Reduced diversity of faecal microbiota in Crohn's disease revealed by a metagenomic approach.

BACKGROUND AND AIM: A role for the intestinal microbial community (microbiota) in the onset and chronicity of Crohn's disease (CD) is strongly suspected. However, investigation of such a complex ecosystem is difficult, even with culture independent molecular approaches. METHODS: We used, for the first time, a comprehensive metagenomic approach to investigate the full range of intestinal microbial diversity. We used a fosmid vector to construct two libraries of genomic DNA isolated directly from faecal samples of six healthy donors and six patients with CD. Bacterial diversity was analysed by screening the two DNA libraries, each composed of 25,000 clones, for the 16S rRNA gene by DNA hybridisation. RESULTS: Among 1190 selected clones, we identified 125 non-redundant ribotypes mainly represented by the phyla Bacteroidetes and Firmicutes. Among the Firmicutes, 43 distinct ribotypes were identified in the healthy microbiota, compared with only 13 in CD (p<0.025). Fluorescent in situ hybridisation directly targeting 16S rRNA in faecal samples analysed individually (n=12) confirmed the significant reduction in the proportion of bacteria belonging to this phylum in CD patients (p<0.02). CONCLUSION: The metagenomic approach allowed us to detect a reduced complexity of the bacterial phylum Firmicutes as a signature of the faecal microbiota in patients with CD. It also indicated the presence of new bacterial species.

A human monoclonal antibody obtained from EBV-transformed B cells with specificity for myosin.

We describe the preparation of a stable human lymphoblastoid cell line obtained during ex vivo studies in which peripheral blood lymphocytes of a Glanzmann's thrombasthenia patient were transformed with Epstein-Barr virus. Somatic hybrids secreted an IgM monoclonal antibody (B7) that reacted with the myosin heavy chain of human platelets by immunoblotting. Flow cytometry showed that B7 barely recognized unstimulated intact platelets, but bound abundantly after permeabilization of fixed cells with Triton X-100. The reactivity of the antibody on thin sections of human myocardium and aorta was studied by immunohistochemistry. B7 specifically stained myosin of myocytes, but there was no labelling of aortic smooth muscle cells. The epitope was conserved in cardiac or skeletal myosin prepared from pig or rabbit. Measurement of the dissociation constant in a competitive ELISA showed that B7 bound with high affinity (10(-8) M). Purified Fab fragments retained their ability to bind to myosin, suggesting that B7 may be useful in the imaging of myocardial necrosis after myocardial infarction, myocarditis, cardiac drug toxicosis or graft rejection. This work also shows that EBV transformation of B cells may uncover naturally occurring autoantibodies which under normal circumstances are inhibited by the immune surveillance system.

Reliability and usefulness of immunofluorescence in heart transplantation.

BACKGROUND AND METHODS: To study the reliability and usefulness of immunofluorescence on heart biopsy specimens for routine monitoring of heart transplant recipients for rejection, frozen sections of 72 consecutive endomyocardial biopsy specimens from 18 heart transplant recipients during the first 6 weeks after transplantation (and later) and from 11 control specimens from donor hearts and other nontransplantation patients were studied. Fifteen patients received OKT3 induction. The diagnosis of vascular (humoral) rejection pattern as defined by Hammond was based on the microvascular deposition of immunoglobulin and C3 or C1q. Echocardiographic data and right-sided heart catheterization were obtained simultaneously. RESULTS: The results showed that immunofluorescence was positive for a vascular rejection pattern in 60% (43 of 72) overall, in heart transplant recipients it was positive in 59% (36 of 61), and in control subjects it was positive in 63% (7 of 11). Most of the patients who had positive immunofluorescence had no hemodynamic compromise. Humoral rejection was not predicted by positive immunofluorescence in our study. We also found no correlation with either positive or negative immunofluorescence for the short-term or long-term outcome during the first 6 weeks after transplantation. CONCLUSION: The usefulness of routine immunofluorescence in all surveillance heart biopsies is questionable as determined by this study.

[2 cases of encephalitis in immunodepressed patients]. / Etude de deux cas d'encéphalite survenue chez des sujets immunodéprimés.

The authors report two new cases of encephalitis occurring in association with immunity deficiency. The first concerns a child suffering from Bruton type congenital hypogammaglobulinemia. Histological examination of the brain under light microscopy shows important lesions of panencephalitis with parenchymal atrophy : the latter was more marked in the cerebellar lamellae. Ultra-structural examination showed numerous nuclear bodies in the nuclei of the macroglial cells and tubular inclusions situated in the cytoplasm of the capillary endothelial cells. The second case is that of a man of 44 years of age who had undergone a kidney transplant for malignant arterial hypertension, who after 3 months immunodepressive treatment suffered from temporo-spatial confusion together with a pyramidal tract syndrome. He died after 3 weeks evolution of the disease. Microscopical examination of the encephalon showed the presence of microglial nodules with aspects of neurophagia, suggestive of a polioencephalitis. These lesions were situated in the temporal cortex and the brain stem. The lymphocytic reaction was very slight in the second case.

[Polyneuritis in chronic lymphoid leukemia. Ultrastructural study of the peripheral nerve (author's transl)]. / Polyradiculonévrite au cours d'une leucémie lymphoïde chronique. Etude ultrastructurale d'un biopsie de nerf périphérique

A 75 years old man had a chronic lymphoid leukemia for 5 years. Within a period of three months he developed a polyneuritis. The biopsy of a peripheral nerve showed a large cellular infiltration by lymphocytes. The ultrastructural study shows in some Schwann cells disruption of the myelin. Their cytoplasm is invaded by a lymphocyte process as in Guillain-Barré. But in this case some of these invading lymphocytes show polygonal structures in their cytoplasm; that is sometimes seen in the proliferating cells of the chronic lymphoid leukemia.

[Leukodystrophy with adrenal insufficiency (adreno-leukodystrophy). A study of 3 familial cases with the ultrastructure of one biopsied case]. / Leucodystrophie avec insuffisance surrénalienne (adréno-leucodystrophie). Etude de trois cas familiaux, avec ultrastructure d'un cas biopsié

Three male siblings in a gypsy family presented with congenital Addison's disease. The youngest showed neurological signs at the age of ten and deteriorated rapidly. The eldest had convulsive symptoms and the third an irregular E.E.G. Investigation of the youngest by means of cerebral biopsy revealed rarefaction of myelin with perivascular sudanophil deposits. An ultrastructural study showed frequent liposfuscin deposits in the astrocytes and oligodendrocytes. In the peripheral nerve there were signs of re-myelinization associated with hypertrophy of Schwann cells. These various aspects have led to these cases of demyelinization associated with adrenal deficiency being classified among the leukodystrophies while some authors have interpreted these lesions as characteristic of Schilder's disease. The fact that in thirty or so cases published, almost half of which are familial, only boys are affected supports the theory of an enzymopathy at the root of both leukodystrophy and adrenal atrophy.

[Peripheral neuropathies i- Waldenstrom's disease. Histological and ultrastructural studies of 5 cases]. / Les neuropathies périphériques de la maladie de Waldenström. Etude histologique et ultrastructurale de 5 cas

In four cases it was the existence of peripheral neuropathy which led to the discovery of macroglobulinemia within an interval from 1 to 11 years. The treatment of the disease also markedly improved the peripheral neuropathy in three cases. Two patients died. Two of the biopsies of the peripheral nerve showed infiltration by atypical lymphocytes. Ultrastructural changes in the five cases were variable. Marked axonal involvement in the two cases with cellular infiltration. In one of these two cases it was combined with segmental demyelinization. This lesion was clearly predominant in the three other cases. In one of them demyelinization occurred in a very peculiar way with dilation of the internal part of the mesaxone going on in the most peripheral layers of the myelin. Discussion of the mechanisms of the involvement of the peripheral nervous parenchyma in Waldenström's disease. Cellular infiltrations, amorphous deposits, changes in the capillaries and macroglobuline may occur.