Major histocompatibility complex class II expression deficiency caused by a RFXANK founder mutation: a survey of 35 patients.

Abstract

Inherited deficiency of major histocompatibility complex (MHC) class II molecules impairs antigen presentation to CD4(+) T cells and results in combined immunodeficiency (CID). Autosomal-recessive mutations in the RFXANK gene account for two-thirds of all cases of MHC class II deficiency. We describe here the genetic, clinical, and immunologic features of 35 patients from 30 unrelated kindreds from North Africa sharing the same RFXANK founder mutation, a 26-bp deletion called I5E6-25_I5E6 + 1), and date the founder event responsible for this mutation in this population to approximately 2250 years ago (95% confidence interval [CI]: 1750-3025 years). Ten of the 23 patients who underwent hematopoietic stem cell transplantation (HSCT) were cured, with the recovery of almost normal immune functions. Five of the patients from this cohort who did not undergo HSCT had a poor prognosis and eventually died (at ages of 1-17 years). However, 7 patients who did not undergo HSCT (at ages of 6-32 years) are still alive on Ig treatment and antibiotic prophylaxis. RFXANK deficiency is a severe, often fatal CID for which HSCT is the only curative treatment. However, some patients may survive for relatively long periods if multiple prophylactic measures are implemented.

Full text

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doi:10.1182/blood-2011-05-352716
Prepublished online September 8, 2011;




Stéphane Blanche, Laurent Abel, Jean-Laurent Casanova, Alain Fischer and Capucine Picard
Bousfiha, Yves Levy, Barbara Lisowska-Grospierre, Danielle Canioni, Julie Bruneau, Marianne Debré,
Monia Ouederni, Quentin B Vincent, Pierre Frange, Fabien Touzot, Sami Scerra, Mohamed Bejaoui, Aziz

founder mutation: a survey of 35 patients
RFXANK
a
Major histocompatibility complex class II expression deficiency caused by
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Major histocompatibility complex class II expression deficiency
caused by a RFXANK founder mutation – a survey of 35 patients
Monia Ouederni1, Quentin B Vincent2,3, Pierre Frange1, Fabien Touzot1, Sami Scerra4,
Mohamed Bejaoui5, Aziz Bousfiha6, Yves Levy4, Barbara Lisowska-Grospierre7 ,
Danielle Canioni2,8, Julie Bruneau2,8, Marianne Debré1, Stéphane Blanche1,2,
Laurent Abel 2,3, Jean-Laurent Casanova1,2,3,9, Alain Fischer1,2,6, Capucine Picard2,3,10

1. Pediatric Hematology-Immunology Unit, Assistance Publique Hôpitaux de Paris
(APHP), Necker Hospital, Paris, France, EU.

2. Paris Descartes University, Sorbonne Paris Cité, France, EU.

3. Laboratory of Human Genetics of Infectious Diseases, Necker Branch, INSERM
U980, Paris, France, EU.

4. Immunology Clinical Unit, Assistance Publique Hôpitaux de Paris (APHP), Mondor
Hospital, Creteil, France.

5. The National Bone Marrow Transplantation Center, Jebel Lakhdar, 1006 Saadoun
Tunis, Tunisia.

6. Clinical Immunology Unit, Department of Pediatrics, CHU Ibn Rochd, King Hassan II
University, Casablanca, Morocco.

7. INSERM U768, Paris, France, EU.

8. Pathology Department, Assistance Publique Hôpitaux de Paris (APHP), Necker
Hospital, Paris, France, EU.

9. St. Giles Laboratory of Human Genetics of Infectious Diseases, Rockefeller Branch,
The Rockefeller University, New York, USA.

10. Study Center for Primary Immunodeficiencies, Assistance Publique Hôpitaux de Paris
(APHP), Necker Hospital, Paris, France, EU.



Correspondence: Capucine Picard, Study Center for Primary Immunodeficiencies,
Assistance Publique Hôpitaux de Paris, Necker Hospital, 149 rue de Sèvres, 75015 Paris,
France, EU telephone: 33 1 44 49 50 88 27, fax: 33 1 42 73 06 40, E-mail:
capucine.picard@inserm.fr

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ABSTRACT
Inherited deficiency of major histocompatibility complex (MHC) class II molecules
impairs antigen presentation to CD4+ T cells and results in combined immunodeficiency
(CID). Autosomal recessive mutations in the RFXANK gene account for two thirds of all
cases of MHC class II deficiency. We describe here the genetic, clinical and immunological
features of 35 patients from 30 unrelated kindreds from North Africa sharing the same
RFXANK founder mutation, a 26 bp deletion (named I5E6-25_I5E6+1), and date the founder
event responsible for this mutation in this population to about 2,250 years ago (95%, CI:
1,750-3,025 years). Ten of the 23 patients who underwent hematopoietic stem cell
transplantation (HSCT) were cured, with the recovery of almost normal immune functions.
Five of the patients from this cohort that did not undergo HSCT had a poor prognosis and
eventually died (at ages of 1 to 17 years). However, 7 patients that did not undergo HSCT
(aged 6 to 32 years) are still alive on immunoglobulin treatment and antibiotic prophylaxis.
RFXANK deficiency is, thus, a severe, often fatal CID, for which HSCT is the only curative
treatment. However, some patients may survive for relatively long periods, provided that
multiple prophylactic measures are implemented.









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INTRODUCTION
Major histocompatibility complex (MHC) class II expression deficiency is a rare
primary immunodeficiency (PID) (MIM 209920). It is inherited as an autosomal recessive
trait. The lack of MHC class II expression leads to impaired antigen presentation by HLA-
DR, DQ and DP molecules on antigen-presenting cells (APC), such as dendritic cells and
macrophages 1. This leads to defective CD4 T-cell development, function and a lack of T
helper cell-dependent antibody production by B cells. Hypogammaglobulinemia and low CD4
T cell counts are found in most patients 2. Impaired T-cell and B-cell immunity results in
susceptibility to a broad range of viral, bacterial, fungal and protozoan infections 3-5.
Infections generally occur in the first year of life and involve the respiratory and
gastrointestinal tracts 2,4,5. Respiratory insufficiency, failure to thrive and organ dysfunctions
ensue, frequently causing death in childhood. Hematopoietic stem cell transplantation (HSCT)
is the only available curative treatment, despite the high risk of toxicity in this particular
setting 4,6-12.

The MHC locus itself is intact in patients with MHC class II expression deficiency.
Impaired transcription of the MHC class II genes has been shown to account for the lack of
expression of DR, DP and DQ MHC class II proteins on APC1. Somatic fusion experiments
led to the description of four complementation groups (A, B, C and D) 1. Four disease-causing
genes were subsequently identified and shown to encode regulatory factors controlling the
transcription of MHC class II genes: CIITA for group A (MIM 600005) 13, RFXANK for group
B (MIM 603200) 14, RFX5 for group C (MIM 601863) 15 and RFXAP for group D (MIM
601861) 16. The RFXANK, RFX5 and RFXAP proteins are three subunits of the ubiquitously
expressed RFX complex, which binds directly to the promoters of all MHC class II genes and
associates with other pleiotropic factors to form the MHC class II enhanceosome. CIITA is an
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inducible factor that controls MHC class II expression by binding the RFX complex and
triggering transcription 17.

Over 150 patients with MHC class II deficiency have been reported to date 2-12,14,15,18-
39. About two thirds of these patients have complementation group B deficiencies, most of the
families being of North African descent (Algeria, Morocco and Tunisia) 2,5,7,9,10,14,18,27-
29,31,32,37. A recurrent mutation of the RFXANK gene, a 26 bp deletion at the boundary between
intron 5 and exon 6 named I5E6-25_I5E6+1 (also known as 752delG-25) has been found in
more than 90% of North African families and more than half of all families with MHC II
deficiency 28,37. Eight other mutations in RFXANK have been described in patients of different
ethnic origins (Netherlands, Italy, France/Spain, Tunisia, Turkey, Saudi Arabia) 14,27-29,31,32,40.
Thirty families with group B MHC class II deficiency investigated at our center carry the
I5E6-25_I5E6+1 mutation. We investigated whether this homozygous genotype conferred a
homogeneous disease phenotype, by studying the clinical and immunological features of a
group of 35 North African patients originating from 30 families and carrying a homozygous
I5E6-25_I5E6+1 deletion.

METHODS
Patients
Thirty-five patients with MHC class II expression deficiency, born between 1970 and
2008, were referred to Necker Hospital in Paris between 1977 and 2010. Diagnosis was based
on personal and family history, and clinical and immunological signs. Informed consent for
participation in the study was obtained from adult patients or from the parents of children in
accordance with the Declaration of Helsinki. All studies were approved by the institutional
review board of Necker Hospital. A homozygous I5E6-25_I5E6+1 deletion in the RFXANK
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gene was clearly demonstrated in all the patients studied 28. This 26 bp deletion encompassing
the intron 5 - exon 6 boundary was detected by polymerase chain reaction (PCR)
amplification with the following primers: Forward: TTg gCA gCA CTg ATA ggg g and
Reverse: CCA gCA gAC ACA gCC AAA AC.

Dating of the mutation
Founder effect analyses were carried out on a subset of 24 available, apparently
unrelated patients (19 from Algeria, 2 from Morocco and 3 from Tunisia). Genotypes were
obtained for more than 250,000 SNPs from the Affymetrix GeneChip Human Mapping 250K
Nsp Array. SNPs with a 100% call rate were scanned for continuous stretches of
homozygosity up- and downstream from the RFXANK locus on chromosome 19p12. Pairwise
comparisons within each mutation group revealed the limits of the longest shared haplotype
and the positions of subsequent recombination breakpoints. The likelihood-based ESTIAGE
method was used to estimate the age of the most recent common ancestor (MRCA) for each
mutation from the observed shared haplotypes and the recombination rates and haplotype
frequencies provided by the HapMap Project 41-43.

Immunological investigations
Lymphocyte subsets were evaluated by routine flow cytometry, with the norms
described by Shearer et al.44. Serum levels of the IgM, A and G and subclasses were assessed
by standard nephelometry techniques. Total IgG antibody (Ab) levels against tetanus toxoid
and diphtheria, titers of antibodies against viruses and bacteria, including multiple
pneumococcal serotypes (23 serotypes), and levels of IgG against H. influenzae PRP antigens
were assessed by standard ELISA techniques. of the expression of MHC class II and class I
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molecules by T and B cells was assessed and in vitro lymphocyte proliferation assays were
carried out with cells from the patients, as previously described2,7.

RESULTS
Epidemiological features and founder effect analysis
We studied 35 patients (16 male and 19 female patients) from 30 kindreds with RFX-
ANK deficiency. This series includes 22 patients described in previous reports (belonging to
19 kindreds) 2,7,10,28,31 and 13 newly diagnosed patients (belonging to 11 kindreds). All of the
patients originated from North Africa (Algeria (71%), Morocco (14%) and Tunisia (14%))
(Table 1). Most patients and their families were living in France, with the exception of three
patients (P28, P29 and P32) living in their countries of origin. In all 30 probands, diagnosis
was based on detection of the homozygous 26 bp deletion (I5E6-25_I5E6+1) in RFXANK
accompanied by defective MHC class II antigen expression on monocytes and B cells (Figure
1a and supplementary figure 1). Five relatives were also found to be homozygous for this
mutation. The parents were consanguineous in 27 of the 30 kindreds. In the remaining three
kindreds, the parents originated from the same country but were not known to be
consanguineous. There were 23 sporadic cases (65%) and 12 (35%) familial cases (7
kindreds). An analysis of Affymetrix 250K Nsp Array data showed that patients carrying the
I5E6-25_I5E6+1 deletion had a common homozygous haplotype around the RFXANK gene.
The largest common haplotype identified was that between patients P25 and P28, extending
over 3.7 megabases (corresponding to 160 SNPs). The smallest common haplotype was that
between P33 and P34, encompassing 470 kb (corresponding to 11 SNPs) (Figure 1b). The
ESTIAGE method estimated the age of the MRCA to 90 generations (95% CI: 68-121
generations). Assuming, a generation time of 25 years, the MRCA of the patients therefore
lived 2,250 years ago (1,700-3,025 years).
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Immunological features
All 35 patients displayed a total absence of MHC class II (HLA-DR) molecule
expression on monocytes (CD14+). In 25 of the 35 patients, no MHC class II molecules were
detected on activated T (CD3+) and B (CD19+) cells either. However, 10 of the 35 patients
(29%) had residual levels of MHC class II molecules on B cells, generally after an infectious
episode (n=8). The percentage of positive cells and the density of MHC class II molecules
was nonetheless lower on positive B-cell populations (less than 6 % of cells displaying MHC
class II expression in nine patients, with a mean of fluorescence intensity (MFI) two orders of
magnitude lower than for the control) than on B cells from healthy controls. One patient (P32)
with concomitant viral infection displayed MHC class II expression on 43% of B cells
(CD19+), with a MFI two orders of magnitude lower than for the control (supplementary
figure 1). The level of HLA class I molecule expression was lower than normal, to various
extents, in 24 cases.

Low absolute CD4+ T-cell counts were found in 33 of the 35 patients (94%) (Figure
2a). CD4+ T-cell count were below 0.5 x 109 cells/l for 22 patients (63%) between the ages of
1 month and 15 years. The five patients studied showed variable decreases in the percentage
of naïve CD4+ T cells. CD8+ T-cell counts were low in eight patients (23%) and high in
another nine patients (26%) (Figure 2b). NK cell (CD3-CD16+ CD56+) counts were low in
three of the 12 patients tested (25%). In vitro proliferative responses of T cells to mitogens
were normal for all patients, but no proliferation in response to antigens was observed. B-cell
immunity was also profoundly impaired. Nineteen of the 24 patients studied (79%) had
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normal numbers of circulating B lymphocytes, whereas the other five (21%) had fewer
circulating B lymphocytes than normal. Before the initiation of immunoglobulin (IgG)
substitution therapy, 27 of 32 patients (84%) had low levels of IgG, 17 had low levels of IgA
(53%) and 21 had low levels of IgM (65%). High IgM levels were found in three patients
(9%). Three of the five patients with normal IgG levels were too young for the assessment of
IgG production, and the other two were only one and two years old. All five patients (P6, P8,
P14, P32 and P35) who had reached puberty had hypogammaglobulinemia. No antibodies
against viruses, bacteria and protein vaccines could be detected in the 20 patients tested.
However, three patients developed antibodies against Streptococcus pneumoniae, probably
reflecting T-independent B-cell immunity: P1 after an acute episode of pneumonia episode
and P34 and P35, four years after immunization with a non conjugated anti-pneumococcal
vaccine. Immunological data are summarized in Table 2.

Clinical features
Median follow-up was five years (range: birth to 32 years) at the time of this report.
Infections were the most prominent clinical feature of the disease, with all patients having had
multiple infections. Mean age at first infection was 4.2 months (range: 1 to 14 months). The
first clinical sign of the disease was gastrointestinal infection (15 cases, 43%) or pulmonary
infection (12 cases, 34%) in most patients. The median age at diagnosis of MHC class II
deficiency was 21 months (range: from birth to 15 years). Supportive treatment based on
intravenous or subcutaneous IgG therapy and antibiotic prophylaxis (trimethoprim-
sulfamethoxazole) was initiated after diagnosis, in all patients. Regular administration of IgG
markedly decreased the frequency of infectious episodes. The most common clinical signs in
the patients are listed in Table 1.

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Gastrointestinal and hepatic signs
Thirty-one patients (89%) had gastrointestinal infections. Diarrhea began during the
first year of life and became protracted after two years in 22 of the patients (63%). All
patients over the age of five years that had not undergone HSCT suffered from protracted (or
recurrent) diarrhea. Failure to thrive (SDS of -5 to -2 at ages of 3 months to 10 years) was
observed in 28 patients (80%). Histological study of intestinal mucosa was performed in 14
cases. In 10 patients, this study revealed various degrees of villous atrophy, frequently
associated with intraepithelial infiltration by lymphocytes, and infiltration by eosinophils,
macrophages and plasma cells in the lamina propria, with no evidence of MHC class II
expression in comparison with one healthy control (Figure 3 a-f). The bacteria most
frequently isolated from stools during episodes of diarrhea included Gram-positive
(Staphylococcus aureus (n=5), Enterococcus spp. (n=4)) and Gram-negative (Escherichia coli
(n=5), Klebsiella pneumonia (n=5), Campylobacter jejuni (n=7), Proteus mirabilis (n=4),
Pseudomonas aeruginosa (n=2), Salmonella enteritidis (n=8), Shigella spp. (n=1),
Clostridium difficile (n=1)) bacteria. Viral infections were also a frequent cause of recurrent
diarrhea (the viruses detected included adenovirus (n=7), rotavirus (n=1), cytomegalovirus
(CMV) (n=1) and enterovirus (n=8) (in inoculated patient with attenuated Poliovirus vaccine
(n=2)). Thirteen patients (37%) displayed chronic oral thrush complicated by esophageal
candidiasis in one patient (P14). Cryptosporidium parvum was recurrently found in the stools
of 12 patients (34%) between the ages of 7 months and 10 years. Five patients displayed
Giardia intestinalis colonization between the ages of one and seven years. Hepatic
abnormalities were also frequent, and 16 patients (46%) had hepatomegaly and/or high serum
levels of liver enzymes. Beyond the age of five years, only three of the 12 patients that did not
undergo HSCT (P6, P29 and P35) had no detectable liver disease. Patient P31 developed
CMV hepatitis at the age of 18 months. In five patients (P1, P2, P3, P8 and P32), hepatic
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involvement resulted from cholangitis caused by C. parvum, diagnosed on the basis of
ultrasound and/or the results of biliary catheterization. Three of the five patients (P1, P2 and
P8) died from liver failure at the ages of 4, 10 and 17 years, respectively.
Respiratory tract infections
Recurrent pneumonia was reported in 29 patients (83%), and occurred before the age
of one year in 22 of these patients (76%). Eight patients (23%) developed acute hypoxic
pneumonia requiring mechanical ventilation between the ages of seven months and five years
(before the initiation of IgG substitution in six of these cases). The pneumonia was caused by
Pneumocystis jirovicii (n=4), CMV (n=2) or adenovirus (n=2) infection and was fatal in one
case (P7). P. jirovicii was detected in three other patients with mild symptoms. Respiratory
syncytial virus (RSV) was identified of two patients (P33 and P22). Ten patients (28%)
developed documented bacterial pneumonia between the age of seven months and 29 years,
caused by S. pneumoniae (n=2), H. influenza (n=4, associated with M. catarrhalis in one
case), S. aureus (n=3, associated with Proteus mirabilis in one case), K. pneumoniae (n=1)
and Mycoplasma pneumonia (n=1). In these ten patients, the incidence of bacterial pneumonia
was 3/100 patients.year in the absence of IgG treatment and 0.6/100 patients.year with IgG
treatment. Ten patients (29%) developed chronic bronchopneumopathy with bronchiectasis
between the ages of one and 12 years (mean 4.5 years). Sputum cultures were positive for M.
catarrhalis (7 cases), S. pneumoniae (8 cases), S. aureus (3 cases), H influenzae (8 cases), P.
mirabilis (1 case), Klebsiella oxytoca (1 case), K. pneumoniae (1 case), P. aeruginosa (3
cases) and M. pneumonia (1 case). Chronic sinusitis was documented in three patients over
the age of three years (P1, P8 and P30). Sixteen patients (46%) had recurrent acute otitis
media beginning before the age of two years, and requiring a trans-tympanic drain in three
patients (P23, P30 and P35). H. influenza was responsible for these infections in three
patients, but the causal pathogens were not documented in the other cases.
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Other infections
Seven patients were vaccinated with BCG and none developed BCGitis. Five patients (14%)
had acute pyelonephritis caused by E. coli (n=2), K. pneumoniae (n=2) or P. aeruginosa
(n=1). Eleven patients (31%) had sepsis due to S. aureus (n=2), S. epidermidis (n=3), S.
pneumonia (n=3) and P. aeruginosa (n=3), occurring after central venous line colonization in
five cases. One case of arthritis was observed. Five patients had central nervous system
infections, P6 presented meningitis caused by Listeria monocytogenes at the age of 26 years
and CMV retinitis at the age of 29 years. Two siblings (P3, P4) developed neurological
manifestations about 6 months after oral immunization with an attenuated live poliovirus
strain. These patients developed slowly progressive muscle weakness without recovery until
their death with recurrent poliovirus identification in their stools (Table 3 and 4). Patient P12
suffered from chronic lymphocytic meningitis caused by Herpes Simplex Virus (HSV) type 2
at the age of 14 months and eventually died from this meningoencephalitis at the age of two
years. P20 had chronic lymphocytic meningitis caused by adenovirus at the age of four years,
which led to death at the age of five years. Cutaneous chickenpox was observed in eight
patients, with a favourable outcome in the absence of treatment in all cases. HSV caused
recurrent gingivostomatitis in five patients, beginning in the first two years of life in all cases.

Other manifestations
Autoimmune hemolytic anemia (AIHA) (with positive Coombs-type IgG and
complement) occurred in three patients, at the ages of 5, 10 and 31 years (P1, P8 and P6,
respectively), requiring treatment with corticosteroids and splenectomy at the age of nine
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years in one patient (P1) and treatment with corticosteroids in the other two cases. Four other
patients (P10, P16, P26 and P27) had peripheral neutropenia with no detectable
autoantibodies. Four children (P2, P8, P15, P18) had transient hypereosinophilia (range: 3 to
10 x 109 cells/l) associated with eczema or cryptosporidiosis. Eczema was observed in a total
of four patients (P9, P15, P18 and P31). Finally, an allergic cutaneous reaction was observed
in four patients (P6 and P23 after amoxicillin treatment and P26 and P35 after trimethoprim-
sulfamethoxazole treatment).

Outcome and treatment
HSCT
HSCT is the only known curative treatment for MHC class II expression deficiency. In
our series, 23 (66%) of the 35 patients underwent HSCT (Figure 4a, Table 3a). The
indications for HSCT mostly related to clinical status (age, infection state). Myeloablative
conditioning regimens combining busulfan with cyclophosphamide were used in all patients
undergoing HSCT. Most patients receiving grafts from an unrelated or HLA-mismatched
donor were given in vivo immunosuppressive treatment (antithymocyte globulin, Campath IG
or anti-LFA1 with or without CD2, depending on the conditioning protocol in routine use at
the time). Ten of the 23 patients who underwent transplantation (43%) are alive and have
been cured, with the recovery of almost normal immune functions (Table 3a). They were
transplanted between the ages of one and nine years (mean age 2.8 years). Follow-up is
currently one to 24 years after HSCT (mean 11.8 years). Complete engraftment was observed
in nine patients and partial engraftment was observed in another. One patient, P33, who
underwent HSCT with cord blood with a single mismatch, currently displays poor
engraftment and little immunological recovery one year after transplantation. Twelve of 23
patients who underwent HSCT (52%) died after transplantation (1 months to 6 years after
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HSCT; mean 1.3 years; Table 3a). These patients underwent transplantation between the ages
of six months and nine years (mean 2.9 years). Nine patients died in the period immediately
following HSCT (1 to 5 months after HSCT), from severe viral and/or fungal infections
(n=6), severe graft-versus-host disease (GVH) (n=1) or both severe viral infections and severe
GVH disease (n=2). An absence of engraftment was noted in three patients receiving haplo-
identical grafts. These patients died between 4.5 and 6 years after HSCT, from liver failure
and sepsis, viral meningoencephalitis, and liver and medullary failure.

Outcome of patients who did not undergo HSCT
Twelve patients did not undergo HSCT (Table 3b). Clinical course was unfavourable
in these patients, with the progression of infectious complications to death in five patients
(42%; range: 1 to 17 years; mean age at death: 7 years; Figure 4b). The causes of death were
severe viral infections before the age of five years, with two cases of viral
meningoencephalitis. P7 died from severe undocumented interstitial pneumonia. Liver failure
was responsible for the death of two patients (P1 and P8) over the age of five years (Table
3b). P8 had severe progressive respiratory, gut and hepatic dysfunctions, with failure to
thrive, resulting in death at the age of 17 years. Seven patients that did not undergo
transplantation (58%) are still alive at time of this report, at a median age of 12 years (range:
6 years to 32 years; mean age: 15 years). All of these patients, except one, are living in France
and have respiratory and/or gastrointestinal and/or hepatic symptoms (Table 3b). Four
patients (P6, P14, P32 and P35) have reached puberty and are still alive on supportive care
(IgG treatment, antibiotic prophylaxis with trimethoprim-sulfamethoxazole; Table 3b).
The oldest of these patients is 32 years old (P6) and has symptoms since the age of
seven months. He was diagnosed with MHC class II deficiency at the age of five years. His
symptoms improved between the ages of 10 and 20 years, with a decrease in the frequency
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and severity of infections. After the age of 20 years, P6 presented symptomatic RSV infection
and recurrent HSV1 infections, cholestasis and moderate cytolysis. A liver biopsy showed
granulomatous lesions associated with moderate portal fibrosis, but no pathogens were
detected. P6 developed meningitis caused by L. monocytogenes at the age of 26 years, CMV
colitis at the age of 27 years and CMV retinitis at of the age 29 years. He had AIHA requiring
steroid treatment at the age of 31 years. He also has recurrent gastroenteritis (C. jejuni, C.
difficile, Salmonella spp.), recurrent gastritis caused by Helicobacter pylori, ENT infection
and bacterial pneumonia (H. influenzae), with chronic bronchopneumonia. He has a
Karnofsky Performance Scale Index of 90%.

The second patient, P14, has had symptoms since the first year of life and was
diagnosed with MHC class II deficiency at the age of two years. He has recurrent ENT
infections (H. influenzae) and bacterial pneumonia (S. pneumoniae), with chronic
bronchopneumonia since the age of one year. He also had CMV, VZV and zoster infections
with a favorable outcome during childhood. He has recurrent bacterial diarrhea caused by
Salmonella spp. and C. jejuni. Since the age of nine years, this patient has suffered from liver
dysfunction related to C. parvum infection. He is now 23 years old and has a Karnofsky
Performance Scale Index of 90%.

The third patient, P32, was diagnosed at the age of 10 years. She is now 12 years old
and has a Karnofsky Performance Scale Index of 60%. She has had recurrent diarrhea and
chronic bronchopneumonia requiring oxygen treatment since the age of one year. Hepatic
involvement due to cholangitis caused by C. parvum was diagnosed at the age of 10 years.
This patient is one of those who displayed residual MHC class II expression on B cells, with
no expression on monocytes. Despite this residual expression, she had severe CD4+ T-cell
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