Case 1

This is a 13 month old female who is referred to you for cold symptoms, fever (T 38.5 degrees) and a persistently low absolute neutrophil count (ANC = WBC x (segs% + bands%)) of 40 (WBC 4.0, 1% bands, 0% segs). Her ESR is 12. She was hospitalized three weeks ago for a pseudomonas external otitis media and neutropenia that was treated with two weeks of intravenous antibiotics. Three weeks ago, her ANC was 342, with a low of 54, and at discharge her ANC was 896 (one week ago). She had been doing well since discharge until her current illness.

Of note, a CBC at birth and at 10 months of age demonstrated normal absolute neutrophil counts. There is no history of increased bacterial or fungal infections. There is no family history of recurrent bacterial infection, neutropenia, immunodeficiency disease, autoimmune disease, or malignancy. There is no history of infant deaths in the family. There has been no history of recent medication use.

Exam: VS are normal. Height and weight are at the 75th percentile for age. Physical examination had no unusual findings. No oral thrush, lymphadenopathy, hepatosplenomegaly, or skin lesions are noted.

She is hospitalized for IV antibiotics (ceftazidime). Blood cultures return negative and she is discharged after 3 days. During this hospital stay she does well. Antineutrophil antibody testing is sent off to a specialized reference laboratory and it returns positive. A bone marrow examination is done (mostly because of parental concern) which shows a normal cellular marrow. During subsequent febrile illnesses, she does well clinically. Three months later, after she initially presented with neutropenia, her ANC improves to 2400. Two months later, it remains over 2000.

Diagnosis: Chronic benign neutropenia of infancy and childhood.

Case 2

This is a 2 year old male who presents with a chief complaint of recurrent skin and soft tissue infections. The infecting organisms were catalase producing bacteria. Three nitroblue tetrazolium (NBT) slide tests sent to a reference laboratory returned normal. Screening tests of humoral, cell mediated, and complement mediated immunity were normal. Referral is now being made to a hematologist during his current hospitalization for the treatment of cervical lymphadenitis and left lower lobe pneumonia with bilateral pleural effusions. The patient's blood samples were sent to a neutrophil research laboratory with results that demonstrated no reduction of NBT (an abnormal result in contradiction to the normal NBT slide test results) and negligible quantitative production of superoxide. His mother demonstrated an intermediate production of superoxide and a decreased number of neutrophils capable of reducing NBT.

Past Medical History: At 2 months of age, he developed a perianal furuncle that was incised and drained because of no response to oral antibiotics. At 5 months of age, he had surgical treatment for multiple perianal fistulas with abscesses. At 7 months of age, he had a left inguinal Klebsiella pneumoniae lymphadenitis that was treated with incision and drainage and oral amoxicillin/clavulanic acid. Two weeks later, a left subauricular lymph node abscess was incised and drained and a persistent perianal fistula received topical treatment with silver nitrate. Pseudomonas aeruginosa grew out of cultures of the neck abscess and the patient was hospitalized for intravenous antibiotic treatment and immunological evaluation. In addition to his subauricular lymphadenitis, he had a left calf cellulitis that grew Serratia marcescens and a left inguinal abscess that grew Staphylococcus epidermidis.

Clinical Course: He completes treatment with 6 weeks of intravenous antibiotics until his ESR decreases <30. However subsequently, he develops a slight limp at which time a large lytic bone lesion is found in the distal left tibia on plain x-rays. Culture of that lesion grows out Staphylococcus aureus after debridement and curettage. He receives 6 more weeks of intravenous antibiotics until his ESR is in the normal range. He is placed on subcutaneous injections of gamma interferon (three times a week) and twice daily doses of oral trimethoprim-sulfamethoxazole and has not required any further hospitalizations for bacterial infections for the last 3 years.

Diagnosis: Chronic granulomatous disease.

Neutrophils (polymorphonuclear leukocytes) represent the first line of active defense against bacterial and fungal invasion for the innate immune system. Though a critical component of the body's immune system, primary disorders of neutrophil function account for only about 18% of all primary immunodeficiencies, with T cell (20-30%) and B cell deficiencies (50%) comprising the majority of defects seen. Despite the relative rarity of primary neutrophil defects, clinical situations in which neutrophil function is decreased, such as prematurity, are commonly associated with increased rates of invasive bacterial infection. Primary deficiencies of neutrophil numbers or function are usually associated with an increased risk of serious, often life-threatening infections. Secondary deficiencies of neutrophil numbers or function are usually markers of systemic disease and tend to be clinically benign.

The most common problem seen by primary care physicians is neutropenia (decreased neutrophil count). Acute inflammatory processes are commonly associated with normal or reactive increases in neutrophil counts. When low neutrophil counts are associated with infection it must be decided whether neutropenia is secondary to the infection, or if an underlying neutropenia contributed to the risk of infection. A key point to remember is that the risk of infection with neutropenia is high when bone marrow production of neutrophils is decreased from either primary or secondary causes.

In general, common disorders are usually benign clinically and occur in children with no significant medical history of bacterial or fungal infections. Rare congenital disorders result in extremely high risks of infection and require specialized laboratory tests to correctly diagnose.

The most common presentation of neutropenia (low neutrophil counts) and neutrophilia (high neutrophil counts) is an acute febrile illness in an otherwise normal child. Serious primary neutropenia or primary disorders of neutrophil function are associated with "frequent" or "atypical" bacterial infections. These important points should be kept in mind: i) An overwhelming, sudden onset of sepsis, as observed in children receiving intensive multi-agent chemotherapy, is rare in most children with neutropenia or defects in neutrophil function. ii) An intact humoral and T-cell immune response and monocytosis compensates somewhat for a decreased neutrophil count or altered neutrophil function. iii) Little or no increased propensity to infection is associated with the most common form of severe neutropenia in children, so-called "chronic benign neutropenia". iv) Neutrophil defects commonly present with frequent, indolent bacterial or fungal infections that are difficult to detect and treat.

While there are subtle differences in the presentation of primary neutrophil disorders, the overlap in presentation is significant and determining the precise defect is impossible on clinical findings alone. With the passage of time it is generally easy to determine that a child's history of infections is abnormal, suggesting a more serious disorder.

DISORDERS OF NEUTROPHIL NUMBERS

Key to the understanding of quantitative abnormalities of neutrophil counts is an understanding of neutrophil distribution. The peripheral neutrophil count reflects the equilibrium between the circulating pool and the marginated pool of neutrophils adherent to vascular endothelium, and a tissue pool. The complete blood count (CBC) only monitors neutrophils in the circulating pool.

NEUTROPHILIA (increased neutrophils): Upon infection and activation of the immune response, neutrophils from the marrow storage pool are released resulting in up to three fold increases in neutrophil counts within four to five hours. More than half of the peripheral granulocytes are attached to the vascular endothelium at any given time point and represent a "marginated" pool that can be released almost immediately at times of stress. Epinephrine mediated "demargination" of neutrophils to the circulating pool is sometimes seen during phlebotomy in an anxious child and can result in spuriously elevated mature neutrophil counts. Among the other causes of neutrophilia are: reactive leukocytosis, ethnic neutrophilia, Pelger Huet, leukemoid reaction, leukoerthyroblastic response, chronic myeloid leukemia and leukocyte adhesion defect.

NEUTROPENIA (decreased neutrophils): Neutropenia is defined by an absolute neutrophil count (ANC) <1,500/cubic mm. ANC = (%bands + %mature neutrophils) X total WBC count. Decreased neutrophil production, storage, or release; redistribution from circulating to marginated pools; or increased destruction explains most cases of neutropenia. The key determinants of infection risk are the adequacy of the bone marrow storage or reserve pool and the general robustness of the immune response. These determinants affect the ability to deliver neutrophils to infected sites and the ability of the immune system to compensate for quantitative deficiencies in neutrophils. Neutropenia discovered during the evaluation of infection is generally a secondary finding and characterizes the general low risk of infection associated with a normal marrow reserve and immune system. Some of the causes of neutropenia are summarized below:

1. Kostman syndrome: primary decrease in bone marrow reserve; AR (autosomal recessive), AD (autosomal dominant), S (sex linked recessive); extremely rare; severe neutropenia in the newborn.

2. Shwachman-Diamond Syndrome: primary decrease in bone marrow reserve; AR, S; extremely rare; steatorrhea from exocrine pancreatic deficiency; metaphyseal dysplasia; 50% survival; 1/3 progress to myelodysplastic syndrome or acute myeloid leukemia; normal sweat chloride.

3. Cyclic neutropenia: primary cyclic (every 21 days) variations in bone marrow reserve; AD, S; 1-2 per million; regularly recurring fever every 21 days with oropharyngeal and skin infections; diagnose with CBCs 2-3 times per week for 8 weeks.

4. Chemotherapy: direct toxicity to neutrophil precursors results in a severe reduction in bone marrow reserve (severity dependent on the intensity of chemotherapy agents used); generally a high risk of infection with poor marrow reserve and generalized suppression of the immune system.

5. Nutritional: protein-calorie malnutrition, B12 deficiency, copper deficiency can result in ineffective myelopoiesis; treat by correcting deficiency.

6. Viral infection: bone marrow suppression from a direct effect of infecting virus or through an immune mechanism.

7. Chronic benign neutropenia of infancy and childhood: normal bone marrow reserve; generally thought to be mediated through an anti-neutrophil antibody; common; median age of detection 8 months, 90% detected before 14 months; most resolve spontaneously within months of diagnosis; no significant propensity to infection; treatment is supportive.

8. Autoimmune neutropenia: generally with normal bone marrow reserve but may be associated with a late maturational arrest; antibody mediated destruction of neutrophils; may have an associated primary autoimmune disorder.

9. Alloimmune neutropenia: normal bone marrow reserve; secondary to maternal anti-neutrophil antibody that has crossed the placenta; resolves by 3-4 months of age; generally supportive care.

10. Drug-induced neutropenia: normal bone marrow reserve but has been associated with a late maturational arrest; antibody or complement mediated neutrophil destruction; treatment consists of stopping unnecessary medications.

11. Infection related neutropenia: normal bone marrow reserve; virus induced anti-neutrophil antibody; parvovirus B19 and HIV can be screened; no treatment generally necessary.

12. Hypersplenism: normal bone marrow reserve; sequestration/possible destruction of neutrophils in the spleen; associated with malaria, TB, neoplasm, collagen-vascular diseases, hemolytic anemia; spherocytes and tailed RBC on blood smear; treat underlying disorder.

13. Pseudo-neutropenia (severe infection): normal bone marrow reserve; generally associated with increases in marginated and tissue pools; mild and spontaneously resolves.

In general, the risk of infection can be related to the bone marrow reserve and overall immune competence of patients with neutropenia. Careful consideration of the overall risk of infection needs to be utilized to determine the appropriate management of these children. Inadequate treatment or follow-up of children with neutropenia and a high risk of infection can be fatal, while over aggressive treatment of a child with a benign neutropenia may result in inappropriate medical care and unnecessary morbidity.

These guidelines can be used in the evaluation and management of the child with neutropenia:

1. Patients with ANCs <500 and fever require prompt evaluation and the rapid initiation of broad spectrum parenteral antibiotics.

2. Empiric parenteral antibiotic therapy (consider ceftazidime, vancomycin, or meropenem) to cover S. aureus, Pseudomonas aeruginosa, Escherichia coli, and Klebsiella species.

3. Consider outpatient management with a parenteral broad spectrum antibiotic (ceftriaxone) if the child is non-toxic, the family and follow up are reliable, and the child has none of the signs of more serious neutropenia syndromes. Followup in an outpatient setting to check the patient's clinical status, check culture results, and administer a second dose of ceftriaxone.

4. DANGER SIGNS: failure to thrive, inflammatory anemia, thrombocytopenia, splenomegaly, lymphadenopathy, joint swelling/bone pain, dysmorphia, recurrent serious infections, fever/infectious symptoms every 21 days, unusual or resistant infections, periodontal disease. If any of these danger signs are present, patients should have more extensive evaluations and a hematology consultation. If no danger signs present themselves, no further testing is needed and parents should be reassured. The most likely diagnosis in the young child is chronic benign neutropenia.

5. Routine use of granulocyte-colony stimulating factor to increase bone marrow production of neutrophils is not indicated for most acquired neutropenias and should be limited to specific disorders where the neutropenia is due to inadequacy of the marrow reserve pool.

DISORDERS OF NEUTROPHIL FUNCTION

Abnormalities in neutrophil numbers are often recognized on a CBC done for other reasons. Children with neutrophil dysfunction must be suspected on clinical grounds, keeping in mind that even the most common primary neutrophil dysfunction syndrome is extremely rare. Children with primary neutrophil function defects usually present with low-grade chronic bacterial or fungal infections. Skin and mucosal infections, lymphadenitis, and abscesses are among those infections commonly seen. These infections tend to be persistent and difficult to resolve with standard treatment. Bacterial sepsis is an unusual finding at presentation.

Children with suspected immunodeficiency should be screened for humoral, cellular, and complement mediated immunity prior to neutrophil function assays. Screening tests routinely available through reference laboratories include: Complete blood count (note that a decrease in neutrophils and platelets/or red blood cells should raise concerns of leukemia or metastatic tumor in the bone marrow), quantitative immunoglobulins (IgG, IgM, IgA, IgE), antibody titers after vaccination (diphtheria, tetanus, and pneumococcus), B-cell counts/phenotype, delayed-hypersensitivity skin tests (Candida, tetanus, or Trichophyton), T-cell counts/phenotype, and total hemolytic complement assay (CH50).

Once a neutrophil defect is suspected and other causes of primary immunodeficiency are ruled out, referral to a specialist familiar with the evaluation of primary neutrophil defects should be made. Anticoagulated samples (usually sodium heparin) should be freshly drawn and transported as quickly as possible to a neutrophil or immunology laboratory. Control samples and parental blood samples are often requested for comparison purposes. Neutrophil function tests generally need to be performed at research laboratories experienced with the assays. NBT testing for reactive oxygen species is sometimes available through reference clinical laboratories, though often times it is unreliable with high false negative rates when done in this setting. General tests of neutrophil function include tests such as modified Boyden chambers (chemotaxis), nitroblue tetrazolium (NBT) slide test (reactive oxygen species), dihydrorhodamine oxidation by flow cytometry (reactive oxygen species), neutrophil receptor quantitation, and phagocytosis/particle ingestion assays.

Characteristic disorders of neutrophil function include:

1. Chronic granulomatous disease: Pathophysiology is impaired respiratory burst (defective NADPH oxidase); inheritance usually X-linked, some autosomal recessive; 1 in 500,000 prevalence, 1 in 2,000,000 live births, two-thirds< 1 year old at diagnosis; most common infections include pneumonia and lymphadenitis; most common infecting organisms (generally catalase negative) S. aureus, S. marcescens, B. cepacia, and Aspergillus sp.; most common clinical findings are lymphadenopathy, hypergammaglobulinemia, hepatomegaly, splenomegaly, anemia of chronic disease, underweight, chronic diarrhea, short stature, gingivitis, and dermatitis; diagnosis made by NBT, flow cytometry, or cytochrome C reduction; treatment includes prevention of infection with daily trimethoprim-sulfamethoxazole and gamma-interferon three times a week; bone marrow transplantation is experimental but curative.

2. Chediak-Higashi syndrome: Pathophysiology is decreased degranulation, chemotaxis and granulopoiesis; inheritance autosomal recessive; rare with 200 cases reported; multisystem disorder with clinical characteristics that include mild coagulopathy, peripheral and cranial neuropathy, hepatosplenomegaly, pancytopenia, partial oculocutaneous albinism, frequent bacterial infections (usually S. aureus), progressive lymphoproliferative syndrome with death by age 20; diagnosis made by neutropenia, giant lysosomes in neutrophils; treatment includes prevention of infection with daily trimethoprim-sulfamethoxazole and daily ascorbic acid; bone marrow transplantation in accelerated phase.

3. Leukocyte adhesion defect I: Pathophysiology is the lack of an adhesion receptor resulting in impaired chemotaxis, adhesion, and phagocytosis; inheritance autosomal recessive; rare with 60 cases reported; clinically characterized by moderate to severe phenotypes with a lack of pus at sites of infection, delayed separation of the umbilical cord, severe periodontitis and/or gingivitis; diagnosis made by the presence of neutrophilia, decreased neutrophil integrin adhesion receptor (CD11b/CD18) by flow cytometry, and impaired chemotaxis; treatment includes prevention of infection with daily trimethoprim-sulfamethoxazole and bone marrow transplantation if severe.

4. Hyper-IgE (Job's) syndrome: Pathophysiology is a variable chemotactic defect; inheritance autosomal dominant; rare with 50 cases reported; clinically characterized S. aureus pneumonia with pneumatoceles, fungal superinfection of lung cysts, recurrent "cold" abscesses that don't respond to antibiotics, eczema, mucocutaneous candidiasis, coarse facial features, and short stature; diagnosis made by an IgE>2,500 and eosinophilia; treatment includes prevention of infection with daily trimethoprim-sulfamethoxazole and bone marrow transplantation if severe.

Children with neutrophil functional defects rarely present with overwhelming bacterial or fungal infections, but more commonly suffer from low grade, chronic infections that may become indolent and impossible to effectively treat. Chronic infection and inflammation associated with deep seated infections leads to a high rate of morbidity and shortened survival. Low grade infections that are neglected can evolve into serious disseminated infections without the appropriate, timely administration of antibiotics or antifungal agents. Adding to this difficulty in clinical monitoring, is an attenuated inflammatory response that often masks serious infection. Referral to subspecialists experienced in the management of children with immunodeficiencies or neutrophil disorders is critical to minimize morbidity and mortality for this population.

Questions

1. The risk of infection with neutropenia is highest when:
. . . . . a. Neutropenia onset is rapid
. . . . . b. Low bone marrow cellularity is present
. . . . . c. Peripheral destruction of neutrophils is occurring
. . . . . d. Anti-neutrophil antibody is present

2. The most common cause of neutropenia is:
. . . . . a. Anti-neutrophil antibody
. . . . . b. Drugs
. . . . . c. Infection
. . . . . d. Bone marrow failure
. . . . . e. Malignancy

3. Neutropenia associated with steatorrhea is most characteristic of:
. . . . . a. Cystic fibrosis
. . . . . b. Kostmann syndrome
. . . . . c. Evan's Syndrome
. . . . . d. Chediak-Higashi Syndrome
. . . . . e. Shwachman-Diamond Syndrome

4. Neutrophil defect associated with increased infections with catalase-negative organisms
. . . . . a. Hyper-IgE syndrome
. . . . . b. Leukocyte adhesion deficiency type I
. . . . . c. Leukocyte adhesion deficiency type II
. . . . . d. Chronic granulomatous disease
. . . . . e. Pseudo-neutropenia

5. Infections in children with defects in neutrophil function are characterized by:
. . . . . a. Decreased inflammation that may mask serious infection
. . . . . b. Indolent, chronic infections
. . . . . c. Bacterial and fungal organisms
. . . . . d. Elevated erythrocyte sedimentation rate
. . . . . e. All of the above

References

1. Boxer LA, Blackwood RA. Leukocyte disorders: quantitative and qualitative disorders of the neutrophil, Part 1. Pediatr Rev 1996;17:19-28.

2. Curnutte J, Coates TD. Disorders of Phagocyte Function and Number. In: Hoffman R, Benz EJ, Shattil SJ, et al (eds). Hematology: Basic Principles and Practice, 3rd edition. 2000, Philadelphia: Churchill Livingstone, pp. 721-728.

3. Dinauer MC, Lekstrom-Himes JA, Dale DC. Inherited Neutrophil Disorders: Molecular Basis and New Therapies. Hematology (Am Soc Hematol Educ Program) 2000:303-318.

4. Holland SM, Gallin JI. Evaluation of the patient with recurrent bacterial infections. Ann Rev Med 1998;49:185-199.

5. Kyono W, Coates TC. A practical approach to neutrophil disorders. Pediatr Clin North Am (In Press), 2002

6. Malech HL, Nauseef WM. Primary inherited defects in neutrophil function: etiology and treatment. Semin Hematol 1997;34:279-290.

7. Woroniecka M, Ballow M. Office evaluation of children with recurrent infection. Pediatr Clin North Am 2000;47:1211-1224., 2000

Answers to questions

1.b, 2.c, 3.e, 4.d, 5.e