Plasma viraemia as a marker of viral replication in HIV-infected individuals.

Two hundred and twenty-eight blood samples from 154 HIV-seropositive subjects were investigated for the presence of HIV in peripheral blood mononuclear cells (PBMC) and plasma (without prior ultracentrifugation or filtration), using normal PBMC as the target for replication. HIV was recovered from PBMC and plasma in 80.5 and 19.5% of the patients, respectively. Plasma viraemia was significantly associated with clinical manifestations of HIV infection, indicating that HIV replication increased as disease progressed. This was confirmed by the statistically significant correlations between plasma viraemia and low CD4 cell counts (P less than 0.01) and low anti-p24 antibody titers (P less than 0.01). On patient follow-up, detection of HIV in plasma was transient. p24 antigenaemia was only detected in 13.6% of cases and was also associated with advanced clinical stages of the disease. When HIV RNA detection by polymerase chain reaction (PCR) was compared with plasma viraemia, HIV RNA was detected in plasma in all symptomatic cases and in 53.8% (seven out of 13) of asymptomatic patients [Centers for Disease Control (CDC) stages II and III], confirming that PCR was far more sensitive than plasma culture. These results indicate that cell-free virus production is associated with the clinical stage of HIV infection and may serve as a marker for disease progression. Detection of HIV RNA by PCR appears to be the most sensitive method to detect viraemia.

HIV detection in culture of peripheral blood mononuclear cells: a relevant in vitro correlate of the antiviral effect of ZDV in vivo.

To better assess the antiviral effect of zidovudine (ZDV) in vivo and understand its limitations, we have studied human immunodeficiency virus (HIV) replication in peripheral blood mononuclear cell (PBMC) cultures from 25 ZDV-treated patients and 20 untreated controls. Three months after initiation of therapy, 9 of the 25 treated cases became negative for HIV isolation (36%). Untreated cases never converted to a negative culture. In patients treated by ZDV and who remained culture positive, the kinetics of HIV replication in PBMC culture was found to vary with time. A statistically significant delay in the production of HIV in PBMC cultures from treated cases could be demonstrated after 3 months of ZDV therapy, when compared with untreated patients. By contrast, in such untreated patients the time required to reach the peak of reverse transcriptase activity in culture decreased during the follow-up period. These changes of in vitro HIV replication kinetics as well as the change to negative culture during ZDV therapy probably reflected the reduction of the number of infected cells in vivo. These results as well as the decrease of p24 antigenemia do indicate that ZDV indeed inhibits HIV replication in vivo. However, the effect of ZDV on HIV replication kinetics in PBMC fails to reach significance at 6 months, suggesting that the antiviral effect of ZDV may decrease over time. Our results suggest that ZDV is most effective when the intensity of HIV replication in vivo is still low.

Comparison of HIV detection by virus isolation in lymphocyte cultures and molecular amplification of HIV DNA and RNA by PCR in offspring of seropositive mothers.

An early and accurate diagnosis of HIV infection is needed in the offspring of seropositive mothers. To this end, we have used two techniques for the direct detection of HIV in 12 newborns tested within 2 weeks after birth and 12 children. HIV isolation was carried out in lymphocyte cocultures and compared with detection of DNA and RNA sequences by molecular amplification using the polymerase chain reaction (PCR). In lymphocyte cocultures, HIV was isolated in 8 of 24 cases (33%), including 3 newborns, 3 symptomatic children, and 2 asymptomatic ones. HIV DNA was detected by PCR in twice as many cases, i.e., in 16/24 cases (66%), including 7/12 newborns, 4/4 symptomatic children, and 5/8 asymptomatic ones, 2 of whom became seronegative, HIV RNA was detected in 10 of 16 cases (60%) with detectable HIV DNA, including all of the cases who had a positive HIV isolation. Only children with clinical or biological signs of HIV infection were positive for HIV RNA. Furthermore, signs of HIV infection appeared within 6 months in three of the four newborns who were positive for HIV RNA at birth. These results indicate that HIV DNA detection by PCR is far more sensitive than HIV isolation in culture for the early diagnosis of HIV infection in offspring of seropositive mothers. HIV RNA detection appears to be a useful prognostic marker since it does correlate with disease progression and may serve as a clue for HIV replication in vivo.

Lack of reactivity of anti-human immunodeficiency virus (HIV) P17/18 antibodies against alpha 1 thymosin and of anti-alpha 1 thymosin monoclonal antibody against P17/18 protein.

The blood rate of alpha 1 thymosin is increased during HIV infection, despite the thymus involution. Anti-alpha 1 thymosin antibodies inhibit HIV replication in vitro. A homology between alpha 1 thymosin and the HIV P17/18 core protein exists and would explain a cross-antigenicity. We have studied the interaction between anti P17/18 antibodies from HIV patients and alpha 1 thymosin and between an anti-alpha 1 thymosin monoclonal antibody and the P17/18 protein. We were unable to confirm any cross-reactivity. During acquired immune deficiency syndrome, a major involution of the thymus appears with a severe depletion of thymocytes and epithelial cells. Certain thymic functions are missing, as corroborated by the reduction of the hormone thymulin in the blood. At the same time, the blood rate of the 2 other hormones (partly of thymic origin), alpha 1 thymosin and beta 4 thymosin is increased. One of the theories explaining this discordance is that patients with acquired immunodeficiency syndrome produce molecules which have a cross antigenicity with these thymic hormones. Sarin et al. have recorded a 50% homology between the C-terminal part (last 18 aminoacids) of alpha 1 thymosin and the part between the 92nd and the 109th aminoacids of the HIV P17/18 protein. The cross reactivity between this P17/18 protein and alpha 1 thymosin would explain the high rates of alpha 1 thymosin found in the radio-immunoassay of sera from patients infected with HIV. Another result of this cross-reactivity is the ability of alpha 1 thymosin antibodies to inhibit HIV replication in the H9 permissive cell line.(ABSTRACT TRUNCATED AT 250 WORDS)

Relevance of the quantitative detection of HIV proviral sequences in PBMC of infected individuals.

Changes in HIV replication during progression of HIV infection were assessed by estimation of the number of peripheral blood mononuclear cells (PBMC) harboring HIV proviral sequences. Samples from 23 patients at different stages of HIV infection were analyzed by quantitative polymerase chain reaction (PCR) using GAG primers. HIV titers in PBMC were also determined by serial dilutions of cells in coculture with phytohemagglutinin-activated normal PBMC. A positive correlation was observed between the number of HIV DNA copies and the HIV titer in PBMC. The PCR test was more sensitive than the coculture technique. The number of HIV copies detected by PCR ranged from 50 to 10,500 per 10(6) PBMC: assuming one copy per cell this implies a frequency of proviral HIV-containing cells of one per 100 to one per 20,000 mononuclear cells. The mean number of HIV DNA sequences in PBMC was significantly lower in asymptomatic patients than in AIDS patients and patients with AIDS-related complex (ARC). In patients who progressed from asymptomatic infection to AIDS, the number of HIV DNA copies in PBMC rose, indicating an increase of HIV replication. These results show that the number of infected PBMC increases during clinical progression. However, some asymptomatic patients had a higher number of HIV DNA copies in their PBMCs suggesting that increased HIV replication precedes the appearance of clinical symptoms.

Hepatitis C: description of a highly sensitive method for clinical detection of viral RNA.

To improve the sensitivity of hepatitis C virus RNA (HCV RNA) detection in serum by 'nested' polymerase chain reaction (PCR), primers belonging to 5' non-coding (5'NC) regions were used to compare the classical phenol/chloroform technique by using the proteinase K and silica gel technique with guanidinium thiocyanate. The silica gel techniques was found to be more efficient and sensitive for the extraction and purification of viral RNA from serum samples. The silica gel technique also avoids contact with hazardous volatile chemicals like phenol and chloroform and provides a better protection for viral RNA. Furthermore, the RNA detection sensitivity was greatly improved by modifying the buffer for reverse transcription and PCR. Using silica gel extraction, and the modified buffer, viral RNA was detected in 699 sera from anti-HCV second generation ELISA positive patients. These sera were distributed in second generation RIBA confirmed, indeterminate and non confirmed groups with PCR positive rates of 71.4%, 45.7% and 15.8%, respectively. Two out of 227 ELISA negative patients showed the presence of HCV RNA in serum. An association between the presence of antibodies against a determined viral peptide and viremia was not detected.

Identification and detection of long incubation non-A, non-B hepatitis virus and associated antigens or antibodies.

Three distinct antigen/antibody systems supposedly associated with an HBV-like virus of non-A, non-B hepatitis have been identified. Because of previously demonstrated cross-reactivity with HBe/3 and HBc antigens and other analogies the following terminology is tentatively used. 1. The previously reported serum antigen has been redesignated non-A, non-B e antigen, since it is equivalent to HBe/3 Ag and cross-reacts with it. Non-A, non-BeAg or Ab were detected in 51/62 post-transfusion and 11/56 sporadic acute non-A, non-B hepatitis cases, and in 12/14 cases affecting staff members. In non-A, non-B chronic persistent or active hepatitis and cryptogenic cirrhosis, the prevalence was similarly high: 14/18, 22/48 and 12/18 respectively. Ten out of 26 implicated blood donors were found positive for non-A, non-BeAg accounting for 7 out of 8 post-transfusion cases. A high prevalence of non-A, non-BeAg was also found in haemophiliacs (11/48) and haemodialysed patients (6/42), whereas anti-non-A, non-Be was respectively detected in 4/48 and 6/42 of these cases. 2. Using immunofluorescence, a second antigen termed non-A, non-BcAg has been identified in liver biopsies from 55/84 non-A, non-B chronic hepatitis or cryptogenic cirrhosis cases. All 8 positive biopsies examined by electron microscopy revealed clusters of 22--25 nm intranuclear particles identical to those described in chimpanzees. Anti-non-A, non-Bc detectable by counter-electrophoresis and indirect immunofluorescence was found in the serum of all patients of which biopsy was positive for non-A, non-BcAg. Anti-non-A, non-Bc was also detected in 5/5 non-A, non-BeAg positive cases of post-transfusion hepatitis, 2--6 weeks after onset end remained positive for the 6 month follow-up period. 3. A third antigen, tentatively designated non-A, non-BsAg, has been found less frequently than non-A, non-BeAg in serum. However, it was detectable in 3/18 and 2/12 washed ultracentrifugation pellets of sera positive for non-A, non-BeAg or anti-non-A, non-Be, respectively.

Costs of nosocomial infection in a neonatal unit.

Hospital costs for 61 neonates with acquired nosocomial infection were compared with 61 matched, uninfected controls. The increase in length of hospital stay (+23 per cent) added to the number of laboratory tests increase total hospitalization costs to 32 per cent. An additional US $1250 for each case of infection is the cost to the Social Security system. The importance of this increase, contrasted with the generally benign nature of the infections studied, justifies the maintenance of high standards of quality in neonatal care.

A rapid and easy PCR-RFLP method for genotyping Serratia marcescens strains isolated in different hospital outbreaks and patient environments in the Lyon area, France.

A new genotyping method for Serratia marcescens is described. This method uses the flagellin gene as target for polymerase chain reaction amplification and Alu I restriction fragment length polymorphism. The strains tested belonged to 13 different hospital clusters of S. marcescens isolated between 1983 and 1988, concerning outbreaks and/or patient environments in different hospital units in Lyon and the Rhone-Alpes region of France. Initially, the classification had been performed by marcescinotyping. These strains were then tested by ribotyping and genotyping of the flagellin gene. Genotyping showed similar classification to ribotyping. The genotyping method is the easiest technique, as reproducible as ribotyping, and with almost the same ability to discriminate different strains. It does not need expensive equipment, is more rapid, and is less labor intensive than ribotyping. With this method, all strains of S. marcescens including sporadic isolates could be amplified and typed. Antibiotic sensitivity determination was found to be a useful complementary and confirmation test for all these typing methods.

[Medical follow-up of patients with positive serology for hepatitis C virus]. / Prise en charge médicale des malades séropositifs pour le virus de l'hépatite C.

OBJECTIVE: A screening campaign requested by the French Health Insurance Fund was performed in the Lyon area to access the prevalence of anti-hepatitis C virus in patients from 271 general practitioners and to observe the follow-up of the patients with had positive serology. A total of 101 patients had already had hepatitis C virus antibodies and 30 patients had newly detected antibodies against hepatitis C virus. All subject received appropriate medical care. The aim of this study was to analyse medical follow-up. METHODS: Follow-up was performed for one year by the patient's general practitioner or a specialist of liver disease. Clinical, biological, histological and virological data were collected and treatment was proposed when appropriate. Information was gathered anonymously by phone. RESULTS: Follow-up was regular for 28 patients (93%). During this follow-up, ALAT levels remained within the normal limit for 13 patients (43%) and 6 of them had undetectable viral RNA by PCR. In the other patients (50%), a liver biopsy was affered to 11 patients (40%), and performed in 9 (30%). Treatment was started in 8 patients. CONCLUSION: The results of this study on medical follow-up are satisfying. The general practitioner plays a key role in the follow-up of patients infected by hepatitis C virus.

[Epidemiology of acute hepatitis among outpatients in the urban community of Lyon, 1983]. / Epidémiologie des hépatites aiguës survenues chez des malades ambulatoires dans la communauté urbaine de Lyon, en 1983.

We conducted in 1983 an hepatitis surveillance programme in collaboration with 93% of the medical analysis laboratories in Lyons' urban area (1,100,000 inhabitants) and diagnosed 1,002 cases of acute hepatitis (incidence : 90.5 cases/10(5) inhabitants; HAV : 50.4 cases/10(5) inhabitants; HBV : 12 cases/10(5) inhabitants; non A non B : 24 cases/10(5) inhabitants; drugs : 4.2 cases/10(5) inhabitants). HBs Ag was undetectable in 12% of acute hepatitis B. Hepatitis A accounted for 91% of children's cases and 48% of cases among adults between 20 and 40 years old. Epidemics were observed among children after summer holidays related to the return of migrants from trips to their native country. For acute hepatitis B and non A non B, classical epidemiological data were observed, but the prevalence of drug addicts and homosexuals was low.

[Viral hepatitis: incidence and epidemiologic pattern in an urban area (author's transl)]. / Hépatites virales: incidence et caractéristiques épidémiologiques dans une agglomération urbaine.

In order to describe the incidence and the epidemiologic pattern of viral hepatitis (V.H.) in a French urban area, with a special attention to ambulatory cases, an epidemiological information system has been developed during one year. The physicians (sample), the medical laboratories and, at a lesser degree, the hospital care units, have taken part in this work. The incidence rate of ambulatory cares has been estimated 85 cases/10(5) persons (69/110), less than previously assumed. Non-B V.H. remains more frequent (52 cases/10(5) persons), but B.V.H. are rather close to them (33 cases/10(5) persons). Large differences exist between geographic areas, without evident rational. Higher incidence rates characterize children (especially 5-9 years) and migrant people (especially from North Africa): those facts are completely account by very high rates within the subgroup of migrant children (473 and 260 cases/10(5) persons). The reported patterns allows us to point on this high risk characteristic of migrant's children and to assume an important modification in the pattern of local inhabitants V.H. in the region. These trends have to be followed by further studies.

[Smoking among students of the CM2) (2d-year intermediate course): evaluation of knowledge and behavior]. / Tabagisme chez des élèves de CM2: évaluation des connaissances et des comportements.

This paper reports the results of a survey conducted on smoking among 1490 eleven years old schoolchildren. They answered to an individual questionnaire before and after they had been informed on smoking by school doctors. In this age group, 26.2% of the children have already had some contact with cigarette, and 0.5% smoke regularly. The household smoking habits have an important influence on the attitude of children toward smoking. Knowledges on smoking dangers have been evaluated before and after information. This survey shows the importance of smoking prevention, especially among young children.

[Hospital infection in the maternity department. 3 years of surveillance in 9,204 deliveries of which 1,333 were cesarean sections]. / Infection hospitalière en maternité. Trois années de surveillance portant sur 9 204 accouchements dont 1 333 césariennes.

Hospital or nosocomial infection, or infection acquired in hospitals, is a health problem in all hospital departments and particularly in the maternity department. We report on a prospective survey of surveillance of hospital-acquired infections both from the mother and the baby's point of view after delivery vaginally or with caesarean carried out at the obstetrical clinic of the Edouard Herriot Hospital in Lyon (France) over three successive years with a series of 9,204 deliveries. The incidence of infection in women who were delivered without caesarean section was 1.37% when urinary tract infections had been excluded but 13% in women who had caesarean sections. Endometritis, skin infections and urinary tract infections were the leading causes. As far as the newborn were concerned, hospital infection ran at about 2.60% and this in the main was due to staphylococcal pustules in the skin. These figures are still too high and prevention should be based on more information given and more care taken by the whole staff of such a hospital.

[Cost of acute hepatitis A in adults in France]. / Coût en France des hépatites A aiguës de l'adulte.

OBJECTIVES: Seroprophylaxy against hepatitis A has been limited to subjects visiting endemic areas or in contact with acutely infected patients due to the short half-life and high cost of the non-specific immunoglobulins. Inactivated vaccines might offer a better solution, but the real cost factors remain unknown. We undertook this study to determine the cost of acute hepatitis A. METHODS: Serum samples from all the non-hospital medical laboratories in the Lyon (France) area were prospectively screened to identify 100 consecutive patients with acute hepatitis (positive for hepatitis A IgM antibodies). A questionnaire was addressed to these 100 subjects 1 year later requesting information on health care expenditures and work loss related to their episode of acute hepatitis A. Imprecise answers were completed by telephone interview. RESULTS: Answers were obtained from 92 subjects. Confirmation was obtained that the 8 other subjects had not died from their episode of acute hepatitis. Only two patients were hospitalized (for 2 and 4 days). Total mean cost (including medical expenditures and work loss was 11,789 French francs (range 1,524-62,117). Medical consultations and prescriptions comprised 20.0% of the total cost. The real mean cost of work loss (8,423 F) was greater than the cost calculated by the national health service (6,500 F). CONCLUSIONS: This evaluation of the real cost of hepatitis A in non-hospitalized patients should contribute to the definition of an adapted policy for the prevention of hepatitis A.

[Nosocomial septicemia and pseudobacteremia caused by Serratia marcescens]. / Septicémies nosocomiales et pseudo-bactériémies à Serratia marcescens.

An epidemiological survey was carried out which included a dual epidemic of septicaemia and pseudo-bacteremia caused by Serratia marcescens. The survey enabled 15 septicaemias and 43 pseudobacteremias to be detected in a regional hospital between March and August, 1983. Two mishandlings were at the origin of the outbreak: citrated tube normally reserved for coagulation tests were severely contaminated by Serratia marcescens, and inaccurate samplings had been made. Once the mechanisms of contamination were found, specific preventive measures put an end to the epidemic. The authors insist on the need for uncontaminated tubes and citrate solutions and for the development of precise sampling methods which are essential to avoid the occurrence of pseudo-bacteremia or septicaemia. It is important to detect such epidemics at an early stage by an efficient control of nosocomial infections, thus avoiding their extension.

[General practice patterns in hepatitis C virus mass screening in the Lyons region]. / Pratique des médecins généralistes en matière de dépistage du virus de l'hépatite C dans la région lyonnaise.

A study on the screening practices for hepatitis C was carried out among a sample of 786 general practitioners in the Lyon region. The sample comprised three groups: 272 doctors who had participated in a study of this screening, 242 doctors who had not responded to the request for participation, and 272 doctors who had refused to participate. Overall, 88.5% of the doctors surveyed had prescribed at least one HCV serology in the last 12 months, 82% had at least one HCV positive patient and of them, 59.3% had partly or completely screened these patients. The only differences between the three groups are related either to their belonging to an HIV or drug addict care network, or to a particular medical practice (study of allergies, sexology, nutrition, etc.). In fact, the first prescribe more among groups at risk and the second prescribe less and have fewer HCV positive patients.