Anti-Interleukin-23 Autoantibodies in Adult-Onset Immunodeficiency.

BACKGROUND: Autoantibodies against interleukin-12 (anti-interleukin-12) are often identified in patients with thymoma, but opportunistic infections develop in only some of these patients. Interleukin-12 (with subunits p40 and p35) shares a common subunit with interleukin-23 (subunits p40 and p19). In a patient with disseminated Burkholderia gladioli infection, the identification of both anti-interleukin-23 and anti-interleukin-12 prompted further investigation. METHODS: Among the patients (most of whom had thymoma) who were known to have anti-interleukin-12, we screened for autoantibodies against interleukin-23 (anti-interleukin-23). To validate the potential role of anti-interleukin-23 with respect to opportunistic infection, we tested a second cohort of patients with thymoma as well as patients without either thymoma or known anti-interleukin-12 who had unusual infections. RESULTS: Among 30 patients with anti-interleukin-12 who had severe mycobacterial, bacterial, or fungal infections, 15 (50%) also had autoantibodies that neutralized interleukin-23. The potency of such neutralization was correlated with the severity of these infections. The neutralizing activity of anti-interleukin-12 alone was not associated with infection. In the validation cohort of 91 patients with thymoma, the presence of anti-interleukin-23 was associated with infection status in 74 patients (81%). Overall, neutralizing anti-interleukin-23 was detected in 30 of 116 patients (26%) with thymoma and in 30 of 36 patients (83%) with disseminated, cerebral, or pulmonary infections. Anti-interleukin-23 was present in 6 of 32 patients (19%) with severe intracellular infections and in 2 of 16 patients (12%) with unusual intracranial infections, including Cladophialophora bantiana and Mycobacterium avium complex. CONCLUSIONS: Among patients with a variety of mycobacterial, bacterial, or fungal infections, the presence of neutralizing anti-interleukin-23 was associated with severe, persistent opportunistic infections. (Funded by the National Institute of Allergy and Infectious Diseases and others.).

Functional neutrophil disorders: Chronic granulomatous disease and beyond.

Since their description by Metchnikoff in 1905, phagocytes have been increasingly recognized to be the entities that traffic to sites of infection and inflammation, engulf and kill infecting organisms, and clear out apoptotic debris all the while making antigens available and accessible to the lymphoid organs for future use. Therefore, phagocytes provide the gateway and the first check in host protection and immune response. Disorders in killing and chemotaxis lead not only to infection susceptibility, but also to autoimmunity. We aim to describe chronic granulomatous disease and the leukocyte adhesion deficiencies as well as myeloperoxidase deficiency and G6PD deficiency as paradigms of critical pathways.

Characterization of the antispike IgG immune response to COVID-19 vaccines in people with a wide variety of immunodeficiencies.

Research on coronavirus disease 2019 vaccination in immune-deficient/disordered people (IDP) has focused on cancer and organ transplantation populations. In a prospective cohort of 195 IDP and 35 healthy volunteers (HV), antispike immunoglobulin G (IgG) was detected in 88% of IDP after dose 2, increasing to 93% by 6 months after dose 3. Despite high seroconversion, median IgG levels for IDP never surpassed one-third that of HV. IgG binding to Omicron BA.1 was lowest among variants. Angiotensin-converting enzyme 2 pseudo-neutralization only modestly correlated with antispike IgG concentration. IgG levels were not significantly altered by receipt of different messenger RNA-based vaccines, immunomodulating treatments, and prior severe acute respiratory syndrome coronavirus 2 infections. While our data show that three doses of coronavirus disease 2019 vaccinations induce antispike IgG in most IDP, additional doses are needed to increase protection. Because of the notably reduced IgG response to Omicron BA.1, the efficacy of additional vaccinations, including bivalent vaccines, should be studied in this population.

Intestinal microbiome and metabolome signatures in patients with chronic granulomatous disease.

BACKGROUND: Chronic granulomatous disease (CGD) is caused by defects in any 1 of the 6 subunits forming the nicotinamide adenine dinucleotide phosphate oxidase complex 2 (NOX2), leading to severely reduced or absent phagocyte-derived reactive oxygen species production. Almost 50% of patients with CGD have inflammatory bowel disease (CGD-IBD). While conventional IBD therapies can treat CGD-IBD, their benefits must be weighed against the risk of infection. Understanding the impact of NOX2 defects on the intestinal microbiota may lead to the identification of novel CGD-IBD treatments. OBJECTIVE: We sought to identify microbiome and metabolome signatures that can distinguish individuals with CGD and CGD-IBD. METHODS: We conducted a cross-sectional observational study of 79 patients with CGD, 8 pathogenic variant carriers, and 19 healthy controls followed at the National Institutes of Health Clinical Center. We profiled the intestinal microbiome (amplicon sequencing) and stool metabolome, and validated our findings in a second cohort of 36 patients with CGD recruited through the Primary Immune Deficiency Treatment Consortium. RESULTS: We identified distinct intestinal microbiome and metabolome profiles in patients with CGD compared to healthy individuals. We observed enrichment for Erysipelatoclostridium spp, Sellimonas spp, and Lachnoclostridium spp in CGD stool samples. Despite differences in bacterial alpha and beta diversity between the 2 cohorts, several taxa correlated significantly between both cohorts. We further demonstrated that patients with CGD-IBD have a distinct microbiome and metabolome profile compared to patients without CGD-IBD. CONCLUSION: Intestinal microbiome and metabolome signatures distinguished patients with CGD and CGD-IBD, and identified potential biomarkers and therapeutic targets.

Small bowel disease prevalence on video capsule endoscopy in chronic granulomatous disease-associated inflammatory bowel disease.

BACKGROUND AND AIM: Patients with chronic granulomatous disease (CGD) may develop inflammatory bowel disease (IBD). Characterization of small bowel disease in this cohort is scarce. Here, we sought to determine the prevalence and characteristics of small bowel disease and evaluate the clinical utility of video capsule endoscopy (VCE) for its diagnosis. METHODS: A retrospective study was performed on patients with CGD who were evaluated for gastrointestinal disease with VCE as a part of ongoing natural history studies at a single academic center. VCEs were reviewed for inflammatory findings and severity of disease utilizing the Capsule Endoscopy Crohn's Disease Activity Index. Radiographic studies and endoscopies performed within 30 days of VCE were compared with small bowel inflammatory findings. RESULTS: Twenty-six VCEs corresponding to 25 patients were found. The majority of patients were male and White; mean age was 28 years old. The majority (85%) demonstrated presence of small bowel inflammatory findings on VCE including strictures, ulcers, erosions, and erythema. Duodenal and ileal inflammatory disease on endoscopy did not correlate with disease on VCE. Moderate-severe colonic disease correlated with moderate-severe disease on VCE. Radiography did not correlate with disease on VCE. Prolonged small bowel transit time correlated with moderate-severe small bowel disease. CONCLUSIONS: Small bowel IBD was highly prevalent in this cohort of patients with CGD. Limitations included small sample size. Given that radiology and duodenal/ileal disease did not correlate with VCE findings, VCE-driven investigation of small bowel disease should be considered in patients with CGD-associated IBD, particularly those with colonic disease.

Self-extinguishing relay waves enable homeostatic control of human neutrophil swarming.

Neutrophils exhibit self-amplified swarming to sites of injury and infection. How swarming is controlled to ensure the proper level of neutrophil recruitment is unknown. Using an ex vivo model of infection, we find that human neutrophils use active relay to generate multiple pulsatile waves of swarming signals. Unlike classic active relay systems such as action potentials, neutrophil swarming relay waves are self-extinguishing, limiting the spatial range of cell recruitment. We identify an NADPH-oxidase-based negative feedback loop that is needed for this self-extinguishing behavior. Through this circuit, neutrophils adjust the number and size of swarming waves for homeostatic levels of cell recruitment over a wide range of initial cell densities. We link a broken homeostat to neutrophil over-recruitment in the context of human chronic granulomatous disease.

Chromosomal microarray analysis supplements exome sequencing to diagnose children with suspected inborn errors of immunity.

Purpose: Though copy number variants (CNVs) have been suggested to play a significant role in inborn errors of immunity (IEI), the precise nature of this role remains largely unexplored. We sought to determine the diagnostic contribution of CNVs using genome-wide chromosomal microarray analysis (CMA) in children with IEI. Methods: We performed exome sequencing (ES) and CMA for 332 unrelated pediatric probands referred for evaluation of IEI. The analysis included primary, secondary, and incidental findings. Results: Of the 332 probands, 134 (40.4%) received molecular diagnoses. Of these, 116/134 (86.6%) were diagnosed by ES alone. An additional 15/134 (11.2%) were diagnosed by CMA alone, including two likely de novo changes. Three (2.2%) participants had diagnostic molecular findings from both ES and CMA, including two compound heterozygotes and one participant with two distinct diagnoses. Half of the participants with CMA contribution to diagnosis had CNVs in at least one non-immune gene, highlighting the clinical complexity of these cases. Overall, CMA contributed to 18/134 diagnoses (13.4%), increasing the overall diagnostic yield by 15.5% beyond ES alone. Conclusion: Pairing ES and CMA can provide a comprehensive evaluation to clarify the complex factors that contribute to both immune and non-immune phenotypes. Such a combined approach to genetic testing helps untangle complex phenotypes, not only by clarifying the differential diagnosis, but in some cases by identifying multiple diagnoses contributing to the overall clinical presentation.

Multiomics integration of 22 immune-mediated monogenic diseases reveals an emergent axis of human immune health.

Monogenic diseases are often studied in isolation due to their rarity. Here we utilize multiomics to assess 22 monogenic immune-mediated conditions with age- and sex-matched healthy controls. Despite clearly detectable disease-specific and "pan-disease" signatures, individuals possess stable personal immune states over time. Temporally stable differences among subjects tend to dominate over differences attributable to disease conditions or medication use. Unsupervised principal variation analysis of personal immune states and machine learning classification distinguishing between healthy controls and patients converge to a metric of immune health (IHM). The IHM discriminates healthy from multiple polygenic autoimmune and inflammatory disease states in independent cohorts, marks healthy aging, and is a pre-vaccination predictor of antibody responses to influenza vaccination in the elderly. We identified easy-to-measure circulating protein biomarker surrogates of the IHM that capture immune health variations beyond age. Our work provides a conceptual framework and biomarkers for defining and measuring human immune health.

Gastrointestinal and Hepatic Manifestations of Chronic Granulomatous Disease.

Chronic granulomatous disease (CGD) is a rare inborn error of immunity, resulting from a defect in nicotinamide adenine dinucleotide phosphate oxidation and decreased production of phagocyte reactive oxygen species. The main clinical manifestations are recurrent infections and chronic inflammatory disorders. Current approaches to management include antimicrobial prophylaxis and control of inflammatory complications. Hematopoietic stem cell transplantation or gene therapy can provide definitive treatment. Gastrointestinal and hepatic manifestations are common in CGD and include structural changes, dysmotility, CGD-associated inflammatory bowel disease, liver abscesses, and noncirrhotic portal hypertension. The findings can be heterogeneous, and the management is complex in light of the underlying immune dysfunction. This review describes the various clinical findings and the latest studies in management of gastrointestinal and hepatic manifestations in CGD, as well as the management experience at the National Institutes of Health.

Human Dectin-1 deficiency impairs macrophage-mediated defense against phaeohyphomycosis.

Subcutaneous phaeohyphomycosis typically affects immunocompetent individuals following traumatic inoculation. Severe or disseminated infection can occur in CARD9 deficiency or after transplantation, but the mechanisms protecting against phaeohyphomycosis remain unclear. We evaluated a patient with progressive, refractory Corynespora cassiicola phaeohyphomycosis and found that he carried biallelic deleterious mutations in CLEC7A encoding the CARD9-coupled, ß-glucan-binding receptor, Dectin-1. The patient's PBMCs failed to produce TNF-α and IL-1ß in response to ß-glucan and/or C. cassiicola. To confirm the cellular and molecular requirements for immunity against C. cassiicola, we developed a mouse model of this infection. Mouse macrophages required Dectin-1 and CARD9 for IL-1ß and TNF-α production, which enhanced fungal killing in an interdependent manner. Deficiency of either Dectin-1 or CARD9 was associated with more severe fungal disease, recapitulating the human observation. Because these data implicated impaired Dectin-1 responses in susceptibility to phaeohyphomycosis, we evaluated 17 additional unrelated patients with severe forms of the infection. We found that 12 out of 17 carried deleterious CLEC7A mutations associated with an altered Dectin-1 extracellular C-terminal domain and impaired Dectin-1-dependent cytokine production. Thus, we show that Dectin-1 and CARD9 promote protective TNF-α- and IL-1ß-mediated macrophage defense against C. cassiicola. More broadly, we demonstrate that human Dectin-1 deficiency may contribute to susceptibility to severe phaeohyphomycosis by certain dematiaceous fungi.

Disseminated Mycoplasma Orale Infection in Patients With Phosphoinositide-3-Kinase Regulatory Subunit 1 Mutations.

Mycoplasma orale is a rare cause of invasive infection in immunodeficient hosts. Phosphatidylinositol 3-kinase, regulatory subunit 1 (PI3KR1) mutations predispose patients to sinopulmonary infections, alongside bronchiectasis autoimmunity and lymphoproliferation. We report 2 cases of PI3KR1 deficiency with invasive M orale and effective treatment options.

Neutralizing GM-CSF autoantibodies in pulmonary alveolar proteinosis, cryptococcal meningitis and severe nocardiosis.

BACKGROUND: Anti GM-CSF autoantibodies (aAb) have been related to acquired pulmonary alveolar proteinosis (PAP) and described in cases of severe infections such as cryptococcosis and nocardiosis in previously healthy subjects. Whether there are different anti-GM-CSF autoantibodies corresponding to these phenotypes is unclear. Therefore, we examined anti-GM-CSF autoantibodies to determine whether amount or neutralizing activity could distinguish between groups. METHODS: Plasma samples gathered in the National Institute of Health from patients with anti GM-CSF aAb and either PAP (n = 15), cryptococcal meningitis (n = 15), severe nocardiosis (n = 5) or overlapping phenotypes (n = 6) were compared. The relative amount of aAb was assessed using a particle-based approach, reported as a mouse monoclonal anti-human GM-CSF as standard curve and expressed in an arbitrary Mouse Monoclonal Antibody Unit (MMAU). The neutralizing activity of the plasma was assessed by inhibition of GM-CSF-induced intracellular phospho-STAT5 (pSTAT5) in monocytes. RESULTS: Anti-GM-CSF aAb relative amounts were higher in PAP patients compared to those with cryptococcosis (mean 495 ± 464 MMAU vs 197 ± 159 MMAU, p = 0.02); there was no difference with patients with nocardiosis (430 ± 493 MMAU) nor between the two types of infections. The dilution of plasma resulting in 50% inhibition of GM-CSF-induced pSTAT5 (approximate IC50) did not vary appreciably across groups of patients (1.6 ± 3.1%, 3.9 ± 6% and 1.8 ± 2.2% in PAP patients, cryptococcosis and nocardiosis patients, respectively). Nor was the concentration of GM-CSF necessary to induce 50% of maximal GM-CSF-induced pSTAT5 in the presence of 10 MMAU of anti-GM-CSF aAb (EC50). When studying longitudinal samples from patients with PAP or disseminated nocardiosis, the neutralizing effect of anti-GM-CSF aAb was relatively constant over time despite targeted treatments and variations in aAb levels. CONCLUSIONS: Despite different clinical manifestations, anti-GM-CSF antibodies were similar across PAP, cryptococcosis and nocardiosis. Underlying host genetics and functional analyses may help further differentiate the biology of these conditions.

Immunogenetics associated with severe coccidioidomycosis.

Disseminated coccidioidomycosis (DCM) is caused by Coccidioides, pathogenic fungi endemic to the southwestern United States and Mexico. Illness occurs in approximately 30% of those infected, less than 1% of whom develop disseminated disease. To address why some individuals allow dissemination, we enrolled patients with DCM and performed whole-exome sequencing. In an exploratory set of 67 patients with DCM, 2 had haploinsufficient STAT3 mutations, and defects in ß-glucan sensing and response were seen in 34 of 67 cases. Damaging CLEC7A and PLCG2 variants were associated with impaired production of ß-glucan-stimulated TNF-α from PBMCs compared with healthy controls. Using ancestry-matched controls, damaging CLEC7A and PLCG2 variants were overrepresented in DCM, including CLEC7A Y238* and PLCG2 R268W. A validation cohort of 111 patients with DCM confirmed the PLCG2 R268W, CLEC7A I223S, and CLEC7A Y238* variants. Stimulation with a DECTIN-1 agonist induced DUOX1/DUOXA1-derived hydrogen peroxide [H2O2] in transfected cells. Heterozygous DUOX1 or DUOXA1 variants that impaired H2O2 production were overrepresented in discovery and validation cohorts. Patients with DCM have impaired ß-glucan sensing or response affecting TNF-α and H2O2 production. Impaired Coccidioides recognition and decreased cellular response are associated with disseminated coccidioidomycosis.

Rank correlation inferences for clustered data with small sample size.

This paper develops methods to test for associations between two variables with clustered data using a U-Statistic approach with a second-order approximation to the variance of the parameter estimate for the test statistic. The tests that are presented are for clustered versions of: Pearsons χ 2 test, the Spearman rank correlation and Kendall's τ for continuous data or ordinal data and for alternative measures of Kendall's τ that allow for ties in the data. Shih and Fay use the U-Statistic approach but only consider a first-order approximation. The first-order approximation has inflated significance level in scenarios with small sample sizes. We derive the test statistics using the second-order approximations aiming to improve the type I error rates. The method applies to data where clusters have the same number of measurements for each variable or where one of the variables may be measured once per cluster while the other variable may be measured multiple times. We evaluate the performance of the test statistics through simulation with small sample sizes. The methods are all available in the R package cluscor.

IFNγ regulates NAD+ metabolism to promote the respiratory burst in human monocytes.

Interferon γ (IFNγ) is an essential and pleiotropic activator of human monocytes, but little is known about the changes in cellular metabolism required for IFNγ-induced activation. We sought to elucidate the mechanisms by which IFNγ reprograms monocyte metabolism to support its immunologic activities. We found that IFNγ increased oxygen consumption rates (OCR) in monocytes, indicative of reactive oxygen species generation by both mitochondria and nicotinamide adenine dinucleotide phosphate (NADPH) oxidase. Transcriptional profiling revealed that this oxidative phenotype was driven by IFNγ-induced reprogramming of NAD+ metabolism, which is dependent on nicotinamide phosphoribosyltransferase (NAMPT)-mediated NAD+ salvage to generate NADH and NADPH for oxidation by mitochondrial complex I and NADPH oxidase, respectively. Consistent with this pathway, monocytes from patients with gain-of-function mutations in STAT1 demonstrated higher-than-normal OCR, whereas chemical or genetic disruption of mitochondrial complex I (rotenone treatment or Leigh syndrome patient monocytes) or NADPH oxidase (diphenyleneiodonium treatment or chronic granulomatous disease [CGD] patient monocytes) reduced OCR. Interestingly, inhibition of NAMPT in healthy monocytes completely abrogated the IFNγ-induced oxygen consumption, comparable to levels observed in CGD monocytes. These data identify an IFNγ-induced, NAMPT-dependent, NAD+ salvage pathway that is critical for IFNγ activation of human monocytes.

Febrile neutropenia management and outcomes in hematopoietic cell transplantation for chronic granulomatous disease.

OBJECTIVE: We analyzed events and therapies related to febrile neutropenia in patients receiving hematopoietic cell transplantation (HCT) for chronic granulomatous disease (CGD). METHODS: Three protocols for HCT were used to extract the relation between conditioning and infectious complications during transplantation for CGD, especially the relation of fever and neutropenia to microbiological events and antibiotic therapy. RESULTS: Sixty-nine recipients received either reduced intensity conditioning with matched related or unrelated donors or conditioning specific to haploidentical-related donors utilizing posttransplant cyclophosphamide. Fever prior to neutropenia was common (52) and in eight recipients, Gram negative bacterial infection occurred prior to neutropenia, and in nine during neutropenia. Alemtuzumab as conditioning was associated with preneutropenic infection. Empiric therapy (noncarbapenem) by institutional guideline was given in 40. Carbapenems were given before neutropenia (8) or as empiric therapy in neutropenia (18), or a switch to a carbapenem (n = 22) occurred in 48 cases. No deaths related to infection associated with neutropenia occurred. CONCLUSION: The management of febrile neutropenia in HCT for CGD led to no deaths related to infection associated with neutropenia. Bacteremias occurred both prior to neutropenia and during neutropenia. Bacteria isolated may have represented the recrudescence of prior infection, representing the population transplanted and the platform for HCT. The treatment of prior infections may have had an influence on the necessity of carbapenem use as either empiric or directed therapy for bacterial infections.

Hyperinflammatory Syndromes After Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) Messenger RNA vaccination in Individuals With Underlying Immune Dysregulation.

The development of effective severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) messenger RNA (mRNA) vaccines has been a significant accomplishment. Adverse events are extremely rare, but continued surveillance is important, especially in at-risk populations. In 5 patients with preexisting immune dysregulation, hyperinflammatory syndromes, including hemophagocytic lymphohistiocytosis, developed after SARS-CoV-2 mRNA vaccination. Early recognition of this rare condition is essential.

Gastrointestinal Computed Tomography Findings in Chronic Granulomatous Disease with Subgroup Clinicopathologic Analysis.

BACKGROUND: Chronic granulomatous disease (CGD) is a rare primary immunodeficiency which can lead to gastrointestinal (GI) complications including inflammatory bowel disease. Radiographic findings in this cohort have not been well described. AIMS: To describe the frequency and spectrum of gastrointestinal abnormalities seen on computed tomography (CT) in patients with CGD and determine whether radiography was predictive of endoscopic or histopathologic inflammatory findings. METHODS: A retrospective review was conducted on 141 consecutive CGD patients seen at the National Institutes of Health between 1988 and 2011. All corresponding CTs were reviewed for gastrointestinal abnormalities including wall thickening. Endoscopic and histopathologic findings were reviewed in subjects with documented endoscopy within 30 days of an imaging study. Findings were compared between patients with and without wall thickening on CT to determine whether bowel wall thickening was predictive of endoscopic or histologic inflammatory findings. RESULTS: Two hundred and ninety-two CTs were reviewed. GI wall thickening was present on CT in 61% of patients (n = 86). Among a subgroup of 20 patients who underwent endoscopy at the time of their imaging, there was a statistically significant correlation between radiographic gastrointestinal wall thickening and endoscopic inflammation in the same intestinal segment (p = 0.035). Additionally, there was a significant correlation between radiographic gastrointestinal wall thickening and inflammatory features on histopathology (p = 0.02). CONCLUSIONS: GI abnormalities are commonly observed on CT in CGD patients. Bowel wall thickening correlates with endoscopic and histopathologic evidence of inflammation. These findings may be used to better facilitate directed endoscopic assessment and histopathologic sampling in patients with CGD.

Safety and Efficacy of Ustekinumab in the Inflammatory Bowel Disease of Chronic Granulomatous Disease.

Chronic granulomatous disease (CGD) is a rare primary immunodeficiency caused by mutations encoding the NADPH oxidase complex.1 Those affected are at increased risk of bacterial and fungal infections and require antimicrobial prophylaxis. Dysregulated inflammation may cause inflammatory bowel disease (IBD), termed CGD-associated IBD or CGD colitis, a distinct entity from Crohn's disease (CD) or ulcerative colitis (UC).

Hematologically important mutations: The autosomal forms of chronic granulomatous disease (third update).

Chronic granulomatous disease (CGD) is an immunodeficiency disorder affecting about 1 in 250,000 individuals. CGD patients suffer from severe, recurrent bacterial and fungal infections. The disease is caused by mutations in the genes encoding the components of the leukocyte NADPH oxidase. This enzyme produces superoxide, which is subsequently metabolized to hydrogen peroxide and other reactive oxygen species (ROS). These products are essential for intracellular killing of pathogens by phagocytic leukocytes (neutrophils, eosinophils, monocytes and macrophages). The leukocyte NADPH oxidase is composed of five subunits, four of which are encoded by autosomal genes. These are CYBA, encoding p22phox, NCF1, encoding p47phox, NCF2, encoding p67phox and NCF4, encoding p40phox. This article lists all mutations identified in these genes in CGD patients. In addition, cytochrome b558 chaperone-1 (CYBC1), recently recognized as an essential chaperone protein for the expression of the X-linked NADPH oxidase component gp91phox (also called Nox2), is encoded by the autosomal gene CYBC1. Mutations in this gene also lead to CGD. Finally, RAC2, a small GTPase of the Rho family, is needed for activation of the NADPH oxidase, and mutations in the RAC2 gene therefore also induce CGD-like symptoms. Mutations in these last two genes are also listed in this article.