have developed a live, single-cycle RV by deletion of the G gene from an RV vaccine vector expressing HIV-1 Gag (SPBN-DeltaG-Gag). Passage of SPBN-DeltaG-Gag on cells stably expressing RV-G allowed efficient propagation of the G-deleted RV. The in vivo immunogenicity data comparing single-cycle RV to a replication-competent control (BNSP-Gag) showed lower RV-specific antibodies; however, the overall isotype profiles (IgG2a/IgG1) were similar for the two vaccine vectors. Despite this difference, mice immunized with SPBN-DeltaG-Gag and BNSP-Gag mounted similar levels of Gag-specific CD8(+) T-cell responses as measured by major histocompatibility complex class I Gag-tetramer staining, gamma interferon-enzyme-linked immunospot assay, and cytotoxic T-cell assay. Moreover, these cellular responses were maintained equally at immunization titers as low as 10(3) focus-forming units for both RV vaccine vectors. CD8(+) T-cell responses were significantly enhanced by a boost with a single-cycle RV complemented with a heterologous vesicular stomatitis virus glycoprotein. These findings demonstrate that single-cycle RV is an effective alternative to replication-competent RV vectors for future development of vaccines for HIV-1 and other infectious diseases.

Antiviral responses following L-leucyl-L-leucine methyl esther (LLME)-treated lymphocyte infusions: graft-versus-infection without graft-versus-host disease.

Although allogeneic hematopoietic progenitor cell transplant (HPCT) is curative therapy for many disorders, it is associated with significant morbidity and mortality, which can be related to graft-versus-host disease (GVHD) and the immunosuppressive measures required for its prevention and/or treatment. Whether the immunosuppression is pharmacologic or secondary to graft manipulation, the graft recipient is left at increased risk of the threatening opportunistic infection. Refractory viral diseases in the immunocompromised host have been treated by infusion of virus-specific lymphotyces and by unmanipulated donor lymphocyte infusion (DLI) therapy. L-leucyl-L-leucine methyl ester (LLME) is a compound that induces programmed cell death of natural killer (NK) cells, monocytes, granulocytes, most CD8(+) T cells, and a small fraction of CD4(+) T cells. We have undertaken a study of the use of LLME-treated DLI following T cell-depleted allogeneic HPCT, specifically to aid with immune reconstitution. In this ongoing clinical trial, we have demonstrated the rapid emergence of virus-specific responses following LLME DLI with minimal associated GVHD. This paper examines the pace of immune recovery and the rapid development of antiviral responses in 6 patients who developed viral infections during the time period immediately preceding or coincident with the administration of the LLME DLI.

Adenovirus DNA polymerase is recognized by human CD8+ T cells.

Donor lymphocytes have potential as a treatment for adenovirus (Ad) disease in haematopoietic stem cell transplant (SCT) recipients, but better understanding of Ad-specific T-cell responses is required. Most healthy adults exhibit memory T-cell responses to hexon, a capsid protein synthesized late after infection. However, since the Ad E3-19k downregulates major histocompatibility complex (MHC) class I molecules, cytotoxic T cells (CTLs) targeted to early viral proteins may be more effective in eliminating Ad-infected cells in vivo. Here we show that Ad-specific CTLs recognize the early region 2 proteins DNA polymerase (Pol) and DNA-binding protein (DBP). Firstly, memory Ad-specific CD8(+) T cells were amplified from healthy donors by in vitro stimulation with Ad-infected dendritic cells and found to exhibit MHC-restricted cytotoxicity to targets expressing Pol and DBP. Secondly, gamma interferon responses to HLA A2-binding motif peptides from Pol and DBP were directly detected in peripheral blood mononuclear cells (PBMCs) from a recently infected normal donor. Peptide-specific CTLs generated to Pol and DBP epitopes were confirmed to exhibit HLA A2-restricted killing of targets expressing Pol or DBP. Lastly, Pol-epitope-specific T cells were detected at similar or higher frequencies than hexon and DBP in three of three SCT recipients recovering from invasive Ad disease. Pol epitopes were well conserved among different Ad serotypes. Therefore, Pol is a promising target for immunotherapy of Ad disease.

Treatment of adenovirus disease in stem cell transplant recipients with cidofovir.

Invasive adenovirus (AdV) disease is fatal in >50% of allogeneic hematopoietic stem cell transplant (SCT) recipients. Treatment with cidofovir may improve outcomes based on in vitro susceptibility data and case reports. Six consecutive cases of invasive AdV disease treated with cidofovir were reviewed among 84 allogeneic adult SCT recipients (incidence, 7.1%). Cidofovir was administered intravenously at 5 mg/kg per dose (1-7 doses). All patients received intravenous immune globulin. Blood AdV DNA levels (viral loads, VLs) were monitored with a real-time quantitative polymerase chain reaction assay. Published reports of cidofovir treatment of AdV disease in SCT recipients were critically reviewed. The primary manifestations of AdV disease were hepatitis (n = 3), colitis (n = 2), and nephritis (n = 1). All patients had detectable AdV VLs, with peak values from 5 x 10(5) to 2 x 10(8) copies/mL. All patients received CD34+ selected grafts (n = 3) and/or had graft-versus-host disease (n = 4) and had CD4 counts <100 cells/mm3. Only 1 of 5 patients (20%) who received >or=2 doses of cidofovir died with active AdV disease. Four patients exhibited improvement within days of treatment with cidofovir as documented by clinical criteria and declines in AdV VLs (without a change in immunosuppression). In contrast, 1 patient treated late after onset of AdV disease died after 1 dose of cidofovir. In our review of 70 published cases treated with >or=2 doses of cidofovir, 13 (19%) died from AdV disease. In conclusion, early treatment of AdV disease with cidofovir inhibits viral replication in vivo and reduces mortality in allogeneic SCT recipients compared with historical data.

Human T-cell responses to vaccinia virus envelope proteins.

One approach for a safer smallpox vaccine is to utilize recombinant subunits rather than live vaccinia virus (VACV). The products of the VACV envelope genes A27L, L1R, B5R, and A33R induce protective antibodies in animal models. We propose that proteins that elicit T-cell responses, as well as neutralizing antibodies, will be important to include in a molecular vaccine. To evaluate VACV-specific memory T-cell responses, peripheral blood mononuclear cells (PBMC) from four VACV vaccinees were tested against whole VACV and the individual envelope proteins A27, B5, L1, and A33, using gamma interferon enzyme-linked immunospot and cytokine flow cytometry assays. PBMC were stimulated with autologous dendritic cells infected with VACV or electroporated with individual VACV protein mRNAs. T-cell lines from all donors, vaccinated from 1 month to over 20 years ago, recognized all four VACV envelope proteins. Both CD4(+) and CD8(+) T-cell responses to each protein were detected. Further analysis focused on representative proteins B5 and A27. PBMC from a recent vaccinee exhibited high frequencies of CD4(+) and CD8(+) T-cell precursors to both B5 (19.8 and 20%, respectively) and A27 (6.8 and 3.7%). In comparison, B5- and A27-specific T-cell frequencies ranged from 0.4 to 1.3% in a donor vaccinated 3 years ago. Multiple CD4(+) and CD8(+) T-cell epitopes were identified from both A27 and B5, using overlapping 15-mer peptides. These data suggest that all four VACV envelope proteins may contribute to protective immunity, not only by inducing antibody responses, but also by eliciting T-cell responses.

Human CD8+ cytotoxic T cell responses to adenovirus capsid proteins.

Adenoviruses (Ads) cause fatal disease in allogeneic stem cell transplant recipients, but there is no established therapy. Ad-specific CD8+ T cells were detected in PBMC from healthy adults at a mean frequency of 77 per 10(5) CD8+ T cells (range 8-260) by interferon-gamma ELISPOT and cytokine flow cytometry assays. CD8+ T cell lines from 7 of 7 donors exhibited MHC-class-I-restricted killing of targets expressing the capsid protein hexon. In contrast, cytotoxicity against the capsid proteins fiber and penton base was weaker or not detected. Two HLA-A2-restricted hexon epitopes and one HLA-B-restricted epitope were identified, all of which are adjacent to or overlap an HLA-DP4-restricted epitope in the highly conserved C-terminus. Thus, hexon is the immunodominant T cell target among capsid proteins and contains multiple C-terminal epitopes conserved among serotypes. These data support evaluation of donor lymphocyte infusions for treatment of Ad disease post-transplant.

Glioblastoma patients exhibit circulating tumor-specific CD8+ T cells.

PURPOSE: There is growing interest in developing cellular immune therapies for glioblastoma multiforme, but little is known about tumor-specific T-cell responses. A glioblastoma multiforme-specific T-cell assay was developed using monocyte-derived dendritic cells to present tumor antigens from the established glioblastoma multiforme cell line U118. EXPERIMENTAL DESIGN: Peripheral blood mononuclear cells (PBMC) and tumor cells were obtained from nine patients with newly diagnosed brain tumors: five glioblastoma multiforme, two oligodendroglioma, one ependymoma, and one astrocytoma. PBMCs were incubated overnight with autologous tumor cells or autologous dendritic cells loaded with a U118 cell lysate, and responses were detected by IFN-gamma ELISPOT and cytokine flow cytometry assays. RESULTS: PBMCs from all glioblastoma multiforme patients exhibited IFN-gamma responses to autologous tumor but not to HLA-mismatched U118 cells. Glioblastoma multiforme-specific IFN-gamma responses were primarily mediated by CD8+ T cells and represented approximately 2% of total CD8+ T cells. Additionally, all glioblastoma multiforme patients responded to autologous dendritic cells loaded with U118 lysate but not with low-grade astrocytoma cell lysates. PBMCs from four patients with other brain tumor types and one normal donor failed to respond to U118 lysate-loaded autologous dendritic cells. These data indicate that the IFN-gamma responses to U118 lysate-loaded autologous dendritic cells are glioblastoma multiforme specific. Moreover, PBMCs stimulated 1 to 2 weeks with U118 lysate-loaded dendritic cells exhibited MHC class I-restricted cytotoxicity against autologous tumor cells. CONCLUSIONS: Glioblastoma multiforme patients exhibit circulating tumor-specific CD8+ T cells that recognize shared tumor antigens from the glioblastoma multiforme cell line U118. These data show that glioblastoma multiformes are immunogenic and support the development of immunotherapy trials.

The impact of adenovirus infection on the immunocompromised host.

Adenovirus (Ad) infections in immunocompromised hosts have increased in frequency as the number of patients with transplants of bone marrow, liver, kidney, heart and other organs increase in number and survive longer. The numbers of such patients have also increased because of the emergence of the HIV epidemic. Ad infections with the 51 different serotypes recognised to date have few pathognomonic signs and symptoms, and thus require a variety of laboratory-based procedures to confirm infection. These viruses have the ability to target various organs with relative serotype specificity and can cause diverse manifestations including serious life-threatening diseases characteristic of the organs involved. Ads have cytolytic and immunoregulatory properties. The clinical dilemma remains the prompt recognition of Ad-related disease, the differentiation of Ad infection from Ad disease and the differentiation from other causative agents. Since the armamentarium of effective antiviral agents available to treat Ads is unproven by controlled trials and the virus is often not acquired de novo, it is difficult to prevent reactivation in immunodeficient hosts or new acquisition from donor organs. Timely discontinuation of immunosuppressive agents is necessary to prevent morbid outcomes. The clinical diseases, diagnostic tests, antiviral agents and biological aspects of the Ads as pathogens in immunocompromised patients are discussed in the context of this review. Some of the newer diagnostic tests are based on the well-studied molecular biology of Ads, which also have been attenuated by selective viral DNA deletions for use as vectors in numerous gene therapy trials in humans.