Maintenance of cytomegalovirus (CMV) latency and host immune responses of long term renal allograft survivors. II. Secondary CMV infections associated with impaired in vitro proliferative responses to mitogens, allogeneic lymphocytes and CMV infected cells.

The relationship between secondary cytomegalovirus (CMV) infections and host general cellular immunocompetence was investigated in 16 renal allograft recipients with minimal immunosuppressive treatment and excellent renal function. Results were compared with 19 CMV seropositive healthy controls. Significantly impaired immune responses were detected in the subgroup of nine recipients who experience at least 2 years before a secondary CMV infection. Their in vitro lymphocyte reactivity (LR) tests to phytohaemagglutinin (PHA, P = 0.01), pokeweed mitogen (PWM, P less than 0.05), microbial antigens (P less than 0.001) and to pooled allogeneic stimulator lymphocytes in the MLC test (P = 0.02) were lower than the controls. The MLC responses, however, increased with graft survival time (r = 0.8810, P = 0.01). This was positively correlated with the virus specific cellular immunity measurable by the LR responses to CMV infected target cells (r = 0.8333, P = 0.02). In contrast, long term renal allograft survivors who maintained their CMV infection in latency after transplantation (n = 7) showed normal responses to PWM, pooled lymphocytes and CMV infected target cells, whereas the responses to PHA and to bacterial antigens were less severely impaired (P less than 0.05 and P less than 0.001, respectively). This study of long term renal allograft survivors shows that a secondary CMV infection has a long lasting negative effect on immunity especially against alloantigens and CMV infected targets. However, in the data presented here it would be as acceptable to suggest that the patients are consistently relapsing with CMV because they initially had poor immune response and not vice versa.     

Attenuated poxvirus expressing three immunodominant CMV antigens as a vaccine strategy for CMV infection.

BACKGROUND: Human cytomegalovirus (CMV) infection is an important risk factor in the post-transplant (Tx) recovery phase for both hematopoietic stem cell Tx (HSCT) and solid organ Tx (SOT) recipients. CMV infection may be prevented or controlled by simultaneously inducing both CMV-specific neutralizing antibody (nAb) and cellular immunity. Soluble (s) UL55 (surface glycoprotein), UL83 (tegument protein) and UL123/e4 (nuclear protein) are immunodominant in eliciting both CMV nAb and cellular immunity. An attenuated poxvirus, modified vaccinia Ankara (MVA) was selected to develop this vaccine strategy in Tx recipients, because of its clinical safety record, large foreign gene capacity, and capability to activate strong humoral and cellular immune responses against recombinant antigens. OBJECTIVES: A subunit vaccine that targets multiple CMV antigens will be used to gain maximal coverage and protective function against CMV infection. rMVA simultaneously expressing sUL55, UL83 and UL123/e4 will be generated, and humoral and cellular immunity it elicits will be characterized, after murine immunization and in vitro to amplify clinical recall responses. STUDY DESIGN: rMVA will be constructed in two steps using UL123/e4-pLW22 followed by sUL55-UL83-pLW51 transfer plasmids. Western blots will be used to characterize expression levels of each antigen. Primary immunity will be evaluated in mouse models, while recall responses to the virally expressed CMV antigens will be assessed in human peripheral blood. RESULTS: We generated CMV-MVA via homologous recombination, and demonstrated high expression levels of sUL55, UL83 and UL123/e4 by Western blot. CMV-MVA immunization potently induced both humoral and cellular immunity to sUL55, UL83 and UL123 after murine immunization, and cellular immunity to UL83 and UL123 by in vitro amplification of T cell recall responses in human PBMC. CONCLUSIONS: rMVA promotes high level expression of three immunodominant CMV antigens, which is reflected in results of immunization studies in which high titers of UL55-specific antibodies and CD4+ T-help are detected, as well as high levels of UL83-specific and moderate levels of UL123-specific CD8+ CTL.     

Cytomegalovirus-specific antibody responses in renal transplant patients with primary and recurrent CMV infections

Cytomegalovirus (CMV) specific immunoglobulin G (IgG) and immunoglobulin M (IgM) antibody responses were measured before and after renal transplantation in 20 patients with primary CMV infection and in 16 patients with recurrent CMV infection. In primary CMV infection IgG antibody titres to late antigen (IgG-LA) measured by indirect fluorescence (IFA) were approximately seven times higher than those obtained by the complement fixation test (CFT). In contrast, in recurrent CMV infection this difference was found to be about twofold. Virus-specific IgM antibody to late antigen (IgM-LA) was detected in 100 percent of patients with primary CMV infection and in only 50 percent of patients with recurrent CMV infection. The IgM-LA titres were highest in primary CMV infection and reached peak levels at approximately 10 weeks post transplantation, whereas in recurrent CMV infection the IgM-LA titres were lower and reached peak levels at three months post transplantation. Moreover, IgM-LA was found to persist in patients from both groups at nine months post transplantation. IgM antibody to early antigen (IgM-EA) was not detected in any patient in this study. However, significant fourfold titre rises in IgG antibody to EA (IgG-EA) were detected in 100 percent of patients with recurrent CMV infection and in 50 percent of patients with primary CMV infection. These results clearly show the difference in antibody responses to the various antigens of CMV in patients with primary and recurrent CMV infection.     

Antibodies against cytomegalovirus-induced early antigens (CMV-EA) in immunosuppressed renal-allograft recipients.

Antibodies against cytomegalovirus-specific early antigens (CMV-EA) were followed in sera, obtained from fifteen immunosuppressed renal-allograft recipients. Eight patients (sixty-two sera) showed seroconversion 47--137 days post-transplantation. Five patients (forty sera) with CMV antibodies at the moment of renal implantation all showed CMV-EA antibody rises. Two patients remained seronegative until 4 years after transplantation. Thus, in immunosuppressed patients, antibodies against CMV-EA remained a high titres during many years (4--8 years) after transplantation as distinct from the apparently transient nature in acutely infected previously healthy adults with CMV mononucleosis or post-perfusion syndrome. These results support the view that CMV-EA antibodies reflect an active viral proliferation in the host.     

Acute cytomegalovirus infection and the host immune response. I. Development and maintenance of cytomegalovirus (CMV) induced in vitro lymphocyte reactivity and its relationship to the production of CMV antibodies

Cytomegalovirus-induced lymphocyte reactivity (CMV-LR) of peripheral blood lymphocytes and antibodies against CMV-early (EA) and late (LA) antigens were studied in eighteen patients with documented acute CMV infections and seventy-six healthy persons. The development of a positive CMV-LR test lagged far behind the appearance of virus-specific antibodies. Positive CMV-LR tests were shown in all fourteen patients who could be tested twice or more. The median value of twenty-two tests in the acute phase (<50 days) was 269 d/min and the median of thirty tests in the post-illness phase (>50 days) was 1301 d/min (P<0·02). Once positive CMV-LR remained so during the follow-up period, up to 479 days after the onset of illness symptoms. In the meantime the LA and EA antibody response remained positive. Only two patients studied once showed negative CMV-LR responses while their serum contained high CMV-EA antibody titres. In CMV-LA seropositive healthy individuals who also possessed CMV-EA antibodies (LA+EA+) CMV-LR was higher (P<0·01) than in the LA seropositives who lacked EA antibodies (LA+EA−). The young LA+EA+ individuals, however, showed better (P<0·02) CMV−LR test results than the aged ones while their CMV antibody levels—especially the EA antibodies (P<0·02)—were lower. This phenomenon of increased CMV antibody production in relation to depressed CMV-LR is possibly caused by age-associated impairment of T lymphocyte function. The combined CMV-LR and EA antibody tests may provide useful means to study the specific immunological host/virus relationship in different clinical situations.     

Cell-mediated immunity to cytomegalovirus in immunocompromised patients: effect of radiation or chemotherapy.

Using the techniques of complement fixation, immunofluorescence, and in vitro lymphocyte transformation (LTF), we studied the humoral antibody and cell-mediated immunity to cytomegalovirus (CMV) in normal subjects, in patients with cancer receiving localized or nonlocalized radiation, and in renal transplant recipients on immunosuppressive chemotherapy. The LTF activity was determined by the whole blood microassay, using four strains of CMV (AD-169, Davis, Veca, and Towne), AD-169 early antigen, and phytohemagglutinin (PAH). The renal transplant subjects manifested significantly depressed LTF responses to PHA and CMV and frequent presence of immunoglobulin M and early antigen-specific antibody response. The depressed LTF response to CMV recovered significantly 2 years after transplantation. The cancer patients were also characterized by a profound drop in LTF responses to PHA and CMV and in immunoglobulin M and early antigen-specific antibody response after nonlocalized radiation. LTF responses to AD-169 and Towne strains were found to be higher than those to Davis and Veca strains. The LTF response to PHA declined with age. However, LTF responses to specific CMV antigens were found to be somewhat increased with advancing age. These observations suggest that transplantation-associated suppression of CMV-specific cell-mediated immunity may improve a few years after transplantation despite continued immunosuppressive therapy.     

Factors influencing the occurrence of active cytomegalovirus (CMV) infections after organ transplantation.

In this study, several factors influencing the occurrence of active CMV infection after organ transplantation (Tx) are analysed. For this purpose, 105 heart, kidney and lung transplant recipients who were CMV-positive or had a CMV-positive donor, were closely monitored for active CMV infection by antigenaemia, cultures, CMV serology and lymphocyte proliferation (LP) to CMV. Univariate and multivariate regression analysis were performed. As pretransplant risk factors the HLA-type and numbers of HLA mismatches between recipients and their donors, and the CMV serology of the recipient and donor were analysed. A new finding was that recipients of donors positive for HLA-B7 were especially at risk for developing active CMV infection (P = 0.03) and CMV disease (P = 0.03). This was not due to increased rejection treatment in these patients. Post-transplant risk factors for development of active CMV infection were absence of detectable cellular immunity to CMV (lymphocyte proliferation) after Tx (P < 0.01) and rejection treatment with OKT3 or ATG (P = 0.05). High levels of IgG anti-CMV did not prevent occurrence of active CMV infection or CMV disease in the CMV+ recipients.     

Evidence for transfer of cellular and humoral immunity to cytomegalovirus from donor to recipient in allogeneic bone marrow transplantation.

In order to study the importance of the immune status of the donor in the development of immunity after allogeneic bone marrow transplantation (BMT), we monitored 23 cytomegalovirus (CMV) antibody-positive BMT recipients for humoral and cellular immunity to CMV, of whom 12 had a CMV antibody-positive and 11 a CMV antibody-negative marrow donor. Lymphocyte proliferation to CMV recovered significantly earlier after BMT in recipients of marrow from a CMV+ donor (10.4 weeks after BMT) compared with the recipients of marrow from CMV- donors (16.7 weeks after BMT, P less than 0.05). This seemed to be specific, as lymphocyte proliferation to phytohaemagglutinin and Candida were not different between the to groups. IgM responses after active infection were seen in both groups, but initial IgG rises without IgM were seen only in recipients of marrow from CMV+ donors (P less than 0.05). Lymphocyte proliferative or humoral immune responses to CMV were not detected in any of the patients in a control group consisting of nine CMV- recipients. These results indicate that T cell memory to CMV is transferred with donor marrow from CMV+ donors, leading in most patients to direct IgG anti-CMV responses and to quicker recovery of cellular immunity to CMV.     

Discordant cellular and humoral immune responses to cytomegalovirus infection in healthy blood donors: existence of a Th1-type dominant response

Previous studies have documented discordant cellular and humoral immune responses to subjects exposed to HIV-1, and that the nature of such responses may determine susceptibility and resistance to disease. We determined whether there is a spectrum of cellular versus humoral immunodominant responses to cytomegalovirus (CMV) infection. Blood samples from 50 healthy blood donors were tested for anti-CMV IgG antibodies and for proliferative responses of peripheral blood mononuclear cells (PBMC) to CMV antigens. Four patterns of immune responses to CMV were found: no detectable response (30%, Ab(-)/Tc(-)), anti-CMV IgG only (28%, Ab(+)/Tc(-)), both anti-CMV IgG and T lymphocyte proliferation to CMV antigens (18%, Ab(+)/Tc(+)), and, interestingly, T lymphocyte proliferation to CMV only (24%, Ab(-)/Tc(+)). To determine whether these immunodominant phenotypes correlate with the ability of PBMC to secrete IL-2 and IFN-gamma in response to CMV antigens, we found that a greater percentage of individuals with a T cell proliferative response to CMV antigens (Ab(-)/Tc(+) and Ab(+)/Tc(+)) responded with increased IL-2 (P = 0.001) and IFN-gamma levels (P = 0.002), compared to those without a proliferative response (Ab(-)/Tc(-) and Ab(+)/Tc(-)). Our data therefore demonstrate that different individuals exhibit different immunodominant patterns of response to CMV. In particular, some individuals who are exposed to CMV fail to develop an antibody response but do develop cellular immunity. Whether these different patterns predict susceptibility or resistance to CMV-induced disease remains to be determined.     

Strain-specific local and systemic cell-mediated immune responses to cytomegalovirus in humans.

Employing the techniques of complement fixation, immunofluorescence, and in vitro lymphocyte transformation assay, the antibody and cell-mediated immunity to cytomegalovirus (CMV) were studied in the serum, peripheral blood lymphocytes, tonsillar lymphocytes, and cord blood lymphocytes. The study population consisted of 32 children undergoing tonsillectomy and adenoidectomy. In the lymphocyte transformation assay, three strains of CMV (AD-169, ADH-1-41, and Davis), herpes simplex type 1, and phytohemagglutinin were employed as antigens. Sixty-five percent of the subjects were found to have CMV-specific antibody activity. The lymphocyte transformation response to phytohemagglutinin was similar in all subjects. No CMV-specific lymphocyte transformation activity was detected in cultures of cord blood lymphocytes. Significant cell-mediated immunity was observed in the tonsillar lymphocytes of 30% (3/10) of the seronegative individuals and in the peripheral blood lymphocytes obtained from one such subject. Over 75% (16/21) of the seropositive subjects demonstrated cell-mediated immunity against one or more strains of CMV in the peripheral blood lymphocytes and tonsillar lymphocytes. In the lymphocyte transformation assay, no cross-reactivity was apparent between CMV and herpes simplex type 1. These studies demonstrate the presence of strain-specific systemic and mucosal cell-mediated immune response to CMV in humans. The frequency and distribution of lymphocyte transformation responses to the three CMV strains suggest antigenic heterogeneity of CMV.     

The contribution of HLA class I antigens in immune status following two doses of rubella vaccination

The variability of humoral and cellular immune responses modulated by human leukocyte antigen (HLA) genes is a significant factor in the protective effect of rubella vaccines. We performed HLA class I typing in a group of 346 healthy schoolchildren and young adults who previously received two doses of measles-mumps-rubella-II vaccine. Rubella virus–specific humoral (serum antibody) immunity and cell-mediated immunity (lymphocyte proliferation) were assessed. Median values for antibody levels and stimulation indices (SI) were 38.63 IU/ml and 2.29 IU/ml, respectively. The alleles that provided suggestive, but not conclusive, evidence of HLA association with rubella seropositivity were HLA-B*2705 (median, 24.68 IU/ml; p = 0.160), B*4501 (median, 61.22 IU/ml; p = 0.098), Cw*0303 (median, 30.34 IU/ml; p = 0.102) and Cw*0704 (median, 26.58 IU/ml; p = 0.144). These alleles approach, but do not achieve, statistical significance. Of all the alleles analyzed, HLA-B*3503 (median SI, 3.00; p = 0.031) and HLA-Cw*1502 (median SI, 3.19; p = 0.035) were positively associated with lymphoproliferative responses to rubella virus antigens, whereas the HLA-B*3901 (SI, 1.34; p = 0.066) allele was negatively associated. This suggests that class I HLA alleles may have limited associations with humoral and cellular immune responses to rubella vaccine. These data may facilitate our understanding of immune response variation in a genetically outbred heterogeneous population.     

Cytomegalovirus-directed lymphocyte reactivity in healthy adults tested by a CMV-induced lymphocyte transformation test.

A cytomegalovirus (CMV) induced lymphocyte tranformation test (LTT) performed as a microtechnique assay is described. Investigations were done in a group of twenty-five healthy adults with normal lymphocyte reactivity to phytohaemagglutinin and previously encountered antigens. Lymphocyte reactivity was established in CMV-seropositive persons using heat-inactivated cell-free CMV strain AD 169 in an optimal stimulating dose of 10(1-5) TCID 50. Positive CMV-LTT results were shown to be induced by purified inactivated CMV. Most persons (twelve out of seventeen) with antibodies against CMV-induced late or virus-structure antigens (LA) showed positive (stimulation index greater than 2-0) CMV-LTT results. Magnitude of CMV-LTT responses was positively related to the CMV-LA Ab titres (P less than 0-01). All persons (six) with antibodies against CMV early or non-structural antigens showed positive CMV-LTT results and highest stimulation rates were found in this group. Generally CMV-seronegative persons were non-responsive. Only in one case (LA Ab titre 10) a borderline positive response (stimulation index 2-3) was seen. The CMV-LTT was shown to have virus-specific properties because it was not related to simultaneously performed LTTs with Epstein-Barr virus and Herpes simplex virus in the same persons. It is concluded that CMV-LTTs are useful virus-specific in vitro techniques for the study of CMV-directed lymphocyte reactivity. Its possible relationship to other cell-mediated immunity tests and its practical applications are discussed.     

Cytomegalovirus (CMV): specific lymphokine production in congenital CMV infection.

The studies reported here were designed to examine cytomegalovirus (CMV)-specific lymphokine production in infants with congenital CMV infection to elucidate the role of the efferent limb of the immune response to this virus. Mononuclear cells (MNC) from adult control donors and congenitally infected infants were stimulated with mitogen (PHA) or antigens (CMV, mumps), and the supernatants were assayed for production of migration inhibitory factor (MIF) and interferon (IFN). Concordance was observed between lymphocyte proliferative responses to CMV antigen and production of MIF in both control donors and congenitally infected infants. PHA-stimulated cultures from both controls and patients resulted in immune-specific IFN-gamma production in all cases. CMV-stimulated MNC from controls produced IFN when lymphocyte proliferation responses to the antigen were present, whereas those with absent proliferation did not produce IFN. CMV-induced IFN was detected in several patients who had reduced lymphocyte proliferative responses to CMV. The predominant species of IFN stimulated by CMV was neutralized by anti-IFN-gamma, suggesting that CMV induced primarily immune-specific IFN-gamma. In some cases, IFN activity was also reduced to a lesser degree by anti-IFN-alpha, suggesting either the presence of IFN-alpha or of cross-reacting antigenic determinants shared by the two species of IFN. Mumps viral antigen induced primarily IFN-alpha regardless of the immunological status of the donor. We conclude that lymphokine production in congenital CMV reflects the state of activation and/or proliferation of CMV-specific T helper cells rather than an intrinsic defect in the efferent limb of the immune response.     

Humoral and Cellular Responses to a Single Dose of Fendrix in Renal Transplant Recipients with Non‐response to Previous Hepatitis B Vaccination

Approximately 70% of kidney transplant recipients are non‐responders to conventional hepatitis B virus (HBV) vaccines. We examined whether Fendrix™, an HBV vaccine containing 3‐O‐desacyl‐4′‐monophosphoryl lipid A (MPL) as adjuvant, could induce HBV immunity in these patients and compared their vaccination efficacy with healthy controls tested previously by the same assays. We selected 35 kidney transplant recipients who had been vaccinated at least thrice against HBV but had never displayed anti‐HBs antibodies. We re‐assessed their anti‐HBs antibody titres and further determined cellular HBV immunity by proliferation assay and interferon (IFN)‐γ ELISpot. Seventeen recipients did neither display humoral nor cellular immunity and could be tested prior to and at month 1 after vaccination. Of note, HLA antigens associated with non‐response to HBV vaccination (HLA‐DRB1*03 and HLA‐DQB1*02) were over‐represented in these 17 recipients. At month 1 after a single vaccination with Fendrix™, we observed a significant increase in anti‐HBs antibodies (P = 0.02). In seven of 17 recipients, we detected anti‐HBs antibodies ≥10 IU/l (10–264), in four HBV‐specific lymphocyte proliferation (stimulation index of 2.6–8.7) and in one specific IFN‐γ responses (12 spots increment). The vaccination response to Fendrix™ was significantly higher (P = 0.035) than the response to HBVaxPro™ in young healthy controls. In summary, the results show that a single vaccination with Fendrix^™ could already induce HBV‐specific humoral and/or cellular responses in ten of 17 kidney transplant patients. Thus, Fendrix™ appears as an efficient vaccine in this patient cohort.     

Cell-mediated and humoral immune responses of renal transplant recipients with cytomegalovirus infections

Cell-mediated and humoral immune responses were determined in immunosuppressed renal transplant recipients. A micro-phytohaemagglutinin (PHA) stimulation test utilizing whole heparinized blood and a macro-PHA test utilizing separated, washed lymphocytes were used to study cell-mediated immunity. Humoral immune status was estimated by determining quantitative immunoglobulins and complement fixing (CF) antibody titres to cytomegalovirus (CMV) infection. Micro-PHA responses were found to be markedly depressed in patients undergoing immunosuppressive therapy and in patients with chronic uraemia. Macro-PHA responses were normal, indicating that serum factors were responsible for the depressed micro-PHA responses. Antibody responses to CMV infections were found to be ten to a hundred times higher than in normal persons. An inverse relationship was demonstrated between micro-PHA responses and peak CF antibody titres to CMV infections. Humoral immune responses appeared to compensate for depressed cell-mediated immunity as measured by the micro-PHA test. Four patients had very low micro-PHA responses, did not respond to their CMV infections with CF antibody, and died of mixed bacterial and viral infections. Serum immunoglobulins of two were studied and were shown to be greater than two standard deviations below the normal mean. These patients appeared more suppressed than other patients receiving similar therapy and thus probably retained higher concentrations of suppressive drugs.     

ELISpot assay as a sensitive tool to detect cellular immunity following influenza vaccination in kidney transplant recipients

Enhanced cellular immunity following influenza vaccination has been undetectable in kidney transplant recipients so far. Protection from influenza is dependent on cellular and humoral immunity. Aim of the study was to investigate immune responses before and after vaccination with influenza A and B antigens in 65 kidney transplant recipients. A significant increase in proliferative responses was only observed towards influenza B (P < 0.0001) by lymphocyte transformation test. The enzyme-linked immunospot (ELISpot) assay was more sensitive and detected significant, 3- to 5-fold increases (P < 0.0001) in interferon-gamma secretion using influenza A and B antigens. Furthermore, influenza antibody titers increased significantly (P < 0.0001). At month 1 post-vaccination 85% of patients displayed specific cellular, and 95% or 92% humoral immunity against influenza A and B, respectively. Thus, applying the sensitive ELISpot assay, influenza-specific cellular immunity could be detected for the first time in kidney transplant recipients after vaccination.     

Clinical significance of CMV-specific T helper responses in lung transplant recipients

Background. Cytomegalovirus (CMV) disease continues to be a major problem for lung transplant patients who generate an inefficient immune response to control this viral infection. Both T helper and cytotoxic T cells are thought to play an important role in prevention and control of CMV disease. We investigated the clinical significance of CMV-specific memory responses in lung transplant recipients. Method. Peripheral blood samples (140) were collected from 99 lung transplant recipients. Patients were grouped according to their pre-transplant CMV serological status as recipient/donor (R−/D+, 25 patients), 28 R+/D+ patients, 35 R+/D− patients and 11 R−/D− patients. Memory responses to CMV whole antigen, 5 CMV proteins, and tetanus toxoid (TT) were measured in a 6-day proliferative assay. Results were expressed as the stimulation index (SI = experimental cpm/background cpm), and were considered positive if the SI was >3 and the cpm values were over 1,000. Results. The frequency of positive CMV memory responses was similar in three groups: 64% for R−/D+, 63% for R+/D+ and 56% for R+/D− except for R−/D− (21%). The memory response to TT was similar for all four groups (70% of samples were positive). The level of responsiveness to individual CMV proteins was much higher in R+/D+ group (65%) than the other two groups (35% for R+/D−, and 31% for R−/D+). We determined the temporal relationship between the presence of CMV-specific memory responses and the diagnosis of CMV disease. In the R−/D+ group, 16 of 17 patients who had CMV disease eventually developed CMV-specific memory. In those patients (n = 3) who failed to develop CMV-specific T helper response for a prolonged time, all had recurrent CMV disease. In the R+/D+ group, 4 of 8 patients with CMV disease exhibited CMV-specific memory responses. Three of 4 patients in whom we observed a persistent absence of CMV-specific memory had multiple episodes of CMV pneumonitis. In the R+/D− group, only one of 4 patients with CMV disease had suppressed CMV-specific memory response after first episode of CMV pneumonitis and had recurrent disease. Conclusion. In lung transplant recipients, the loss or persistent lack of CMV-specific memory following infection was associated with chronic CMV disease. These data suggest that monitoring T helper memory responses following primary CMV infection or after augmented immunosuppression for treatment of rejection may identify those patients at risk for morbidity associated with recurrent CMV disease.     

Cytomegalovirus infection in infancy: virological and immunological studies

Immunological and virological studies on 18 infants with cytomegalovirus (CMV) infection were performed. Eleven of these infants were studied on multiple occasions over a period of 1 year. The patients were divided into three clinical groups based on the probable time of infection and the resulting variation in clinical presentation. General parameters of cell-mediated immunity as determined by E-rosette formation and lymphocyte proliferative responses to mitogens and antigens were found to be normal. Quantitation of CMV excretion in urine, CMV-specific immunofluorescent (IF) and complement-fixing (CF) antibody titres and CMV-specific cell-mediated immune responses were done on all patients at approximately monthly intervals. Throughout the study period all patients continued to excrete CMV despite the presence of high antibody titres to the virus. CMV-specific lymphocyte proliferative responses were absent or diminished in 15 of the 18 patients. The immunological and virological status of all patients was similar regardless of the clinical manifestation of infection.     

Poor memory B cell generation contributes to non-protective responses to DTaP vaccine antigens in otitis-prone children

We recently identified a cohort of children with recurrent episodes of acute otitis media (AOM) who fail to generate protective antibody titres to otopathogens and several vaccine antigens. In this study we determined the antibody levels against DTaP vaccine antigens, diphtheria toxoid (DT), tetanus toxoid (TT) and acellular pertussis toxoid (PT) in sera from 15 stringently defined otitis-prone (sOP) children and 20 non-otitis-prone (NOP) children. We found significantly lower concentrations of immunoglobulin (Ig)G antibodies against vaccine antigens in the serum of sOP children compared to age-matched NOP children. To elucidate immunological cellular responses to the vaccines in these children, we investigated memory B cell responses to DTaP vaccination. We used fluorescently conjugated vaccine antigens to label antigen receptors on the surface of memory B cells and examined the frequency of antigen-specific CD19⁺ CD27⁺ memory B cells in the peripheral blood. sOP children showed a significantly lower percentage of antigen-specific CD19⁺ CD27⁺ memory B cells than NOP children. We also found a linear correlation between the frequencies of memory B cells and circulating IgG titres for DT, TT and PT proteins. To our knowledge, this is the first study to show significant differences in memory B cell responses to DTaP vaccine antigens and their correlation with the circulating antibodies in young children with recurrent AOM.     

Defective anticarbohydrate antibody responses to naturally occurring bacteria following bone marrow transplantation.

The long-term recipients of allogeneic bone marrow transplantation (BMT) are at an increased risk of death due to bacterial infections. We evaluated the anticarbohydrate antibody responses of BMT recipients to a naturally occurring bacterial carbohydrate, polyribose phosphate (PRP). The recipients of autologous BMT achieved protective anti-PRP levels (>100 ng/mL) by 3 years after transplantation, with a pattern consistent with a recapitulation of the ontogeny of anticarbohydrate antibody responses. None of the six recipients of unrelated BMT who were off immunosuppressive therapy had protective anti-PRP levels, though their response to a protein antigen (tetanus toxoid) was normal. Of 48 recipients of histocompatible BMT, 22 (46%) had protective anti-PRP antibody levels, whereas 13 (27%) recipients who were >3 years post-BMT did not have protective levels. Therefore, all unrelated recipients and a significant proportion of histocompatible recipients without clinical graft-vs.-host disease had persistent and prolonged defects in their capacity to produce antibodies to naturally occurring bacterial carbohydrate antigens. These results suggest that allogeneic BMT recipients should be longitudinally evaluated for their anticarbohydrate antibody responses and that patients with defective antibody responses should receive prophylactic antibiotics or replacement immunoglobulin therapy or both to reduce their risk of late bacterial infections.