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Transmission of Ehrlichia risticii, the agent of Potomac horse fever, using naturally infected aquatic insects and helminth vectors: preliminary report.

Ehrlichia risticii, the agent of Potomac horse fever (PHF), has been recently detected in trematode stages found in snail secretions and in aquatic insects. Based on these findings, horses could conceivably be exposed to E. risticii by skin penetration with infected cercariae, by ingestion of infected cercariae in water or via metacercariae in a second intermediate host, such as an aquatic insect. In order to test this hypothesis, horses were challenged with infectious snail secretions and aquatic insects collected from a PHF endemic region in northern California. Two horses stood with their front feet in water harbouring E. risticii-infected cercariae, 2 horses drank water harbouring E. risticii-infected cercariae, and 6 horses were fed pools of different aquatic insects harbouring E. risticii-infected metacercariae. In this preliminary study, only the one horse infected orally with mature caddisflies (Dicosmoecus gilvipes) developed the clinical and haematological disease syndrome of PHF. The agent was isolated from the blood of the infected horse in a continuous cell line and identified as E. risticii by characterisation of the 16S rRNA gene. Therefore, E. risticii is maintained in nature in a complex aquatic ecosystem and transmission to horses can occur through accidental ingestion of insects such as caddisflies containing infected metacercariae. At present, the small number of horses used in this study does not exclude other insects and free trematode stages as potential sources of infection.

Animals↗

Transformation systems in insects.

Genetic transformation is an important technology that provides unique opportunities to find, isolate, and analyze genes, as well as to create organisms with unique functional characteristics. Insect biologists have been developing genetic transformation technologies that rely extensively on transposable elements. A number of class II transposable elements isolated originally from insects have been converted into broad host range insect gene vectors. Class II transposable elements are particularly amenable to gene vector development, although they suffer from some limitations such as low rates of recombination. Use of these gene vectors requires the physical introduction of the vectors into developing insect embryos by microinjection. Microinjection methods vary to accommodate the unique physical and developmental characteristics of the target insects. All methods rely on the use of fine glass needles in conjunction with micromanipulators and a microscope. A serious constraint on the use of existing systems can be the inefficiency of successfully delivering the gene vectors to the germ cells of the developing embryo. The general method for vector delivery to insect germ cells is described, as well as variations that are useful under some conditions.

Animals↗

Control of malaria vectors with the insect growth regulator pyriproxyfen in a gem-mining area in Sri Lanka.

The study was conducted in eight adjacent villages in central Sri Lanka where there are many shallow pits dug by gem miners that fill with water. These become breeding places of the main malarial vector Anopheles culicifacies, and of the second most important vector Anopheles subpictus, but not of Anopheles varuna, the third most important vector. With the help of local volunteers, data on the adult populations of these three species was collected by various standard methods, and data on the incidence of malaria cases was collected by two clinics set up for the project and through the existing hospitals. Prevalence of malaria infection in symptom-less people was investigated by mass blood surveys. On the basis of a year's pre-intervention data the villages were stratified into four with high levels of malaria transmission and four with lower transmission. Within each stratum two villages were randomly assigned for mosquito control by treating all the gem pits, as well as river bed pools, with a granular formulation of the insect growth regulator pyriproxyfen at a target dose of 0.01 mg a.i./litre. The intervention caused significant reductions in the adult populations of An. culicifacies and An. subpictus. Similarly, incidence of malaria was reduced in the intervention villages to about 24% (95% c.l. 20-29%) of that in the controls. Prevalence of parasitaemia also declined significantly. It is concluded that in this situation where, with active community participation, the breeding sites of the main vectors could be located; vector control by a highly active and persistent insect growth regulator can be a very effective means of malaria control.

Adolescent↗

Aquatic insects as a vector for Mycobacterium ulcerans.

Mycobacterium ulcerans is an emerging environmental pathogen which causes chronic skin ulcers (i.e., Buruli ulcer) in otherwise healthy humans living in tropical countries, particularly those in Africa. In spite of epidemiological and PCR data linking M. ulcerans to water, the mode of transmission of this organism remains elusive. To determine the role of aquatic insects in the transmission of M. ulcerans, we have set up an experimental model with aquariums that mimic aquatic microenvironments. We report that M. ulcerans may be transmitted to laboratory mice by the bite of aquatic bugs (Naucoridae) that are infected with this organism. In addition, M. ulcerans appears to be localized exclusively within salivary glands of these insects, where it can both survive and multiply without causing any observable damage in the insect tissues. Subsequently, we isolated M. ulcerans from wild aquatic insects collected from a zone in the Daloa region of Ivory Coast where Buruli ulcer is endemic. Taken together, these results point to aquatic insects as a possible vector of M. ulcerans.

Animals↗

Characterization of constitutive and putative differentially expressed mRNAs by means of expressed sequence tags, differential display reverse transcriptase-PCR and randomly amplified polymorphic DNA-PCR from the sand fly vector Lutzomyia longipalpis.

Molecular studies of insect disease vectors are of paramount importance for understanding parasite-vector relationship. Advances in this area have led to important findings regarding changes in vectors' physiology upon blood feeding and parasite infection. Mechanisms for interfering with the vectorial capacity of insects responsible for the transmission of diseases such as malaria, Chagas disease and dengue fever are being devised with the ultimate goal of developing transgenic insects. A primary necessity for this goal is information on gene expression and control in the target insect. Our group is investigating molecular aspects of the interaction between Leishmania parasites and Lutzomyia sand flies. As an initial step in our studies we have used random sequencing of cDNA clones from two expression libraries made from head/thorax and abdomen of sugar fed L. longipalpis for the identification of expressed sequence tags (EST). We applied differential display reverse transcriptase-PCR and randomly amplified polymorphic DNA-PCR to characterize differentially expressed mRNA from sugar and blood fed insects, and, in one case, from a L. (V.) braziliensis-infected L. longipalpis. We identified 37 cDNAs that have shown homology to known sequences from GeneBank. Of these, 32 cDNAs code for constitutive proteins such as zinc finger protein, glutamine synthetase, G binding protein, ubiquitin conjugating enzyme. Three are putative differentially expressed cDNAs from blood fed and Leishmania-infected midgut, a chitinase, a V-ATPase and a MAP kinase. Finally, two sequences are homologous to Drosophila melanogaster gene products recently discovered through the Drosophila genome initiative.

Animals↗