Identification and first report of Inonotus (Phellinus) tropicalis as an etiologic agent in a patient with chronic granulomatous disease.

Although isolates of filamentous basidiomycetes can usually be recognized in a clinical laboratory setting, identification is problematic, as they seldom exhibit diagnostic morphological features formed in nature. This paper is the first report of Inonotus (Phellinus) tropicalis inciting human disease and describes the methods used to support the identification.

Antibody responses to bacteriophage phi X-174 in human subjects exposed to the antarctic winter-over model of spaceflight.

BACKGROUND: It has been proposed that exposure to long-term spaceflight conditions (stress, isolation, sleep disruption, containment, microbial contamination, and solar radiation) or to ground-based models of spaceflight will alter human immune responses, but specific antibody responses have not been fully evaluated. OBJECTIVE: We sought to determine whether exposure to the 8-month Antarctic winter-over model of spaceflight would alter human antibody responses. METHODS: During the 1999 Australian National Antarctic Research Expeditions, 11 adult study subjects at Casey, Antarctica, and 7 control subjects at Macquarie Island, sub-Antarctica, received primary and secondary immunizations with the T cell-dependent neoantigen bacteriophage phi X-174. Periodic plasma samples were analyzed for specific antibody function. RESULTS: All of the subjects from Casey, Antarctica, cleared bacteriophage phi X-174 normally by 1 week after primary immunization, and all had normal primary and secondary antibody responses, including immunologic memory amplification and switch from IgM to IgG antibody production. One subject showed a high normal pattern, and one subject had a low normal pattern. The control subjects from Macquarie Island also had normal immune responses to bacteriophage phi X-174. CONCLUSIONS: These data do not support the hypothesis that de novo specific antibody responses of subjects become defective during the conditions of the Antarctic winter-over. Because the Antarctic winter-over model of spaceflight lacks the important factors of microgravity and solar radiation, caution must be used in interpreting these data to anticipate normal antibody responses in long-term spaceflight.

Prospective 5-year study of peripheral blood CD4, CD8, and CD19/CD20 lymphocytes and serum Igs in children born to HIV-1 women. The P(2)C(2) HIV Study Group.

BACKGROUND: Peripheral blood CD4(+) and CD8(+) T cells, CD19(+)/20(+) B cells, and serum Igs are known to be altered by the progression of pediatric HIV-1 infection, but their evaluation as predictors of survival needs further definition. OBJECTIVE: To determine the natural history of these immune factors and their importance in predicting survival, we studied 298 HIV-1 vertically infected (HIV-1(+)) children over a 5-year period. METHODS: These immune factors and serum HIV-1 RNA levels were measured in two groups: (1) a birth cohort of children enrolled up to age 28 days postnatally, including 93 HIV-1(+) and 463 HIV-1 uninfected infants (HIV-1(-)), and (2) an older cohort of 205 HIV-1(+) children enrolled after the age of 28 days, who were classified as survivors or nonsurvivors. RESULTS: In the birth cohort HIV-1(+) children had significantly lower CD4(+) T-cell counts, higher CD8(+) T-cell counts, and lower CD19(+)/20(+) B-cell counts and higher IgG, IgA, and IgM levels than HIV-1(-) children. In the older cohort survivors had significantly higher CD4(+) and CD8(+) T-cell and CD19(+)/CD20(+) B-cell counts and higher IgG, lower IgA, and lower IgM levels than did nonsurvivors. In univariable analysis factors affecting survival in the older cohort were baseline CD4(+) and CD8(+) T-cell and CD19(+)/20(+) B-cell counts and IgG and HIV-1 RNA levels (all P <.05). In multivariable analysis high baseline CD4(+) T-cell count and low baseline HIV-1 RNA load remained important. CONCLUSION: The longitudinal mean profiles of CD4 and CD8 T-cell and CD19/20 B-cell counts and serum IgG levels helped to describe the natural progression of HIV-1 disease in children. However, only baseline CD4 T-cell count independently predicted survival.

Shipment impairs lymphocyte proliferative responses to microbial antigens.

Lymphocyte proliferation assays (LPAs) are widely used to assess T-lymphocyte function of patients with human immunodeficiency virus infection and other primary and secondary immunodeficiency disorders. Since these assays require expertise not readily available at all clinical sites, specimens may be shipped to central labs for testing. We conducted a large multicenter study to evaluate the effects of shipping on assay performance and found significant loss of LPA activity. This may lead to erroneous results for individual subjects and introduce bias into multicenter trials.

Evaluation of immune survival factors in pediatric HIV-1 infection.

Peripheral blood CD4+ and CD8+ T cells, CD19+/20+ B cells, and serum immunoglobulins (Igs) have been implicated as survival factors for pediatric HIV-1 infection. To determine which of these immune factors might be important in predicting survival, we studied HIV-1 vertically infected (HIV-1+) children over a 5-year period. Peripheral blood lymphocytes and Igs were measured in 298 HIV-1+ children, who were classified as survivors or nonsurvivors, and in 463 HIV-1 vertically exposed and noninfected (HIV-1-) children. Measurements of other possible survival factors were included in this study: albumin, hemoglobin, lactic dehydrogenase (LDH), and HIV-1 RNA levels. Survivors had significantly higher CD4+ T-cell, CD8+ T-cell, and CD19+/CD20+ B-cell counts and serum IgG levels, but lower serum IgA and IgM levels than nonsurvivors. Serum albumin and blood hemoglobin levels were higher, but serum LDH and HIV-1 RNA levels were lower in the survivors compared to nonsurvivors. In univariable analysis, factors affecting survival were baseline CD4+ T-cell and CD8+ T-cell counts, IgG, albumin, hemoglobin, LDH, and HIV-1 RNA (all p < 0.001). In multivariable analysis, high baseline CD4+ T-cell count, IgG and albumin levels, and low baseline HIV-1 RNA load remained important factors for survival. Serum IgG level has been identified as an immune factor that independently predicts survival, in addition to the already established CD4+ T-cell count. The HIV-1 RNA and serum albumin levels also predicted survival.

Congenital heart disease in mice deficient for the DiGeorge syndrome region.

The heterozygous chromosome deletion within the band 22q11 (del22q11) is an important cause of congenital cardiovascular defects. It is the genetic basis of DiGeorge syndrome and causes the most common deletion syndrome in humans. Because the deleted region is largely conserved in the mouse, we were able to engineer a chromosome deletion (Df1) spanning a segment of the murine region homologous to the human deleted region. Here we describe heterozygously deleted (Df1/+) mice with cardiovascular abnormalities of the same type as those associated with del22q11; we have traced the embryological origin of these abnormalities to defective development of the fourth pharyngeal arch arteries. Genetic complementation of the deletion using a chromosome duplicated for the Df1 DNA segment corrects the heart defects, indicating that they are caused by reduced dosage of genes located within Df1. The Df1/+ mouse model reveals the pathogenic basis of the most clinically severe aspect of DiGeorge syndrome and uncovers a new mechanism leading to aortic arch abnormalities. These mutants represent a mouse model of a human deletion syndrome generated by chromosome engineering.

Incidence of presumed cytomegalovirus retinitis in HIV-infected pediatric patients.

INTRODUCTION: Large population studies of adult patients suggest an incidence of cytomegalovirus (CMV) retinitis as high as 19% to 20% as a late complication of adult HIV infection. We conducted this prospective study of a large cohort of HIV-infected children to determine the incidence of CMV retinitis in HIV-infected children. METHODS: From January 1984 to August 1997, 173 HIV-infected children were followed up for an average of 55.3 months (13-164 months). The patients were seen in the Department of Pediatrics at least once every 6 months. Ophthalmologic examinations were initiated when a patient's CD4 count dropped below 50 or sooner if required for ophthalmologic or other indications. Ophthalmologic examination was then repeated every 6 months. RESULTS: A total of 116 (67%) of 173 patients underwent ophthalmologic examination. Four (3.4%) of 116 patients had CMV retinitis at a mean time of 17.3 months (8-38 months) after their CD4 counts dropped below 20. None of the 4 patients with CMV retinitis had subjective visual complaints despite advanced retinitis. Three patients had bilateral and 1 patient had unilateral CMV retinitis. CONCLUSIONS: CMV retinitis occurred infrequently in HIV-infected pediatric patients and was diagnosed only in patients with a CD4 count below 20. Routine ophthalmologic screening examinations may not be necessary in pediatric patients until the CD4 count is below 20. Because children may not complain of decreased vision, at-risk children should undergo frequent ophthalmologic examination.

A comparison of peripheral blood coculture versus 18- or 24-month serology in the diagnosis of human immunodeficiency virus infection in the offspring of infected mothers. Women and Infants Transmission Study.

The Women and Infants Transmission Study (WITS) has established virologic definitions of human immunodeficiency virus (HIV)-infected and uninfected children that have been widely used but never formally compared with serologic definitions of infection. Data from the offspring of HIV-infected women in the WITS with frequent HIV cultures during the first year of life and with HIV serology at 18 and/or 24 months of age were analyzed. Seventy-seven infants were HIV-infected and 430 uninfected by both virologic and serologic criteria. Thirteen were virologically infected (> or = 2 positive cultures) but either seronegative or serologically indeterminate. All but 1 of these had clinical HIV disease at the time of analysis. In this pediatric cohort, children defined as infected by virologic criteria often (13/90) had negative or indeterminate serology despite symptoms of HIV disease. Results suggest that serology at 18-24 months has high specificity but poor sensitivity. It should not be considered the reference standard in identifying HIV infection in perinatally exposed children.

Long-term results of triple-drug-based immunosuppression in nonneonatal pediatric heart transplant recipients.

BACKGROUND: Few reports document long-term results of pediatric cardiac transplantation in which triple therapy (cyclosporine, azathioprine, and corticosteroids) was the mainstay of immunosuppression. This report details a single center's pediatric transplant experience and analyzes the relative contributions of selected pre/posttransplant risk factors on long-term morbidity and mortality. METHODS: Retrospective data were collected for all non-neonatal pediatric transplant recipients including: presenting diagnosis, cardiac hemodynamics (particularly pulmonary vascular resistance index), donor ischemic time, occurrence of postoperative infections, episodes of allograft rejection, incidence of posttransplant lymphoproliferative disease or coronary artery disease (CAD), and overall survival. Analysis of single variables and a Cox-proportional hazards model were utilized to determine the impact of pre/posttransplant risk factors on long-term survival. RESULTS: From 1984 to 1995, 64 patients (mean age, 8.3 years), 46 of whom had cardiomyopathy and 18 who had inoperable complex congenital heart disease, underwent cardiac transplantation and received triple-drug immunosuppression. Orthotopic transplantation was performed unless the pulmonary vascular resistance index remained >6 um2 (despite use of pulmonary vasodilator). One patient required heterotopic transplantation. Average donor ischemic time was 217 min. An average of 1.2 rejection episodes/patient occurred (average follow-up period: 50 months). No patient developed posttransplant lymphoproliferative disease, but 22 patients (34%) developed CAD. Overall survival was 80%, 60%, and 57% at 1, 5, and 10 years, respectively. Of outcome variables analyzed, rejection frequency was significantly increased in patients who subsequently developed CAD, but the presence of CAD was not significantly correlated with mortality. CONCLUSION: Triple-drug-based immunosuppressive maintenance therapy in pediatric heart transplant recipients results in good long-term graft survival.

Complete DiGeorge syndrome: persistence of profound immunodeficiency.

OBJECTIVE: DiGeorge syndrome is characterized by developmental defects of the heart, parathyroid glands, and thymus. The objective of this study was to determine whether T-cell function spontaneously improves in patients with DiGeorge syndrome who have profoundly depressed T-cell proliferative responses to mitogens at presentation, regardless of the T-cell count. STUDY DESIGN: We conducted a retrospective chart review of eight patients with DiGeorge syndrome who had no proliferative responses to mitogens on presentation. RESULTS: Despite lack of responsiveness of the patients' peripheral blood lymphocytes to mitogens, T cells were occasionally detected, and the patients' cells often responded to IL-2 and in mixed lymphocyte reactions. Unresponsiveness to mitogens and clinical immunodeficiency persisted without immune-based therapy. One patient is alive and well after immunoreconstitution from thymic transplantation. The others either died early of complications of their disease such as gastroesophageal reflux with aspiration (2 patients) or infection (2 patients) or died after attempts at immunorestorative therapy with IL-2, thymus transplantation, or bone marrow transplantation (3 patients). CONCLUSION: Eight patients with DiGeorge syndrome who were first seen with no mitogen responsiveness did not improve spontaneously. We recommend HLA-identical bone marrow transplantation or thymic transplantation for these patients as soon as the diagnosis is confirmed.

Objective measures of allergic disease in children with human immunodeficiency virus infection.

BACKGROUND: Available information suggests that IgE levels are elevated in adults infected with human immunodeficiency virus (HIV), and that increased IgE levels correlate with allergic disease, with decreased CD4 counts, and with a poor prognosis. Data with respect to these factors in children are scant. OBJECTIVE: We investigated whether serum IgE levels are elevated in children with HIV and, if so, whether the serum IgE level correlates with the degree of immunodeficiency and/or objective indicators of allergic disease. METHODS: Serum IgE levels, CD4 counts, absolute eosinophil counts, and immediate hypersensitivity skin test (IHST) results were collected from 43 children with symptomatic HIV infection (mean age 7.2 years). Associations between serum IgE levels, CD4 counts, and eosinophil counts were investigated by multiple stepwise linear regression analysis. Data were stratified according to IHST positivity, and analysis of variance was used to compare mean values for age, CD4 counts, IgE levels, and eosinophil counts between the two groups. RESULTS: Serum IgE values were elevated more than 2 SDs above control age-matched mean values in 17 of 43 patients (40%). IHST results were positive in 12 of 43 patients (28%). CD4 counts were less than 200/mm3 in 17 of 43 patients (40%). Stepwise linear regression failed to demonstrate any correlation between serum IgE levels and either CD4 or eosinophil counts. With data divided into two groups according to IHST results (positive vs negative), analysis of variance failed to reveal significant differences between means for patient age, CD4 counts, IgE levels, or eosinophil counts. CONCLUSIONS: Our findings confirm that serum IgE levels are increased in children infected with HIV, just as in adults. However, an elevated serum IgE level did not correlate with allergic disease as measured by IHST results and eosinophil counts, nor with the degree of immune dysfunction as approximated by CD4 counts. The mechanism and significance of elevated serum IgE levels remain unclear in children with HIV, and warrant further investigation.

Mutation analysis of IL2RG in human X-linked severe combined immunodeficiency.

Severe combined immunodeficiency (SCID) is a syndrome of profoundly impaired cellular and humoral immunity. In humans, SCID is most commonly caused by mutations in the X-linked gene IL2RG, which encodes the common gamma chain, gamma c, of the leukocyte receptors for interleukin-2 and multiple other cytokines. To investigate the frequency and variety of IL2RG mutations that cause SCID, we analyzed DNA, RNA, and B-cell lines from a total of 103 unrelated SCID-affected males and their relatives using a combination of molecular and immunologic techniques. Sixty-two different mutations spanning all eight IL2RG exons were found in 87 cases, making possible correlations between mutation type and functional consequences. Although skewed maternal X chromosome inactivation, single-strand conformation polymorphism, mRNA expression, and cell surface staining with anti-gamma c antibodies were all helpful in establishing IL2RG defects as the cause of SCID, only dideoxy fingerprinting and DNA sequence determination each detected 100% of the IL2RG mutations in our series. Abnormal gamma c chains may be expressed in the lymphocytes of as many as two thirds of patients with X-linked SCID. Specific mutation diagnosis thus remains technically challenging, but it is important for genetic counseling and perhaps for helping to select appropriate subjects for retroviral gene therapy trials, This is a US government work. There are no restrictions on its use.

Outcomes among pediatric heart transplant recipients.

Postoperative cytomegalovirus prophylaxis with cytomegalovirus immunoglobulin or ganciclovir has decreased the incidence of cytomegalovirus disease in cytomegalovirus-negative recipients of cytomegalovirus-positive donor organs. In adults, these drugs have also been used to treat recipients who developed symptomatic cytomegalovirus disease. This report describes outcomes of predominantly cytomegalovirus-negative pediatric cardiac transplant recipients of cytomegalovirus-positive donor organs who received cytomegalovirus immunoglobulin plus ganciclovir as cytomegalovirus prophylaxis, as well as results of this combination therapy when used to treat cytomegalovirus disease. We reviewed the records of children who received donor hearts at our institution between 1989 and 1994. Cytomegalovirus-negative patients who received cytomegalovirus-positive donor organs were given prophylaxis consisting of ganciclovir (5 mg/kg every 12 hours for 14 days, followed by maintenance dosage of 5 to 6 mg/kg every day for 14 days) plus 7 scheduled cytomegalovirus immunoglobulin infusions. Cytomegalovirus infection was documented by culture, polymerase chain reaction, and cytomegalovirus immunoglobulin M seroconversion of a 4-fold or greater rise in cytomegalovirus immunoglobulin G titers. After infection, patients were diagnosed with cytomegalovirus disease when they developed clinical symptoms. These episodes were treated with cytomegalovirus immunoglobulin infusions plus ganciclovir (5 mg/kg every 12 hours) until symptoms resolved. Of 40 cardiac transplant recipients, 10 cytomegalovirus-negative and 9 cytomegalovirus-positive patients received cytomegalovirus-positive donor organs. Five patients (3 of whom were seronegative and had received dual-agent prophylaxis) developed cytomegalovirus disease, which resolved with dual-agent therapy. During an average 15-month follow-up period, no significant morbidity or mortality was attributable to cytomegalovirus disease. Post-transplant dual-therapy cytomegalovirus prophylaxis appears to be as safe and effective in children as in adults, when our results are compared with the published results of studies in adults. Dual-agent treatment eradicated symptoms among patients who developed cytomegalovirus disease. This regimen may allow safer use of the cytomegalovirus-positive donor pool for pediatric recipients.

Early detection of HIV in infants: aspects of design and analysis for diagnostic test studies.

OBJECTIVE: To review investigations on the early detection of HIV infection in infants to determine adherence to traditional methods of study design and analysis for evaluating new laboratory tests. DATA SOURCES: A National Library of Medicine (MEDLINE) search was conducted to identify such investigations through 1993. Cited references in identified manuscripts were also considered. The search was restricted to investigations of human subjects and those published in the English language. STUDY SELECTION: Final inclusion criteria included (1) report of the age and human immunodeficiency virus (HIV) infection status of the subjects at the time of the diagnostic testing, and (2) presentation of data allowing confirmation of presented analyses and additional analyses. DATA EXTRACTION: Criteria for judging the investigations included (1) whether the criteria used to determine the positive and negative test results were defined; (2) whether the necessary sample size for the study was calculated; (3) whether the patients studied were representative of the patients to whom the test would be applied; (4) whether a gold standard evaluation was performed; (5) whether the outcomes included in the analyses were independent; (6) whether the test characteristics were properly analyzed; and, (7) whether confidence intervals were presented. DATA SYNTHESIS: An informative presentation of a diagnostic test should include as a minimum the seven criteria listed above. Only 21 of 36 (58%) of the studies incorporated at least three of the criteria. CONCLUSIONS: There is a wide variation in the manner in which investigations of diagnostic tests are conducted and the results reported. Increased awareness and use of standard study designs and analyses will allow the application of metanalyses. Such analyses will help guide the direction taken for finding and establishing early diagnostic procedures for HIV infection at birth or during infancy.

Postpericardiotomy syndrome in pediatric heart transplant recipients. Immunologic characteristics.

Clinical features of postpericardiotomy syndrome (PPS) occur in pediatric heart transplant recipients despite immunosuppression, which raises questions about the mechanism of PPS. We studied the clinical and immunologic characteristics of 15 pediatric heart transplant patients, ages 1.1 to 17.8 years (mean, 7.5 years); 7 had clinical evidence of PPS (PPS+), and 8 were without clinical features of PPS (PPS-). Indicators of PPS included fever, irritability, pericardial friction rub, leukocytosis without other cause, and pericardial effusion. The onset of PPS was from 9 to 26 postoperative days (mean, 16 days). Immunosuppressive regimens were comparable up to the day of PPS diagnosis in PPS+ patients, and up to day 16 in PPS- patients (average onset of PPS in PPS+ patients). No differences were found between groups with respect to weight-adjusted dosages of cyclosporin A, azathioprine, or corticosteroids. Mean cyclosporin A levels in PPS+ and PPS- patients were 142 +/- 88 ng/mL (mean +/- standard deviation) and 265 +/- 122 ng/mL (p = 0.045), respectively. Echocardiographic data on 3 PPS+ patients within 1 day of PPS diagnosis revealed pericardial effusions ranging from 5 to 24 mm. No data were available on the remaining 4 PPS+ patients. Minimal pericardial effusions (< 10 mm) were seen in 4 PPS- patients during a comparable time period. One PPS- patient required pericardiocentesis. Endomyocardial biopsy rejection grade did not differ between groups. Means pretransplant soluble interleukin-2 receptor levels did not differ between PPS+ and PPS- patients (758 +/- 410 vs 653 +/- 270 IU/mL); nor did the PPS+ pretransplant levels differ from levels obtained 1 or 2 months postoperatively (700 +/- 437 and 751 +/- 367 IU/mL, respectively). Although pretransplant percentages of the standard T-cell (CD2, CD3, CD4, CD8) and B-cell (DR and CD19) markers differed from post-transplant values, the changes could be explained by the immunosuppressive regimen and did not differ between PPS+ and PPS- patients. In the PPS+ patients, however, there were significant increases in the proportion of activated helper T cells (CD4+/25+) and cytotoxic T cells (Leu-7+/CD8+) following heart transplantation in comparison with pretransplant levels. We speculate that these changes in activation marker in PPS+ patients suggest a possible role for cell-mediated immunity in the pathogenesis of PPS in this group of patients.

Sensitivity of immune complex-dissociated p24 antigen testing for early detection of human immunodeficiency virus in infants.

Several investigators have suggested that early diagnosis of human immunodeficiency virus (HIV) infection in infants could be accomplished with a modified, more-sensitive, acid-dissociated p24 antigen enzyme-linked immunosorbent assay (ELISA) technique (p24 antigen immune complex dissociation [ICD]). We compared detection of HIV infection by HIV culture, PCR, and p24 antigen ICD assays in 46 infants by using samples collected independently. The detection sensitivity of the p24 antigen ICD assay was 0% with cord blood samples (2 HIV-positive infants), 38% with plasma samples from infants under 3 months of age (8 HIV-positive infants), and 58% overall (12 HIV-positive infants). By contrast, the sensitivities of HIV culture and PCR were 50% for cord blood samples, 75% for plasma samples from infants under 3 months of age, and 83% overall. These results indicate that the p24 antigen ICD does not offer the sensitivity necessary for this assay to be used as an indicator of HIV infection in infants.

A comparative study of human immunodeficiency virus culture, polymerase chain reaction and anti-human immunodeficiency virus immunoglobulin A antibody detection in the diagnosis during early infancy of vertically acquired human immunodeficiency virus infection.

The infection status of 91 infants born to mothers with human immunodeficiency virus (HIV) infection was determined. Twenty-eight (31%) infants had confirmed HIV infection and 63 (69%) had seroreverted to HIV and lack evidence of infection. During the first 6 months of life HIV culture had a sensitivity and specificity for diagnosis of HIV infection of 80 and 100%, respectively. False negative HIV cultures were observed in only 7 of 35 specimens, 6 from among the 12 infected infants tested at birth. The sensitivity and specificity of polymerase chain reaction (PCR) detection of HIV were 95 and 93% respectively. A single false negative PCR test result was observed among the 19 tests performed on specimens from HIV-infected infants. False positive PCR test results were observed occasionally throughout the first 6 months of life. Detection of HIV-specific IgA antibody lacked diagnostic sensitivity; positive test results were observed in only 53% of specimens obtained from infected infants. Culture and PCR detection offer excellent sensitivity and specificity for diagnosis of HIV infection during the first 6 months of life; however, false-negative HIV cultures sometimes are observed, particularly during the newborn period, and either false negative or false positive PCR test results may be noted occasionally. For purposes of clinical decision-making, any positive test result should be confirmed with a second HIV culture or PCR test performed on a separate blood specimen.

Application of carrier testing to genetic counseling for X-linked agammaglobulinemia.

Bruton X-linked agammaglobulinemia (XLA) is a phenotypically recessive genetic disorder of B lymphocyte development. Female carriers of XLA, although asymptomatic, have a characteristic B cell lineage-specific skewing of the pattern of X inactivation. Skewing apparently results from defective growth and maturation of B cell precursors bearing a mutant active X chromosome. In this study, carrier status was tested in 58 women from 22 families referred with a history of agammaglobulinemia. Primary carrier analysis to examine patterns of X inactivation in CD19+ peripheral blood cells (B lymphocytes) was conducted using quantitative PCR at the androgen-receptor locus. Obligate carriers of XLA demonstrated > 95% skewing of X inactivation in peripheral blood CD19+ cells but not in CD19- cells. Carrier status for mothers of isolated affected males could be assessed in 10 of 11 families: 7 women showed skewing, and 3 did not. Five carriers were found in six families in which there were no living affected males. Among all those tested, one individual's carrier status was considered to be indeterminate and five women were noninformative for the carrier test. Results obtained by the carrier test were congruent with linkage analysis (where applicable) using the RFLPs DXS178 and DXS94 and two newly developed polymorphic microsatellite markers, DXS178CA and DXS101AAT. Refinements in techniques for primary carrier testing and genetic mapping of XLA now make possible an ordered approach to diagnosis, prenatal diagnosis, and genetic counseling.