A comparative proteomic analysis of Gluconacetobacter diazotrophicus PAL5 at exponential and stationary phases of cultures in the presence of high and low levels of inorganic nitrogen compound.

A proteomic view of G. diazotrophicus PAL5 at the exponential (E) and stationary phases (S) of cultures in the presence of low (L) and high levels (H) of combined nitrogen is presented. The proteomes analyzed on 2D-gels showed 131 proteins (42E+32S+29H+28L) differentially expressed by G. diazotrophicus, from which 46 were identified by combining mass spectrometry and bioinformatics tools. Proteins related to cofactor, energy and DNA metabolisms and cytoplasmic pH homeostasis were differentially expressed in E growth phase, under L and H conditions, in line with the high metabolic rate of the cells and the low pH of the media. Proteins most abundant in S-phase cells were stress associated and transporters plus transferases in agreement with the general phenomenon that binding protein-dependent systems are induced under nutrient limitation as part of hunger response. Cells grown in L condition produced nitrogen-fixation accessory proteins with roles in biosynthesis and stabilization of the nitrogenase complex plus proteins for protection of the nitrogenases from O(2)-induced inactivation. Proteins of the cell wall biogenesis apparatus were also expressed under nitrogen limitation and might function in the reshaping of the nitrogen-fixing G. diazotrophicus cells previously described. Genes whose protein products were detected in our analysis were mapped onto the chromosome and, based on the tendency of functionally related bacterial genes to cluster, we identified genes of particular pathways that could be organized in operons and are co-regulated. These results showed the great potential of proteomics to describe events in G. diazotrophicus cells by looking at proteins expressed under distinct growth conditions.

Passive haemagglutination tests using purified antigens covalently coupled to turkey erythrocytes.

Passive haemagglutination tests have been developed by covalent coupling purified antigens to turkey red blood cells. Circulating antibodies can be assessed in 20 minutes using one drop of blood. False positive reactions are avoided by using highly purified antigens; sensitized erythrocytes are stable in the absence of freeze-drying and blood samples can be preserved on paper discs. This method, applied to the determination of circulating tetanus (T) and diphtheria (D) antibodies and titres compared to other in vivo or in vitro methods, gave good correlation. The titration of circulating antibodies can be applied in emergency care units and field trials to establish whether the individuals are adequately protected. Results of surveys by several health care centres have shown that tetanus immune coverage was insufficient in France. The decrease of both T and D immune coverage with age has been established. The antibody response of pregnant women, vaccinated with two different adsorbed T toxoids exhibiting a low and a high titre as expressed in international immunizing units (I.I.U.), was studied. No significant difference in circulating antibody titres was obtained after the first injection of either vaccine, but titres after second injection were much higher for the vaccine having a low value expressed in I.I.U. The activity of commercial and reference T vaccines can be evaluated in mice after immunization and titration of the antitoxin levels. This simple method is much easier than the official evolution of immunodeficiency in certain diseases. The passive haemagglutination test has also been used to measure anti-HBs and anti-gp 160 antibodies.

Rabies and brucellosis immunization status and adverse reactions to rabies vaccines in veterinary students.

Rabies pre-immunization has been recommended for high risk professions, including veterinarians. Cell-cultured rabies vaccines have considerably reduced the risk of post-vaccination neurological reactions found in earlier vaccines. However, some adverse reactions have been reported with Human Diploid Cell Vaccines. 329 French veterinary students were surveyed about their rabies and brucellosis vaccination status, the occurrence of adverse reactions to rabies vaccine, and their antibody titer monitoring practices. Questions also were asked to determine if mandatory rabies pre-exposure immunization upon entry to veterinary school motivated students to maintain their rabies pre-exposure vaccination. The overall vaccination rate was 98.5% for rabies and 17% for brucellosis. 19% of the rabies vaccinated students reported some form of adverse reaction, whatever the vaccine brand used, but not experienced systemic allergic reaction. Adverse reactions were twice more frequent in female than male students and were more frequent after primary series than revaccination series (Relative Risk = 1.76). Despite the mild reactions encountered, rabies pre-exposure vaccination has been well-accepted by French veterinary students. In contrast, vaccination against brucellosis was not as well-accepted for prophylaxis.

[Vaccination against hepatitis B virus at the Lyon Pasteur Institute. A seven-year evaluation]. / Vaccination contre le virus de l'hépatite B à l'Institut Pasteur de Lyon. Bilan de sept ans.

Results of immunization against hepatitis B among Pasteur Institute staff members are reported. Prior to immunization, 439 subjects were tested for hepatitis B virus (HBV) markers, including HBs antigen, anti-HBs antibody, and anti-HBc antibody (Ausria, Ausab, Corab assays; Abbott). Forty-seven subjects tested positive for anti-HBs antibody. 317 subjects negative for all the HBs markers studied were given three intramuscular doses of Hevac B (Pasteur vaccins) at one-month intervals. Anti-HBs antibodies were assayed after the third injection with the following results: mean titer, 1,454 mIU/ml, standard deviation, 5,349 mIU/ml, and range, 4 to 41,100 mIU/ml. Anti-HBs titers above 10 mIU/ml were found in 879.4% of subjects. Non-responders and weak responders (anti-HBs titer under 10 mIU/ml) were given a fourth dose of vaccine. Ultimately, after the last (third of fourth) injection 97.6% of subjects had protective antibody titers. No case of HBV infection was seen during the seven-year follow-up period.

[Antitetanus vaccination by Polan T in the elderly: followed of the immune response by Vaccitest T after vaccination and booster]. / Vaccination antitétanique par Polan T dans une population âgée: suivi de la réponse humorale par le Vaccitest T après vaccination et rappel.

Tetanus vaccine activity without mineral adjuvant (Polan T) has been measured in a population of aged people (36 females and 11 males-mean age = 80.18 years). Titrations by hemagglutination of sensitized turkey red cells are performed before and after primo-immunization and first booster. This research gives knowledges regarding to the antitetanus response in elderly, regarding to the efficiency of the Polan T vaccine in this population uneasy to immunize and regarding to the quality of the simple and cheap assay for antitetanus antibodies titration.

[Biologic risk]. / Le risque biologique.

In the biotechnology development, the biohazard become an important field of environmental medicine. After evaluation of biological agents, the laboratory organization in different security steps in achieved. One chapter "Health and Security" must be included in every scientific or industrial scheme of procedures. One example is given with the schedule for Hepatitis B and AIDS prevention.

Influence of food on the bioavailability of diltiazem and two of its metabolites following the administration of conventional tablets and slow-release capsules.

The influence of food on the bioavailability of a conventional tablet and of a slow-release capsule of diltiazem was investigated in two separate groups of 24 healthy volunteers in two open crossover studies. Diltiazem, as a conventional tablet (2 x 30 mg, first group) or as a slow-release capsule (120 mg SR, second group), was administered in a fasting condition and 30 min after a breakfast of 784 kcal (23 per cent proteins, 55 per cent lipids, and 22 per cent of carbohydrates). Multiple blood samples were withdrawn during the next 24 h and diltiazem, desmethyldiltiazem, and deacetyldiltiazem were assayed by HPLC. Neither the rate of absorption, assessed by the rate constant of absorption, the peak plasma concentration, and the time required to reach the peak, nor the amount of drug reaching the systemic circulation, assessed by the area under the plasma concentration time curve (AUC infinity) were influenced by food, and that independently of the formulation. Compared to the fasting experiment, food did not affect either the rate of formation or the AUC infinity of desmethyldiltiazem or deacetyldiltiazem. The results of the present study show that the relative bioavailability of the single dose of diltiazem administered as a slow-release capsule is significantly higher (69 per cent) than that estimated after the administration of diltiazem in a conventional tablet. It was concluded that food does not influence the bioavailability of diltiazem administered as a conventional tablet or as a slow-release formulation.

Human monoclonal antibodies with a protective activity against tetanus toxin.

The antibodies and their protective activity in response to tetanus toxoid in man were studied by producing human antitetanus monoclonal antibodies after transformation of peripheral blood lymphocytes with Epstein Barr virus. Two human monoclonal IgG that reacted with the heavy chain of the toxin were obtained. One of them binds the COOH-terminal moiety and the other the NH2-terminal moiety. Only the NH2-terminal specific monoclonal antibody neutralized toxin in mouse, but in doses approximately 100-fold higher than those of a polyclonal antiserum. However, the association of these 2 antibodies was protective with doses lower than necessary for the monoclonal antibodies alone. To replace the polyclonal antibodies used, a good protection could be achieved by mixed human monoclonal antibodies against different epitopes of tetanus toxin.

Myasthenia gravis after general anesthesia and hepatitis B vaccine.

A 48-year-old man presented with the first symptoms of myasthenia gravis one month after a general anesthesia and a second dose of hepatitis B plasma vaccine. Whether either event may have acted as a nonspecific challenge to the patient's immune system is speculative, but the case is described to discover similar observations, if any.

[The phenomenon of "early and stimulated death" in mice infected and then vaccinated against rabies]. / Le phénomène de "mort précoce et stimulée" chez la souris infectée puis vaccinée contre la rage.

Live (diluted) and inactivated rabies vaccines of low antigen content induce early and enhanced death in mice, inoculated before vaccination with a wild type of rabies virus. Such vaccines, which neither induce interferon nor protect, produce a low level of antibodies which appear later than with vaccines of higher antigenicity. It is recommended to examine rabies vaccines not only by the usual (pre-exposure) potency test-NIH test or modified NIH test (one vaccination), but also by a post-exposure potency test.

Combined tetanus-rabies vaccination.

Studies in mice have shown that Calcium Phosphate adsorbed tetanus toxoid (IPADT) can be used as a vehicle for freezedried rabies vaccine. Trials were undertaken in human volunteers and patients receiving a post-exposure treatment using the same vaccines to evaluate tolerance and antibody response to both vaccines.

Novel assay and pharmacokinetics of levamisole and p-hydroxylevamisole in human plasma and urine.

A new gas chromatographic method was developed for the quantification of levamisole in human plasma and urine, using a nitrogen-phosphorus flame ionization detector. The adsorption of the drug onto glass was prevented by treating the glassware with a siliconizing agent. The sensitivity of the assay was 10 ng ml-1 and as low as 2 ng ml-1 can be detected in plasma. The urinary metabolite p-hydroxylevamisole was analysed by high performance liquid chromatography with ultraviolet detection. The sensitivity of this assay was 0.50 micrograms ml-1. Plasma and urinary concentrations of levamisole were determined in 10 healthy volunteers including seven men and three women following the administration of a single 150 mg dose of levamisole. Levamisole was rapidly absorbed (tmax 1.5 h), giving a peak plasma concentration of 716.7 +/- 217.5 ng ml-1. The plasma elimination half-life of levamisole was 5.6 +/- 2.5 h. Only 3.2 +/- 2.9 per cent of the oral dose was recovered as unchanged drug in the urine, suggesting the importance of clearance of levamisole by routes other than the kidney, and most probably by hepatic metabolism. The urinary concentrations of p-hydroxylevamisole were determined before and after hydrolysis of the urine samples with beta-glucuronidase, and the level of conjugation of the metabolite with glucuronic acid was then estimated. Cumulative recovery of the metabolite accounted for 1.6 +/- 1.1 per cent and 12.4 +/- 5.5 per cent of the oral dose of levamisole before and after hydrolysis, respectively, indicating that p-hydroxylation is a relatively important route of metabolism of levamisole, and that the p-hydroxylated metabolite is excreted mainly in conjugation with glucuronic acid. Except for the absorption rate of levamisole which is approximately twice as rapid in women as in men, there is no marked difference in the pharmacokinetics of levamisole between healthy men and women.

[Activities of the diagnosis centre of the Institut Pasteur de Lyon 1976-1980]. / Bilan d'activité du centre de diagnostic de l'Institut Pasteur de Lyon de 1976 à 1980.

A report is presented of five years of work in the Rabies Diagnosis Centre at Lyon. This Centre, supported by the Ministry of Health at the Institut Pasteur of Lyon, investigates the diagnosis in animals of having contaminated humans. During these five years 4012 samples from animals were analysed, 179 of which being diagnosed as rabid (i.e. 4.53%). Four samples from humans were also analysed, one of which being positive. Data collected concerning rabid animals are compared to national or regional data. Studies of various methods used (immunofluorescence test, inoculation in mice, histology--Dr. J.-A. Grimaud and Dr. M. Chevallier) for diagnosis allowed evaluation of each of them. Some of these analyses were completed by bacteriological studies (Dr. E.-N. Guillermet). Besides this diagnosis the Centre takes part in epidemiological study of the disease, maintains a virus collection and allows diagnosis of other zoonosis or anthropozoonosis useful for veterinarians and for better protection of human health.