Annia Alba

Facilitated invasion of an overseas invader: human mediated settlement and expansion of the giant African snail, Lissachatina fulica , in Cuba

The giant African snail, Lissachatina fulica, is considered one of the most invasive species worldwide, acting as a crop pest and diseases vector. It was first detected in Cuba in 2014 and is dispersing throughout Havana. We mapped 34 sites in the vicinity of Havana to assess its spread and analysed ecological (forestation and humidity) and anthropogenic (pollution and religious sites) factors in relation to the presence/absence of the snails using multivariate correspondence analysis. There were 14 sites at which the snail was present and where religious rituals of the Yoruba creed, an African rooted religion, were observed. No other variables showed significant relationships. This indicates that the rituals may be a major factor in the dispersal of the snail in Havana and more widely in Cuba. In light of this an outreach program with key Yoruba leaders may help in slowing the dispersal of the snail within Cuba, once the threats posed by this species are known.

Detection and identification of Leishmania spp.: application of two hsp70-based PCR-RFLP protocols to clinical samples from the New World

Leishmaniasis is highly prevalent in New World countries, where several methods are available for detection and identification of Leishmania spp. Two hsp70-based PCR protocols (PCR-N and PCR-F) and their corresponding restriction fragment length polymorphisms (RFLP) were applied for detection and identification of Leishmania spp. in clinical samples recruited in Colombia, Guatemala, and Honduras. A total of 93 cases were studied. The samples were classified into positive or suspected of leishmaniasis according to parasitological criteria. Molecular amplification of two different hsp70 gene fragments and further RFLP analysis for identification of Leishmania species was done. The detection in parasitologically positive samples was higher using PCR-N than PCR-F. In the total of samples studied, the main species identified were Leishmania panamensis, Leishmania braziliensis, and Leishmania infantum (chagasi). Although RFLP-N was more efficient for the identification, RFLP-F is necessary for discrimination between L. panamensis and Leishmania guyanesis, of great importance in Colombia. Unexpectedly, one sample from this country revealed an RFLP pattern corresponding to Leishmania naiffi. Both molecular variants are applicable for the study of clinical samples originated in Colombia, Honduras, and Guatemala. Choosing the better tool for each setting depends on the species circulating. More studies are needed to confirm the presence of L. naiffi in Colombian territory.

A multiplex PCR for the detection of Fasciola hepatica in the intermediate snail host Galba cubensis.

Fasciolosis is a snail-borne trematode infection that has re-emerged as a human disease, and is considered a significant problem for veterinary medicine worldwide. The evaluation of the transmission risk of fasciolosis as well as the efficacy of the strategies for its control could be carried out through epidemiological surveillance of the snails that act as intermediate hosts of the parasites. The present study aimed to develop the first multiplex PCR to detect Fasciola hepatica in Galba cubensis, an important intermediate host of the parasite in the Americas and especially in the Caribbean basin. The multiplex PCR was optimized for the amplification of a 340 bp fragment of the second internal transcribed spacer (ITS-2) of F. hepatica rDNA, while another set of primers was designed and used to amplify a conserved segment of the nuclear 18S rDNA of the snail (451 bp), as an internal control of the reaction. The assay was able to detect up to 100 pg of the parasite even at high concentrations of snail DNA, an analytical sensitivity that allows the detection of less than a single miracidium, which is the minimal biological infestation unit. A controlled laboratory-reared G. cubensis - F. hepatica system was used for the evaluation of the developed multiplex PCR, and 100% sensitivity and specificity was achieved. This assay constitutes a novel, useful and suitable technique for the survey of fasciolosis transmission through one of the main intermediate hosts in the Western hemisphere.

Fasciola hepatica - Pseudosuccinea columella interaction: effect of increasing parasite doses, successive exposures and geographical origin on the infection outcome of susceptible and naturally-resistant snails from Cuba

BackgroundPseudosuccinea columella is one of the most widespread vectors of Fasciola hepatica, a globally distributed trematode that affects humans, livestock and wildlife. The exclusive occurrence in Cuba of susceptible and naturally-resistant populations to F. hepatica within this snail species, offers a fascinating model for evolutionary biology, health sciences and vector control strategies. In particular, resistance in P. columella is characterized by the encapsulation of the parasite by host’s immune cells and has been experimentally tested using different Cuban F. hepatica isolates with no records of successful infection. Here, we aimed to explore for the first time, the effect of different parasite doses, successive exposures and different parasite origins on the infection outcomes of the two phenotypes of P. columella occurring in Cuba.MethodsTo increase the chances for F. hepatica to establish, we challenged Cuban P. columella with increasing single parasite doses of 5, 15 or 30 miracidia and serial exposures (three-times) of 5 miracidia using a sympatric F. hepatica isolate from Cuba, previously characterized by microsatellite markers. Additionally, we exposed the snails to F. hepatica from different geographical origins (i.e. Dominican Republic and France). Parasite prevalence, redial burden and survival of snails were recorded at 25 days post-exposure.ResultsNo parasite development was noted in snails from the resistant populations independent of the experimental approach. Contrastingly, an overall increase in prevalence and redial burden was observed in susceptible snails when infected with high miracidia doses and after serial exposures. Significant differences in redial burden between single 15 miracidia and serial 3 × 5 miracidia infected snails suggest that immune priming potentially occurs in susceptible P. columella. Compatibility differences of allopatric (Caribbean vs European) F. hepatica with susceptible snails were related to the geographical scale of the combinations.ConclusionsHere, the effectiveness of P. columella resistance to F. hepatica does not decline with increasing parasite doses, successive infection or different geographical origins of parasite isolates, while presenting new evidence for specificity for infection in susceptible P. columella snails. Understanding the peculiarities of the P. columella-F. hepatica interaction and the extent of the resistant phenotype is crucial for an effective parasite control and for developing alternatives to tackle fasciolosis transmission.

Updated distribution and experimental life-history traits of the recently invasive snail Lissachatina fulica in Havana, Cuba

The giant African snail Lissachatina fulica has been reported invading Cuba since 2014 and is now well established in areas of Havana and several nearby regions. This invasive species is of major concern to health authorities given its role as an important vector of parasites such as Angiostrongylus cantonensis, the causative agent of eosinophilic meningoencephalitis in humans. Here, we update the distribution within Cuba. We also report on our studies of experimental life-history traits to assess the population dynamics and potential for spread of this species in Cuba. The experimental population had a very low probability of dying at first age intervals (Type I survival curve) with a life expectancy of 71 weeks. During our experiment, sexual maturity was reached after 22 weeks (individuals of 70 mm shell length) and eggs were laid regularly (mean batch size: 188 ± 111.79 SD). We failed to detect any deviations from the mortality curve and individuals reached an average of 77.3 mm (shell length) and weighted 57.7 g after one year. Predicted curve models indicate that snails reaching their average lifespan of five years should attain 10-12 cm (shell length) and weight 160 g. The spreading of this invasive and vector snail has been tracked for four years in Cuba showing a steady increase of invaded localities. How fast and how far this species develops in Cuba is unknown but the life history parameters indicated in this paper show that it has a large potential to invade all areas of Cuba quickly unless a systematic abatement strategy is developed.

Isolation of 1E4 IgM Anti-Fasciola hepatica Rediae Monoclonal Antibody from Ascites: Comparison of Two Purification Protocols

Purification of immunoglobulin M (IgM) antibodies could be challenging, and is often characterized by the optimization of the purification protocol to best suit the particular features of the molecule. Here, two different schemes were compared to purify, from ascites, the 1E4 IgM monoclonal antibody (mAb) previously raised against the stage of redia of the trematode Fasciola hepatica. This immunoglobulin is used as capture antibody in an immunoenzymatic assay to detect parasite ongoing infection in its intermediate hosts. The first purification protocol of the 1E4 mAb involved two chromatographic steps: an affinity chromatography on a Concanavalin A matrix followed by size exclusion chromatography. An immunoaffinity chromatography was selected as the second protocol for one-step purification of the antibody using the crude extract of adult parasites coupled to a commercial matrix. Immunoreactivity of the fractions during purification schemes was assessed by indirect immunoenzymatic assays against the crude extract of F. hepatica rediae, while purity was estimated by protein electrophoresis. Losses on the recovery of the antibody isolated by the first purification protocol occurred due to protein precipitation during the concentration of the sample and to low resolution of the size exclusion molecular chromatography step regarding this particular immunoglobulin. The immunoaffinity chromatography using F. hepatica antigens as ligands proved to be the most suitable protocol yielding a pure and immunoreactive antibody. The purification protocols used are discussed regarding efficiency and difficulties.

Changes in Structure and Composition of Two Communities of Rocky Shores Molluscs Exposed to Different Human Impact in Playa Jibacoa, Cuba

Rocky shore molluscs are highly relevant in keeping shore’s dynamics and ecological balance of beaches. The knowledge regarding species distribution patterns is important in understanding how environmental and anthropogenic factors may influence the structure of these communities. This study aimed to explore changes in structure and composition of littoral molluscs in two sites of Playa Jibacoa, Cuba with different human use. Fortyfour species of littoral molluscs were identified. The supralittoral zones exhibited the lowest diversity whereas the interlittoral and sublittoral the highest. These findings point the latter as the most vulnerable zones to human perturbation. Species were more abundant and evenly distributed in the site with less human activity. This effect showed a seasonal pattern since in both sites these variables were more affected in the summer probably because of an increased attendance of tourists to the beach. Species were mostly associated with the type of substrate and the stratum than to any other variable. Significant changes in diversity were observed between seasons with a decrease in summer.