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Background Anopheles funestus is a major vector of malaria in sub-Saharan Africa. In order to apply effective control measures against this vector, it is necessary to understand the underlying physiological factors that play a critical role in its development, reproduction, fertility and susceptibility to insecticides. One enzyme family involved in the above mentioned biological pathways is the esterases. The aim of this study was to analyse esterase activity levels at different ages during the life-span of adult Anopheles funestus Giles in order to better understand the complex biological processes in this species. Methods Isoenzyme electrophoresis (IEE) was used to examine the esterase activity in laboratory colonised An. funestus adults aged between 2 h (h) and 30 days post eclosion as well as in wild An. funestus adults aged between 2 h and 15 days post eclosion. Esterase activity was quantified by densitometry analysis of the IEE gels. Esterases were classified according to their activity inhibition by organic phosphates, eserine sulphate and sulphydryl reagents. Results Nine esterases IEE profiles were common to both the laboratory colonised and wild An. funestus adults. These esterases were further divided into acetylesterases, arylesterases, carboxylesterases and acetylcholinesterase. The activity level of certain specific esterases was primarily influenced by age and/or gender. Conclusions The information from this study contributes towards the general understanding of esterase enzyme activity variation in adults of a major malaria vector An. funestus. This variation likely carries physiological and adaptive significance and may influence specific characteristics, such as reproductive fitness and insecticide resistance that are epidemiologically important.

A parasitological study was conducted on 66 dogs seroreactive for Leishmania captured as a control measure of visceral leishmaniasis in the State of Rio de Janeiro, Brazil. Biological samples from different anatomical sites were collected during autopsy of the animals and cultured on biphasic medium (NNN/Schneider). The Leishmania isolates were characterized by isoenzyme electrophoresis. Leishmania was isolated from 80.3% of the animals: 12 animals with Leishmania (Viannia) braziliensis isolated exclusively from cutaneous lesions, 39 with L. (L.) chagasi isolated from different sites in the same animal, and 2 with simultaneous isolation of L. (V.) braziliensis from cutaneous lesions and L. (L.) chagasi from different sites. Isolation in culture revealed the absence of Leishmania parasites in 13 animals. The results obtained confirm the existence of mixed infections in dogs in Rio de Janeiro and indicate the need to complement the investigation of seroreactive dogs using methods for the parasitological diagnosis and identification of Leishmania species.

INTRODUCTION: Visceral leishmaniasis presents urban behavior in some Brazilian cities, with domestic dogs as the main infection source. In Cuiabá, MT, canine visceral leishmaniasis was diagnosed and characterized as recommended by the Ministry of Health. METHODS: Biological samples from suspected canine carriers were analyzed by the isoenzyme electrophoresis technique. The 6PGDH enzyme and reference strain IOC/L0566 (MHOM/BR/1975/M2903) of Leishmania (Leishmania) infantum was used as one of the controls. RESULTS: Electrophoresis analysis revealed that the canine isolates belonged to the species L. (L.) infantum. CONCLUSIONS: The authors emphasize the importance of species characterization, particularly in areas of mixed infection like Cuiabá.

A fortuitously discovered case of feline leishmaniosis is reported. The parasites were found in the skin and the bone marrow of a domestic female cat that spontaneously died after a few weeks of evolution. Serological tests for FeLV, FIV and PIF virus detection gave negative results. By using Western blot serology, a characteristic pattern of leishmaniosis was obtained and by performing an isoenzyme electrophoresis, a Leishmania infantum MON-1 strain was identified. The same zymodeme is implicated in most of the canine and human leishmaniosis in Southern Europe. A study on the prevalence of asymptomatic feline leismaniosis is foreseen.

Fourteen Giardia duodenalis isolates were examined using the RAPD (random amplified polymorphic deoxyribonucleic acid) technique. Simple reproducible polymorphisms were generated using 3 different RAPD primers. The results generated by each primer were very similar and were significantly correlated with each other. These data were then compared to existing isoenzyme electrophoresis data on the same isolates. The RAPD data divided the isolates into 10 groupings or rapdemes while the isoenzyme data divided them into 10 similar zymodemes. Both methods grouped 4 isolates (BAH42, BAH44c9, BAH12c9 and BAH39c7), which comprised a phenotypically heterogeneous assemblage with respect to growth rate and metabolism, into similar groupings. The 2 methods were significantly correlated (P

Isoenzyme electrophoresis using 13 enzyme systems was applied to 31 Australian and 7 Swiss isolates of Giardia of human, cat, cattle, dog, sheep and rat origin. The Portland (ATCC No. 30888) reference strain was also included. The 39 isolates were divided into 22 different zymodemes. These consisted of 19 zymodemes containing the P1 and Australian isolates and three zymodemes containing Swiss isolates only. Differences in enzyme profiles between zymodemes was measured by euclidean distance and it was found that Australian isolates of Giardia exhibited more variation than the Swiss isolates. Relationships between zymodemes determined by clustering analysis are discussed with particular reference to the zoonotic potential of Giardia.

Summary: The accurate identification of the causative organisms of disease is fundamental to the study of epidemiology. Hence molecular tools are now widely used to detect and distinguish pathogens, and have greatly improved our understanding of epidemiology. David Godfrey pioneered the use of molecular markers in the epidemiology of African trypanosomiasis, thus enabling the light of reliable evidence to shine on this previously problematic and controversial subject area. From the early 1970’s David’s group employed first isoenzyme electrophoresis and subsequently DNAbased characterization methods to aid identification of trypanosomes collected from a range of endemic countries across Africa. These investigations had a major impact on our understanding of the zoonotic nature of human trypanosomiasis in Africa and of the genetic diversity of African trypanosomes.

The limited usefulness of traditional taxonomic methods combined with the discovery of a Strongyloides parasitic in man in Papua New Guinea (PNG) that resembles S. fuelleborni, a parasite of man and other primates in tropical Africa, has precipitated the need to apply new methods to the taxonomy of the genus. In this study isoenzyme electrophoresis has been used on Strongyloides isolates of many different origins. Cluster analysis of the data suggested that existence of three main groups within the material considered, consisting of (1) isolates of S. stercoralis, (2) isolates from PNG domestic animals, and (3) isolates from PNG man and African non-human primates. The taxonomic implications of these groups are considered.

Free-living amoebae (FLA) are distributed ubiquitously in aquatic environments with increasing importance in hygienic, medical and ecological relationships to man. In this study, water samples from Belgian industrial cooling circuits were quantitatively surveyed for the presence of FLA. Isolated, thermotolerant amoebae were identified morphologically as well as using the following molecular methods: enzyme-linked immunosorbent assay and isoenzyme electrophoresis and PCR. Thermophilic amoebae were present at nearly all collection sites, and the different detection methods gave similar results. Naegleria fowleri was the most frequently encountered thermotolerant species, and concentrations of thermotolerant FLA were correlated with higher temperatures.

Forty human strains of Leishmania infantum isolated in 1985-99 from 17 patients with repeated cutaneous, mucosal or visceral leishmaniasis episodes in Catalonia (Spain) were examined by isoenzyme electrophoresis. Six zymodemes were revealed: MON-1, MON-24, MON-28, MON-29, MON-33 and MON-34. In 2 patients 2 different zymodemes were identified in consecutive episodes.

We phenotypically characterized 43 leishmanial parasites from cutaneous leishmaniasis by isoenzyme electrophoresis and the indirect immunofluorescence antibody test (23 McAbs). Identifications revealed 11 (25.6%) strains of Leishmania (V .) braziliensis , 4 (9.3%) of L . (V .) shawi shawi , 7 (16.3%) of L . (V .) shawi santarensis , 6 (13.9%) of L . (V .) guyanensis and L . (V .) lainsoni , 2 (4.7%) of L . (L .) amazonensis , and 7 (16.3%) of a putative hybrid parasite, L . (V .) guyanensis /L . (V .) shawi shawi . McAbs detected three different serodemes of L . (V .) braziliensis : I-7, II-1, and III-3 strains. Among the strains of L . (V .) shawi we identified two populations: one (7 strains) expressing the B19 epitope that was previously considered to be species-specific for L . (V .) guyanensis . We have given this population sub-specific rank, naming it L . (V .) s . santarensis . The other one (4 strains) did not express the B19 epitope like the L . (V .) shawi reference strain, which we now designate as L . (V .) s . shawi . For the first time in the eastern Brazilian Amazon we register a putative hybrid parasite (7 strains), L . (V .) guyanensis /L . (V .) s . shawi , characterized by a new 6PGDH three-band profile at the level of L . (V .) guyanensis . Its PGM profile, however, was very similar to that of L . (V .) s . shawi . These results suggest that the lower Amazon region – western Para state, Brazil, represents a biome where L . (V .) guyanensis and L . (V .) s . shawi exchange genetic information.

Thirty isolates of Giardia duodenalis from humans and felines were compared by isoenzyme electrophoresis. Using 10 enzyme systems, 13 different zymodemes were distinguished. The majority of zymodemes could be divided into two groups: one group comprising human and feline isolates with worldwide geographic distribution; the other group containing human isolates restricted to Western Australia. A number of isolates showed multiple-banded patterns and the genetic significance of these findings is discussed. The marked heterogeneity of G. duodenalis demonstrated in this study is considered in relation to the epidemiology of giardiasis. The findings are consistent with felines serving as a reservoir of infection to humans.

Blood from 115 camels in Kuwait was examined for blood parasites. Two camels of a local herd (1.7%) were found to be infected with Trypanosoma (Trypanozoon) evansi and three camels (2.6%) with microfilarial nematodes. The Trypanosoma stocks isolated from these two camels were screened for isoenzyme patterns of 10 enzymes using thin-layer starch-gel electrophoresis. The results revealed that these two stocks were identical to camel stocks of T. evansi from certain countries in Africa, as well as to two stocks isolated from dogs in Kuwait. This is the first record of Trypanosoma (Trypanozoon) evansi isolates and microfilariae from camels in Kuwait.

SUMMARYA total of 114 stocks of Trypanosoma congolense originating from Kenya, Uganda, Tanzania, Zambia and Sudan, but including, for comparison, stocks from The Gambia, Liberia, Ivory Coast, Nigeria and Cameroun, were compared by isoenzyme electrophoresis for 6 enzymes. The zymodemes were grouped, both from a dendrogram and using a cladistic method, after calculating the dissimilarity, or distance, between profiles. Previous observations are broadly confirmed, the zymodemes clustering separately according to geographical origin and ecological zone. Thus, one group was composed almost entirely of East African stocks, and another of stocks from both East and West Africa, although each group was of savanna origin. A third group was composed of stocks from the humid, rain-forest zones of West Africa, and was particularly characterized by isoenzyme variants of superoxide dismutase and glucose-phosphate isomerase. Two stocks from the Kenyan coast formed a markedly separate group, which may be taxonomically distinct.

Six strains of Leishmania isolated from wild mammals and humans on the Pacific Coast of Ecuador were identified by isoenzyme electrophoresis and by their reactivity patterns to a cross-panel of specific monoclonal antibodies using a radioimmune binding assay. Single isolates from Sciurus vulgaris. Potos flavus, and Tamandua tetradactyla were identified as Leishmania amazonensis. Three other strains, isolated from cutaneous lesions of humans, were identified as Leishmania panamensis.

Cultures of old, well documented strains of Entamoeba histolytica (NIH:200, HK9 etc.) have been compared with “E. histolytica-like” strains and, in turn, with E. moshkovskii, E. invadens and E. chattoni. They have been grouped according to their isoenzyme patterns. The “E. histolytica-like” organisms give patterns similar to E. moshkovskii and both of these are different from E. histolytica Groups I to IV. E. invadens and E. chattoni have easily distinguished characteristic isoenzyme bands. “E. polecki” is indistinguishable from E. histolytica.

The substrate specificity of aspartate aminotransferase (ASAT, E.C. 2.6.1.1.) from Leishmania was examined following observations of artefacts on gels stained for alanine aminotransferase (ALAT, E.C. 2.6.1.2.) after thin-layer starch-gel electrophoresis. Leishmanial ASAT acted on L-aspartate, L-alanine, L-tryptophan and L-tyrosine. Interpretation of ALAT zymograms must thus take into account the presence of interfering ASAT bands, and the need is emphasized for rigorous controls in isoenzyme electrophoresis.

Guinea-pigs were inoculated with Entamoeba histolytica strains isolated from cases of amoebic liver abscess, amoebic dysentery and asymptomatic cyst passers and the strains were classified in zymodemes by isoenzyme electrophoresis. It was observed that certain non-pathogenic zymodemes were potentially pathogenic for the guinea-pigs. The relationship of zymodemes and virulence in laboratory animals is discussed.

112 Leishmania isolates, obtained in French Guiana from human lesions, phlebotomine sandflies and wild mammals, were characterized by isoenzyme electrophoresis. Leishmania braziliensis guyanensis and L. mexicana amazonensis were found parasitizing different natural hosts. L.b. guyanensis was the dominant species (103 isolates) responsible for most of the human lesions (96·7%). Based on variations observed in 2 enzymes, 3 distinct zymodemes were distinguished within the L.b. guyanensis taxon.

Trypanosoma evansi was seen in blood samples taken randomly from both wild and semi-captive capybaras caught in Mato Grosso do Sul State, Brazil and in sick dogs brought into local veterinary clinics. Trypanosome stocks from capybaras and dogs were significantly different in their patterns of growth in mice, while the trypanosomes from dogs were mostly dyskinetoplastic. By isoenzyme electrophoresis all the trypanosomes were of the most common type of T. evansi found elsewhere.

The frequency of hybrid formation between two Trypanosoma brucei clones during cyclical transmission through Glossina morsitans centralis was analyzed. In two independent experiments, teneral G. m. centralis were infected with an equal mixture of two T. brucei clones showing different homozygous isoenzyme patterns for isocitrate dehydrogenase (ICD; E.C.1.1.1.42) and alkaline phosphatase (AP; E.C. 3.1.3.1). Trypanosomes were cyclically transmitted to mice from 23 infective flies and the subsequent bloodstream-form populations were characterized by isoenzyme electrophoresis. Heterozygous patterns for ICD and AP indicated that hybrid formation occurred in at least 9 of the 23 vectors. There was further evidence that extrusion of hybrid parasites with saliva from a single fly was not necessarily continuous but could alter over time with the occurrence of either or both of the homozygous parental clones.

Isoenzyme electrophoresis of 7 different enzyme systems was used to compare 24 strains of Naegleria fowleri and 6 strains of N. gruberi. The 30 strains could be grouped into 4 distinct categories based upon zymogram patterns. No interstrain band variation in all enzyme systems was demonstrated in pathogenic strains of N. fowleri. Three nonpathogenic high temperature-tolerant strains of Naegleria had similar zymograms. Four of the 5 remaining nonpathogenic Naegleria strains had no interstrain band variation. Based upon zymograms, the 22 pathogenic strains constitute a homogenous species. Similarly the high temperature-tolerant nonpathogenic strains formed a cohesive group. The remaining nonpathogenic strains could be separated into 2 groups.

This study presents the results of the characterisation of 17 strains ofLeishmania by isoenzyme electrophoresis from a focus of leishmaniasis in southern Spain: two from human visceral leishmaniasis, four from human cutaneous leishmaniasis and 11 from sandflies. The 17 strains are grouped in 6 zymodemes characterised by their variability as regards to the electrophoretic mobility of the enzymes MDH, G6PD, NP and ME. Thus, we confirm the high intraspecific variability ofLeishmania (L.) infantum in a focus of southern Spain, as already suggested by previous studies. Zymodemes GR-15 and GR-17 are also described for the first time in Spain, and they characteristically possess the same relative electrophoretic mobility in the enzyme ME (93). Sixteen zymodemes of theL. infantum complex found in southern Spain were numerically analysed on the basis of the enzymatic profiles of 122Leishmania strains characterised from this area.