Nine more cases of human infections by Dirofilaria repens diagnosed in Italy by morphology and recombinant DNA technology.

Dirofilariosis is a mosquito-borne disease of carnivors due, in Italy, to Dirofilaria repens and D. immitis species which are sympatric in many areas. Nine more cases of human infection are here reported, detected in Italian patients living in northern regions (where D. immitis and D. repens are largely present), in Tuscany (where D. repens is more frequent than D. immitis), and in Sicily (where D. immitis is present only sporadically). The nematodes surgically removed (7 female and 2 male specimens) have been identified by a PCR-based method and by morphology as D. repens. Morphological, clinical, and epidemiological data of these human infections are discussed.

Localization of HLA-A2.1-restricted T cell epitopes in the circumsporozoite protein of Plasmodium falciparum.

Localization of human MHC class I-restricted T cell epitopes in the circumsporozoite (CS) protein of the human parasite Plasmodium falciparum is an important objective in the development of antimalarial vaccines. To this purpose, we synthesized a series of overlapping synthetic 20-mer peptides, spanning the entire sequence of the 7G8 CS molecule except for the central repeat B cell domain. The P.f.CS peptides were first tested for their ability to bind to the human MHC class I HLA-A2.1 molecule on T2, a human cell line. Subsequently, the use of a series of shorter peptide analogues allowed us to determine the optimal A2.1 binding sequence present in several of the 20-mers. Binding P.f.CS peptides were further tested for their capacity to activate PBL from HLA-A2.1+ immune donors living in a malaria-endemic area. Specific IFN-gamma production was detected in the supernatant of cultures of PBL from exposed individuals. Cytotoxic T cell lines and clones were derived from the PBL of one responder, and their activity was shown to be HLA-A2.1-restricted and specific for the peptide 334-342 of the CS protein. In addition, double transgenic HLA-A2.1 x human beta 2-microglobulin mice were immunized with peptide 1-10 of the CS protein. T cells derived from immune lymph nodes displayed a peptide-specific HLA-A2.1-restricted cytolytic activity after one in vitro stimulation.

Sensitivity to antimalarial drugs by Plasmodium falciparum in Goundry, Oubritenga province, Burkina Faso.

Plasmodium falciparum susceptibility to chloroquine in vivo and to chloroquine, mefloquine, quinine, amodiaquine and sulfadoxine/pyrimethamine in vitro was investigated in children living in Goundry village, Oubritenga Province (Burkina Faso) in November 1992. An extended WHO in vivo field test was used, with follow-up on days 2, 4, 7, 14, 21 and 28 after treatment with 25 mg chloroquine per kg body weight given over 3 days, in children from 2 to 8 years old with P. falciparum monospecific infection, asexual parasitaemia > 800 parasites/microliter of blood and negative Bergqvist urine tests. At the same time, the in vitro response was assessed using WHO standard test kits. Out of the 71 in vivo responses examined, 50 (70.4%) were classified as resistant to chloroquine at RI (43.6%) or RII (26.8%) levels. There were no RIII responses. Out of the 43 isolates tested for chloroquine susceptibility in vitro, 32 (74.4%) were resistant to the drug with mean EC50 and EC99 values of 1.41 mumol and 10.96 mumol/l of blood, respectively. Resistance to sulfadoxine/pyrimethamine in vitro was observed in one out of 19 tested cases, with mean EC50 and EC99 values of 0.00002 mumol and 35.05 mumol/l of blood, respectively. All isolates were inhibited by mefloquine at 12.8 mumol/l of blood, quinine at 51.2 mumol/l of blood and amodiaquine at 0.4 mumol/l of blood, indicating full sensitivity to these 3 drugs. The present study demonstrates the high prevalence of chloroquine-resistant strains of P. falciparum in the study area of Burkina Faso and indicates that isolates resistant to sulfadoxine/pyrimethamine may also be present.

Chloroquine and sulfadoxine/pyrimethamine sensitivity in Burkina Faso. In vivo sensitivity of Plasmodium falciparum to chloroquine and sulfadoxine/pyrimethamine in Burkina Faso.

Plasmodium falciparum susceptibility to chloroquine and sulfadoxine/pyrimethamine was investigated in children living in an urban district of Ouagadougou, Burkina Faso, in August 1992. A modified WHO Standard Field in vivo test was used, with follow-up on days 2, 4, 7, and 14 after treatment with 25 mg chloroquine per kg body weight given over 3 days, or with standard doses of sulfadoxine/pyrimethamine in children from 2 to 8 years old with P. falciparum monospecific infection, asexual parasitaemia > 800/microliters of blood and negative Bergqvist and Lignin urine tests. Out of a total of 515 children screened, 152 were selected and randomized into treatment with chloroquine (86 children) or sulfadoxine/pyrimethamine (66 children). With the chloroquine regimen, parasitaemia did not clear by day 7 in 7 cases (8.1%), and there was recurrence of parasitaemia in 21 subjects (24.4%) on day 14, while all the children who had received sulfadoxine/pyrimethamine were fully sensitive on day 7 and day 14. It can be concluded that sulfadoxine/pyrimethamine is a good alternative for the treatment of falciparum malaria in areas of Burkina Faso with established chloroquine resistance.

In vivo sensitivity of Plasmodium falciparum to halofantrine hydrochloride in Burkina Faso.

Plasmodium falciparum susceptibility to halofantrine hydrochloride was investigated in a small village near Ouagadougou, Burkina Faso, where the parasite was known to be chloroquine resistant. An in vivo test was carried out in July 1992 at the beginning of the rainy season in children ranging in age from two to eight years with P. falciparum monospecific infections, asexual parasitemia greater than 800/microliters of blood, and a negative result on a Bergqvist urine test for 4-aminoquinolines. Among 206 children screened, 74 were selected for study. Blood samples were collected on days 0, 2, 4, 7 and 14, and 100 microscopic fields of thick and thin blood smears were examined for parasite density and species identification. Halofantrine hydrochloride was administered under supervision at the standard dose of 24 mg/kg as 8 mg/kg given three times at 6-hr intervals with an observation period of 1 hr after each 8-mg/kg dose. Parasitemias cleared in all 74 cases by day 7, but there was a recurrence of parasitemia in six subjects (8.1%) on day 14. A second course of therapy with halofantrine resulted in prompt clearance of parasitemias in all of these children. The drug was well-tolerated and the hematologic and biochemical indices were not adversely affected by treatment.

Susceptibility of Plasmodium falciparum to chloroquine and mefloquine in Ouagadougou area (Burkina Faso).

Plasmodium falciparum susceptibility to chloroquine in vivo and to chloroquine and mefloquine in vitro was investigated in children living in Ouagadougou area (Burkina Faso) in October 1991. The 7-day WHO in vivo field test was used, with follow-up on days 2, 4, 7 after treatment with 25 mg base of chloroquine per kg body weight given over 3 days, on children aged 2-8 years with monospecific P. falciparum infection (parasite density higher than 800 asexual parasites/microliters of blood), and negative Bergqvist urine tests. At the same time, the in vitro response was assessed using WHO standard test kits. Chloroquine treatment in vivo resulted in parasite clearance in 47 subjects (92.2%) within 7 days (S/RI responses). Parasitaemia did not clear in 4 cases (7.8% of RII responses). There were no RIII responses. The sensitivity study in vitro showed a low degree of chloroquine resistance in 2 out of 12 isolates tested and a mean 50% effective dose (EC50) and EC99 of 0.12 mumol and 1.47 mumol/litre of blood, respectively. All isolates tested were inhibited by mefloquine at 1.6 mumol/litre of blood, indicating full sensitivity. The present study demonstrates that first-line treatment with chloroquine is still satisfactorily effective in the study area of Burkina Faso.

A national survey of the prevalence of chloroquine resistant Plasmodium falciparum in Burkina Faso.

Plasmodium falciparum susceptibility to chloroquine was investigated in 10 areas of Burkina Faso in the rainy seasons in 1990-1991. The 7-days in-vivo test was carried out from August to November on children aged 2-8 years with monospecific P. falciparum infection (asexual parasitaemia > 800 microliters-1 of blood), axillary temperature < 37.5 degrees C, and a negative Bergqvist urine test for 4-aminoquinolines. Among 2190 children screened, 366 were selected. Blood samples were collected on days 0, 2, 4 and 7 by finger-prick, and 100 microscopic fields of thick and thin smears were examined for parasite density and species identification. Chloroquine was given under supervision at the standard dose of 25 mg kg-1 over three days (days 0, 1 and 2) with an observation period of one hour after treatment. Parasitaemia did not clear in 63 cases (17.2%) with a 13.4% RII response and 3.8% RIII response. The results do not seem to indicate a decline in the sensitivity of P. falciparum to chloroquine in Burkina Faso during the past two years.

[In vivo chemosensitivity of Plasmodium falciparum to chloroquine in Burkina Faso: development of resistance 1988-1990]. / Surveillance de la chimiosensibilite in vivo de Plasmodium falciparum a la chloroquine au Burkina Faso: evolution de la resistance 1988-1990.

A study of Plasmodium falciparum sensitivity to chloroquine was carried out in 1988 and 1990 in 5 localities, representatives of different climatic areas of Burkina Faso. The 7-day in vivo standard test performed in 1988 showed a total clearance failure of 25%. No significant difference with 1990 data was found, except for an increase of the resistance in the area of Fada N'Gourma, close to the border with Benin, Niger and Togo.

In vitro immune recognition of synthetic peptides from the Plasmodium falciparum CS protein by individuals naturally exposed to different sporozoite challenge.

The impact of duration and intensity of sporozoite challenge on the in vitro cell immune response to synthetic peptides of the circumsporozoite (CS) protein of Plasmodium falciparum was investigated in residents of a malaria endemic area in Burkina Faso (West Africa). Lymphocyte proliferation and interferon-gamma (IFN-gamma) production were used to assess immune recognition of synthetic peptides corresponding to the polymorphic Th2R and Th3R regions, to the conserved CS.T3 sequence and to NANP and degenerate NVDP repeats. Immune responses were measured in adults and children from a village where they received more than 100 sporozoite inoculations per year and in adults living in a town, exposed to a 10-100 times lower challenge. A lifetime intense exposure apparently increased the ability to proliferate in response to most peptides in the rural adults, who all produced antibodies to NANP repeats. Surprisingly, cell cultures from these subjects seldom contained appreciable levels of IFN-gamma. In the urban adults, possibly due to the moderate challenge they are exposed to, significant differences in the proliferative potentials of the peptides could be detected. The highest stimulation indices were obtained with the genetically unrestricted CS.T3 peptide. Remarkably, proliferative responses to Th2R and Th3R appeared to be correlated with the humoral response to the CS protein, indicating a T helper significance of the epitopes. The differing proliferative potential of the polymorphic epitopes in the urban adults suggests that polymorphism might delay the development of immune responsiveness under conditions of sporadic transmission. The children from the highly malarious village displayed the lowest proliferative scores, accompanied by a high prevalence of antibodies to NANP repeats. On the basis of these findings, the hypothesis is proposed that a pure B cell reactivity to NANP repeats could ontogenetically precede the mounting of a conventional T-B cooperative immune response.