Antiprotozoal and molluscicidal activities of five Brazilian plants

Leishmaniasis, Chagas' disease and schistosomiasis (bilharzia) are parasitic diseases with wide distribution on the American continent, affecting millions of people. In the present study, biological assays for antiprotozoal and molluscicidal activities were carried out with ethanolic extracts of plant species from the Brazilian part of the Upper Paraná River. Crude extracts were obtained by percolation with absolute ethanol from the leaves of Cayaponia podantha Cogn., Nectandra falcifolia (Nees) Castiglioni and Paullinia elegans Cambess., as well as from the aerial parts of Helicteres gardneriana St. Hil. & Naud. and Melochia arenosa Benth., all belonging to genera used in folk medicine. Trypanocidal activity of plants was assayed on epimastigote cultures in liver infusion tryptose. Anti-leishmanial activity was determined over cultures of promastigote forms of the parasite in Schneider's Drosophila medium. Microscopic countings of parasites, after their incubation in the presence of different concentrations of the crude extracts, were made in order to determine the percentage of growth inhibition. C. podantha and M. arenosa, at a concentration of 10 µg/mL, showed 90.4 ± 11.52 and 88.9 ± 2.20% growth inhibition, respectively, of epimastigote forms of Trypanosoma cruzi, whereas N. falcifolia demonstrated an LD50 of 138.5 µg/mL against promastigote forms of Leishmania (Viannia) braziliensis. Regarding molluscicidal activity, the acute toxicity of the extracts on Biomphalaria glabrata was evaluated by a rapid screening procedure. M. arenosa was 100% lethal to snails at 200 µg/mL and showed an LD50 of 143 µg/mL. Screening of plant extracts represents a continuous effort to find new antiparasitic drugs.

Cytoadhesion of Plasmodium falciparum-infected erythrocytes and the infected placenta: a two-way pathway

Malaria is undoubtedly the world's most devastating parasitic disease, affecting 300 to 500 million people every year. Some cases of Plasmodium falciparum infection progress to the deadly forms of the disease responsible for 1 to 3 million deaths annually. P. falciparum-infected erythrocytes adhere to host receptors in the deep microvasculature of several organs. The cytoadhesion of infected erythrocytes to placental syncytiotrophoblast receptors leads to pregnancy-associated malaria (PAM). This specific maternal-fetal syndrome causes maternal anemia, low birth weight and the death of 62,000 to 363,000 infants per year in sub-Saharan Africa, and thus has a poor outcome for both mother and fetus. However, PAM and non-PAM parasites have been shown to differ antigenically and genetically. After multiple pregnancies, women from different geographical areas develop adhesion-blocking antibodies that protect against placental parasitemia and clinical symptoms of PAM. The recent description of a new parasite ligand encoded by the var2CSA gene as the only gene up-regulated in PAM parasites renders the development of an anti-PAM vaccine more feasible. The search for a vaccine to prevent P. falciparum sequestration in the placenta by eliciting adhesion-blocking antibodies and a cellular immune response, and the development of new methods for evaluating such antibodies should be key priorities in mother-child health programs in areas of endemic malaria. This review summarizes the main molecular, immunological and physiopathological aspects of PAM, including findings related to new targets in the P. falciparum var gene family. Finally, we focus on a new methodology for mimicking cytoadhesion under blood flow conditions in human placental tissue.

Genetic polymorphisms in vitamin D receptor, vitamin D-binding protein, Toll-like receptor 2, nitric oxide synthase 2, and interferon-γ genes and its association with susceptibility to tuberculosis

Mycobacterium tuberculosis kills more people than any other single pathogen, with an estimated one-third of the world's population being infected. Among those infected, only 10% will develop the disease. There are several demonstrations that susceptibility to tuberculosis is linked to host genetic factors in twins, family and associated-based case control studies. In the past years, there has been dramatic improvement in our understanding of the role of innate and adaptive immunity in the human host defense to tuberculosis. To date, attention has been paid to the role of genetic host and parasitic factors in tuberculosis pathogenesis mainly regarding innate and adaptive immune responses and their complex interactions. Many studies have focused on the candidate genes for tuberculosis susceptibility ranging from those expressed in several cells from the innate or adaptive immune system such as Toll-like receptors, cytokines (TNF-α, TGF-β, IFN-γ, IL-1b, IL-1RA, IL-12, IL-10), nitric oxide synthase and vitamin D, both nuclear receptors and their carrier, the vitamin D-binding protein (VDBP). The identification of possible genes that can promote resistance or susceptibility to tuberculosis could be the first step to understanding disease pathogenesis and can help to identify new tools for treatment and vaccine development. Thus, in this mini-review, we summarize the current state of investigation on some of the genetic determinants, such as the candidate polymorphisms of vitamin D, VDBP, Toll-like receptor, nitric oxide synthase 2 and interferon-γ genes, to generate resistance or susceptibility to M. tuberculosis infection.

In vitro larvicidal activity of geraniol and citronellal against Contracaecum sp (Nematoda: Anisakidae)

Human infection with fish parasites can result from the ingestion of incompletely cooked or raw fish, giving origin to parasitic diseases such as anisakiasis, caused by parasites of the Anisakidae family. The present study assessed the in vitro larvicidal effect of two monoterpene compounds, geraniol and citronellal, against Contracaecum sp (Nematoda: Anisakidae). Four hundred live larvae of Contracaecum sp obtained from "traíra" fish (Hoplias malabaricus, Bloch, 1974) were analyzed on 40 Petri dishes (10 larvae each) with the compounds to be tested. The final concentrations tested for each compound were 250, 125, 62.5, and 31.2 µg/mL and the evaluation was carried out at five different times (2, 4, 8, 24, and 48 h). The larvicidal action of geraniol and citronellal was statistically superior (P < 0.005) to the control (1% ethanol) at concentrations of 250 and 31.2 µg/mL (geraniol) and 250, 125, and 62.5 μg/mL (citronellal). However, no larvicidal activity was observed at concentrations of 125 and 62.5 µg/mL for geraniol and 31.2 µg/mL for citronellal. When the larvicidal action of geraniol was compared to that of citronellal, the former was found to be statistically superior (P < 0.05) to the latter at concentrations of 250 and 31.2 μg/mL. On the other hand, citronellal was statistically superior (P < 0.005) to geraniol at concentrations of 125 and 62.5 μg/mL. The larval mortality rate in terms of time (hours) was higher for geraniol with the passing of time at the 250 μg/mL concentration. At this concentration (in 48 h) the best larvicidal effect was observed with 90% lethality. The larvae were considered to be dead using no motility and loss of structural integrity as parameters. The data indicate that natural terpene compounds should be more explored for antihelminthic activity and can be useful for other studies about anisakiasis treatment.

Novel regulatory SNPs in the promoter region of the TNFRSF18 gene in a Gabonese population

Parasites are accountable for driving diversity within immune gene families. We identified and investigated regulatory single nucleotide polymorphisms (SNPs) in the promoter regions of the tumor necrosis factor receptor superfamily member 18 (TNFRSF18) gene by direct sequencing in a group of male Gabonese individuals exposed to a wide array of parasitic diseases such as malaria, filariasis and schistosomiasis. Two new promoter variants were identified in 40 individuals. Both novel variants were heterozygous and were linked to SNP #rs3753344 (C/T), which has been described. One of the SNP variants (ss2080581728) was close to the general transcription factor site, the TATA box. We further validated these new promoter variants for their allelic gene expression using transient transfection assays. One new promoter variant with two base changes (C/T - ss2080581728/rs3753344) displayed an altered expression of the marker gene. Both novel variants remained less active at the non-induced state in comparison to the major allele. The allele frequencies observed in this study were consistent with data for other African populations. The detection and analysis of these human immune gene polymorphisms contribute to a better understanding of the interaction between host-parasite and expression of Treg activity.

Association of MICA gene polymorphisms with liver fibrosis in schistosomiasis patients in the Dongting Lake region

Major histocompatibility complex class I chain-related A (MICA) is a highly polymorphic gene located within the MHC class I region of the human genome. Expressed as a cell surface glycoprotein, MICA modulates immune surveillance by binding to its cognate receptor on natural killer cells, NKG2D, and its genetic polymorphisms have been recently associated with susceptibility to some infectious diseases. We determined whether MICA polymorphisms were associated with the high rate of Schistosoma parasitic worm infection or severity of disease outcome in the Dongting Lake region of Hunan Province, China. Polymerase chain reaction-sequence specific priming (PCR-SSP) and sequencing-based typing (SBT) were applied for high-resolution allele typing of schistosomiasis cases (N = 103, age range = 36.2-80.5 years, 64 males and 39 females) and healthy controls (N = 141, age range = 28.6-73.3 years, 73 males and 68 females). Fourteen MICA alleles and five short-tandem repeat (STR) alleles were identified among the two populations. Three (MICA*012:01/02, MICA*017 and MICA*027) showed a higher frequency in healthy controls than in schistosomiasis patients, but the difference was not significantly correlated with susceptibility to S. japonicum infection (Pc > 0.05). In contrast, higher MICA*A5 allele frequency was significantly correlated with advanced liver fibrosis (Pc < 0.05). Furthermore, the distribution profile of MICA alleles in this Hunan Han population was significantly different from those published for Korean, Thai, American-Caucasian, and Afro-American populations (P < 0.01), but similar to other Han populations within China (P > 0.05). This study provides the initial evidence that MICA genetic polymorphisms may underlie the severity of liver fibrosis occurring in schistosomiasis patients from the Dongting Lake region.

Heparan sulphate, its derivatives and analogues share structural characteristics that can be exploited, particularly in inhibiting microbial attachment

Heparan sulphate (HS) and the related polysaccharide, heparin, exhibit conformational and charge arrangement properties, which provide a degree of redundancy allowing several seemingly distinct sequences to exhibit the same activity. This can also be mimicked by other sulphated polysaccharides, both in overall effect and in the details of interactions and structural consequences of interactions with proteins. Together, these provide a source of active compounds suitable for further development as potential drugs. These polysaccharides also possess considerable size, which bestows upon them an additional useful property: the capability of disrupting processes comprising many individual interactions, such as those characterising the attachment of microbial pathogens to host cells. The range of involvement of HS in microbial attachment is reviewed and examples, which include viral, bacterial and parasitic infections and which, in many cases, are now being investigated as potential targets for intervention, are identified.

Experimental models in vaccine research: malaria and leishmaniasis

Animal models have a long history of being useful tools, not only to test and select vaccines, but also to help understand the elaborate details of the immune response that follows infection. Different models have been extensively used to investigate putative immunological correlates of protection against parasitic diseases that are important to reach a successful vaccine. The greatest challenge has been the improvement and adaptation of these models to reflect the reality of human disease and the screening of vaccine candidates capable of overcoming the challenge of natural transmission. This review will discuss the advantages and challenges of using experimental animal models for vaccine development and how the knowledge achieved can be extrapolated to human disease by looking into two important parasitic diseases: malaria and leishmaniasis.

Effects of a hydroxy-cinnamoyl conjugate of spermidine in the neuromuscular junction of crayfish /

N'-coumaroyl spermidine (NlCSpd) is a plant derived chemical which is proposed to belong to a class of low molecular weight neuroactive substances called phenolic polyamines. NlCSpd is stnicturally similar to glutamate receptor blocking toxins found in certain spiders and wasps, such as JSTX-3 and NSTX-3 found in Nephila spiders. The goal of the present study was to determine if plant-derived phenolic polyamines act like other structurally related chemicals found in Arthropod venoms, such as JSTX-3, and whether they can be classified in the same pharmacological group as the spider and wasp toxins. A comparison was made to determine the relative potencies of various phenolic polyamines fi-om plants and insect venoms. This comparison was done by measuring the effect of various concentrations ofNlCSpd on the amplitude of excitatory postsynaptic potentials (EPSPs) elicited in muscle of the crayfish Proccanbarus clarkii. NlCSpd was also tested on L-glutamate induced potentials to determine if a postsynaptic component to sj^naptic block occurs. NlCSpd and an analogue with an a longer polyamine chain, NlCSpm, blocked EPSPs in a dose dependent manner, NlCSpd having an IC50 of lOOnM. NlCSpd also blocked L-glutamate induced potentials. The two main components of the NlCSpd molecule alone are insufficient for activity. NlCSpd acts postsynaptically by interfering with crayfish glutamatergic synaptic transmission, likely blocking glutamate receptors by interacting with the same site(s) as other phenolic polyamines. Certain moieties on the polyamines molecule are necessary for activity while others are not.

Synthesis of 4-hydroxycinnamic amides of di-, tri-, and tetraamines : potential insect toxins /

The monoconjugates of phenolic acids (i.e. coumaric acid) with polyamines such as spermidine and spermine are strikingly similar to some toxins from spiders and predatory wasps. Many plants contain phenolic acid polyamine conjugates and there is some reliable information supporting their roles as plant defense chemicals. Eleven monoacylated compounds of diamines, triamines, tetraamines and oxa-polyamine amines were prepared in three to seven steps: 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 and 32. The synthesis proceeds through stepwise construction of the polyamine backbone (as in 62 and 72), followed by protection and deprotection steps of the amino functions. Desymmetrization of readily available and prepared symmetrical polyamines is a key step in the synthesis. The protecting groups employed were tert-butoxycarbonyl (BOC) and trifluoroacetyl (TFA) group which were removed under different conditions: acid and base respectively. Deprotection and refunctionalization of the polyamine reagent demonstrated the versatility of these systems for N-acylation.

Host-parasite relationships in mycoparasite /

A mycoparasite, Piptocephalis virginiana ^ shows a resemblance to fungal parasites of higher plants in the fine structure of hyphae and haustoria. The morphology and fine structure of host and parasitic fungi have been described. The mode of penetration of the host cell, Choanephora cucurbitarum , probably involves mechanical forces. Although the presence of cell wall degrading enzyme was not detected by conventional techniques, its role in penetration can't be ruled out. A collar around the haustorial neck is formed as an extension of the host cell wall. No papilla was detected although appressorixim was seen during penetration. The young haustorium is enclosed in highly invaginating plasmalemma of the host cell and n\imerous cisternae of endoplasmic reticulum. Appearance of an electron—dense sheath around the mature haustorium seems to coincide with the disappearance of cisternae of endoplasmic reticulum from the host cystoplasm in the vicinity of the haustorium. The role of host cytoplasm particularly of endoplasmic reticulum in the development of the sheath is discussed. Extensive accumulation of spherosomes-like bodies, containing lipids, is found in haustorium, parasite and host hypha. Electron microscope revealed the parasiticculture spore has more lipid content than the axenic culture spore of P. virginiana . The biochemical and cytochemical tests also support these results. The mature spore of C. cucurbitarum possesses a thick three-layered cell wall, different from the hyphal wall. Its germination is accompanied by the formation of an elastic thin inner layer which surrounds the emerging germ tube and the growing hypha. High resolution autoradiography showed that H N-acetyl-glucosamine , a precursor of chitin, was incorporated preferentially in the thin inner layer of the spore wall and also in the cell wall of the growing hypha. When the label was fed to the infected cells, at different intervals after inoculation, grains were observed on the sheath which developed around the haustorium of P. virginiana , 30 hours after inoculation. The significance of these results in relation to the origin and composition of the sheath is discussed.

Investigations on the host-parasite interface of a mycoparasite system /

The cell wall composition of Choanephora cucur - bitarum and the host-parasite interface, after infection with Piptocephalis virginiana , were examined in detail. The cell walls of C_. cucurbitarum were determined to be composed of chitin (17%), chitosan (28.4%), neutral sugars (7.2%),uronic acid (2.4%), proteins (8.2%) and lipids (13.8%). The structure of hyphal walls investigated by electron microscopy of shadowed replicas before and after alkali-acid hydrolysis, showed two distinct regions: microfibrillar and amorphous. The microfibrils which were composed of mainly chitin, were organized into two distinct layers: an outer, thicker layer of randomly orientated microfibrils and an inner, thin layer of parallel microfibrils.Electronmicrographs of the host-parasite interface of C_. cucurbitarum and the mycoparasite , P_. virginiana , 30 h following inoculation, showed that the sheath zone has a similar electron density to that of the host cell wall. The sheath was not present around the young (18 h old) haustorium. High-resolution autoradiographs of infected host hyphae showed that radioactive N-acetyl-D-glucosamine , a precursor of chitin, was incorporated preferentially in the host cell wall and sheath zone. Cell fractionation of label fed hyphae showed that 84% of the label was present in the cell wall and specifically in the chitin portion of the wall. The antifungal antibiotic, Polyoxin D, a specific inhibitor of the enzyme, chitin synthetase, suppressed the incorporation of the label in the cell wall and sheath zone and resulted in a decrease in electron density of the developing sheath. The significance of these results is discussed in the light of host resistance.

The nesting biology of Ceratina (Hymenoptera: Apidae) in the Niagara Region : new species, nest site selection and parasitism

One of the most common bee genera in the Niagara Region, the genus Ceratina (Hymenoptera: Apidae) is composed of four species, C. dupla, C. calcarata, the very rare C. strenua, and a previously unknown species provisionally named C. near dupla. The primary goal of this thesis was to investigate how these closely related species coexist with one another in the Niagara ~ee community. The first necessary step was to describe and compare the nesting biologies and life histories of the three most common species, C. dupla, C. calcarata and the new C. near dupla, which was conducted in 2008 via nest collections and pan trapping. Ceratina dupla and C. calcarata were common, each comprising 49% of the population, while C. near dupla was rare, comprising only 2% of the population. Ceratina dupla and C. near dupla both nested more commonly in teasel (Dipsacus sp.) in the sun, occasionally in raspberry (Rubus sp.) in the shade, and never in shady sumac (Rhus sp.), while C. calcarata nested most commonly in raspberry and sumac (shaded) and occasionally in teasel (sunny). Ceratina near dupla differed from both C. dupla and C. calcarata in that it appeared to be partially bivoltine, with some females founding nests very early and then again very late in the season. To examine the interactions and possible competition for nests that may be taking place between C. dupla and C. calcarata, a nest choice experiment was conducted in 2009. This experiment allowed both species to choose among twigs from all three substrates in the sun and in the shade. I then compared the results from 2008 (where bees chose from what was available), to where they nested when given all options (2009 experiment). Both C. dupla and C. calcarata had the same preferences for microhabitat and nest substrate in 2009, that being raspberry and sumac twigs in the sun. As that microhabitat and nest substrate combination is extremely rare in nature, both species must make a choice. In nature Ceratina dupla nests more often in the preferred microhabitat (sun), while C. calcarata nests in the preferred substrate (raspberry). Nesting in the shade also leads to smaller clutch sizes, higher parasitism and lower numbers of live brood in C. calcarata, suggesting that C. dupla may be outcompeting C. calcarata for the sunny nesting sites. The development and host preferences of Ceratina parasitoids were also examined. Ceratina species in Niagara were parasitized by no less than eight species of arthropod. Six of these were wasps from the superfamily Chalcidoidea (Hymenoptera), one was a wasp from the family Ichneumonidae (Hymenoptera) and one was a physogastric mite from the family Pyemotidae (Acari). Parasites shared a wide range of developmental strategies, from ichneumonid larvae that needed to consume multiple Ceratina immatures to complete development, to the species from the Eulophidae (Baryscapus) and Encyrtidae (Coelopencyrtus), in which multiple individuals completed development inside a single Ceratina host. Biological data on parasitoids is scarce in the scientific literature, and this Chapter documents these interactions for future research.

Soil-Transmitted Helminth Infections, Nutrition and Growth in School-age Children from Rural Communities in Honduras

Background: Soil-transmitted helminth (STH) infections are endemic in Honduras but their impact on children’s health is not well studied. Objectives: To evaluate the prevalence and intensity of STH infections and their association with nutrition and growth in a sample of Honduran children. Methodology: A cross-sectional study was done among Honduran rural school-age children in 2011. Blood and stool samples and anthropometric measurements were obtained to determine nutritional status, STH infection and growth status, respectively. Results: The STH prevalence among 320 studied children was 72.5%. Prevalence by species was 30%, 67% and 16% for Ascaris, Trichuris and 16% hookworms, respectively. High intensity infections were associated with decreased growth scores but regardless of intensity, co-infections negatively affected growth indicators. Conclusions: The health burden of STH infections is related to high parasitic load but also to the presence of low-intensity concurrent infections. The synergistic effects of polyparasitism in underprivileged children warrants more attention.

Soil-Transmitted Helminths, Poverty, and Malnutrition in Honduran Children Living in Remote Rural Communities

Soil-transmitted helminth (STH) infections are endemic in Honduras, but their prevalence according to the levels of poverty in the population has not been examined. The present cross-sectional study is aimed to determine the role of different levels of poverty in STH prevalence and infection intensity as well as the potential associations of STH infections with malnutrition and anemia. Research participants were children attending a medical brigade serving remote communities in Northern Honduras in June 2014. Demographic data were obtained, and poverty levels were determined using the unsatisfied basic needs method. STH infections were investigated by the Kato-Katz method; hemoglobin concentrations were determined with the HemoCue system; and stunting, thinness, and underweight were determined by anthropometry. Data were analyzed using descriptive statistics and univariate and multivariable logistic regression models. Among 130 children who participated in this study, a high prevalence (69.2%) of parasitism was found and the poorest children were significantly more infected than those living in less poor communities (79.6% vs. 61.8%; P = 0.030). Prevalence rates of Trichuris trichiura, Ascaris lumbricoides, and hookworms were 69.2%, 12.3%, and 3.85%, respectively. In total, 69% of children had anemia and 30% were stunted. Households’ earthen floor and lack of latrines were associated with infection. Greater efforts should be made to reduce STH prevalence and improve overall childhood health, in particular, among the poorest children lacking the basic necessities of life.