Fatal Granulomatous Disease Research Articles

AUSTWICKIA CHELONAE IN A WILD GOPHER TORTOISE (GOPHERUS POLYPHEMUS) AND EVIDENCE OF POSITIVE SELECTION ON THE DIPHTHERIA-LIKE TOXIN GENE.

Austwickia (Dermatophilus) chelonae is a filamentous, Gram-positive Actinobacteria in the Dermatophilaceae family. It has caused fatal granulomatous disease in diverse captive reptile species on three continents, but its presence in wild or free-ranging populations was unknown. An adult female gopher tortoise (Gopherus polyphemus) was presented euhydrated, but cachectic and infested with ticks, with two firm, encapsulated masses over the cranioventral neck and right stifle. The tortoise had moderate nonregenerative anemia and evidence of inflammation; plasma biochemistry data was within normal limits. Fine needle aspirate of the neck lesion revealed abundant necrosis and aggregates of cocci. Computed tomography delineated the masses and revealed an additional mass adjacent to the left zygomatic bone. After surgical excision, histology identified chronic granulomas with intralesional filamentous bacteria. Pan-bacterial 16S rRNA PCR and sequencing of the masses identified A. chelonae. Despite treatment with oxytetracycline and ceftazidime, the tortoise deteriorated and was euthanatized. An esophageal lesion consistent with A. chelonae was seen on postmortem examination, although it was determined that the tortoise ultimately succumbed to fungal pneumonia caused by Metarhizium robertsii, an entomopathogenic biotoxin sprayed as insect control. This case reveals A. chelonae is present in free-ranging chelonians in North America. This organism produces a toxin gene similar to diphtheria toxin, one of the most potent known biotoxins, which has not been previously identified outside the genus Corynebacterium. Novel PCR primers were designed for the toxin and rpoB genes, which were amplified and sequenced from two cases and compared with two available genomes. Selection analysis revealed that the toxin gene is under positive selection, which implies it interacts significantly with the immune system, making it a good candidate for immunodiagnostic test development.

STUDY OF THE EFFECT OF ACTIVE INFECTIONS ON DIHYDRORHODAMINE (DHR) TEST IN CHRONIC GRANULOMATOUS DISEASE PATIENTS (CGD)

Chronic granulomatous disease (CGD) is the most common type of primary innate immune deficiency caused by defects in the nicotinamide adenine dinucleotide phosphate (NADPH) oxidase enzyme in the neutrophils. (1) It’s considered as a fatal granulomatous disease of childhood as most patients with CGD die by 10 years of age. (2) Any mutation at one of the genes encoding for glycoproteins that are forming the five subunits of the NADPH oxidase enzyme leads to CGD. If the mutation leaves some residual NADPH oxidase activity intact the clinical expression of the disease is attenuated and prognosis is better. (3) CGD can be treated by conventional treatment, stem cell transplantation or genetic therapy. (4) The (DHR) test was approved to be the gold standard for diagnosis of CGD. (5) Aim of the work The aim of the present study is to detect the effect of active infections on the interpretation of DHR test results in chronic granulomatous disease patients and patients with active infections with no evidence of CGD.

Chronic Granulomatous Disease: Lessons from a Rare Disorder

Chronic granulomatous disease (CGD) is a rare primary immunodeficiency with X-linked or autosomal recessive inheritance involving defects in genes encoding phox proteins, which are the subunits of the phagocyte NADPH oxidase. This results in failure to produce superoxide anion and downstream antimicrobial oxidant metabolites and to activate antimicrobial proteases. Affected patients are susceptible to severe, life-threatening bacterial and fungal infections and excessive inflammation characterized by granulomatous enteritis resembling Crohn's disease and genitourinary obstruction. Early diagnosis of CGD and rapid treatment of infections are critical. Prophylaxis with antibacterial and mold-active antifungal agents and the administration of interferon-γ has significantly improved the natural history of CGD. Currently, the only cure is allogeneic hematopoietic cell transplant (HCT), although there remains controversy as to which patients with CGD should get a transplant. Allele-based HLA typing of alternative donors, improved supportive care measures, and use of reduced toxicity conditioning have resulted in event-free survival (EFS) of at least 80% even with an unrelated donor and even better when the patient has no active infections/inflammation. Gene correction of CGD would eliminate the risks of graft-versus-host disease (GVHD) and the immunoablative chemotherapy required for allogeneic HCT. Based on gene therapy trials in patients with SCID-X1, ADA-SCID, and the early experience with CGD, it is clear that at least some degree of myeloablation will be necessary for CGD as there is no inherent selective growth advantage for gene-corrected cells. Current efforts for gene therapy focus on use of lentivector constructs, which are thought to be safer from the standpoint of insertional mutagenesis and more efficient in transducing hematopoietic stem cells (HSCs).

Infantile Onset Panniculitis with Uveitis and Systemic Granulomatosis: immunohistochemical findings

Infantile-Onset Panniculitis with Systemic Granulomatosis is a recently described clinicopathologic entity, considered part of the spectrum of pediatric granulomatous inflammatory diseases. Through the International Registry of Pediatric Granulomatous Arthritis (PGA), we now identified 5 children with this disorder, all manifesting from very young age panniculitis, fever, hepatosplenomegaly, arthritis, uveitis and acute phase response. Underlying infections, immune deficiency and autoimmune disease, were excluded. No CARD15 or CIAS1 mutations were found. Histologically, the subcutaneous nodules showed a non-vasculitic non-cytophagic lobular panniculits. Giant and epitheloid cell granulomas were found in liver (pt 1, 5), synovium (pt 2), lymph node (pt 3, 5), colon (pt 3), subcutaneous fat (pt 3), dermis and lung (pt 4). Immunohistochemical study of the granulomas revealed the presence of abundant CD68+ macrophages, numerous CD4+ T lymphocytes and few CD8+ cells. TNF stainings were only weakly positive, conversely abundant IL-6 staining was apparent, especially in the corona of lymphocytes. Despite steroid and cyclosporin treatment, the course was progressive in patient 1 with severe lung involvement and death from respiratory insufficiency at age 14. In patients 2, 3, 4 and 5, therapy with anti-TNF MoAbs allowed better disease control. Infantile onset Panniculitis with Systemic Granulomatosis may be a potentially fatal granulomatous disease in children. Although the response to anti-TNF MoAbs in four of our patients is of note, the paucity of TNF with overwhelming presence of IL-6 in situ in granulomas suggest the implication of alternative immune-inflammatory pathways. The role of the Th17 pthway is currently under investigation

Disorders of cells of the myelo-monocyte cell lineage and of the complement system: molecular basis and clinical aspects

Congenital disorders of phagocytes and of the complement system are considered to be part of the immunodeficiency syndromes in man. Phagocytes, i.e. granulocytes, monocytes/macrophages, and complement factors, actually act in concert with antigen specific T and B lyrnphocytes and their products in a non-specific way to induce effective immunity. Defects of phagocytes and deficiency of complement therefore results in defective inflammatory responses and more or less severe and recurrent microbial infections. In the 1950s the Iirst congenital phagocyte disorder (fatal granulomatous disease of childhood) was described by the pediatrician Dr. Charles Janeway. Deficiency of a single complement factor, namely Cl-esterase inhibitor, was first described in 1962.

Disorders of phagocyte function.

Over a century ago Metchnikoffformulated the phagocytic theory of host defenses. In experiments with marine invertebrates, Metchnikoff noted that certain strains of the invading fungus were ingested and destroyed by phagocytes. Other strains were not attacked, and a disseminated disease then developed (1). Metchnikoff predicted that abnormalities of phagocytic cells would compromise host defenses. A century later this concept was commonly demonstrated by the development of over­ whelming bacterial and fungal infection in conditions resulting in severe quantitative or functional deficiencies of circulating neutrophils. Beginning with Janeway's description in 1954, of a fatal granulomatous disease of childhood (2), several intrinsic disorders of phagocytic cells have been characterized. In the past 15 years, application of new techniques and intense interest in the biology of phagocytic cells have greatly promoted knowledge of factors regulating leukocyte function. The present discussion is not intended to be an exhaustive review of phagocyte defects but is focused on well-characterized and historically important disorders and emphasizes recent advances in our knowledge of the underlying defects. For a detailed discussion of the evaluation and management of patients with phagocyte defects, the reader is referred to several recent reviews (3-6).

Immunopathological aspects of leishmaniasis

Our present understanding of the immunopathology of leishmaniasis has improved tremendously owing to immunological investigations of past years in patients as well as in experimental models. A general concept has emerged which helps in understanding the various forms of cutaneous and visceral leishmaniasis. In this concept, one first has to take into consideration the generic background. While immune response genes certainly are relevant, not only for the different degrees of susceptibility, but also for the balance between the T and B cell responses, genes which do not belong to the histocompatibility complex may also be involved. In particular, the primary killing capacity of macrophages for parasites depends on genes controlling the phagocytic function of the cells. A non-susceptible animal strain just does not permit theLeishmania to multiply within a macrophage. Human genetically determined conditions such as fatal granulomatous disease and myeloperoxidase deficiency illustrate how the phagocytic function can be controlled by genes not belonging to the MHC. While the overall outcome of leishmaniasis can now be quite well explained with the assumption that the T cell response represents the primary defence mechanisms with the antibody response being indispensable for the successful elimination ofLeishmania, many specific problems still need further investigation. In particular, it is not yet known whether antibody-mediated cellular cytotoxicity contributes to the successful elimination ofLeishmania. Further studies should also be carried out on the possible inhibitory function of antibody excess on T cell cytotoxicity. Finally, kala-azar is another example of a disease where the major signs and symptoms can be explained in terms of serum sickness-type mechanisms.

Capacity of nitroblue tetrazolium-reduction of neutrophilic granulocytes (author's transl)

A modification of the NBT-reaction is described which allows a differentiation of the NBT uptake and NBT-reduction. By means of this method it has been shown that the capacity of NBT-reduction of neutrophil granulocytes taken from hematologically healthy persons is independent of the probands' age and, moreover, of the presence of acute infections and their etiology. It has been shown, that the increase in the NBT-reduction of neutrophilic granulocytes of newborns and persons with acute bacterial infections results from an increased NBT-ingestion. To grive an example of a strongly reduced capacity of NBT-reduction of the neutrophilic granulocytes 2 patients suffering from fatal granulomatous disease in childhood have been examined.

Dermatologic manifestations of fatal granulomatous disease of childhood

Dermatologic manifestations of fatal granulomatous disease of childhood

Impaired lymphocyte transformation and chromosomal abnormalities in fatal granulomatosus disease of childhood.

SummaryA previously described case of fatal granulomatous disease of childhood is presented here as certain previous findings could not be confirmed and since two unusual features were noted. Upon the basis of impaired leucocyte function in the father of this patient a new mode of genetic transmission for the disease was proposed meanwhile we have been unable to confirm these original findings (11). This case in itself was interesting since, firstly an impaired in vitro lymphocyte transformation was noted and this later related to the presence of a serum inhibitory factor. Secondly, chromosomal abnormalities were recorded in peripheral blood cells of both patient and his mother and this might be considered evidence to favour a sex‐linked transmission of the disease in this case.

Ultrastructure of the Specific Granules of Human Neutrophil Granulocytes

Electron microscopic studies of circulating neutrophil granulocytes of a boy suffering from Fatal Granulomatous Disease (FGD) showed two abnormal kinds of cytoplasmic granules, both larger than normal granules. One was characterized by a homogeneous matrix of very low electron density, the other contained one or more crystalloid inclusions of larger size than found in normal neutrophils.In neutrophil granulocytes from adults with long‐standing bacterial infection, by light microscopy showing toxic granulation, the granules displayed the same ultrastructural abnormalities as in FGD.It is concluded that the ultrastructure of the granules of neutrophils in FGD reflects non‐specific toxic granulation. The functional defect of the granules in FGD has no known morphological counterpart.

Defective Bactericidal Activity of Monocytes in Fatal Granulomatous Disease

Abstract The bactericidal activity of monocyte-rich cell suspensions obtained from the peripheral blood of patients with fatal granulomatous disease (FGD), their mothers and normal controls was studied. Defective killing of Staphylococcus aureus, Strain 502A, by monocyte-rich cell suspensions from children with FGD was found when compared with monocyte-rich suspensions from normal controls. This defect, less severe, was also observed in the mothers of affected patients. In this selected population of mothers, the monocyte defect varied in parallel with the previously described neutrophil defect.

The pattern of genetic transmission of the leukocyte defect in fatal granulomatous disease of childhood.

The leukocyte-phagocytic function test which was found to be abnormal in boys with fatal granulomatous disease of childhood has been found to be abnormal to an intermediate extent in their mothers. Nine of nine mothers were shown to be abnormal, whereas none of eight fathers and none of five healthy brothers exhibited a defect. 10 of 16 female siblings were abnormal to the same degree as their mothers, as were all three maternal grandmothers available for study. Assuming that this intermediate functional defect represents the heterozygous state, the nine family pedigrees are entirely compatible with the concept that the trait is transmitted on the X-chromosome.A tetrazolium dye-phagocytosis histochemical test was also abnormal in the carrier females and provided independent confirmation of the selection of the female siblings suspected of being carriers for the trait. In addition, this procedure gives indirect evidence that the gene in question is subject to the random inactivation that appears to affect many X-linked genes in mammalian females. The family members were also studied with two of the metabolic assays that have been shown to be abnormal in the cells of affected boys. One assay, the oxidation of the first carbon of glucose-1-(14)C by the isolated leukocytes, was significantly abnormal in the cells of carrier females. The other assay, the oxidation of formate-(14)C by leukocytes of heterozygotes was not significantly different from control values. The practical problem of diagnosing patients would appear to be best solved with a tetrazolium dye procedure, whereas the more subtle abnormality in carrier females is best detected with the leukocyte function test. Improved methods for the function test are being developed.

Fatal granulomatous disease.

SummaryTwo brothers with Fatal granulomatous disease of childhood (FGD) are presented. Both patients exhibited pronounced food allergy. Otherwise, the course of the disease was typical of FGD. From infancy recurrent infections, notably suppurative lymphadenitis, and a fatal outcome before puberty.Microscopic examination of affected organs revealed granulomas with tuberculoid structure; the macrophages contained a characteristic yellow‐brown lipid pigment.A defective function of the neutrophil granulocytes has been demonstrated in FGD. The phagocytic capacity is intact but phagocytized bacteria are not killed since phagocytosis does not induce liberation of catabolic enzymes from the neutrophil granules. In one of our patients, defective degranulation and vacuolization of the cytoplasm of neutrophils was demonstrated in vitro following phagocytosis of staphylococci. However, exposure to sodium fluoride or kleb‐siella toxin did induce degranulation and vacuolization of the defective granulocytes.

A NEWLY DEFINED X-LINKED TRAIT IN MAN WITH DEMONSTRATION OF THE LYON EFFECT IN CARRIER FEMALES

Fatal granulomatous disease of childhood, which has been defined as a clinical entity for ten years, can now be further defined in terms of the functional, metabolic, and histochemical capacities of the isolated leucocytes of the affected male children. We find that the disease is genetically determined, being transmitted in a sex-linked manner, and that a combined phagocytosis/histochemical procedure demonstrates two populations of cells in the isolated leucocytes of the carrier females.