Detection of yellow fever viral RNA by nucleic acid hybridization and viral antigen by immunocytochemistry in fixed human liver.

Histopathologic examination of liver from patients with yellow fever is often not diagnostic. We therefore compared 2 virus-specific assays applicable to fixed liver, in situ nucleic acid hybridization and an immunocytochemical [alkaline phosphatase-antialkaline phosphatase (APAAP)] technique. Yellow fever structural gene sequences were detected by use of 35S-labeled negative-sense RNA probe (but not by immunocytochemistry) in 11 of 17 livers from children with fatal illness during the 1965 epidemic in Senegal. These fixed liver samples had been stored at ambient temperatures for 23 years. Both techniques were diagnostic on tissues collected 15-37 months before testing. Immunocytochemistry is a practical procedure for rapid specific diagnosis of liver stored for months, whereas RNA-RNA hybridization is a sensitive technique which can be applied to material stored for years.

Comparison of the enzyme-linked immunosorbent assay (ELISA) with standard tests used to detect yellow fever virus antibodies.

The enzyme-linked immunosorbent assay (ELISA) was used to detect antibodies to yellow fever virus in 110 sera from patients living in an epidemic yellow fever area. The results were then compared with those obtained with the hemagglutination-inhibition (HI), complement-fixation (CF), neutralization (NT), and indirect immunofluorescence (IFA) tests. This ELISA, which used a type-specific antigen, showed the same results as the NT test and was found to be more sensitive and more specific than the HI and CF tests.

Yellow fever in the Gambia, 1978--1979: entomological aspects and epidemiological correlations.

An entomological survey was conducted in the Gambia in January 1979, during the last phase of a yellow fever (YF) outbreak which began during the previous rainy season. In the dry conditions which prevailed in January, Aedes aegypti was the only YF vector present. Two YF virus strains were isolated from females of this mosquito species caught in a village of western Gambia, where active human cases were documented. The ae. aegypti breeding sites were exclusively of the domestic type. Larval indices varied greatly from place to place, but generally appeared to correlate with the incidence of disease. A better understanding of the conditions that prevailed at the onset and during the early phase of the epidemic will require further entomological investigations. Nevertheless, it appears probable that initial transmission as by sylvatic vectors such as the Ae. furcifer-taylori group and possibly others such as Ae. luteocephalus, Ae. metallicus, and Ae. vittatus. As the outbreak progressed, interhuman transmission by Ae. aegypti also occurred, and this mixed epidemiological pattern later gave way to transmission by Ae. aegypti only when sylvatic vector populations declined in the dry season. We speculate that a prolongation of the rainy season during 1976--1978 was important in the origin of the outbreak. The relationship of this epidemic to the established focus of sylvatic YF in southeastern Senegal is discussed. The Gambian outbreak is considered the result of a recent northwesterly extension of the YF Emergence Zone.

Yellow fever in the Gambia, 1978--1979: epidemiologic aspects with observations on the occurrence of orungo virus infections.

An epidemic of yellow fever (YF) occurred in the Gambia between May 1978 and January 1979. Retrospective case-finding methods and active surveillance led to the identification of 271 clinically suspected cases. A confirmatory or presumptive laboratory diagnosis was established in 94 cases. The earliest serologically documented case occurred in June 1978, at the extreme east of the Gambia. Small numbers of cases occurred in August and September. The epidemic peaked in October, and cases continued to occur at a diminishing rate through January, when a mass vaccination campaign was completed. The outbreak was largely confined to the eastern half of the country (MacCarthy Island and Upper River Divisions). In nine survey villages in this area (total population 1,531) the attack rate was 2.6--4.4%, with a mortality rate of 0.8%, and a case fatality rate of 19.4%. If these villages are representative of the total affected region, there may have been as many as 8,400 cases and 1,600 deaths during the outbreak. The disease incidence was highest in the 0- to 9-year age group (6.7%) and decreased with advancing age to 1.7% in persons over 40 years. Overall, 32.6% of survey village inhabitants had YF complement-fixing (CF) antibodies. The prevalence of antibody patterns indicating primary YF infection decreased with age, in concert with disease incidence. The overall inapparent:apparent infection ratio was 12:1. In persons with serological responses indicating flaviviral superinfection, the inapparent:apparent infection ratio was 10 times higher than in persons with primary YF infection. Sylvatic vectors of YF virus, principally Aedes furcifer-taylori and Ae. luteocephalus are believed to have been responsible for transmission, at least at the beginning of the outbreak. Eighty-four percent of wild monkeys shot in January 1979 had YF neutralizing antibodies, and 32% had CF antibodies. Domestic Aedes aegypti were absent or present at very low indices in many severely affected villages (see companion paper). In January, however, aegypti-borne YF 2.5 months into the dry season was documented by isolation of YF virus from a sick man and from this vector species in the absence of sylvatic vectors. Thus, in villages where the classical urban vector was abundant, interhuman transmission by Ae. aegypti occurred and continued into the dry season. A mass vaccination campaign, begun in December, was completed on 25 January, with over 95% coverage of the Gambian population. A seroconversion rate of 93% was determined in a group of vaccinees. This outbreak emphasizes the continuing public health importance of YF in West Africa and points out the need for inclusion of 17D YF vaccination in future programs of multiple immunication.

Removal of gp160 induced bio-hazards for a safe AIDS vaccine candidate.

In the first AIDS vaccine trial, immunizing preparations were based on HIV-1 Env protein (gp160). Immunogenic properties of gp160 which trigger both a humoral and cellular immune response have since justified its use in various vaccine programs, both past and present. Many reports however have underlined deleterious effects on the immune system--anti-HIV-1 enhanced antibodies, anti-CD4 autoantibodies, and inhibition of T cell activation by HIV-1--particularly associated with the Env protein. The present study shows that gp160 presented in a biologically inactivated but immunogenic form, as used in our trial, could avoid these complications. Bio-hazards associated with gp160 which indeed could be removed by appropriate treatment of the native protein, should be taken into consideration in AIDS vaccine programs.

Field trials with Vectolex (Bacillus sphaericus) and Vectobac (Bacillus thuringiensis (H-14)) against Anopheles gambiae and Culex quinquefasciatus breeding in Zaire.

Under field conditions in Kinshasa, Zaire, an aqueous suspension of Bacillus thuringiensis (H-14), Vectobac (12-AS), lost most of its larvicidal activity at all concentrations after 48 h against Culex quinquefasciatus breeding in polluted gutter water and Anopheles gambiae breeding in clear water irrigation ponds. However, good control of Cx. quinquefasciatus was obtained using a granular formulation of B. sphaericus, Vectolex-G (ABG-6185), at concentrations of 10-30 kg/ha. High concentrations of Vectolex-G gave excellent control of An. gambiae breeding in irrigation ponds. The Vectobac-G was less active against An. gambiae than Vectolex-G, in spite of good dispersion of Vectobac-G particles.

Impact of deltamethrin-impregnated bednets on biting rates of mosquitoes in Zaire.

In a rural area of Zaire, the whole population of a village was protected by deltamethrin-impregnated mosquitoes bednets. A similar village was observed as a control. Biting rates for mosquitoes were recorded in both villages. The principal man-biting species were Mansonia africana, Mansonia uniformis, and Aedes aegypti. In the village protected by the impregnated mosquito bednets, the number of Mansonia bites was reduced 96% indoors and at a lesser rate outdoors. Biting rates of Ae. aegypti dropped to 0 indoors, but the outdoor biting rate remained unchanged. It is concluded that the reduction in mosquito bites is not only caused by the repellent action of the deltamethrin but also by a reduction in mosquito numbers.

Efficacy of Bacillus sphaericus against the malaria vector Anopheles gambiae and other mosquitoes in swamps and rice fields in Zaire.

The microbial control of Anopheles gambiae and other mosquitoes with a granular formulation of Bacillus sphaericus (Vectolex) was evaluated in rice fields and swamps, located around the suburban region of Kingabwa-village in Kinshasa, Zaíre. Ten treatment cycles with 15-day intervals were carried out with the same application rate, 10 kg/ha, during the dry season (May to September 1991). The treatments reduced larval populations of An. gambiae by 98% after 48 h, but repetitive applications were required every 15 days to maintain control. The persistence of B. sphaericus spores was more apparent in rice fields than in swamps. A significant reduction in nuisance biting by Culex quinquefasciatus and Mansonia uniformis was observed. For An. gambiae, a decrease of 13.6% in human biting was noted during the post-treatment period. The entomological inoculation rate was reduced from 0.238 to 0.143. The efficacy of B. sphaericus does not appear to offer outstanding potential for control of An. gambiae in rice fields and swamps and seems to be limited due to different factors tied to ecology and natural conditions in the fields.

[Rabies virus study by the plaque forming system. A simplified technique]. / Etude du virus rabique par la méthode des plages. Technique simplifiée.

The author describes here a simplified and economical plaque forming system for rabies virus study. It is achieved in three steps (cells preparation and infection, apply of overlay, staining). It presents more advantages than previous techniques: time as well as culture media and handling spearing, easy and durable reading. Cells prepared with di-ethyl-amino-ethyl-dextran are added to equal quantities of diluted virus (for titration) or to virus-serum mixing (for seroneutralization) and overlaid after 4 hours with a carboxymethylcellulose medium. Reading is carried out after 6 days incubation and staining with amido black. Plaques obtained are to 2-3 mm in diameter, regularly reproductible, clear and easy to read. This technique enables titration and seroneutralization and even a rabies virus cloning.

[The culicidian fauna and its nuisance in Kinshasha (Zaire)]. / La faune culicidienne et sa nuisance à Kinshasa (Zaïre).

A public health study to inventory bloodsucking mosquitoes was conducted in Kinshasa and its regions. 20 culicidian species were represented by 7 Anopheles, 6 Culex, 5 Aedes and 2 Mansonia. In general, the number of bites/man/night (b/m/n) was in average 141.5 of Cx. quinquefasciatus, 60.1 of Cx. antennatus, 21.1 of M. africana, 16.3 of An. gambiae and 7.7 of M. uniformis. The nuisance is different from region to other. Culex quinquefasciatus is the most abundant and aggressive species in the urban area with an average of 400 b/m/n. In certain suburban zones, Culex antennatus is predominant, with aggression on the order of 176.5 b/m/n. The Anopheles gambiae complex is predominant in the semi-rural zone at the periphery of the city with 26.05 b/m/n. Other mosquitoes are equally implicated in their aggression and nuisance to man, in particular, Mansonia africana, Mansonia uniformis and Aedes aegypti. A geographical distribution map has been established for the preponderant species in Kinshasa.

[The Anopheles fauna and the transmission of human malaria in Kinshasa (Zaire)]. / La faune anophélienne et la transmission du paludisme humain a Kinshasa (Zaïre).

A longitudinal epidemiological study of malaria and its vectors was conducted in Kinshasa. 264 night-bite collections on human bait (1,056 man nights) and 384 collections of the house-resting fauna were carried out from April 1989 to October 1990. The anophelian fauna was identified and inventoried, 7 Anopheles species were found: Anopheles gambiae, An. funestus, An. paludis, An. hancocki, An. counstani, An. brunnipes, and An. nili. A single species, An. gambiae s. l. is responsible for the transmission of malaria, it represents 93.27% of the anopheline fauna. The average number of anophele bites man day was 16.28 bites/man/night, it varied between 1 b/m/n in urban area to 26.05 b/m/n in semi-rural area. The average of the sporozoite index for An. gambiae was 3.3%, but it varied from 0% in the urban area to 6.52% in the semi-rural area. The entomological inoculation rate (h) was 197 infective bites per year. This rate fluctuated from 1 infective bite each 128 nights in urban area to 1.7 infective night-bite in semi-rural area. Other epidemiological index were also determined: the level of daily survival rate (p = 8.75 days), the vectorial capacity of 17.97 and the Macdonald's stability 3.5 bites on man taken by a vector during its entire lifetime.

[Seroprevalence of rubella in Kinshasa (Zaire)]. / Séroprévalence de la rubéole a Kinshasa (Zaïre).

A serosurvey of rubella was carried out by haemagglutination inhibition and IgM assay among 106 newborn infants (91% positive); 101 suckling infants aged 9-18 months (32.7% positive); 100 children aged 2-4 (58% positive); and 100 young girls 9-11 (68% positive), while 93% of mothers showed the presence of protective antibodies. These figures indicated that large numbers of women old enough to bear children are susceptible to infection with rubella, at least early in life. A vaccination programme is therefore recommended for one year-old children of both sexes and again for young girls prior to puberty.

[Laryngeal histoplasmosis due to "Histoplasma duboisii" (author's transl)]. / Histoplasmose laryngée à "Histoplasma duboisii".

Primitive detection of the Histoplasma duboisii accompanied with proteiform manifestations, is rare in the superior air ways. Its localisation at the larynx level is an exception and makes the very interest of our following reported observation. In front of this hard diagnostic, only the pathogenic agent visualisation gives evidence. Among serological techniques, immuno-fluorescence and E.L.I.S.A. are very helpful. Therapeutic possibilities have been improved by Miconazole which both associates efficiency and inocuity.

[Randomization of a population for a clinical trial of immunization against the human immunodeficiency virus (HIV)]. / Randomisation d'une population pour un essai clinique d'immunisation contre le virus de l'immunodeficience humaine (VIH).

Serum HIV antibodies has been investigated in different subpopulation from four different regions in Zäire by Elisa (Elavia Pasteur) and Western Blot. Seropositive prevalence differed from 2.4% (rural population) to 12.5% (urban population) according to the regions. When the group with 2.4% migrated to the area with 12.5% positives, after 8-12 months the number of seropositives in this group raised to 8%, showing an increase of 5.6% within one year. Such population is suitable for a large scale clinical trial (with anti-AIDS vaccine) to be performed on individuals with high risk of natural infection.

PIP: Major obstacles to development of a vaccine against the HIV infection have apparently been resolved by utilizing viral signals not directly from the virus or its products, but from membranes of infected cells. Evaluation of the level of protection provided by the vaccine requires a large scale clinical trial in a population with a high rate of infection. Screening studies have been conducted in different areas and in different subgroups in Zaire in order to identify such a group. A population living or working at a site some 30 km east of Kinshasa was studied between December 1986-March 1988, and a parallel study was conducted of persons residing in Kinshasa. Groups from 2 rural regions were also studied. The 1554 persons screened were divided into 8 groups based on their residence histories. Serum HIV antibodies were assessed by ELISA and Western Blot. The 4 groups composed of men living and working in Kinshasa or within a suburban radius of 30 km had a seroprevalence rate of 12.67%, with no significant difference by residence. A population of 136 pregnant women in the same locations had a seroprevalence rate of 12.5%, for a sex ratio of 1.1. A group of 71 persons studied in a provincial city of southern Zaire who had spent time in Kinshasa several years previously had a seroprevalence of 4.23%. A group of new arrivals to the Kinshasa vicinity from the provinces who had never previously resided in Kinshasa had a seroprevalence of 2.45%. The final group was also composed of new arrivals from the same provinces with no previous urban experience. Screening after 8-12 months in the city showed that their seropositive rate had increased from 2.45% to 8.00%. This increase of 5.6% within 1 year indicates that this group would be appropriate for a large scale clinical trial on individuals with high risk of natural infection.

[Immunization against the human immunodeficiency virus in Zaire]. / Immunisation contre le virus de l'immunodéficience humaine au Zaïre.

The first experimental immunization of human against the AIDS retrovirus HIV-1 was started in a series of HIV seronegative healthy volunteers in november 1986. Priming used a vaccinia virus recombinant (V25) expressing Gp 160 env determinants of HTLV III B at the surface of infected cells. This priming which induced a weak immune reaction was performed on HIV seronegative French and Zaïrian individuals living in Zaïre (Kinshasa). These results prompted to boost the primary immune response. Four different protocols were used: slow drip intravenous infusion with paraformaldehyde fixed autologous cells infected with V25 (first protocol), repeated scarification with V25 for the second protocol. The third protocol used scarifications with fragment of Gp 120 env protein, and the fourth protocol used intramuscular injections of purified autologous cell membrane infected with V25. Results of the immune reaction obtained after these boosts: The three last protocols showed a cell mediated immunity (CMI) that not significantly enhanced in comparison with CMI obtained after V25 priming alone. Moreover, the sera showed low and variable neutralizing antibodies titers one to four months after boosting. By contrast boosting with V25 infected fixed cells (D.Z. individual) provide strong humoral and cellular group specific anamnestic immune response. Indeed, high levels of antibodies to viral envelope and neutralizing antibodies against divergent HIV-1 strains were observed. Group specific CMI and cell mediated cytotoxicity were enhanced by boosts. Skin-tests showed high mediated and delayed hypersensitivity to GP 160 in vivo. For the first time, these results show that an immune stage against HIV can be obtained in a man.

PIP: The 1st experimental immunization of humans against the AIDS retrovirus HIV-1 was begun in November 1986 among a group of HIV-seronegative healthy volunteers. A priming, involving a vaccine virus recombinant (V25) expressing Gp 160 env determinants of HTLV-III B at the surface of the infected cells was utilized. This priming, which induced a weak immune reaction, was performed on HIV-seronegative French and Zairian individuals living in Kinshasa, Zaire. These results prompted a boost to the primary immune response. 4 different protocols were used: the slow drip intravenous infusion with paraformaldehyde-fixed autologous cells infected with V25; repeated scarification with V25 for the 2nd protocol; scarifications with fragments of Gp 120 env protein; and intramuscular injections of purified autologous cell membrane infected with V25. The results of the immune reaction obtained after these boosts indicated the following: The last 3 protocols showed a cell- mediated immunity (CMI) that did not significantly enhance in comparison with CMI obtained after V25 priming alone. Moreover, the sera showed low and variable neutralizing antibody titers 1-4 months after boosting. By contrast, boosting with V25 infected fixed cells (D.Z.) provided strong humoral and cellular group specific anamnestic immune responses. Indeed, high levels of antibodies to viral envelope and neutralizing antibodies against divergent HIV-1 strains were observed. Group- specific CMI and cell mediated cytotoxicity were enhanced by boosts. Skin tests showed high mediated and delayed hypersensitivity to Gp 160 in vivo. For the 1st time, these results show that an immune state against HIV can be obtained in a man. (author's modified)

[Minor arboviral diseases in Central and West Africa (author's transl)]. / Les arboviroses mineures en Afrique centrale et occidentale.

Many arboviruses can cause febrile illness in man, with or without rash, quite apart from yellow fever and the aetiological agents of haemorrhagic fevers. Exanthema are one of the commonest signs. Neuro-vascular attacks frequently occur and in some cases meningitic complications can happen. Diagnosis is often difficult: viraemia is of short duration and complement fixing antibodies do not appear consistently in convalescent serum. In Central and West Africa, 19 different arboviruses have been implicated either by isolation or by serological conversion. Some were isolated from man for the first time.

[Yellow fever in Senegal from 1976 to 1980 (author's transl)]. / La fièvre jaune au Sénégal de 1976 à 1980.

The isolation of a YF strain in Kedougou (eastern region) in December 1976 from Aedes gr. furcifer taylori lead to a mass vaccination campaign in Senegal. In 1977, 67 strains were isolated from different mosquitoes (essentially A. luteocephalus and A. gr. furcifer taylori). Among these strains, 3 were isolated from males and this very important phenomenon might explain the virus conservation during the dry season. In 1978, 55 strains were isolated from mosquitoes of the same species, but not from males, and five from monkeys (E. patas and C. aethiops). But no increase of morbidity or mortality was observed in humans. In December 1978, a YF outbreak occurred in the Gambia with 271 suspect cases and 63 deaths; 2 strains were isolated from A. aegypti. In 1979, the epidemic seemed to have disappeared. Up to now, one single strain has been isolated from mosquitoes. But danger remains, and 3 YF human cases with 2 deaths were observed in Sine-Saloum, among French unvaccinated tourists.

[Sylvatic yellow fever in Africa recent advances and present approach (author's transl)]. / La fièvre jaune selvatique en Afrique: donnees récentes et conceptions actuelles.

Recent works carried out in west and central Africa resulted in numerous yellow fever virus isolations from sylvatic mosquitoes in the forest-savanna transitional zone. This virus was also obtained from monkeys, whereas studies on sequence and duration of the observed epizootics permitted a previsional approach of the yellow fever mechanisms in the same belt, the epidemiological importance of which was stressed ("emergence zone"). In the same course of research and publications, the prevalent part of the involved mosquitoes as virus-reservoirs was constantly emphasized ("reservoir-vector"). Recent investigations on transovarial transmission and yellow fever isolations from male mosquitoes caught in the field, provide decisive support to such a conception. It can explain that epizootics may be locally observed several years in succession, despite the fact that yellow fever virus circulation seems to be fundamentally of a dynamic character. Yellow fever virus was recently obtained from ticks and tick-eggs.