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Biomedical subjects

F H Collins

Publications and source records attributed to F H Collins.

At least 109 records · Page 6Linked to original sources
PubMed1990

Plasmodium falciparum infection rates in Anopheles gambiae, An. arabiensis, and An. funestus in western Kenya.

Mosquitoes collected monthly for 1 year from human habitations in the Kisumu area of western Kenya were identified by morphological characters as Anopheles gambiae Giles sensu lato (An. gambiae s.l.) or An. funestus. Of the mosquitoes collected, 7,244 (67%) of the An. gambiae s.l. and 8,511 (87%) of the An. funestus were tested by enzyme-linked immunosorbent assay (ELISA) for the presence of Plasmodium falciparum circumsporozoite (CS) protein. ELISA positivity rates were 8.2% for An. gambiae s.l. and 6.1% for An. funestus. Both An. gambiae and An. arabiensis were detected among 432 ELISA-positive and 668 ELISA-negative An. gambiae s.l. identified to species with a ribosomal DNA probe. The species-specific infection rates were calculated to be 9.6% for An. gambiae and 0.4% for An. arabiensis. These results confirm that An. gambiae and An. funestus are the primary malaria vectors in western Kenya and that An. arabiensis is a relatively minor vector.

Animals↗
PubMed1989

Site-specific ribosomal DNA insertion elements in Anopheles gambiae and A. arabiensis: nucleotide sequence of gene-element boundaries.

The nucleotide sequence of the junctions between the 28S ribosomal gene and site-specific insertion elements from two sibling mosquito species, Anopheles gambiae and A. arabiensis, is reported. In both species, elements insert at the same point within the 28S gene, but this site is 634 basepairs (bp) 3' of the R1 (Type I) insertion site in Drosophila melanogaster. The two mosquito elements each have poly A tails and a polyadenylation signal, but the extreme 3' and 5' ends show no other similarity to each other or to any other insertion element. In both mosquito species, identical target site duplications of 17 bp are generated. The sequence TNTCCCTNT found in this duplication is also found in the 14 bp target site duplications that flank R1 elements in D. melanogaster. Another sequence in this duplication, GGGATAACT, is very similar to the sequence GGGAGTAACT found in the 24 base sequence required by the Bombyx mori R2 endonuclease.

Animals↗
PubMed1989

Structure of rDNA in the mosquito Anopheles gambiae and rDNA sequence variation within and between species of the A. gambiae complex.

The structure of the rDNA repeating unit of Anopheles gambiae (Diptera: Culicidae) was determined by restriction endonuclease mapping and hybridization analyses on four independent clones obtained from a genomic library of a colony (G3) from the Gambia (West Africa). rDNA gene coding sequences are conserved, but much intragenomic and intraspecific (geographic) variation occurs in the intergenic spacer. Hybridization of subclones from spacer and coding sequences to genomic DNA that was isolated from single mosquitoes from laboratory colonies of four other A. gambiae complex species reveals conservation of coding sequences but concerted evolution in the intergenic spacers.

Africa, Western↗
PubMed1989

Microgeographic variation in rDNA intergenic spacers of Anopheles gambiae in western Kenya.

The genetic population structure of Anopheles gambiae (Diptera: Culicidae) in western Kenya was investigated by hybridizing a rapidly evolving rDNA intergenic spacer sequence to restriction endonuclease digests of genomic DNA extracted from single mosquitoes from seven localities. Significantly different distributions of restriction fragment arrays were obtained from field sites less than 10 km apart, which suggests restricted gene flow and a subdivided population structure. Eight of twenty-one possible comparisons between pairs of populations yielded significant differences. An eastern Kenya coastal population did not share its restriction fragment arrays with any of the western populations, suggesting that isolation by distance can be complete on a relatively small geographic scale (700 km).

Animals↗
PubMed1989

A general system of resistance to malaria infection in Anopheles gambiae controlled by two main genetic loci.

Genetic analysis of a system of Plasmodium refractoriness in Anopheles gambia suggests that the joint action of 2 unlinked genetic loci substantially controls expression of the susceptible and refractory phenotypes. One genetic component, here named Pif-B (for Plasmodium infectivity factor), is closely linked or identical to a polymorphic autosomal esterase locus which can be visualized by gel electrophoresis. This locus exerts the major controlling effect on susceptibility to Plasmodium cynomolgi B. The other genetic component is independent of esterase and exerts major control over refractoriness to the P. cynomolgi Ceylon strain parasite. Genetic assortment of the esterase-independent component suggests that it is controlled by 1 principal locus, here named Pif-C. The 2 genetic components of Plasmodium refractoriness appear to contribute to the same phenotype through physiologically independent means.

Animals↗
PubMed1989

Association of a Plasmodium-refractory phenotype with an esterase locus in Anopheles gambiae.

In a genetically selected strain of the malaria vector Anopheles gambiae, most species of Plasmodium parasites are surrounded by melanized capsules and killed in the wall of the mosquito midgut. Genetic studies demonstrate a significant association between the refractory response to Plasmodium cynomolgi B strain and the Est A allele at an autosomal esterase locus. Increased susceptibility to parasites is associated with an alternate Est C allele. Lines selected to be homozygous for the Est A and Est C alleles show enhanced levels of refractoriness and susceptibility when compared to the unselected parental stock. Expression of the refractory phenotype is reduced much more than the susceptible phenotype at high parasite density, suggesting that refractoriness may be due to a positive effector.

Alleles↗
PubMed1989

Ultrastructural localization of phenoloxidase in the midgut of refractory Anopheles gambiae and association of the enzyme with encapsulated Plasmodium cynomolgi.

A melanogenic enzyme, phenoloxidase, was localized ultrastructurally in the midgut epithelia of 2 strains of Anopheles gambiae, a refractory strain that melanotically encapsulates Plasmodium cynomolgi ookinetes on the midgut, and a susceptible strain that does not. Midguts were incubated with either dopa or dopamine, and the resultant electron-dense product of phenoloxidase activity was localized on the basal lamina (BL) and cellular basal membrane labyrinth (BML) in uninfected mosquitoes of both strains. In infected refractory mosquitoes, the reaction products still were observed on the BL and BML but were especially dense in the BML of midgut cells near encapsulated ookinetes and in the capsule itself. In infected susceptible mosquitoes, phenoloxidase localization was reduced or absent in the BL and BML and was not observed near parasites. Phenylthiourea (PTU) inhibited the phenoloxidase reaction, indicating that the reaction product deposited in the absence of PTU resulted from enzyme activity and not autooxidation of the substrates. It is concluded that higher levels of phenoloxidase in the refractory strain following a blood meal may contribute to the ability to encapsulate ookinetes.

Animals↗
PubMed1989

A nitrocellulose membrane-based ELISA for the detection of Plasmodium infections in mosquitos.

A nitrocellulose (NC) membrane was evaluated as a solid-phase support for the detection of malaria-infected mosquitos using monoclonal antibodies (MAb) with a laboratory model based on Plasmodium inui and Anopheles dirus. MAbs produced against sporozoites of the N34 strain of P. inui, and selected by immunofluorescence assay and the circumsporozoite precipitin test, were used. A one-site indirect NC-ELISA that used unlabelled MAb and enzyme-labelled anti-mouse IgG was developed. Its sensitivity was about 200 sporozoites and it reliably detected one infected mosquito in a pool of 20. This indirect NC-ELISA has the advantage that it does not require direct conjugation of the MAb to an enzyme or biotin. In the direct one-site NC-ELISA, which is also reported, the relatively simple biotinylation procedure was an alternative to the enzyme- or radiolabelled MAbs. The NC-ELISAs were simple and rapid. Furthermore, the indirect NC-ELISA can be used to detect sporozoite antigen localized in various body sectors of mosquitos.

Animals↗
PubMed1988

The genetics and expression of an esterase locus in Anopheles gambiae.

The main polymorphic system of esterase isoenzymes in adults of the G3 laboratory strain of Anopheles gambiae consists of two to five major bands of activity per individual. The bands are designated 5S, 5F, 13, 14, and 15. In genetic crosses, the genes which coded for the bands assorted as three codominant alleles, Est A, Est B, and Est C, at a single autosomal locus. Homozygotes for the Est C allele were significantly underrepresented among backcross progeny. The developmental pattern of esterase expression was examined. Esterase gene expression in embryos was first detectable between 2 and 12 hr after oviposition. The initiation or termination of expression of some of the bands corresponded to boundaries between developmental stages. Most of the esterase fractions were not specifically localized within the tissues tested, with the exception of a series of bands which were restricted largely to adult male testes.

Animals↗
PubMed1988

Monoclonal antibody-based enzyme-linked immunosorbent assay (ELISA) for detection of Plasmodium malariae sporozoites in mosquitoes.

A monoclonal antibody specific for a repeated epitope of the circumsporozoite protein of Plasmodium malariae sporozoites has been used to develop a two-site, single antibody-based enzyme-linked immunosorbent assay that can detect P. malariae sporozoites in mosquitoes. The assay uses a purified monoclonal antibody produced against sporozoites of the Uganda I/CDC strain of P. malariae to capture the antigen and the same monoclonal antibody labeled with horseradish peroxidase as the detector. Sporozoites have been detected in laboratory-infected mosquitoes stored at room temperature in the presence of a desiccant for as long as 18 months. The detection limit of the assay is approximately 50 P. malariae sporozoites per test well. Cross-reaction has not been observed with mosquitoes infected with P. falciparum, P. vivax, or P. ovale sporozoites.

Animals↗
PubMed1988

Comparison of DNA probe and cytogenetic methods for identifying field collected Anopheles gambiae complex mosquitoes.

A recently developed DNA probe method was compared with the standard cytogenetic method for identifying the species of individual mosquitoes in the Anopheles gambiae complex. The complex consists of 6 morphologically indistinguishable sibling species that include the major African malaria vectors. Half-gravid, field collected mosquitoes were split into 2 portions: the abdomen was preserved for ovarian nurse cell cytotaxonomy and the head/thorax portion was desiccated for DNA extraction. Cytogenetic examination of the Kenya specimens showed 88 An. gambiae and 108 An. arabiensis. The Zimbabwe specimens consisted of 6 An. gambiae and 55 An. Quadriannulatus. All samples of the 3 species were polymorphic for the major chromosomal inversions previously recorded in field specimens from eastern and southern Africa, indicating that the collections reflected natural levels of intraspecific variation in the field populations sampled. Approximately 97% of the cytologically identified mosquitoes were also identified to species by the DNA probe method, and in every case the DNA probe and cytogenetic methods of species identification produced concordant results.

Animals↗
PubMed1988

Ultrastructure of the encapsulation of Plasmodium cynomolgi (B strain) on the midgut of a refractory strain of Anopheles gambiae.

Using transmission electron microscopy, we investigated the encapsulation of the simian malaria parasite, Plasmodium cynomolgi, in a refractory strain of the mosquito, Anopheles gambiae. After the ookinete penetrates the mosquito midgut epithelium and lodges between the basal membrane and the basal lamina, an electron-dense, melanin-like substance begins to coalesce around the parasite. Completely encapsulated parasites were found as early as 16 hr after the blood meal. Granules of the melanin-like substance often appeared to condense onto the parasite from the fluid in the extracellular spaces of the basal membrane labyrinth. Melanin granules also appeared to condense from the hemolymph onto the basal lamina underlying the parasite. In addition, groups of tubules, vesicles, and membranous whorls often were found in midgut cells that were located next to or were enclosing parasites. These structures were unusually electron-dense, and may have been associated with melanization. Hemocytes rarely were observed near completed capsules and neither hemocytes nor their remnants were components of the capsules. During later stages of encapsulation, parasites appeared abnormal and often were infiltrated with melanin. Although late-stage capsules were usually located basally, completed capsules enclosed by membranes were occasionally observed near the apical border of the midgut. Other capsules associated with cellular debris, were found in the lumen of the midgut from 1 to 6 days after the blood meal.

Animals↗
PubMed1987

Detection of antibodies in human sera to the repeating epitope of the circumsporozoite protein of Plasmodium falciparum using the synthetic peptide (NANP)3 in an enzyme-linked immunosorbent assay (ELISA).

An enzyme-linked immunosorbent assay (ELISA) was developed to detect antibody in human sera to a synthetic peptide, Asn-Ala-Asn-Pro (NANP)3, derived from the repeating amino acid sequence found in the surface circumsporozoite protein of Plasmodium falciparum sporozoites. One hundred four sera from U.S. residents were used to determine a cut-off value for reactivity. Test sera were considered reactive when the absorbance was greater than that at the 95th percentile of the control sera. Sera from 112 Kenyans living in an area of holoendemic malaria transmission were tested. Of the total number of sera, 65% had detectable antibody to (NANP)3. The percentage of reactive sera increased from 41% in sera from children under 4 years of age to 85% in sera from adults 20 to 39 years of age. The high exposure to malaria parasites of the Kenyans was reflected in indirect fluorescent antibody assay titers to blood stage P. falciparum parasites. All of the Kenyan sera had antibody present at titers greater than 1:256.

Adolescent↗
PubMed1987

Age-specific prevalence of antibody to a synthetic peptide of the circumsporozoite protein of Plasmodium falciparum in children from three villages in Kenya.

The presence of antibody to the repeating epitope of the circumsporozoite protein of Plasmodium falciparum was determined in children 1 month to 10 years old from three villages in western Kenya using the synthetic peptide (PNAN)5 in an enzyme-linked immunosorbent assay. The percentage of antibody-positive children increased with age and differed in the three villages. The village with the lowest percentage of antibody-positive children had the lowest percentage of infections as determined by detection of blood stage parasites. The villages also differed in the age at which antibody first appeared. In one village, only 12% of the children had antibody by the age of 5; while in the other two villages, 60% and 73% had antibody by 4 years of age.

Age Factors↗
PubMed1987

Assessment of a synthetic DNA probe for Plasmodium falciparum in African blood specimens.

Synthetic DNA oligomers homologous to 21-base long repetitive sequences of Plasmodium falciparum DNA were labeled with 32P using T4 kinase, and were hybridized with purified DNA and with processed blood samples from Africa. The sequence PFR1, its antiparallel oligomer PFR1R, and PFR1 covalently attached to biotin hybridized similarly to P. falciparum DNA. One-microliter aliquots of blood from Zaire spotted on prewet nylon filters and hybridized with PFR1 gave detectable autoradiogram signals from samples with parasitemias as low as 1,000 parasites/mm3. Blood lysis and protein digestion followed by alkylation allowed dot-blot processing of larger aliquots of blood. After hybridization with PFR 1 and autoradiography, 26 samples were scored positive visually, compared with 34 scored positive by microscopy. The effective sensitivity for processed 10-microliter samples was about 500 parasites/mm3. Signals from hybridized probes were quantitated by liquid scintillation counting and densitometry, and were proportional to the amounts of purified P. falciparum DNA applied to the filter. Autoradiogram signals also were roughly proportional (correlation coefficient, r = 0.77) to the number of parasites/mm3 of blood from field samples as determined by microscopic examination.

Animals↗
PubMed1987

A ribosomal RNA gene probe differentiates member species of the Anopheles gambiae complex.

A 0.59 kilobase DNA fragment cloned from an rDNA cistron of the mosquito Anopheles gambiae can be used as a probe to differentiate between A. gambiae, A. arabiensis, and A. melas, three morphologically identical sibling species in the A. gambiae complex which otherwise can be reliably distinguished only by polytene chromosome banding patterns. Although all are important (and often sympatric) African malaria vectors, their relative roles in malaria transmission have thus far been difficult to assess. The probe, an EcoRI-SalI fragment from the 3' end of the 28S beta coding region of the cistron, is present in all three species, but the species differ uniquely with respect to the location of an EcoRI site in the nontranscribed spacer (NTS) downstream of the fragment. We have routinely used the probe to identify A. gambiae complex mosquitoes to species on the basis of genomic DNA extracted from individual air dried specimens. A single mosquito abdomen provides more than sufficient DNA for the assay, and neither eggs nor a bloodmeal in the abdomen interfere with DNA yield. Moreover, the DNA extraction procedure does not degrade the bloodmeal IgG, so the residual protein pellet can be used to identify the mosquito bloodmeal source. Since the rDNA cistron organization as detected by the probe does not differ between male and female mosquitoes, the probe can be used for either sex. Preliminary experiments show that the probe is equally useful for mosquito larvae and pupae.

Animals↗
PubMed1987

Infection of chimpanzees with Nigerian I/CDC strain of Plasmodium ovale.

Seven splenectomized chimpanzees were infected with the Nigerian I/CDC strain of Plasmodium ovale. Two of the animals had no history of previous malarial infection whereas three had been infected with P. vivax, one with P. malariae, and one with P. vivax and P. malariae. The two animals with no previous malarial experience had maximum parasitemias of 88,700 and 127,000 per mm3 while the other animals had maximum parasitemias ranging from 10,100 to 60,600 per mm3. Anopheles freeborni, An. dirus, An. stephensi, and An. gambiae were readily infected via membrane feeding on heparinized blood obtained from these chimpanzees during the ascending phases of their primary attacks. The parasitemias in the chimpanzees with previous malarial experience were transient.

Animals↗