[Why could gut microbiota become a medication?]. / Pourquoi la flore intestinale a-t-elle vocation à devenir médicament ?

The gut microbiota (or gut flora) is a set of bacteria living in symbiosis with the host. Strictly associated with the intestinal tract and interacting with it, the gut microbiota is not a tissue nor an organ, but a supra-organism. A disruption of dialogue between bacteria and human cells is a risk factor or a possible cause of various diseases. The restoration of this dialogue, thanks to the transfer of the gut microbiota of a healthy individual to a patient whose balance of gut flora has been broken, is a new therapeutic approach. If its exact effect still eludes scientific understanding, its clinical benefit is well established for an indication, and is recently being tested for many others. The proven contribution of gut microbiota in the human physiological balance calls for intensifying research throughout the world about the state of knowledge and technologies, as well as on the legal and ethical dimension of fecal microbiota transfer. This didactic paper updates the questions in relation with this therapeutic act.

Effect of indinavir on HIV-related wasting.

OBJECTIVE: To study the effect of the protease inhibitor indinavir on body weight and body composition of subjects with HIV-related wasting. DESIGN: Prospective measurement of body weight in patients who had wasting and were treated with indinavir. A subgroup of 16 representative patients also underwent a metabolic study that included measurements of body composition (skinfolds and bioelectrical impedance) and food intake. Seven from this subgroup who did not have chronic diarrhoea also underwent indirect calorimetry for measurement of resting energy expenditure; the nine patients with wasting and chronic diarrhoea had measurements of faecal losses and intestinal permeability using the lactulose-mannitol test. SETTING: A tertiary care university hospital. PATIENTS: Two hundred and fourteen HIV-infected patients with wasting (less than 95% of usual body weight) had their body weight measured at day 0; 186 patients had a second body weight measurement within the first 100 days of treatment, and 160 patients were weighed a third time, at a median of 176 days. RESULTS: Body weight increased significantly (P < 0.0001) during treatment, whatever the degree of weight loss at baseline. After a median of 176 days on treatment, body weight had increased in 119 out of the 160 patients followed (74.4%; mean weight gain, 6.3+/-SD 3.8 kg; range, 1-18 kg), had not changed in 13 (8.1%) and had fallen in 28 (17.5%; mean weight loss, 4.2+/-3.0 kg; range, 1-12 kg), relative to baseline. Overall, 119 out of the 214 patients (55.6%) from the initial population gained weight. Fat mass, fat-free mass and body cell mass increased significantly in the 16 patients who underwent metabolic studies, together with energy, protein and lipid intake. In the patients with chronic diarrhoea, intestinal permeability improved but there was no change in intestinal losses. In patients who had wasting but not chronic diarrhoea, resting energy expenditure did not change significantly. Body weight changes correlated with changes in the CD4+ cell count (r = 0.882; P = 0.00001) and, to a lesser extent, with changes in the viral load (r = -0.466; P = 0.047). CONCLUSION: Indinavir significantly improved the nutritional status of these patients with HIV-related wasting.

Determination of anti-Cryptosporidium coproantibodies by time-resolved immunofluorometric assay.

The role of the mucosal immune response against Cryptosporidium has been suggested by studies on the therapeutic effects of hyperimmune colostrum. In order to study the intestinal response to this infection, we have developed a sandwich-type time-resolved immunofluorometric assay for the determination of anti-Cryptosporidium coproantibodies. This assay has the inherent sensitivity of an immunoassay without the problems due to background responses from other biological compounds, and is thus suitable for faecal samples. The intra-assay coefficients of variation (5.1%, 4.6%, and 5.8% for immunoglobulins (Ig) A, M and G respectively), inter-assay coefficients of variation (9.4%; 10.5% and 12.2% for IgA, IgM and IgG, respectively) and specificity (100% for all 3 isotypes) were all satisfactory. Using this assay to study 12 patients with the acquired immune deficiency syndrome (AIDS) who were infected with cryptosporidiosis, we found a marked elevation of anti-Cryptosporidium IgA and IgM coproantibody titres relative to 18 healthy control values, but no correlation with the gravity of the infection in terms of oocyst shedding. These results suggest that a non-protective mucosal immune response develops to Cryptosporidium in AIDS patients.

Differential measurement of monomeric and polymeric IgA by one-directional immunoelectrodiffusion.

A technique to specifically quantify monomeric IgA and total IgA in colostrum has been developed using a modified one-dimensional immunoelectrophoretic assay. This method employed electrophoresis in antibody-containing polyacrylamide-agarose gel in the presence of a gel barrier which blocks polymeric IgA. The addition of PEG (polyethylene-glycol 6000) to the anodic gel increased the sharpness of the peaks, the height of which was proportional to the antigen concentration. This method proved to be sufficiently simple, precise, reproducible (CV less than 3%) and linear (from 20-300 mg/l) to measure the monomeric IgA: total IgA ratio rapidly (14 +/- 4.5% for 20 samples in duplicate). Immunoelectrodiffusion studies confirmed that human colostral and serum IgA standards could be used to determine directly monomeric IgA, total IgA and polymeric IgA levels (by difference) rather than to apply correction factors to estimate these IgA levels.

Characterisation of the main molecular forms of human fecal immunoglobulin A.

The molecular composition of fecal IgA is poorly documented, although it is of theoretical and practical importance to determine the different forms of IgA in faeces. Two main molecular forms were isolated by successive steps of ion-exchange chromatography and gel-filtration. The first consisted of secretory IgA dimers dissociating into slightly lower molecular mass forms under the influence of the electric field during electrophoresis. The other contained cleaved-IgA complexed with alpha 1-antitrypsin, that is considered to be a serum origin marker. These results confirm that secretory IgA are relatively resistant to digestive enzymes in vivo, and suggest that alpha 1-antitrypsin-bound fragments originate from serum IgA monomers. Analysis of the proportions of these forms may be of value in the investigation of gut diseases.

Effect of thiol derivatives on mixed mucus and blood clots in vitro.

The disintegrating effect of three reducing thiol derivatives: [sodium mercaptoethane sulphonate (Mesna), N-acetyl-L-cysteine (NAC) and dithio-1,4-threitol (DTT)] was investigated in vitro upon blood clots formed in the absence or in the presence of tracheobronchial secretions and compared with the effect of iso-osmotic saline solution. The amounts of haemoglobin released from the clots after 30 min incubation and the initial rates of haemoglobin release were compared for the different products at different concentrations. All three reducing agents showed some ability to disintegrate mixed clots to an extent depending on their concentration. After 30 min incubation, statistical analysis showed a highly significant difference in favour of Mesna at the three concentrations used, i.e. 0.1, 1.0 and 10 mmol/1. The initial rate of haemoglobin release in presence of Mesna was at all concentrations significantly higher than that of NAC or DTT. The effects on normal blood clots were much less pronounced. The effectiveness of Mesna in splitting up mixed blood and mucus clots in the management of patients who had inhaled blood is discussed.

Low levels of pancreatic elastase 1 in stools of preterm infants.

The amount of faecal pancreatic enzyme elastase 1 was significantly lower in 42 preterm newborns than in 12 full term babies at day 2 (89 (3-539) v 354 (52-600) microg/g, p<0.0007) and day 5 (164 (3-600) v 600 (158-600) microg/g, p<0.05) and correlated positively with total nutrient intake during the first week of life in preterm infants. This should probably be taken into account during early feeding.

In vitro excystation of Giardia from humans: a scanning electron microscopy study.

The in vitro excystation of Giardia lamblia on cysts isolated from human feces was studied. After purification by sucrose gradient, cysts were incubated in a pepsin-acid solution, then placed in a modified HSP3 medium where excystation occurred within a few minutes. The excystation procedure was studied by continuous observations by light microscopy and sequential observations by scanning electron microscopy (SEM). The in vitro excystation was stopped at timed intervals during incubation by addition of a large amount of 1% glutaraldehyde. The excystation process began by the cyst wall opening at one pole. Flagella protruded rapidly, the parasite emerged progressively from the cyst envelope, posterior end first, the empty cyst collapsed and shrank. Although flagella emerging from the organism were distinguishable, the cell body had not yet shown all the morphological features of the G. lamblia trophozoite. A radical rearrangement of the organism occurred gradually: initially oval in shape, the parasite became round, then elongated, flattened, and underwent cytokinesis. The daughter trophozoites acquired their typical morphological features: the shape, the adhesive disc with the C-shaped structure distinctly visible on the ventral surface, and the definite placement of the flagella. These observations obtained on G. lamblia by SEM were comparable to those obtained with G. muris.

[Protein-losing enteropathy syndrome caused by a localization of Kaposi's sarcoma in AIDS]. / Syndrome d'entéropathie exsudative dû à une localisation d'un sarcome de Kaposi au cours du SIDA.

A 35-year-old HIV positive patient with skin and gastrointestinal localization of a Kaposi's sarcoma was admitted for severe diffuse edema. Increased alpha-I-antitrypsin clearance (150 ml/24 h) allowed us to confirm the diagnosis of protein loosing enteropathy resulting from sarcoma infiltration in the stomach, the duodenum and the entire small bowel. At autopsy, ileal ulcerations were found. Gastrointestinal involvement occurring during Kaposi's sarcoma is common and usually symptomless. The discovery of a protein loosing enteropathy in our patient suggests that this gastrointestinal involvement could play a role in the hypoalbuminemia often found in these patients.

[Detection of protein p53 in the stool of patients with colorectal cancer]. / Détection de la protéine p53 dans les selles de malades atteints de cancer colo-rectal.

BACKGROUND AND AIMS: Mutations of TP53, a tumor suppressor gene, are found in 60% to 70% of colorectal cancers. These mutations usually induce an overexpression caused by modification of the p53 protein conformation. The aim of this study was to investigate whether stool specimens of patients with colorectal cancer contain increased amounts of p53 protein. METHODS: p53 protein was measured using a sandwich enzyme immunoassay in the stool specimens of 52 patients: 25 with colorectal cancer, 4 with colorectal adenomas and 23 apparently free of gastrointestinal disease. Results were expressed as pg/mg of total protein. The presence of fecal occult-blood was searched using Hemoccult II and Hemolex (an immunochemical assay for human hemoglobin). RESULTS: Median concentrations of stool p53 protein were 16.6 pg/mg (range: 0-591 pg/mg) in patients with colorectal cancers, 39.1 pg/mg (range: 5-72 pg/mg) in patients with adenomas and 5.9 pg/mg (range: 0-65 pg/mg) in control subjects. Resection of colorectal cancers caused a marked decrease of stool p53 protein concentrations. When the cut-off value for stool p53 protein was set at 60 pg/mg of fecal protein (concentrations over the 95th percentile), the positivity of the assay was independent of tumor size and Astler-Coller stage, but weakly associated with rectal location of cancer. The sensitivity of stool p53 protein for colorectal cancer was 44%, and the specificity was 96%. In contrast, the sensitivity of Hemoccult II and Hemolex tests was 48% and 44%, whereas their specificity was 91% and 96%, respectively. CONCLUSION: The detection of p53 protein is achievable in stool, but this assay is not more efficient than fecal occult blood tests for detection of colorectal cancer.

[Effect of fasting and refeeding on the adaptation of the small intestine in rats. A model for physiopathologic studies]. / Effet du jeûne et de la réalimentation sur l'adaptation de l'intestin grêle chez le rat. Un modèle d'études physio-pathologiques.

A chronological study was carried out on 50 male Wistar rats (350 g) to determine the effects of 3 days of fasting and 16 h to 9 days of refeeding on the morphology of jejunal and ileal mucosa (villus, crypt and enterocyte heights; number of mitosis), on some aspects of their functional adaptation (sucrase, maltase, protein) and on nitrogen and lipid absorptions. Three days of fasting resulted in weight loss (12 p. 100), in a jejunal mucosa atrophy (villus height: 376 +/- 18 vs. 492 +/- 4 microns in controls; enterocyte height: 31 +/- 2 vs. 41 +/- 0.3 micron in controls) and a decrease in disaccharidases activities (sucrase: 927 +/- 90 vs. 3,363 +/- 21 mU/10 cm length in controls). No change in ileal mucosa morphometry was noticed. Ad libitum refeeding caused a rapid and progressive weight gain, a jejunal morphometric regrowth identical to control values at 16 h (villus height: 521 +/- 20, enterocyte height 42 +/- 0.9 microns), and maximum at 40 h of refeeding (villus height: 601 +/- 5 microns). Disaccharidases adaptation was delayed, with a maximum at 64 h of refeeding (sucrase: 3,524 +/- 56 mU/10 cm length). Despite a 30 p. 100 increase of food consumption over the whole study (45 p. 100 during the first 16 h of refeeding), nitrogen and lipid absorption coefficients remained identical to those found in controls with an increased nitrogen balance of 70 p. 100 at 16 h and 54 p. 100 at 40 h refeeding, as compared to controls.(ABSTRACT TRUNCATED AT 250 WORDS)

A high-yield outbred suckling mouse model of cryptosporidiosis.

Outbred suckling mice (NMRI strain) were used as hosts. They were initially inoculated with oocysts of human origin, and subsequently with parasites recovered from the mouse ileal mucosa. Cryptosporidia were counted in an aliquot of whole-ileum homogenate. Parasite load was expressed as cryptosporidia per centimeter of ileum. Serial passage of C. parvum in NMRI mouse litters led to a gradual amplification of parasite burden relative to animals initially inoculated with the human isolate.

[Faecal occult blood testing: comparison of a latex agglutination test (Hemolex) and an immunoturbidimetric test (QuikRead FOB)]. / Recherche de sang occulte dans les selles: analyse comparative d'un test d'agglutination de particules de latex sensibilisées (Hémolex) et d'un test immunoturbidimétrique (QuikRead FOB).

UNLABELLED: Occult blood detection is the most prescribed faecal examination. AIM: To compare results obtained with the latex agglutination test Hémolex LA (Orion diagnostica, Finlande) with those given by an immuno-turbidimetric test which allows an automatic reading (QuikRead FOB, Orion diagnostica, Finlande). MATERIAL AND METHODS: this prospective study was carried out in 140 patients. The reference method was the latex agglutination test, Hemolex LA performed on stool extract obtained through weighting samples. On the base of the results, samples were separated into 2 groups: positive (n = 45) and negative (n = 95). As the QuikRead FOB test indicated a stool extract obtained through a sampling set, such an extraction was performed before Hemolex LA et QuikRead FOB testing. RESULTS: all the 95 samples from the negative group gave similar results with the 3 methods. In contrast, 12/45 of the positive samples gave conflicting results, 11 results were negative with the 2 tests performed on stool extract obtained via sampling set, 1 result was negative with the QuikRead FOB method only. DISCUSSION: analytical performance were similar with the 2 methods and discrepancies observed wi-thin the positive group were mainly related to the extraction method.

Detection of occult blood in stools: comparison of three gaiac tests and a latex agglutination test.

The detection of occult blood in the stools is the only simple screening method for colorectal cancer. The aim of this study was to compare the results obtained with the new Hemolex kit (Orion diagnostica, Fumouze France) with those given by three gaiac tests--Hemoccult (Smithkline diagnostics), Hemofec (Boehringer Mannheim) and Hemopreuve (Fumouze) of 165 stools from patients without special diet. Seventy-one patients with at least two positive gaiac tests or a positive Hemolex test underwent colonoscopy followed, if negative, by fibroscopy: 28 had lesions of the lower digestive tract and five of the upper digestive tract. Sensitivity, specificity and negative and positive predictive value were of 70, 98, 91 and 92% respectively for Hemolex; 82, 74, 94 and 44% for Hemoccult; 94, 67, 98 and 42% for Hemopreuve and 91, 73, 97 and 46% for Hemofec. The results obtained in this study confirm the value of the Hemolex test for the detection of human occult blood in the stools whereas the gaiac tests used are influenced by dietary components (unless restricted), explaining their poor positive predictive value. In conclusion, due to their good negative predictive values, the authors recommend that screening for colorectal tumours should be based on the use of two or three gaiac tests which should be confirmed, when positive, by an immunological test for human hemoglobin.

Biochemical and metabolic peculiarities of some parasites availing as targets for therapy.

Biochemical and metabolic peculiarities of some parasites involved in their interactions with their hosts are reviewed according to (1) carbohydrate metabolism comprising glycolysis, Pasteur effect, CO2 fixation and electron transport system; (2) amino acid and protein metabolism ; (3) purine and pyrimidine nucleotides metabolism. These peculiarities are becoming targets for treatment without affecting the host.

[Element of digestive pathophysiology and fecal analysis]. / Elément de physiopathologie digestive et coprologie fonctionnelle.

Fecal analysis includes qualitative and quantitative studies which allows quantification and labelling of numerous pathophysiologic phenomenona. Malabsorption and over-absorption of water and electrolytes give rise to six types of watery diarrheas, and two types of constipations; malabsorption of nutriments and maldigestion of food, give rise to two types of fatty and nitrogenous diarrheas with metabolic consequences. Fecal analysis often discriminates organic from non-organic diseases and brings informations on increase or decrease of caloric losses, to the nutritionist. Microscopic observations which requires a high degree of competence and experience, allows the recognition of malabsorption/maldigestion phenomenona, of fortuitous presence of parasites and a good interpretation of a fecal file.

[New fecal markers: recent developments and perspectives]. / Les nouveaux marqueurs disponibles en coprologie fonctionnelle et perspectives.

Fecal occult blood testing is the most widely prescribed screening test for colorectal cancer. Recent development of immunological tests has increased specificity. Fecal DNA analysis opens up a new field for early detection of this widespread neoplasia. Inflammatory bowel disease is another important area where the development of fecal markers provides an interesting alternative to the gold standard but costly and invasive endoscopic investigations with histological analysis of biopsy specimens. Fecal TNFalpha and calprotectin can now be proposed to distinguish organic from non-organic intestinal disease, so select candidates for further investigations, and to assess disease activity. Measurement of fecal elastase provides real progress in screening for exocrine pancreatic insufficiency in patients with malabsorption syndrome. The development of non-invasive fecal markers is thus of increasing interest, providing data about the entire gastrointestinal tract useful for screening and individual patient management.

[Quality control in coprology]. / L'assurance de qualité en coprologie fonctionnelle.

Quality control in medical laboratories was defined in guidelines for good execution of laboratory analyses issued by the French health authorities in 1994. Application of these guidelines is difficult in coprology because the sample is a complex heterogeneous matrix which varies with disease, surgery, food intake, and treatment. In addition, commercial quality control kits are not available for stool biochemical analyses and a national quality control program has not been established. We thus developed our own fecal quality control technique using pooling lyophylized stool samples. Manual or partially automated methods are used in coprology, leading to a long pre-analysis phase which is not always taken into account in quality control. This implies the need for complementary tools to insure the quality of coprology analyses. For example, semi-quantitative microscopic lipid analysis can be used as an internal standard for a given specimen. Quality assurance also involves a post-analytical phase where results obtained for a given specimen are compared with other available data and interpreted in light of the patient's clinical and therapeutic status. This quality assurance strategy enables accurate reliable results useful for long-term patient management.

[Cryptosporidium parvum: functional study of the intestinal malabsorption syndrome]. / Cryptosporidium parvum: étude fonctionnelle du syndrome de malabsorption intestinale.

Cryptosporidiosis is an important cause of diarrhea associated with growth retardation in children and severe malnutrition in immunocompromised patients. The pathophysiology is poorly understood. In the suckling rat model, we show that C. parvum infection impairs net electrogenic transport across the ileal mucosa without involvement of prostaglandins, as well as trans- and paracellular permeability and leucine and glutamate absorption. These results provide evidence for the development of an intestinal malabsorptive syndrome during cryptosporidiosis. Unspecific process such as villous atrophy and inflammatory cytokines secretion should be regarded as possible mediators of this syndrome. However, specific mechanisms have to be considered since C. parvum induces a rearrangement of the host enterocyte cytoskeleton which might impaired intracellular trafficking thus reducing the membrane expression of nutrient transporters. Infection and malnutrition are known to be tightly associated, making each other worse. As no specific efficient therapy exists, cryptosporidiosis-induced malnutrition must be taken into account when establishing therapeutic scheme.