Immune Work-Up for the General Pediatrician Fatima Gutierrez M.D.

Goals and objectives Learn when to Suspect an Immune Deficiency Important Aspects of History Taking Physical Exam Clues Initial Laboratory Work-up When to get an Immune Consult Overview of Specialized Tests

When to suspect: 8 or more new ear infections within one year 2 or more serious sinus infections within 1 year Two or more months on antibiotics with little effect 2 or more pneumonias within 1 year Failure of an infant to gain weight or grow normally

When to suspect Recurrent, deep skin or or organ abscesses Persistent thrush in mouth or elsewhere on skin, after age 1 Need for IV antibiotics to clear infections 2 or more deep-seated infections Family history of Primary Immunodeficiency

Zebras among us Sinusitis/Pneumonia Low Antibody levels CF Allergic Rhinitis Anatomy Low Antibody levels Protein-losing entropathy, nephropathy Severe eczema Burns 2° Immunodeficiency Malnourishment Diabetes Sickle Cell

Basic immunology Primary Immunodeficiencies Can Be Categories by their Defect: Cellular Immunity (T-Cells) Humoral Immunity (B-Cell) Phagocytes Complement

T- cells Plasma cells Immunoglobulins B-lymphocytes Bone Marrow Stem Cell T- cells

T-Cell Defects T-Cells directly Attack Foreign Antigens Opportunistic infections Candida sp, Pneumocystis jiroveci Persistent Thrush Diarrhea/Malabsorption Poor Growth/Failure to Thrive

B-Cell Defect Unable to make specific antibodies Multiple / Severe Bacterial Infections Persistent Upper Respiratory Tract Infections Streptococcus, Haemophilis Usually seen after 6 months when maternal antibodies lost Growth usually normal unless patient with chronic infections

Phagocyte defects Natural Killer Cells – directly attack cells infected with Virus Phagocytes – ingest and kill microorganisms Chronic Granolomatous Disease – - Infections caused by catalase + organisms LAD No Pus Gingivitis/Periodontitis Skin Infections, Liver, GI Tract

Complement defects Composed of 30 proteins Function in ordered integrated fashion to defend against infection and produce inflammation C5- C8 deficiency associated with autoimmune C1-C4 deficiency Associated with Rheumatic Disorders Pyogenic Infections C5-C9 Complement deficiency Neiserria Infections Basic Screen – CH50 – measures function of alternate pathway

Stats: Immunodeficiency Primary Secondary 1:500 – 1:100,000 - All 1:500 IgA def 1:100,000 SCID 1:200,000 CGD Secondary 1:300 - HIV 1:5 “unaware” of their HIV

Stage 1: History & Exam History Infection History Sex Consanguinity History of Miscarriage/ Death in Infants Primary Immune Deficiency in Family History of Autoimmune/Connective Tissue d/o Environment (Smoke, allergies) Delayed Cord Separation (LAD) HIV risk factors Passive cigarrette smoke inhalation can predispose to otitis media, pneumonia, bronchitis. Allerigies can worsen congestion, predispose to sinusitis and otitis media

Physical exam Eczema , Wiskott Aldrich or Hyper IgE? Wiskott Aldrich Thrombocytopenia Small platelets Hyper IgE -Job Syndrome Skin infections – Staph Sinusitis Coarse Facies Retain Primary Teeth Lack of tonsils/lymph nodes – X-linked aggamaglobulinemia Lack of Pus- Leukocyte Adhesion Disorder Chronic Eczema – Wiscott Aldrich, Hyper IgE

Physical Exam: Conical Teeth Look at Mother Incongentia Pigmenti

NemO: Not just a fish Nuclear Factor – kB Essential Modulator X-Linked Defect in Cellular Immunity Variable Immunoglubulin levels Normal B-cell numbers Recurrent bacterial sinopulmonary and oportunistic Infections Ectodermal Dysplasia Conical Teeth Treatment Stem Cell Transplant

STAGE 1: important info in the CBC Lymphocyte count Absolute Lymphocyte count Lymphopenia Screen for B and T cell Deficiencies Lymphocytosis Omenn Syndrome, Leukocyte Adhesion Deficiency Neutrophil Count Leukocyte Adhesion Deficiency - >100,000/mm3 Platelet Size and Number Wiskott Aldrich

ALC below 2000 abnormal in Infants! WBC total x %lymphocytes = Absolute Lymphocyte Count (ALC) Normal absolute lymphocyte count 10th- 90th percentiles (#/μL) by age 0-3months 3400-7600 6-12months 3900-9000 1-2 years 3600-8900 Look at normal values based on Age!!

Stage 1: Immunoglobulins IgG, IgM, IgA, IgE IgG in young infants less reliable –largely maternal Lab Variations! Age Adjusted Normals Low IgG in Infant Hypogammaglobulinemia of Infancy Undetectable IgA Primary Immune Deficiency High Levels IgM + Absence of other Ig Hyper IgM IgE Atopic Disease or Parasitic Illness, Hyper IgE

Immunoglobulin Development IgG nadir for infants is age 3 months to 1 year of life Premature infants will lack adequate maternal IgG Note in IgG at approximately 6 months of life (maternal vs infant sources) IgM production starts immediately after birth IgA rate of synthesis is slowest

X-Ray THYMUS! May shrink in response to stress/surgery/infe ction Thymus continues to grow until teens Largest size relative to body weight at birth No Thymus

DelayeD Hypersensitivity SkIN Test T-cell Response Common Antigens Candida Trychophyton Tetanus Diphtheria PPD Anergy (no response) Immune suppr. Meds- Steroids Infants < 6 months / 2yrs Malnutrition Collagen Vascular Disease Fever- Leukocytosis Reaction measured at 48 hours High False negative

HIV TestIng HIV – DNA when IgG unreliable ELISA tests IgG Infants have Maternal IgG Severe Hypogammaglobulinemias Liver Disease Guidelines : www.aidsinfo.nih.gov

Abnormal Labs or Concerning clinical Picture: Immunology consult

Measurement of Specific T, B, NK and phagocytic cells Phenotyping Measurement of Specific T, B, NK and phagocytic cells

Lymphocyte studies

The Case of SCID NK Cells Tcells- B-Cells

4 fold increase of baseline 1 month after vaccine given Humoral Immune Panel Tests your Body’s Ability to Make Antibodies – Majority of Vaccines activate B-cells in the presence of T-cell Tests for Common Vaccines Diphtheria Tetanus Streptococcus Majority of Vaccines T-Dependent Normal Response Diphtheria > 0.1 Tetanus > 0.1 Or… 4 fold increase of baseline 1 month after vaccine given

Humoral Immune Panel Streptococcus – Note minimal response Serotypes in Prevnar: 4, 6B, 9V, 14, 18C, 19F, 23F Serotypes in PCV13 Serotypes 1, 3, 4, 5, 6A, 6B, 7F, 9V, 14, 18C, 19A, 19F, 23F Test not accurate if patient received IVIG (Wait 3-4 weeks) Bacteriophage X1074 if patients have recenlty received IVIG

lymphocyte proliferation Assays: (Mitogens and Antigens) Measures the ability of lymphocytes to respond to mitogens and Specific antigens Strongly Mitogenic on T-Cells - PHA (Phytohemagluttin) - ConA (Concanavalin-A) Stimulates T&B cells (Mitogen - Pokeweed Minimal Response Assay working well Phytohemagluttin- isolated from red kidney bean ConA- from jack beans Pokeweed mitogen from root Antigens: - Candida -Tetanus - Dipthera *Prior exposure required for response !! > 2yrs age

Take Home message Early Recognition of Primary Immune Deficiency – Many Primary Immune Deficiencies are Lethal! Recognize Lab Value Diversity Don’t forget to look for HIV

References Noroski MD, L., Shearer MD, W. Short Screening for Primary Immunodeficiencies in the Clinical Immunology Laboratory. Clinical Immunology and Immunopathology. Vol. 86, No3, March pp. 2237-245, 1998 Fleisher, Shearer, Kotzin, Schroder. Clinical Immunology – Principles and Practice 2nd Ed. Rich, R. Schmitz PhD, Folds PhD. Clinical and Laboratory Assesment of Immunity. Journal of Allergy and Immunology 2003: 111:S702-11 American Academy of Allergy Asthma and Immunology – www. aaai.org Immune Deficiency Foundation – primaryimmune.org Jeffrey Modell Foundation www. jfmworld.org