Antitumor and Antiviral Activity of Colombian Medicinal Plant Extracts

Extracts of nine species of plants traditionally used in Colombia for the treatment of a variety of diseases were tested in vitro for their potential antitumor (cytotoxicity) and antiherpetic activity. MTT (Tetrazolium blue) and Neutral Red colorimetric assays were used to evaluate the reduction of viability of cell cultures in presence and absence of the extracts. MTT was also used to evaluate the effects of the extracts on the lytic activity of herpes simplex virus type 2 (HSV-2). The 50% cytotoxic concentration (CC50) and the 50% inhibitory concentration of the viral effect (EC50) for each extract were calculated by linear regression analysis. Extracts from Annona muricata, A. cherimolia and Rollinia membranacea, known for their cytotoxicity were used as positive controls. Likewise, acyclovir and heparin were used as positive controls of antiherpetic activity. Methanolic extract from Annona sp. on HEp-2 cells presented a CC50 value at 72 hr of 49.6x103mg/ml. Neither of the other extracts examined showed a significant cytotoxicity. The aqueous extract from Beta vulgaris, the ethanol extract from Callisia grasilis and the methanol extract Annona sp. showed some antiherpetic activity with acceptable therapeutic indexes (the ratio of CC50 to EC50). These species are good candidates for further activity-monitored fractionation to identify active principles.

Study of Schistosoma mansoni isolates from patients with failure of treatment with oxamniquine

After three successive treatments with oxamniquine the continuing elimination of Schistosoma mansoni eggs was observed in patients, who came from various regions of Brazil, with different clinical forms of schistosomiasis. The objective of the present study was to determine the experimental behaviour of five different S. mansoni isolates in Swiss Webster mice that were submitted to treatment with the same drug. The experimental group with failure of treatment showed higher mean number of surviving male worms when it was compared to the group without failure of treatment. These date suggest the possibility of resistance to oxamniquine.

Studies on the virulence and attenuation of Trypanosoma cruzi using immunodeficient animals

Tissue invasion and pathology by Trypanosoma cruzi result from an interaction between parasite virulence and host immunity. Successive in vivo generations of the parasite select populations with increasing ability to invade the host. Conversely, prolonged in vitro selection of the parasite produces attenuated sublines with low infectivity for mammals. One such subline (TCC clone) has been extensively used in our laboratory as experimental vaccine and tested in comparative experiments with its virulent ancestor (TUL). The experiments here reviewed aimed at the use of immunodeficient mice for testing the infectivity of TCC parasites. It has not been possible to obtain virulent, revertant sublines by prolonged passaged in such mice.

Characteristics of the bological cycle of Lutzomyia evandroi Costa Lima & Antunes, 1936 (Diptera: Psychodidae) under experimental conditions

Lutzomyia evandroi Costa Lima and Antunes, 1936 is found in Rio Grande do Norte, northeastern Brazil, in areas of visceral and mucocutaneous leishmaniasis and follows the same geographic distribution of L. longipalpis. The biological cycle, oviposition, morphological and behavioral characteristics of the species were studied under experimental conditions. The average number of eggs per wild caught female varied from 21 to 50 eggs along the year, with a peak occurring between January and March and another in August, with oviposition lasting for 4 to 12 days. The mean larval phase was 24 days. Ovipositing rates were influenced by rainfall and temperature indexes, with an increase of eggs per oviposition at the beginning and at the end of the rainy season, and a decrease at the peak of the rainy season.

Cytotoxic and Antiviral Activities of Colombian Medicinal Plant Extracts of the Euphorbia genus

Forty-seven plant extracts of 10 species of the genus Euphorbia (Euphorbiaceae) used by Colombian traditional healers for the treatment of ulcers, cancers, tumors, warts, and other diseases, were tested in vitro for their potential antitumour (antiproliferative and cytotoxic) and antiherpetic activity. To evaluate the capacity of the extracts to inhibit the lytic activity of herpes simplex virus type 2 (HSV-2) and the reduction of viability of infected or uninfected cell cultures, the end-point titration technique (EPTT) and the MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] colorimetric assay were used, respectively. The therapeutic index of the positive extracts for the antiviral activity was determined by calculating the ratio CC50 (50% cytotoxic concentration) over IC50 (50% inhibitory concentration of the viral effect). Five of the 47 extracts (11%) representing 3 out of 10 Euphorbia species (30%) exhibited antiherpetic action; the highest activity was found in the leaf/stem water-methanol extracts from E. cotinifolia and E. tirucalli. The therapeutic indexes of these two plant species were > 7.1; these extracts exhibited no cytotoxicity. Six extracts (13%) representing 4 plant species (40%) showed cytotoxic activity. The highest cytotoxicity was found in the dichloromethane extract obtained from E. cotinifolia leaves and the CC50 values for the most susceptible cell lines, HEp-2 and CHO, were 35.1 and 18.1 µg/ml, respectively.

A technique for preparing polytene chromosomes from Aedes aegypti (Diptera, Culicinae)

Polytene chromosome preparations were obtained from larval, pupal and adult female Malpighian tubules of Aedes aegypti. The Malpighian tubules of the pupae (0-4 h old) from larvae reared at 20ºC provided the best cytogenetic analysis. The interaction of nucleic acids and proteins that influence the spreading of the chromosomes could be reduced with the preparation technique of the sheets submitted to a stronger treatment starting with the hypotony of tissue and successive bathings with acetic acid. A simple technique should facilitate molecular cytogenetics used in the location of resistance and vector competence genes.

Chagas disease: current epidemiological trends after the interruption of vectorial and transfusional transmission in the Southern Cone countries

Chagas disease, named after Carlos Chagas who first described it in 1909, exists only on the American Continent. It is caused by a parasite, Trypanosoma cruzi, transmitted to humans by blood-sucking triatomine bugs and by blood transfusion. Chagas disease has two successive phases, acute and chronic. The acute phase lasts 6 to 8 weeks. After several years of starting the chronic phase, 20% to 35% of the infected individuals, depending on the geographical area will develop irreversible lesions of the autonomous nervous system in the heart, esophagus, colon and the peripheral nervous system. Data on the prevalence and distribution of Chagas disease improved in quality during the 1980's as a result of the demographically representative cross-sectional studies carried out in countries where accurate information was not available. A group of experts met in Brasília in 1979 and devised standard protocols to carry out countrywide prevalence studies on human T. cruzi infection and triatomine house infestation. Thanks to a coordinated multi-country program in the Southern Cone countries the transmission of Chagas disease by vectors and by blood transfusion has been interrupted in Uruguay in1997, in Chile in 1999, and in 8 of the 12 endemic states of Brazil in 2000 and so the incidence of new infections by T. cruzi in the whole continent has decreased by 70%. Similar control multi-country initiatives have been launched in the Andean countries and in Central America and rapid progress has been recorded to ensure the interruption of the transmission of Chagas disease by 2005 as requested by a Resolution of the World Health Assembly approved in 1998. The cost-benefit analysis of the investments of the vector control program in Brazil indicate that there are savings of US$17 in medical care and disabilities for each dollar spent on prevention, showing that the program is a health investment with good return. Since the inception in 1979 of the Steering Committee on Chagas Disease of the Special Program for Research and Training in Tropical Diseases of the World Health Organization (TDR), the objective was set to promote and finance research aimed at the development of new methods and tools to control this disease. The well known research institutions in Latin America were the key elements of a world wide network of laboratories that received - on a competitive basis - financial support for projects in line with the priorities established. It is presented the time line of the different milestones that were answering successively and logically the outstanding scientific questions identified by the Scientific Working Group in 1978 and that influenced the development and industrial production of practical solutions for diagnosis of the infection and disease control.

Arthropod and filarioid parasites associated with wild rodents in the northeast marshes of Buenos Aires, Argentina

During 1995, 16 species of arthropods and 2 species of filarioids, totaling 1 287 specimens were collected from 64 wild rodents captured in the Hudson Natural Reserve, Buenos Aires, Argentina. Infestation parameters and indexes were analyzed. Host specific richness was S = 6, diversity H = 1.48, and relative density RDI = 40%. High values of parasite species richness and diversity were found on Oligoryzomys delticola (S = 9; H = 1.40), Oxymycterus rufus (S = 9; H = 1.37), and Oligoryzomys flavescens (S = 9; H = 1.28), followed by Scapteromys aquaticus (S = 6; H = 0.17), and Akodon azarae (S = 4; H = 1.20). Deltamys kempi was infested only by Androlaelaps rotundus. O. delticola and O. flavescens showed the highest similarity index (O = 74.19%), followed by O. flavescens with S. aquaticus, as a result of historical processes and shared microhabitats. Considering arthropods-filarioids associations, significant affinity was observed in Litomosoides bonaerensis with Hoplopleura travassosi, Laelaps paulistanensis, and Gigantolaelaps wolffsohni.

Strongyloides venezuelensis alkaline extract for the diagnosis of human strongyloidiasis by enzyme-linked immunosorbent assay

The present study was conducted to detected IgG antibodies using Strongyloides venezuelensis alkaline extract for the diagnosis of human strongyloidiasis by the enzyme-linked immunosorbent assay (ELISA). Sera from 90 subjects were analyzed (30 with strongyloidiasis, 30 with other parasites and 30 healthy individuals). Results were expressed in antibody titers, which were considered as positive when titer was > 80. Sensibility and specificity of the assay were 100% and 96.7%, respectively. It can be concluded that the heterologous alkaline extract could be employed in ELISA as a diagnostic aid in human strongyloidiasis, due to its advantages as easiness of obtaining, practicability in preparing, and high indexes of sensitivity and specificity.

Insecticide and community interventions to control Triatoma dimidiata in localities of the State of Veracruz, Mexico

Three different interventions to control Triatoma dimidiata in the State of Veracruz were implemented: X-1 = whole dwelling spraying, X-2 = middle wall spraying, X-3 = household cleaning. Cyfluthrin was sprayed 3 times with 8 month intervals. After each spraying, insects were collected and sent to the laboratory to be recorded and to determine genus and species of the adult triatomine bugs, and nymphs were counted. Trypanosoma cruzi presence was determined. With X-1, the infestation, colonization, and natural infection indexes were reduced to 0% in the 3 localities, with respect to t0. With X-2, the infestation index was reduced to 10% at t3 in 3 localities; the colonization index was reduced to 0% in only 1 locality at t3, and the natural infection index was reduced to 0% at t3. With X-3 the 3 indexes were not effectively reduced but they decreased with respect to the baseline study. Insecticide application to the whole dwelling is a more efficient intervention than its application to only the lower half of the walls and to the cleaning of houses.

Life tables and reproductive parameters of Lutzomyia spinicrassa (Diptera: Psychodidae) under laboratory conditions

Lutzomyia spinicrassa is a vector of Leishmania braziliensis in Colombia. This sand fly has a broad geographical distribution in Colombia and Venezuela and it is found mainly in coffee plantations. Baseline biological growth data of L. spinicrassa were obtained under experimental laboratory conditions. The development time from egg to adult ranged from 59 to 121 days, with 12.74 weeks in average. Based on cohorts of 100 females, horizontal life table was constructed. The following predictive parameters were obtained: net rate of reproduction (8.4 females per cohort female), generation time (12.74 weeks), intrinsic rate of population increase (0.17), and finite rate of population increment (1.18). The reproductive value for each class age of the cohort females was calculated. Vertical life tables were elaborated and mortality was described for the generation obtained of the field cohort. In addition, for two successive generations, additive variance and heritability for fecundity were estimated.

Evaluation of methoprene effect on Aedes aegypti (Diptera: Culicidae) development in laboratory conditions

Several Brazilian Aedes aegypti populations are resistant to the larvicidae temephos. Methoprene, that inhibits adult emergence, is one of the alternatives envisaged by the Brazilian Dengue Control Program (PNCD). However, at Brazil vector infestation rates are measured through larvae indexes and it has been claimed that methoprene use in the field could face operational problems. In order to define a standardized protocol, methoprene effect was evaluated in laboratory conditions after continuous exposure of larvae (Rockefeller strain) to a methoprene formulation available to the PNCD. Methoprene-derived mortality occurs mainly at the pupa stage and pupa development is inversely proportional to methoprene concentration. Number and viability of eggs laid by treated and control females are equivalent. A methoprene dose-dependent delay in the development was noted; however, b correlations were found for total mortality or adult emergence inhibition if data obtained when all control mosquitoes have emerged are compared to data obtained when methoprene-treated groups finish development. The cumulative record of total methoprene-induced mortality at the time control adults emerge is proposed for routine evaluation of field populations. Mortality of all specimens, but not of larva, could account for adult emergence inhibition, confirming the inadequacy of larvae indexes to evaluate methoprene effect.

Monitoring of resistance to the pyrethroid cypermethrin in Brazilian Aedes aegypti (Diptera: Culicidae) populations collected between 2001 and 2003

Resistance to cypermethrin of different Aedes aegypti Brazilian populations, collected at two successive periods (2001 and 2002/2003), was monitored using the insecticide-coated bottles bioassay. Slight modifications were included in the method to discriminate between mortality and the knock down effect. Although this pyrethroid was recently started to be used in the country to control the dengue vector, a decrease in susceptibility was noted between both periods analyzed, particularly in the city of Rio de Janeiro. The results indicate that resistance is due at least in part to a target site alteration.

Dispersal pattern of the sand fly Lutzomyia neivai (Diptera: Psychodidae) in a cutaneous leishmaniasis endemic rural area in Southeastern Brazil

The dispersal pattern of the sand fly Lutzomyia neivai was studied through mark-release-recapture experiments in an American cutaneous leishmaniasis endemic rural area in Southeastern Brazil. Over 6500 specimens were marked with fluorescent powder and released in forest edge and peridomicile habitats from August to November 1999, February and April 2000. Recapture attempts were made using Shannon and CDC traps up to eight successive nights after releases. A total of 493 (7.58%) specimens were recaptured. The number of recaptured males and females of L. neivai in CDC traps was not affected by the distance between the trap and the release points. Approximately 90% of males and females recaptured in CDC traps were caught up to 70 m from the release points. The maximum female flight range recorded was 128 m. The average flight range per day was less than 60 m for males and females. Of the flies released in forest edge, approximately 16% of the recaptured females were caught in Shannon traps in the peridomicile habitat. The results indicate that the movements of L. neivai are spatially focal and the possibility of dispersion from forest to peridomicile habitat may be an important way of contracting leishmaniasis in dwellings.

Ticks, ivermectin, and experimental Chagas disease

Following an infestation of dogticks in kennels housing dogs used for long-term studies of the pathogenesis of Chagas disease, we examined the effect of ivermectin treatment on the dogs, ticks, trypanosome parasites, and also on triatomine vectors of Chagas disease. Ivermectin treatment was highly effective in eliminating the ticks, but showed no apparent effect on the dogs nor on their trypanosome infection. Triatominae fed on the dogs soon after ivermectin treatment showed high mortality, but this effect quickly declined for bugs fed at successive intervals after treatment. In conclusion, although ivermectin treatment may have a transient effect on peridomestic populations of Triatominae, it is not the treatment of choice for this situation. The study also showed that although the dogticks could become infected with Trypanosoma cruzi, this only occurred when feeding on dogs in the acute phase of infection, and there was no evidence of subsequent parasite development in the ticks.