Genetic diversity of a population of Schistosoma haematobium derived from schoolchildren in east central Zimbabwe.

To characterize the extent of genetic diversity of Schistosoma haematobium within and among its definitive host (intra- and interhost parasite diversity), 133 individual isolates from 25 infected schoolchildren were compared using randomly amplified polymorphic DNA markers. With 4 primers, 53 unambiguous loci were identified, and of these, 22 were polymorphic. Mean heterozygosity in the population was 0.116 +/- 0.043. Analysis of molecular variance showed the majority of variance occurred within, rather than between, hosts. Frequencies of certain alleles segregated the parasite population into 13 distinct clusters of associated genotypes, with 4 of these first appearing 10 mo after the initial survey. Considering the level of diversity within this limited geographical area and the possibility of rapid turnover of genotypes, parasite variance may impact acquired immunity and clinical outcome of the infection.

Environmental and geographical factors contributing to watershed contamination with Cryptosporidium parvum oocysts.

Cryptosporidium parvum is a waterborne parasite which infects cattle and produces life-threatening zoonosis in people with impaired immune systems. Digital maps of 100-year floodplain boundaries, land use/cover, and livestock operations were used to select and characterize cattle farms in the floodplain area in Lancaster County, Pennsylvania, U.S.A. Over 21% of the cattle farms were located within 100-year floodplain boundaries. On average, a single farm comprised 12.8 ha of pasture (including buildings and farmyard) at risk of inundation. In all farms cattle had unlimited access to the creek. Manure samples collected from closed-in calf pens, cow/heifer yard runoff, and cattle paths through the creek were tested for C. parvum. On 64% of the farms (n=50) at least one sample was positive for C. parvum, and 44% of the farms had oocysts in all manure samples. Concentration varied from 90 to 371 oocysts/g and was significantly higher (P<0.02) in calf samples than in manure from cow and cow/heifer.

Recovery of avian schistosome cercariae from water using penetration stimulant matrix with an unsaturated fatty acid.

Avian schistosome cercariae that emerge from aquatic snails can penetrate human skin causing cercarial dermatitis resulting in serious skin disease in sensitized and immunocompromised people. A trap developed for Schistosoma mansoni cercariae was tested for recovery of avian schistosome cercariae. A matrix with an unsaturated fatty acid, linoleic acid stimulates attachment and penetration of Trichobilharzia spp. cercariae, and the immobilized larvae can be subsequently visualized. The number of trapped cercariae exceeded by 3 to 7 times the number of larvae expected on the surface of the trap, based on their random distribution in the water. Recognition, attachment, and penetration of Trichobilharzia spp. cercariae led to injection of more secretory products into the stimulant matrix than by Schistosoma mansoni cercariae. This method can assist in the identification of waters infected with avian schistosome cercariae so that human exposure to these parasitic larvae can be minimized.

Post treatment sensitivity studies with the ParaSight-F test for malaria diagnosis in Zimbabwe.

The post-treatment diagnostic performance of the Plasmodium falciparum histidine-rich protein (HRP-II) antigen detection test, ParaSight-F test, was assessed on 55 falciparum malaria cases treated with chloroquine during in vivo drug sensitivity studies. The post-treatment sensitivity of the test remained high, except for an insignificant decline on day 1. However, specificity dropped sharply by day 1, subsequently increasing linearly with time to satisfactory values by day 10. As expected, from its inverse relationship to specificity, the false positive rate was high on day 1 and decreased linearly to low level by day 10. The temporary increase in false positive rate-following treatment was due to persistent parasite antigen, rather than subpatent parasitaemia. Thus findings showed that positive readings by the test within 10 days post-treatment may occur in cured cases and will not necessarily imply treatment failure. Furthermore it will be important to take patient antimalarial history into consideration during routine usage of the test for malaria diagnosis. The trend of Youden's J-index for the ParaSight-F test showed that from 10 days post-treatment, the test was generally reliable, with positive readings indicating active infection. It was concluded that the ParaSight-F test was not only valuable at confirming malaria diagnosis on clinical cases in seasonal transmission areas, but had potential for application to detect recrudescent infections within 2 weeks of chloroquine treatment.

Social and cultural factors affecting rates of regular retreatment of mosquito nets with insecticide in Bagamoyo District, Tanzania.

Insecticide-treated mosquito nets have an impact on mortality and morbidity in young children under controlled conditions. When integrated into larger control programs, there is the danger that rates of regular retreatment of the nets with insecticide will drop, greatly limiting their effectiveness as a public health intervention. In Bagamoyo District, Tanzania, rates of retreatment dropped significantly when payment for the insecticide was introduced. A series of neighbourhood (hamlet) meetings were held in all study villages to discuss people's concerns about the insecticide and ways to increase rates of retreatment. Although changes were made in the procedure for retreatment, rates of retreatment remained lower than expected and showed marked variation within as well as between villages. We then conducted unstructured key informant interviews as well as informal discussions in a village with strong variation between different sectors of the villages in rates of retreatment. While logistical problems were most frequently cited as reasons not to bring nets for retreatment, political and social divisions within the community provided a better explanation. This is borne out by the low response to rearrangements in logistics which made retreating the nets significantly easier for households, and the higher response when changes were made in the channels of communication as well as the logistic features. It is clearly more difficult for villagers to appreciate the benefits of the insecticide than those of the nets. Great emphasis needs to be placed on the insecticide and its beneficial effects from the outset for any large-scale programme to be sustainable.

Trial of the ParaSight-F test for malaria diagnosis in the primary health care system, Zimbabwe.

Rapid diagnosis of Plasmodium falciparum malaria remains one of the main limitations to prompt treatment. Diagnosis based on clinical symptoms is decidedly unreliable, especially in areas of seasonal transmission like Zimbabwe. In view of this, the Plasmodium falciparum histidine rich protein (HRP-II) antigen detection assay (ParaSight-F test) was tried at 10 health centres in 3 malaria endemicity zones of Zimbabwe, as a malaria diagnostic tool for primary health care. Parasitological evaluations were conducted using thick and thin film microscopy as gold standard, and ease of test operation and practicability to nurses were ascertained by questionnaire. The sensitivity of the test did not vary substantially by endemicity zone and was approximately 93%. Specificities were 85, 72 and 92% in the hyperendemic, mesoendemic and hypoendemic zones, respectively. Positive predictive values varied considerably with endemicity, the lowest being in the hypoendemic zone (56%). However, negative predictive values did not change significantly, with a mean of 94%. It was found that the ParaSight-F test reduced mistreatment for malaria, relative to clinical diagnosis, by up to 81%, especially in the hypoendemic region. Test acceptability evaluations were good.

Schistosoma mansoni: the immune response against cercarial glycocalyx.

Schistosoma mansoni cercarial glycocalyx was separated and purified by Sephacryl-300 SR. It was found to stimulate the humoral immune response in mice injected with it. Antiglycoalyx antibodies raised in CD/1 mice were found to be cytotoxic to schistosomula in vitro. But conversely, no protective effect was demonstrated in vivo. Eosinophil-mediated cytotoxicity was found to have no effector function in the murine immune response against schistosomes. A monoclonal antiglycocalyx IgM was prepared during our study. It was found to have no cytotoxic effect on schistosomules in vitro. However, it was found to have an inhibitory activity blocking the cytotoxic effect of other antiglycocalyx isotypes in the immune mouse. The contradiction between the result of antiglycocalyx antibody-mediated cytotoxicity obtained in vivo and that obtained in vitro is in itself revealing and suggests that the effect is crucially dependent upon factors as yet poorly understood.

Treatment practices for degedege, a locally recognized febrile illness, and implications for strategies to decrease mortality from severe malaria in Bagamoyo District, Tanzania.

Malaria remains one of the chief causes of mortality among young children in sub-Saharan Africa. Verbal autopsies for cases of childhood mortality in Bagamoyo District, Tanzania demonstrated that degedege, a locally defined illness of children characterized by fever and convulsions, is frequently treated by traditional healers. To investigate this further, an ethnographic study was carried out in one village that included in-depth interviews with 14 traditional healers and 3 focus groups with parents. Parents and traditional healers were unanimous in their conviction that degedege requires traditional treatments, at least initially, and that these treatments are effective. While traditional healers do refer cases that are not improving to the District Hospital, this frequently occurs late in the course of the illness, after one or more stages of traditional treatments. The prognosis will thus be poor for those children who are suffering from severe malaria. Consideration should be given to enlisting the support of traditional healers in efforts to improve treatment for severe malaria, including teaching them how to distinguish febrile convulsions from cases of severe malaria.

The polymerase chain reaction method as a tool for identifying members of the Anopheles gambiae complex (Diptera:Culicidae) in northeastern Tanzania.

Polymerase chain reaction (PCR) primers developed at the Centers for Disease Control in Atlanta for the identification of members of the Anopheles (Cellia) gambiae Giles complex were tested on material collected in the Bagamoyo and Muheza districts of northeastern Tanzania. Part of the sample from Bagamoyo was chromosomally identified and correlated with the PCR identifications. This sample contained 170 Anopheles arabiensis, 328 An. gambiae, and 58 Anopheles merus, of which 121, 237, and 54 specimens, respectively, were identified with both PCR and chromosomes. Three specimens identified chromosomally as An. merus gave only the PCR fragment characteristic for Anopheles quadriannulatus, but on retesting gave the correct result. The Muheza sample consisted of 771 An. arabiensis, 852 An. gambiae, 43 An. merus, and 4 specimens producing the fragment characteristic for An. quadriannulatus. Because An. quadriannulatus has never been recorded from mainland Tanzania and due to the high number of specimens that produced no result (193), it is probable that DNA degradation led to misidentification of An. merus specimens as An. quadriannulatus. The overall probability of correct identification by PCR was 99.685% at first testing, which compares favorably with other genetic methods currently in use.

Local terminology for febrile illnesses in Bagamoyo District, Tanzania and its impact on the design of a community-based malaria control programme.

This paper reviews results of several ethnographic studies that have examined the issue of local terminology for malaria in Africa, then presents findings from an on-going study in Bagamoyo District, Tanzania. The study used a mixture of qualitative and quantitative interview methods to examine local perceptions of malaria and malaria treatment practices. Although the local term homa ya malaria or malaria fever appeared on the surface to correspond closely with the biomedical term malaria, significant and often subtle differences were found between the two terms. Of perhaps greatest importance, common consequences of malaria in endemic areas such as cerebral malaria in young children, severe anaemia and malaria in pregnancy were not connected with homa ya malaria by many people. A set of guidelines are described that were used to determine how best to promote acceptance and use of insecticide-impregnated mosquito nets, given these results. It is demonstrated that the position of the term used to denote malaria in the local taxonomy of febrile illnesses has important implications for the design of health education interventions.

Malaria infection potential of anopheline mosquitoes sampled by light trapping indoors in coastal Tanzanian villages.

Anopheline mosquito populations were studied during 1992 in seven villages south of Bagamoyo, coastal Tanzania, prior to malaria control intervention using insecticide treated bednets. To collect mosquitoes, CDC light traps were used in ten houses per village fortnightly for 12 months. Anopheles females were identified and checked by ELISA for the presence of malaria sporozoite antigen and source of bloodmeal. An.funestus peaked in June-July after the long rains. Three members of the An.gambiae complex had different seasonality: An.arabiensis, An.gambiae and small numbers of An.merus were collected. In most villages transmission was extremely high and perennial with the entomological inoculation rate reaching three to eleven infective bites per person per night in July and persisting at around 0.1 and 1 for most of the remainder of the year. Sporozoite infection rates within the An.gambiae complex ranged from 2% to 25%, with the peaks in January and July following the two rainy periods. An.funestus showed a similar pattern. The light traps were reliable, simple to operate, and proved to be satisfactory to study the mosquito vector population.

Comparison of sampling anopheline mosquitoes by light-trap and human-bait collections indoors at Bagamoyo, Tanzania.

The mosquito sampling efficiency of CDC miniature light-traps, relative to night-biting collections, was evaluated indoors at two sites in coastal Tanzania. We found that the total number of anophelines captured overnight by light-traps (hung beside a bednet in use) was 1.23 times the number of anophelines captured by human-bait collections. This relationship was not affected significantly by changes in the mosquito density, order of trapping method, date of sampling, or number of household occupants. Malaria sporozoite rates were twice as high among mosquitoes captured by light-trap as compared to those captured by night-biting collection. This was attributed to the tendency of light-traps to capture a larger proportion of gravid mosquitoes, which also had high sporozoite rates. The differences in sporozoites rates according to abdominal stage indicates that unfed mosquitoes captured by light-traps may define more precisely the human-biting activity and sporozoite rates as seen by night-biting collections. Our study shows that light-traps, when used in combination with night-biting collections, can be an effective and sensitive means for measuring human-biting activity and the sporozoite rate.

A chemokinetic response in Schistosoma mansoni cercariae.

Schistosoma mansoni cercariae have been shown to aggregate in the presence of glass slides treated with clear nail varnish and linoleic acid. In choice chambers cercariae move toward the stimulant, but this behavior is not seen when linoleic acid is omitted. After 30-45 min, the cercariae were concentrated near the end of the choice-chamber containing the linoleic acid slide. When the cercariae were added in the center of the choice chamber, they formed a diffuse cloud that dispersed slowly in both directions in the absence of linoleic acid. Cercariae aggregating in the vicinity of a stimulant surface are not immediately stimulated to commence penetration; this appears to be time and dose related.

Extraction of Haemoproteus columbae (Haemosporina: Haemoproteidae) antigen from rock dove pigeons (Columba livia) and its use in an antibody ELISA.

Erythrocytic stages of Haemoproteus columbae were extracted from the cytoplasm of nucleated red blood cells (RBC) of Rock dove pigeons (Columba livia) using cationic detergent (N,N',N'-polyoxyethylene(10)-N-tallow-1,3-diaminopropane [EDTA-20]) and discontinuous Percoll gradient density. Crude RBC extract (CRBCE) antigen was prepared. Parasitized RBCs were more resistant to EDTA-20 action than unparasitized cells. An enzyme-linked immunosorbent assay (ELISA) was developed for detection of anti-H. columbae immunoglobulins in 30 wild-captured C. livia. Whole blood, serum, and dried blood on filter paper gave similar results; the latter was selected for sampling convenience. Optimal antigen concentration was 5 micrograms/ml, and anti-H. columbae immunoglobulins were detectable at a 10(-4.11) dilutions. The binding efficacy of anti-chicken IgG to the pigeon immunoglobulins was significantly higher than anti-duck IgG or anti-turkey IgG. Parasitemia by Giemsa-stained thin blood smears ranged from 20.0 to 47.5%, mean = 32.4 +/- 8.3%; 17 of 30 birds had multiply infected RBCs with a mean parasitemia of 2.4 +/- 1.1%, range 0.7-4.9%; 27 birds were positive by the ELISA. No clinical signs of infection were observed. ELISA absorbance values were not correlated with the level of parasitemia in individual birds. All pigeons were negative for anti-Plasmodium relictum and anti-P. elongatum immunoglobulins as determined by ELISA. The pigeons were not subclinically infected with Plasmodium spp. as determined by inoculation of domestic ducklings with blood from dexamethasone-immunosuppressed pigeons.

Seasonal variation in the perceived risk of malaria: implications for the promotion of insecticide-impregnated bed nets.

Bed nets (mosquito nets), impregnated every 6 months with pyrethroid insecticides, are a simple, low-cost malaria control method well suited to conditions in sub-Saharan Africa. As large seasonal variations in levels of net usage may seriously limit the potential impact of the nets on malaria transmission, a study was conducted on local definitions of seasons, perceptions of seasonal variation in mosquito populations and incidence of febrile illnesses in Bagamoyo District, Tanzania, to aid in the design of a communication strategy for promoting sustained use of the nets. Both the diagnosis and treatment of febrile illnesses are affected by what season people think it is, by what illnesses they think are common in each season, and also by their perceptions of how abundant mosquitoes are. During dry seasons when mosquitoes are scarce and malaria is thought to be unlikely, it will be difficult to attain high rates of net usage. It will be necessary to develop locally-appropriate messages and communication materials that explain how it is possible that malaria can be a threat even when mosquitoes are few. Cultural consensus analysis was found to be a particularly valuable tool for understanding the reasons behind large variations in local perceptions of seasonality.

Chloroquine resistant Plasmodium falciparum in coastal Tanzania. A challenge to the continued strategy of village based chemotherapy for malaria control.

An in vivo study to assess Plasmodium falciparum sensitivity to chloroquine was conducted in two villages of the Bagamoyo District, Tanzania in December 1992. The WHO standard field test (7 days) and the extended test (28 days) were carried out on symptom free children. The presence of chloroquine resistance was confirmed with 59% of infections being found resistant. Fifty-three percent were RI, 2% were at RII and 4% at RIII levels of resistance. Dosage was 25 mg/kg chloroquine base delivered over three days.