Taenia saginata and T. hydatigena: intramuscular vaccination of calves with oncospheres

This is a preliminary attempt to immunize calves against bovine cysticercosis (Cestoda) by intramuscular injection of artificially hatched homologous or heterologous oncospheres. In the first two experiments five calves were vaccinated with the oncospheres of Taenia saginata. These calves, together with five controls, were subsequently challenged orally with the eggs of T. saginata. At autopsy all the vaccinated animals harbored a colony of living cysticerci at the site of injection. Three of these calves were completely protected against the oral challenge, and in the other two, only one, and two cysticerci, respectively were found in the sites of election. The five controls harbored 1154, 1680, 820, 960, and 1820 living cysticerci, respectively, in the sites of election.     

Effects of Bayluscide on Bithynia siamensis goniomphalos, the first intermediate host of the human liver fluke, Opisthorchis viverrini, in laboratory and field trials

The molluscicidal effects of Bayluscide (niclosamide) were investigated on Bithynia siamensis goniomphalos, the first intermediate host of human liver fluke, Opisthorchis viverrini. Lethal concentrations of 50% (LC₅₀) and 95% (LC₉₅) against young and adult males were 0.38 and 0.80, 0.42 and 0.86 ppm, respectively. The LC₅₀ and LC₉₅ against young and adult females were 0.42 and 0.86, 0.46 and 0.97 ppm, respectively. No significant differences in mortality rate between sexes or snail size (p > 0.05) was detected. Bayluscide-related tissue damage in B. siamensis goniomphalos included detachment of cilia of the epithelial layer of the digestive tract and decreased number of calcium cells. In tests of lethal concentrations of Bayluscide on non-target animals, no lethal effect was observed on Filopaludina martensi martensi (Viviparous snail) but high mortality rates were recorded in Puntius gonionotus fingerling, Ricefish (Oryzias mekongensis) and shrimp (Macrobrachium lanchesteri), but lower in guppy fish (Poecilia reticulata) after 24 h exposure. For field trials, sufficient Bayluscide was sprayed in 3 roadside ditches to result in final concentrations of 5, 10 or 20 ppm, with mortality rates on B. siamensis goniomphalos of 10.94, 20.00 and 31.25%, respectively. Non-target snails died in small numbers but no effect was observed in other aquatic vertebrate animals. Field trials of Bayluscide on B. siamensis goniomphalos revealed low mortality rates, suggesting the need for application methods of higher efficacy or that Bayluscide is not suitable for application to operculate snails or snails which are able to escape by burying in mud.     

Echinococcus granulosus: the establishment of the metacestode is associated with control of complement-mediated early inflammation

In this work we studied the evolution of early inflammation, complement activation and parasite survival/death along the establishment phase of Echinococcus granulosus metacestode. Using a chamber model of infection in mice, we examined cell infiltration and C3 deposition on individual parasites during their development from protoscoleces to cystic forms. We found that the intensity of the initial inflammation decreased around undamaged but not around damaged parasites: at 43dpi undamaged parasites were mostly associated with poor/mild inflammation while damaged parasites with a strong inflammation. In addition, whereas complement activation participated in the induction of early inflammation, the deposition of C3 on undamaged parasites progressively diminished. Interestingly, we observed some parasites in pre-cystic stage with no/poor C3 deposition at 3dpi. Overall, these results indicated that the establishment and survival of the hydatid cyst is associated with the control of complement and, consequently, of local inflammation at the initial phases of infection.     

Involvement of the fungal nuclear migration gene nudC human homolog in cell proliferation and mitotic spindle formation.

Essential genes which are required for normal nuclear migration and play a role in developmental processes have been isolated from model genetic organisms. One such gene is nudC (nuclear distribution C), which is required for positioning nuclei in the cytoplasm of the filamentous fungus Aspergillus nidulans and for normal colony growth. This gene is highly conserved, structurally and functionally, throughout evolution and the human homolog, HnudC, has been cloned. To study the function of nudC in higher eukaryotic cells, HnudC was downregulated by developing triple ribozyme constructs, consisting of two cis-acting ribozymes which liberate an internal trans-acting ribozyme targeted to HnudC. Efficient cleavage sites in HnudC mRNA were identified using a library selection technique and HnudC-targeted internal ribozymes were cloned into a triple ribozyme cassette. Triple ribozyme constructs were subcloned into an ecdysone-inducible expression vector and stably transfected into human embryonic 293 cells. Muristerone A induced expression of the HnudC ribozyme and produced specific reduction of HnudC mRNA. Downregulation of HnudC mRNA resulted in significant inhibition of cell proliferation in clones expressing the HnudC-targeted triple ribozyme, which was not observed in uninduced cells or cells transfected with vector alone. In induced cultures, many mitotic cells demonstrated defects in spindle architecture during mitosis. The most common defect observed was multiple mitotic spindle poles rather than the expected bipolar structure. These data demonstrate the fundamental importance of HnudC in eukaryotic cell proliferation and a functional role for HnudC in spindle formation at mitosis.     

High allozyme diversity and unidirectional linear migration patterns within a population of tetraploid Isoetes sinensis, a rare and endangered pteridophyte

Isoetes sinensis (Isoetaceae), an aquatic quillwort which occurs only in two fragmented sites of China as an allotetraploid, is critically endangered. Genetic variation among eight subpopulations of I. sinensis was examined in the Xin’an River (119°14′–15′E, 29°28′N) by using allozyme polymorphism. Eighteen loci of 10 enzyme systems were examined and used for the analysis of population genetic parameters. As expected for allotetraploids, fixed heterozygosity was found at four loci. A high level of genetic diversity was observed in the population, with mean number of alleles per loci of 1.8, and mean percentage of polymorphic loci of 55.6%, which were much higher than the average values in fern species. The genetic variation within each subpopulation was not positively correlated with its size, which may be explained by high gene flow (Nm = 2.57), clonal reproduction and fixed heterozygosity of allopolyploid. The I. sinensis population contained high clonal diversity (PD = 0.39, D = 0.95), indicating the successful seedling recruitment of the population. Significant positive relationship was detected between clonal diversity and the size of subpopulation. Partitioning the genetic diversity indicated that 91.1% of the genetic variation was within subpopulations and only 8.9% existed among subpopulations. The migration pattern of I. sinensis along the Xin’an River is best explained by a source–sink model, but with unidirectional gene flow among subpopulations underlined by hydrochoric force. The results were then discussed in relation to both in situ and ex situ conservation efforts of the population.     

The effects of host age and superparasitism by the parasitoid, Microplitis rufiventris on the cellular and humoral immune response of Spodoptera littoralis larvae

To study the dynamics of stage-dependent immune responses in Spodoptera littoralis (Boisd.) larvae (Lepidoptera: Noctuidae), single and superparasitism experiments were carried out using the parasitoid Microplitis rufiventris Kok. (Braconidae: Hymenoptera). Compared to younger (preferred) host larvae, the older (non-preferred) host larvae displayed a vigorous humoral response that often damaged and destroyed the single wasp egg or larva. Superparasitism and host age altered both the cellular and humoral immune responses. Younger host larvae showed a stronger encapsulation response compared to older host larvae. Moreover encapsulation rates in younger hosts (e.g., second instar) decreased with increasing numbers of parasitoid eggs deposited/larvae. In older larvae, the encapsulation rate was low in fourth, less in fifth and absent in sixth instar hosts. Conversely, the order and magnitude of the cellular immune response in S. littoralis hosts were highest in second instar larvae with the first instar larvae being a little lower. The immune response steadily decreased from the third through to the fifth instar and was least obvious in the sixth instar. In contrast, the general humoral immune response was most pronounced in sixth instar larvae and diminished towards younger stages. The results suggest that both cellular and humoral responses are stage-dependent. Wasp offspring in younger superparasitized host larvae fought for host supremacy with only one wasp surviving, while supernumerary wasp larvae generally survived in older superparasitized larvae, but were unable to complete development. Older instars seem to have a method for immobilizing/killing wasp larvae that is not operating in the younger instars.     

大棚模拟条件下角倍蚜春季迁飞数量动态及其与气象因子的关系

2009-2011年,在四川峨眉对大棚模拟条件下角倍蚜春季的迁飞数量及其与气象因子的关系进行观察分析,结果表明春迁蚜的迁飞活动与天气条件密切相关,在迁飞期内,当日平均气温≥9℃、最高气温≥13℃、平均相对湿度≤80％,春迁蚜开始迁飞;当日平均气温≥15 ℃、最高气温≥22 ℃、日照时数≥7 h、平均相对湿度≤65％,春迁蚜大量迁飞.在迁飞初期和中期,春迁蚜迁飞数随气温升高而增加.当天气条件适宜时,春迁蚜日迁飞数量的变化为单峰型,前期和末期低、中期高;当迁飞期内遭遇激烈降温,日迁飞数量波动较大,呈双峰或多峰型,同时迁飞期延长、迁飞率和单位面积迁飞量下降.相关分析与主成分分析结果:春迁蚜日迁飞数与平均气温、日最高气温和日照时数呈极显著正相关(P＜0.001),与平均相对湿度、最小相对湿度呈显著负相关(P＜0.05),与日最低气温和降水量相关性不显著(P＞0.1);表明温度和湿度是影响春迁蚜迁飞的关键因子,日照次之.     

The relationships between the burden of adult parasites, host age and the microfilarial density in human onchocerciasis

We investigate the relationship between the microfilarial density in the skin and the burden of adult female Onchocerca volvulus by analysing pre-control nodulectomy data which allow for a direct approach, independent of exposure. The data of 169 patients in Burkina Faso and 182 patients in Liberia represent savannah and forest onchocerciasis in West Africa, respectively. Whereas in Burkina Faso, a saturating relationship between microfilarial density and worm burden suggests the operation of density-dependent processes within human hosts, the Liberian data show a linear relationship implying no density dependence. The differences may derive from differences between both parasite strains, i.e. the savannah or the forest strain of O. volvulus. Consistently for both parasite strains and independent of the worm burden, the microfilarial density increases with host age emphasising the concept of the acquisition of immunological tolerance. In male hosts in Liberia, the microfilarial density increases stronger with the worm burden than in female hosts, whereas such sex-specific differences cannot be found in Burkina Faso. In the methodological part of this investigation, we suggest the beta-distribution to be most appropriate for describing variability in microfilarial densities and we present an approach to consider the uncertainty in the adult parasite burden which cannot be determined precisely in helminth infections. Implications of density dependence are discussed with respect to immunological processes in the human host and with respect to the success of control programs. The relationships described show that regulatory processes between the parasite and the human host are multi-dimensional, operating within a high degree of biological variability.     

The ecology of cutaneous leishmaniasis in Nizzana, Israel: infection patterns in the reservoir host, and epidemiological implications.

We conducted an extensive interdisciplinary study in an emerging focus of cutaneous leishmaniasis in the Western Negev Desert of Israel between July 1998 and February 2000. The aims of the this study were to determine (1) the reservoir hosts, (2) the distribution of the pathogen within the host range, (3) the associations of host, vector, and pathogen within defined habitats, (4) the demographic distribution of the pathogen within the host populations, and (5) to apply the newly acquired epizootiological data to explain morbidity patterns in humans. Fourteen square (60 m width) sampling plots were delimited in three types of habitats each with a different kind of substrate: loess, sand, and sand-loess ecotone. Rodents and sand flies were trapped and several environmental variables were measured. Leishmania infections in rodents were detected microscopically in stained smears of ear tissue and by a Leishmania-specific polymerase chain reaction. Results indicate that, contrary to previous reports, Psammomys obesus and not Meriones crassus is the main reservoir host in the region. Additional rodents (12 Gerbillus dasyurus and two M. crassus) were also found positive for Leishmania DNA. Prevalence of Leishmania infections amongst P. obesus was highest in loess habitats (65%), intermediate in the sandy-loess ecotone (20%), and 0% in the sandy habitats. Psammomys obesus individuals in the loess habitat of the Nizzana ruins were larger, on average (probably older), than those in the sandy habitat of the Mt. Keren junction. Sand fly density was positively correlated to soil moisture being higher in the relatively humid plots of Nizzana ruins and much lower in the drier sandy soil of Mt. Keren. Elucidation of fundamental ecological factors affecting this disease has helped explain an apparent discrepancy between the distribution of the disease in the zoonotic system and among humans.     

Echinococcus granulosus: variability of the host-protective EG95 vaccine antigen in G6 and G7 genotypic variants

Cystic hydatid disease in humans is caused by the zoonotic parasite Echinococcus granulosus. As an aid to control transmission of the parasite, a vaccine has been produced for prevention of infection in the parasite’s natural animal intermediate hosts. The vaccine utilizes the recombinant oncosphere protein, EG95. An investigation into the genetic variability of EG95 was undertaken in this study to assess potential antigenic variability in E. granulosus with respect to this host-protective protein. Gene-specific PCR conditions were first established to preferentially amplify the EG95 vaccine-encoding gene (designated eg95-1) from the E. granulosus genome that also contains several other EG95-related genes. The optimized PCR conditions were used to amplify eg95-1 from several parasite isolates in order to determine the protein-coding sequence of the gene. An identical eg95-1 gene was amplified from parasites showing a G1 or G2 genotype of E. granulosus. However, from isolates having a G6 or G7 genotype, a gene was amplified which had substantial nucleotide substitutions (encoding amino acid substitutions) compared with the eg95 gene family members. The amino acid substitutions of EG95 in the G6/G7 genotypes may affect the antigenicity/efficacy of the EG95 recombinant antigen against parasites of these genotypes. These findings indicate that characterization of eg95 gene family members in other strains/isolates of E. granulosus may provide valuable information about the potential for the EG95 hydatid vaccine to be effective against E. granulosus strains other than the G1 genotype.     

Response of two species of Indo-Pacific corals, Porites cylindrica and Stylophora pistillata, to short-term thermal stress: The host does matter in determining the tolerance of corals to bleaching

The role of both host and dinoflagellate symbionts was investigated in the response of reef-building corals to thermal stress in the light. Replicate coral nubbins of Stylophora pistillata and Porites cylindrica from the GBR were exposed to either 28 °C (control) or 32 °C for 5 days before being returned to an ambient reef temperature (28 °C). S. pistillata was found to contain either Symbiodinium genotype C1 or C8a, while P. cylindrica had type C15 based on ITS genotyping. Analysis of the quantum yield of photosystem (PS) II fluorescence of the symbionts in P. cylindrica showed that light-induced excitation pressure on the C15 Symbiodinium was significantly less, and the steady state quantum yield of PSII fluorescence at noon (ΔF/Fm′) greater, than that measured in C1/C8a Symbiodinium sp. from S. pistillata. Immunoblots of the PS II D1 protein were significantly lower in Symbiodinium from S. pistillata compared to those in P. cylindrica after exposure to thermal stress. The biochemical markers, heat-stress protein (HSP) 70 and superoxide dismutase (SOD), were significantly greater in P. cylindrica before the experiment, and both species of coral increased their biosynthesis of HSP 70 and SOD when exposed to thermal stress. Concentrations of MAAs, glycerol, and lipids were not significantly affected by thermal stress in these experiments, but DNA damage was greater in heat-stressed S. pistillata compared to P. cylindrica. There was minimal coral mucus, which accounts for up to half of the total energy budget of a coral and provides the first layer of defense for invading microbes, produced by S. pistillata after heat stress compared to P. cylindrica. It is concluded that P. cylindrica contains a heat resistant C15 Symbiodinium and critical host proteins are present at higher concentrations than observed for S. pistillata, the combination of which provides greater protection from bleaching conditions of high temperature in the light.     

Functional response and progeny production of the Madeira mealybug parasitoid, Anagyrus sp. nov. nr. sinope: The effects of host and parasitoid densities

We tested the hypothesis that control of an herbivorous pest would be improved by providing floral resources for adult natural enemies. The herbivore was euonymus scale, Unaspis euonymi (Comstock) (Homoptera: Diaspididae), a serious pest of woody ornamental plants. The experimental landscape consisted of 3 × 3 m plots, each containing a central bed of Euonymus fortunei (Turcz.) that was infested with the scale. Floral resource plants were cultivars of four species that overlapped in bloom periods to provide a continuous supply of floral resources during summer: Trifolium repens L., Euphorbia epithymoides L., Coreopsis verticillata L. var. ‘Moonbeam,’ and Solidago canadensis L. var. ‘Golden Baby.’ Plots contained either low or high densities of all four species, or no resource plants. Densities of euonymus scale were typically lower in plots containing resource plants than in plots without them. Parasitism by Encarsia citrina (Craw.) (Hymenoptera: Aphelinidae) was rarely influenced by the experimental treatments, flower biomass, whole-plant biomass, or scale density, but in some cases was inversely correlated with density of scales within a generation and in the subsequent generation. Parasitism occasionally reduced densities of scales in plots containing resource plants, but this effect apparently was related to vegetative, not floral qualities of plants. A steady increase in parasitism rate over the three-year course of the experiment across the entire landscape was associated with decreasing density of scales, suggesting a numerical response by the parasitoid population. These findings suggest that the parasitoid is capable of effectively controlling euonymus scale in ornamental landscapes where environmental conditions are favorable.     

The CCAAT-binding complex of eukaryotes: evolution of a second NLS in the HapB subunit of the filamentous fungus Aspergillus nidulans despite functional conservation at the molecular level between yeast, A.nidulans and human

The heterotrimeric CCAAT-binding complex is evolutionarily conserved in eukaryotic organisms, including fungi, plants and mammals. In the filamentous fungus Aspergillus nidulans, the corresponding complex was designated AnCF (A.nidulans CCAAT-binding factor). AnCF consists of the subunits HapB, HapC and HapE. All three subunits are necessary for DNA binding. HapB contains two putative nuclear localisation signal sequences (NLSs) designated NLS1 and NLS2. Previously, it was shown that only NLS2 was required for nuclear localisation of HapB. Furthermore, HapC and HapE are transported to the nucleus only in complex with HapB via a piggy back mechanism. Here, by using various GFP constructs and by establishing a novel marker gene for transformation of A.nidulans, i.e. the pabaA gene encoding p-aminobenzoic acid synthase, it was shown that the HapB homologous proteins of both Saccharomyces cerevisiae (Hap2p) and human (NF-YA) use an NLS homologous to HapB NLS1 for nuclear localisation in S.cerevisiae. Interestingly, for A.nidulans HapB, NLS1 was sufficient for nuclear localisation in S.cerevisiae. In A.nidulans, HapB NLS1 was also functional when present in a different protein context. However, in A.nidulans, both S.cerevisiae Hap2p and human NF-YA entered the nucleus only when HapB NLS2 was present in the respective proteins. In that case, both proteins Hap2p and NF-YA complemented, at least in part, the hap phenotype of A.nidulans with respect to lack of growth on acetamide. Similarly, A.nidulans HapB and human NF-YA complemented a hap2 mutant of S.cerevisiae. In summary, HapB, Hap2p and NF-YA are interchangeable. Because the A.nidulans hapB mutant was complemented, at least in part, by both the human NF-YA and S.cerevisiae Hap2p this finding suggests that the piggy-back mechanism of nuclear transport found for A.nidulans is conserved in yeast and human.     

Allopatric distribution of juvenile red-legged banana prawns (Penaeus indicus H. Milne Edwards, 1837) and juvenile white banana prawns (Penaeus merguiensis De Man, 1888), and inferred extensive migration, in the Joseph Bonaparte Gulf, northwest Australia

During October to December 1997, we trawled estuarine habitats in the Joseph Bonaparte Gulf (JBG) to determine the distribution of juvenile red-legged banana prawns, Penaeus indicus (H. Milne Edwards, 1837) and white banana prawns, Penaeus merguiensis (de Man, 1888). We made 229 beam-trawls at 185 sites, mostly over a 100-m path (3-min duration). A Global Positioning System (GPS) receiver was used to verify our location. During October to December 1998, we intensively resampled three of the rivers that were sampled in 1997 to confirm the gulf-wide distribution of P. indicus and P. merguiensis and to investigate the microhabitat use of P. indicus. We chose previously sampled and new sites in Forsyth Creek (eastern JBG), the Lyne River (Cambridge Gulf), and the Berkeley River (western JBG). We made 249 trawls at 21 sites, mostly over 100 m.Juvenile banana prawns were abundant in eastern JBG, Cambridge Gulf and western JBG. They were not abundant in southern JBG, although fewer trawls were made there, due to its inaccessibility. In eastern JBG and Cambridge Gulf, over 96% and 73% (respectively) of juvenile banana prawns were P. indicus and they were more abundant there than in the western JBG. Conversely, in the western JBG over 93% of the juvenile banana prawns were P. merguiensis and they were more abundant than in the eastern JBG and Cambridge Gulf. The Lyne River in the northwestern Cambridge Gulf seems to be the transition zone; both P. indicus and P. merguiensis are equally abundant.P. indicus are most abundant on the mangrove-lined muddy banks of waterways within mangrove forests, similar habitats to P. merguiensis. Within these habitats, they were most abundant in gutters and small creeks, rather than rivers and large creeks. Few P. indicus or P. merguiensis were caught in 100 m² trawls undertaken midriver (on the channel bottom and on emergent banks), although these habitats may be only 100 m from the mangrove-lined habitats. In all creek and river habitats, both species are most catchable at low tide (irrespective of daylight or darkness) when they move out of the mangrove forests and accumulate in the remnant water bodies.The offshore fishery for P. indicus is in northwestern JBG in waters 50-80 m deep, about 300 and 200 km, respectively, from where juveniles are abundant in their extensive inshore habitats in east JBG and in Cambridge Gulf, demonstrating a geographical separation of the juvenile and adult phases. Postlarval P. indicus, spawned offshore, must use tides and currents to travel south and east to reach nursery habitats. Emigrant subadults must migrate north and west, across relatively shallow inshore sand substrates (30-40 m deep) to reach their offshore habitats.     

The biflagellate spermatozoa of Colostethus marchesianus (Melin, 1941) (Anura, Dendrobatidae) from the type locality and of Colostethus sp. (aff. Marchesianus.) from a different locality: A scanning and transmission electron microscopy analysis

Scanning and transmission electron microscopy and fluorochrome staining with DAPI were used to study the spermatozoa of Colostethus marchesianus from the type locality and Colostethus sp. (aff. marchesianus) from a different locality. The sperm cell of these species resembles those of most neobatrachian species, except for the presence of two complete flagella, a character previously described only for four unrelated anuran species belonging to the families Leptodactylidae, Rhacophoridae and Dendrobatidae. Although very similar, the spermatozoa described here showed slight differences in the length of their nucleus, in the extent of the subacrosomal cone over the nucleus, and in the curvature of the axial fiber seen in cross section. The absence of a juxtaxonemal fiber and the presence of a comma shaped axial fiber seem to be common characteristics of dendrobatid spermatozoa. The minor differences found in our study may indicate that the taxa studied here are distinct species. The evolutionary significance of spermatozoa biflagellarity is still unclear, although the presence of this trait in unrelated groups suggests independent origins.     

The unusual canangafruticosides A-E: Five monoterpene glucosides, two monoterpenes and a monoterpene glucoside diester of the aryldihydronaphthalene lignan dicarboxylic acid from leaves of Cananga odorata var. fruticosa

From the leaves of Cananga odorata var. fruticosa, five unusual monoterpene glucosides, named canangafruticosides A-E (1-5), along with two unusual non-glucosidic monoterpenes (6, 7) were isolated. An aryldihydronaphthalene-type lignan dicarboxylate (8) was also isolated, with two moles of canangafruticoside A (1) on its ester moiety. This lignan also showed strong blue fluorescence emission under basic conditions. The structures of these compounds were elucidated by means of spectroscopic methods, with their absolute configurations determined by application of the modified Mosher’s method to a compound chemically derived from canangafruticoside E.