[Epidemics of Ebola haemorrhagic fever in Gabon (1994-2002). Epidemiologic aspects and considerations on control measures]. / Les epidémies de fièvre hémorragique due au virus Ebola au Gabon (1994 - 2002): aspects epidémiologiques et réflexions sur les mesures de contrôle.

Based on the description of the four Ebola haemorrhagic fever epidemics (EHF) occurred in Gabon between 1994 and 2002, the authors are considering the cultural and psycho-sociological aspects accounting for the difficulty to implement control measures. On the whole, the result of these raging epidemics came up to 207 cases and 150 dead (lethality: 72%). Analysing precisely the aspects of the third epidemic and pointing up the possible factors explaining its spreading far beyond its epicentre, the authors bring about the limits of measures not always understood by local populations. The discussion will deal with the possibilities of a better surveillance, a quick management of intervention means including a regional permanent pre-alert and taking into account the issue raised by the possible Ebola virus endemic.

In vitro inactivation of Chlamydia trachomatis and of a panel of DNA (HSV-2, CMV, adenovirus, BK virus) and RNA (RSV, enterovirus) viruses by the spermicide benzalkonium chloride.

Kinetics of inactivation by the detergent spermicide benzalkonium chloride (BZK) of Chlamydia trachomatis and of a panel of DNA viruses [herpes simplex virus hominis type 2 (HSV-2), cytomegalovirus (CMV), adenovirus (ADV) and BK virus (BKV)] and RNA [respiratory syncytial virus (RSV) and enterovirus (ENV)] were established in accordance with a standardized in vitro protocol. After a 5 min incubation, inactivation of >95% of HSV-2 and CMV was obtained at a concentration of 0.0025% (w/v) (25 Ig/L); concentrations as low as 0.0005%, 0.0050% and 0.0125%, induced a 3.0 log10 reduction in infectivity of HSV-2 and CMV, RSV and ADV, respectively. After a 60 min incubation, concentrations of 0.0125% and 0.050% provided a 3.0 log10 reduction in infectivity of ENV and BKV, respectively. These features indicate that sensitivity to BZK was very high (HSV-2 and CMV) or high (RSV) for enveloped viruses, intermediate (ADV) or low (ENV and BKV) for non-enveloped viruses. Furthermore, BZK had marked antichlamydial activity, showing >99% killing after only a 1 min incubation at a concentration of 0.00125%. BZK demonstrates potent in vitro activity against the majority of microorganisms causing sexually transmitted infectious diseases, including those acting as major genital cofactors of human immunodeficiency virus transmission. These attributes qualify BZK as a particularly attractive candidate for microbicide development.

Protection of cynomolgus macaque against cervicovaginal transmission of SIVmac251 by the spermicide benzalkonium chloride.

We evaluated the potential effectiveness of a spermicide cationic surfactant, benzalkonium chloride (BZK), to prevent the transmission of simian immunodeficiency virus (SIV) after intravaginal inoculation in 12 cynomolgus macaques. The inoculation procedure involved deposition of 6.7 ivag-AID50 of cell-free SIVmac251 into the receiving vagina, four times over a 2-week period, at the end of the luteal phase of the menstrual cycle. Six randomly selected females received vaginally foam containing BZK (7.37%, wt/wt) before each inoculation (BZK group), whereas the remaining were not pretreated (control group). In controls, 5 animals presented persistent SIV infection and 1 had a transient viremia. The number of uninfected animals was higher in the BZK group (6 of 6) than in controls (0 of 6). These findings demonstrate that BZK placed in the vaginal receptacle prior to SIV inoculation provides a significant protection in vivo. The wide spectrum of antimicrobial activities of BZK (including HIV) in addition to its efficiency to block the transmucosal passage of SIV in the macaque model qualifies this drug as an attractive topical microbicide to prevent sexually transmitted infections in humans.

In vivo semen-associated pH neutralization of cervicovaginal secretions.

Physiological cervicovaginal acidity can partly inactivate human immunodeficiency virus (HIV). Basic semen components should be able to partially neutralize in vivo cervicovaginal pH. The goals of the study were to evaluate the relationship between cervicovaginal pH and presence of semen components in sexually active African women and to assess whether vaginal douching with water performed just after sexual intercourse could significantly reduce semen components and restore physiological cervicovaginal pH. Cervicovaginal secretion (CVS) from 56 heterosexual African women (19 to 45 years old), living in Bangui, Central African Republic, were evaluated for pH, semen components (prostatic acid phosphatase [PAP] and prostatic specific antigen [PSA]), cellularity, and hemoglobin at inclusion and after vaginal douching with 100 ml of water by using a bock. Before douching, semen components were found in 46 of 56 CVS (82%). The mean vaginal pH was 5.2 (range, 3.6 to 7.7), and concentrations of both PAP and PSA correlated positively and strongly with cervicovaginal pH (P < 0.001). After douching, semen components were found in 35 of 56 CVS (62%) (P = 0.03). Cervicovaginal PAP and PSA levels were significantly decreased (respectively, P < 0.0001 and P < 0.01; PAP, -72%; PSA, -87%), as was the total cell count (-60%; P < 0.0001). Furthermore, in CVS previously positive for both PAP and PSA, the mean vaginal pH was significantly decreased (6.5 versus 5.3, P < 0.01); no genital bleeding was observed. Frequent persistence of semen in CVS from heterosexually active African women leads to a shift from acidity to neutrality that could favor male to female HIV transmission. Vaginal douching provides significant elimination of semen after sexual intercourse; it should be considered for study as a supplementary means for the prevention of heterosexual HIV transmission.

[In vivo neutralization of physiological cervicovaginal acidity by sperm nitrogen bases]. / Neutralisation in vivo de l'acidité cervicovaginale physiologique par les bases azotées du sperme.

Cervicovaginal secretions (CVS) from 46 heterosexual African women, attending the National Reference Center for Sexually Transmitted Diseases and AIDS of Bangui, Central African Republic, were investigated, at inclusion and after vaginal douching with water, in order, i) to determine the relationship between cervicovaginal pH and the presence of semen components [prostatic acid phosphatase (PAP) and prostatic specific antigen (PSA)] in sexually active African women; ii) to assess whether vaginal douching performed after sexual intercourse could efficiently eliminate semen components and restore cervicovaginal acid pH. At inclusion, semen components were found in 41 CVS (89%); the mean cervicovaginal pH was 6.12 (range, 3.86 to 8.33); concentrations of both PAP and PSA correlated positively and strongly with cervicovaginal pH (p < 0.001). After douching, semen components were found in only 31 CVS (67%) (p < 0.03); vaginal PAP and PSA levels were significantly decreased (p < 0.0001); PAP: -21%; PSA: -36%). Frequent persistence of semen in cervicovaginal secretions from heterosexually active African women leads to a shift from acidity to neutrality, that could favor male-to-female HIV transmission.

Isolation and phylogenetic characterization of Ebola viruses causing different outbreaks in Gabon.

Three outbreaks of Ebola hemorrhagic fever have recently occurred in Gabon. Virus has been isolated from clinical materials from all three outbreaks, and nucleotide sequence analysis of the glycoprotein gene of the isolates and virus present in clinical samples has been carried out. These data indicate that each of the three outbreaks should be considered an independent emergence of a different Ebola virus of the Zaire subtype. As in earlier Ebola virus outbreaks, no genetic variability was detected between virus samples taken during an individual outbreak.

[Overproduction of tumor necrosis factor-alpha in African patients with HIV-1 infection, cachexia and low tri-iodothyronine syndrome]. / Surproduction de tumor necrosis factor alpha chez les malades africains infectes par le VIH-1, cachectiques et ayant un syndrome de tri-iodothyronine basse.

Sick euthyroid syndrome characterized by low triiodothyronine (T3) levels is observed in advance stages of HIV infection. The purpose of this prospective study was to determine if proinflammatory cytokines play and role in the pathogenesis of this syndrome in HIV-1-infected patients. Serum levels of tumor necrosis factor-alpha (TNF-alpha) and interleukin (IL)-1 beta were measured in 40 African patients presenting HIV-1 infection associated with low T3 levels in 20 cases (group I) and normal or elevated T3 levels in 20 cases (group II). Elevation of serum TNF-alpha levels was more common and mean serum TNF-alpha level was significantly higher in group I than group II (116 +/- 39 versus 3.05 +/- 0.04 pg/ml; p < 0.01). Serum IL-1 beta levels were not significantly different between the two groups. These findings are consistent with previous experimental data and suggest that sick euthyroid syndrome in cachectic HIV-1 infected patients may be due to overproduction of TNF-alpha.

Comparison of cervicovaginal humoral immunity in clinically asymptomatic (CDC A1 and A2 category) patients with HIV-1 and HIV-2 infection.

Paired sera and cervicovaginal secretions (CVS) from 11 HIV-1- and 11 HIV-2-infected women, all clinically asymptomatic (CDC A1 and A2 categories), were analyzed for total IgG, IgA, albumin (HSA), IgG, and IgA antibodies to env-encoded surface glycoproteins of HIV-1 (gp160) and of HIV-2 (gp105), by comparison to 15 age-matched healthy controls. Secretion rates of IgG and IgA into CVS were evaluated by calculation of their relative coefficients of excretion (RCE) by reference to HSA. Cervicovaginal production of anti-HIV antibodies was evaluated by comparison between specific antibody activities of IgG and of IgA to HIV in CVS were, respectively, 6- and 4-fold increased, whereas the secretion rate of total IgG was 2.1-fold increased and that of total IgA was 2.5-fold reduced. In contrast, total IgG and IgA as well as their secretion rates were normal in HIV-2-infected women. In HIV-1- but not in HIV-2-infected women, HSA levels in cervicovaginal washings were twofold increased, demonstrating alteration of the mucosal barrier in HIV-1 infection. In HIV-1-infected patients, IgG and IgA to gp160 were detected in all sera and CVS. In HIV-2-infected patients, IgG to gp105 was detected in all sera and CVS, whereas IgA to gp105 could be detected in only half of sera and one-third of CVS. Cross-reactivity by IgG and/or IgA to HIV-1 or HIV-2 against the surface glycoprotein of the other HIV type was observed in sera as well as in CVS, and more frequently in HIV-2- than in HIV-1-infected women. Finally, the mean specific activities of IgG and of IgA to gp160 or gp105 were higher in CVS than in sera, evidencing a possible local synthesis of both isotypes in HIV-1 as well as in HIV-2 infections. As early as the asymptomatic stages, HIV-1 affects the cervicovaginal mucosa more than HIV-2 does, suggesting higher viral replication within the female genital tract in HIV-1 infection than in HIV-2 infection.

Cervicovaginal overproduction of specific IgG to human immunodeficiency virus (HIV) contrasts with normal or impaired IgA local response in HIV infection.

Paired sera and cervicovaginal secretions (CVS) from 30 women infected with human immunodeficiency virus (HIV) type 1 (before AIDS) were analyzed for IgG and IgA antibodies to HIV and for IgG, IgA, and human serum albumin. Subjects were compared with 30 aged-matched healthy controls. In HIV-infected women, cervicovaginal immunoglobulins were markedly increased, and IgG predominated. An increased immunoglobulin transudation was implicated, since cervicovaginal albumin levels were 2.3-fold above those of normal controls. Furthermore, IgG excretion by reference to albumin was increased 1.9-fold, whereas the IgA secretion tended to decrease, suggesting a possible enhanced local IgG synthesis. Mean IgG and IgA anti-HIV antibody titers were, respectively, 30- and 12-fold higher in serum than in CVS, but their mean specific activities were higher in CVS than in serum, suggesting a local synthesis of both isotypes. The IgA antibody response to HIV remained poor compared with the strong IgG response.

Proinflammatory cytokine expression in cervicovaginal secretions of normal and HIV-infected women.

Proinflammatory cytokines may stimulate replication and spread of HIV. To evaluate to what extent the female genital tract represents a source of these cytokines, we determined TNF-alpha, IL-1 beta and IL-6 concentrations in paired serum and cervicovaginal washings from 45 HIV-negative and 50 HIV-positive women, and then we looked for the relevant mRNAs in cervicovaginal secretions by RT-PCR. Cytokines were detected by ELISA in cervicovaginal fluid from most healthy women. Cervicovaginal washing levels of TNF-alpha were increased above the control value +2 SD in 11/50 HIV-positive women, those of IL-1 beta in 13/50, and those of IL-6 in 14/50. The prevalences of TNF-alpha, IL-1 beta and IL-6 increase and their levels in cervicovaginal washings were significantly higher in the 20 patients at stage IV than in the 30 patients at earlier stages of the disease. In HIV-infected patients, serum and cervicovaginal washing levels correlated positively for TNF-alpha and IL-6, but not for IL-1 beta. Nine of 15 cytokine mRNAs determinations were positive in HIV-infected women versus 1 of 15 in controls (P < 0.01). These findings could be relevant to bidirectional heterosexual transmission of HIV.

Human immunodeficiency virus type 1 p24 antigen in cervicovaginal secretions.

The failure to detect human immunodeficiency virus (HIV) antigen in cervicovaginal secretions (CVS) from HIV-infected women could be due in part to an in situ formation of immune complexes involving HIV antigens and cervicovaginal anti-HIV antibodies. CVS from 30 HIV-1-infected heterosexual women were assayed for p24 antigen detection by ELISA before and after acid disruption of immune complexes. Before acid treatment, free p24 antigen was detected in only 1 sample of CVS, whereas after acid dissociation, 4 (13.5%) of 30 samples had detectable and neutralizable p24 antigen. Adsorbent capacities of the CVS for recombinant p24 antigen, evaluated in an in vitro model, depended on both titer and avidity of cervicovaginal antibodies to HIV. In conclusion, local antibodies to HIV are able in vivo to bind HIV antigen within CVS and to participate in the immune exclusion of free virus particles or proteins.

Local synthesis of IgG antibodies to HIV within the female and male genital tracts during asymptomatic and pre-AIDS stages of HIV infection.

Paired sera and cervicovaginal secretions or seminal fluids, obtained from HIV-1-infected, clinically asymptomatic women (n = 41) and men (n = 12), were investigated in order to test the hypothesis of a local synthesis of IgG to HIV in the female and male reproductive tracts. Anti-gp41 + p24 IgG was evaluated by an IgG immunocapture assay, and anti-gp160 IgG by an indirect ELISA. Estimation of anti-HIV IgG-specific activities was carried out after ponderal determination of total IgG and evaluation of anti-HIV IgG activity. IgG to gp41 + p24, as well as IgG to gp160, were specifically detected in all sera, cervicovaginal secretions, and seminal fluid samples from all tested HIV-1-infected subjects. The mean specific activities of IgG to gp41 + p24 in cervicovaginal secretions and in seminal fluids were about 33-fold (in women) and 16-fold (in men) that of the corresponding sera; similarly, the mean specific activities of IgG to gp160 in genital secretions were about 17-fold (in women) and 10-fold (in men) that of the corresponding sera. IgGs to HIV are constantly detected in genital secretions from HIV-1-infected subjects, and appear to be largely synthesized in situ within the genital tract of both genders.

Dilution assessment of cervicovaginal secretions obtained by vaginal washing for immunological assays.

Local immunological defense mechanisms in the cervicovaginal mucosa currently remain incompletely defined, especially from a quantitative point of view. Addition of an inert substance, lithium chloride (LiCl), into the washing buffer used to carry out the vaginal washing for collecting cervicovaginal secretions and measurement of its concentration with a flame absorption spectrophotometer, before and after the specimen is sampled, permits the quantification of the volume of cervicovaginal secretions collected and the approximation of the dilution factor of a soluble component introduced by the washing. Lithium, at a concentration of 10 mM, gives the best precision of measurement and has no effect on the results of the immunoassays. In a population of 27 nonpregnant women (age range, 18 to 45 years), the volume of cervicovaginal secretions collected by vaginal washing with 3 ml of LiCl-phosphate-buffered saline was 12% +/- 3.2% (mean +/- standard deviation) of the total volume and showed large interindividual variations (range, 5.6 to 18.8%); the mean dilution factor of a soluble component from the vaginal secretions was 9.9% +/- 2.8% (range, 6.3 to 18.8%). According to the date of the menstrual cycle, the mean volume of collected cervicovaginal secretions was significantly increased in the luteal phase in comparison with the follicular phase; conversely, the mean dilution factor of a soluble component was more important in the follicular than in the luteal phase. These features strengthen the need to quantify accurately the dilution factor introduced by vaginal washing when studying cervicovaginal immunity.

Detection of seminal antibodies to human immunodeficiency virus in vaginal secretions after sexual intercourse: possible means of preventing the risk of human immunodeficiency virus transmission in a rape victim.

Detection of semen anti-human immunodeficiency virus (HIV) antibodies within the cervicovaginal secretions from a non-HIV-infected woman who has had a recent sexual intercourse with an HIV-infected man is theoretically possible since the seminal fluid from all HIV-infected men contains a high titer of IgG antibodies to HIV. We report the case of an HIV-seronegative African woman whose cervico-vaginal secretions contained IgG antibodies to HIV, including antibodies to HIV-env-encoded glycoproteins. This woman had also detectable prostatic specific antigens and acid phosphatase in her cervico-vaginal secretions, establishing the persistence of semen. In order to confirm whether anti-HIV antibodies in seminal fluid could be detected in vitro when mixed with cervico-vaginal secretions, 10(-1) to 10(-6) 10-fold dilutions of seminal fluid from HIV-1-seropositive donors were realized with a pool of HIV-negative cervico-vaginal secretions as diluent. Six commercial enzyme immunoassays or rapid tests were compared for semen anti-HIV detection in the secretions. At a 10(-1) dilution of the mixture, all assays were markedly positive for all tested semens and the greatest dilutions of seminal fluid showing positivity ranged from 10(-3) to 10(-5). The IgG immunocapture assay appeared to be the most sensitive test. The rapid tests permitted the detection of semen IgG antibodies to HIV at dilutions ranging from 10(-1) to 10(-3) suggesting their potential value in emergency situations.

[Alternative strategy for the diagnosis of HIV infection in sub-saharan Africa. Value of the sequential combination of the ELISA test and a 2nd generation rapid test]. / Stratégie alternative pour le diagnostic de l'infection par le VIH en Afrique subsaharienne. Intérêt de la combinaison séquentielle d'un test ELISA et d'un test rapide de 2e génération.

The "conventional" algorithm for HIV testing based on the confirmation of all positive anti-HIV screening reactions by Western blot requires sufficient laboratory facilities and is expensive, that limits its use in developing countries, such as in subsaharian Africa. The apparition of second and third generation screening ELISA which are very sensitive and specific, as well as the development of rapid tests which are simple, visually read, and sufficiently sensitive and specific, has permitted the design of "alternative" strategy for HIV testing utilizing the association of 2 ELISA and/or rapid tests, in order to limit the use of a confirmatory assay. Alternative strategies are less expensive, yield generally very high sensitivity and specificity, and have proved to be valuable for African countries. In this study, 5 alternative strategies, using different associations of two second generation screening tests, one classical ELISA (Genelavia mixt) and one rapid test (Test PACK HIV-1/HIV-2 AB) have been retrospectively evaluated in the field in Bangui, Central African Republic, with a panel of 130 sera (prevalence of HIV infection: 42.7%). The strategy using two sequential screening tests (Test Pack HIV-1/HIV-2 AB following by Genelavia mixt) with the confirmation of discordant results by Western blot permitted to diagnose HIV-1 infection in Bangui with a sensitivity, a specificity and a positive predictive value of 100%, and to reduce the cost of more than 50% in comparison with the conventional strategy. Such an alternative strategy could be useful for the individual notification of HIV serology in Bangui.