Ribeiroia marini: pathogenesis and larval trematode antagonism in the snail, Biomphalaria glabrata

Larval trematode antagonism between Ribeiroia marini and Schistosoma mansoni was studied in the snail Biomphalaria glabrata. A laboratory-raised Puerto Rican strain of B. glabrata was exposed to single and double infections with given numbers of: (1) embryonated eggs of R. marini from laboratory rats, and (2) miracidia of S. mansoni from mice. Snails were maintained in outside environmental tanks in San Juan, Puerto Rico and larval trematode interactions were examined in a series of five experiments. Snails of all sizes were highly susceptible to single infections with R. marini. Rediae and cercariae caused extensive damage to the digestive gland and ovotestis resulting in premature death of snails. Heavily infected snails were castrated and stopped laying eggs. Snails infected first with S. mansoni were only partly susceptible to superinfection with R. marini given on Day 23. In a reverse experiment, snails infected first with R. marini were only partly susceptible to a second infection with S. mansoni given on Day 23. In simultaneous exposures, snails developed double infections (22%) with R. marini dominant and S. mansoni sporocyst and cercaria production reduced. While R. marini is not a strong direct antagonist against established S. mansoni infections, it has several attributes as a possible biological control agent: hardy eggs easily produced in rats; high infectivity to snails of all ages; and ability to castrate and prematurely kill B. glabrata. The R. marini-rat system described here provides a convenient laboratory and field model for the study of intrasnail trematode antagonism and biological control.     

Schistosoma mansoni: lectin-dependent cytotoxicity of hemocytes from susceptible host snails, Biomphalaria glabrata

Normally benign hemocytes from a strain (M-line) of the snail, Biomphalaria glabrata, susceptible to Schistosoma mansoni, became cytotoxic toward the sporocyst stage if the parasite was first treated with the lectin, concanavalin A. Concanavalin A binding was inhibitable with alpha-methyl mannoside and killing was dose-dependent. Maximal levels of concanavalin A-induced cytotoxicity were comparable with levels observed when hemocytes from a resistant snail strain (13-16-R1) encountered untreated sporocysts. Induction of the cytotoxic response did not occur if hemocytes alone were pretreated with the lectin. A unique method incorporating ultraviolet microscopy and the vital fluorescent dye, eosin Y, was used for discriminating between live and dead sporocysts. This model may prove useful in understanding mechanisms used by invertebrate effector cells in recognition and killing of invading organisms.     

Heritability of sex tendency in a harpacticoid copepod,Tigriopus californicus

Abstract.— Systems with genetic variation for the primary sex ratio are important for testing sex-ratio theory and for understanding how this variation is maintained. Evidence is presented for heritable variation of the primary sex ratio in the harpacticoid copepod Tigriopus californicus. Variation in the primary sex ratio among families cannot be accounted for by Mendelian segregation of sex chromosomes. The covariance in sex phenotype between full-sibling clutches and between mothers and offspring suggests that this variation has a polygenic basis. Averaged over four replicates, the full-sibling heritability of sex tendency is 0.13 ± 0.040; and the mother-offspring heritability of sex tendency is 0.31 ± 0.216. Genetic correlations in the sex phenotype across two temperature treatments indicate large genotype-by-temperature interactions. Future experiments need to distinguish between zygotic, parental, or cytoplasmic mechanisms of sex determination in T. californicus.     

The spatial population structure of a harpacticoid copepod community in spring

The spatial population patterns of an assemblage of meiobenthic harpacticoid copepod species were analysed for two sets of samples resulting from a systematic survey of an intertidal, estuarine, sandy beach in spring. The fauna was characterised by mesopsammic species, the 6 most abundant of which showed a great degree of interassociation, such that they could not be considered to distribute themselves independently. Populations of each of the 6 species were overdispersed, such that a negative binomial distribution adequately fitted all population samples. Spatial segregation of closely related species was indicated during the late spring sample, so leading to a patchy population distribution. It is proposed that during early spring, during periods of extreme disturbance in the seasonal r-selecting environment, the species populations do not interact, but that during periods of less extreme perturbation an interactive community evolves.     

The morphology of a harpacticoid copepod gut: A review and synthesis

The digestive tract of a harpacticoid copepod, Tigriopus californicus (Baker), was studied by using techniques of light and electron microscopy. Four cell types could be distinguished: type 1, an embryonic cell which will replace cells worn away or lost during secretion; type 2, a cell which synthesizes and secretes proteins and also plays a role in lipid absorption; and types 3 and 4, two cell types which absorb lipids. From the abundance of each cell type, the length of microvilli, the development of basal plasma membrane (PM), and luminal projections, the following conclusions were made. (1) The midgut caecum absorbs digested nutrients. (2) The anterior midgut absorbs nutrients and more importantly functions in merocrine and exocrine secretion. The presence of concretions in cell types 2 and 3 in the anterior midgut suggests that these tissues contribute in excretion, and in water and/or ion regulation. (3) The posterior midgut absorbs nutrients and contributes some holocrine secretion.     

Selective attraction and reproductive performance of a harpacticoid copepod in a response to biofilms

The relationship between monobacterial films and the preference of harpacticoid copepods for such films was investigated using still water multiple-choice assays with natural biofilm and sterile conditions as controls. Adult Schizopera sp. were most attracted by a heterogeneous natural biofilm, followed by monospecies-biofilms of Rhodovulum sp., Vibrio proteolyticus, and Flexibacter sp. The preferred bacterial films stemmed from different phylogenetic and physiological groups. The results indicated that the harpacticoid Schizopera sp. was effectively and differentially attracted by bacterial films. Since bacteria constitute a substantial portion of the organic carbon available at the sea bottom as nutritive sources for harpacticoid copepods, we subsequently examined the influence of 9 bacterial strains and a natural biofilm as a nutrient source on the growth and reproductive performance of ontogenetic stages (nauplii and copepodids) of Schizopera sp. The food value of bacterial strains was assayed in terms of life table data that provided growth parameters. All variables were affected by the type of food offered. A diet on Rhodovulum sp. resulted in optimal growth performance of nauplii and copepodids demonstrating that bacteria can be used as a sole diet to support postembryonic development. The present study is the first to link behavioral preferences to bacterial biofilms with life history parameters when cultivating harpacticoid copepods on the same bacterial strains as the only diet. This study revealed a discrepancy between the biofilm favored (natural biofilm) and the one leading to maximal reproductive performance (monobacterial film of Rhodovulum sp. MB253) as indicated by major life table data as net reproductive rate (R_o), mean generation time (T_m), and capacity for increase (r_c).     

Bioassay evidence for the transmission of WSSV by the harpacticoid copepod Nitocra sp

White Spot Syndrome Virus (WSSV) is now one of the most devastating and virulent viral agents threatening the penaeid shrimp culture industry and has been responsible for serious economic losses for shrimp farms worldwide. One remarkable characteristic of WSSV is its wide reservoir range, which contributes to its wide geographical distribution. Among epizootiological surveys, there is substantial evidence for WSSV-positive copepods found in shrimp farming ponds. Therefore, copepods are suspected to be the vector of WSSV. In the present study, nested-PCR analysis showed positive results in the harpacticoid copepod Nitocra sp. exposed to WSSV by virus-phytoplankton adhesion route. Oral route and intramuscular injection were used to test the pathogenicity of WSSV isolated from the WSSV-positive Nitocra sp. For the oral route of infection, Marsupenaeus japonicus postlarvae were fed with WSSV-positive copepods. The shrimp postlarvae in the infected treatment became WSSV-positive and occurred 52.50+/-5.00% mortality which was significant higher (P <0.05) than that in the control treatment (20.00+/-0.00%) when postlarvae were fed with WSSV free copepods. In the intramuscular injection challenge, M. japonicus juveniles were injected with the copepods inoculum extracted from the WSSV-positive Nitocra sp., and showed 72.50+/-9.57% mortality which was also significant higher (P <0.05) than that in the control treatment (22.50+/-5.00%) when juveniles were received mock injection of a tissue homogenate prepared from WSSV-negative Nitocra sp. Based on these laboratory challenge studies, it was confirmed that the copepods can serve as a vector in WSSV transmission.     

The response of a harpacticoid copepod community to sediment disturbance in a semi-enclosed lagoon

The harpacticoid fauna of a sandy sediment in the Sluice Dock of Ostend, Belgium, was dominated by a single species, Canuella perplexa, that attained an average annual density of over 400 ind. 10 cm^–2 in 1976 and 1977, whereas other species had a total mean density of less than 10 ind. 10 cm^–2. In the summer of 1979 a dystrophic crisis with abundant growth of Ulva occurred and in March 1980 the lagoon was drained. After refilling of the lagoon, the sediment was immediately colonized by good swimming harpacticoids, that attained densities of 200 ind. 10 cm^–2. Three months after refilling Canuella perplexa reappeared and the system returned to its original state within one month.     

Resistance to schistosome infection in Biomphalaria glabrata induced by gamma radiation

Two strains of Biomphalaria glabrata were studied with respect to the effects of ionizing radiation on their susceptibility to Schistosoma mansoni infection. Gamma radiation at levels of 3.5 and 5 krad did not induce susceptibility in the resistant S-3 strain, but was found to initiate resistance in the susceptible PR-1 strain. In an attempt to understand the induced resistance in irradiated snails, histopathologic examinations and analyses of snail hemolymph were performed. Results indicated that miracidia invading irradiated snails were quickly surrounded and encapsulated by amoebocytes. Similarly, alterations in the hemolymph of irradiated snails suggested that radiation induced aging. It is suggested that radiation-altered snails may be of value in studying the defense mechanisms of these organisms.     

Autecology of an intertidal Alaskan harpacticoid copepod Apolethon sp.

The life history and ecology of a harpacticoid copepod, Apolethon sp., were investigated for one year in the intertidal zone of a subarctic bay in southeastern Alaska. Core samples were collected biweekly at the 0 m tidal level from March, 1992 through March, 1993; zonation and vertical distribution were determined from additional samples. Large seasonal variations in density of Apolethon sp. parallelled that of the total harpacticoid community, with highest densities (124.3±12.5 10 cm^–2; mean ± standard error) in March and lowest densities in May and October (7.6±4.5). The sex ratio of Apolethon sp. varied directly with population density, varying from <0.1 to 1.2 (male:female) during the study period. The species was restricted to a relatively narrow vertical band within the intertidal zone, having its greatest density at – 1 m and being rare above + 1 m tidal height. Adult Apolethon sp. bear conspicuous ovoid mucin structures located laterally on the genital double-somite. The mucin structures are unique to the species; several putative functions are discussed.     

Prey selection by molluscivorous cichlids foraging on a schistosomiasis vector snail,Biomphalaria glabrata

Summary This paper considers prey size selection by four molluscivorous cichlids feeding on the intermediate host snail of Schistosoma parasites, Biomphalaria glabrata. Haplochromis ishmaeli obtains its prey by crushing the snails between the pharyngeal jaws, whereas H. xenognathus, H. sauvagei and Macropleurodus bicolor apply both pharyngeal crushing and oral shelling. The fishes crushed significantly more snails with the highest reward in biomass per second of crushing. Oral shelling occurred far less often than pharyngeal crushing. Encounter rates with prey showed significant variations between different size classes of prey. The fish have no overall knowledge of snail availability in a tank. The probability that a snail will be eaten at encounter, calculated from the number encountered and the number eaten, reflects the prey size preference of the fish. Those snails with the highest biomass/crushing-time ratio had the highest probability of being crushed; observed and predicted prey size preferences corresponded well. Although for oral shelling the potential reward in biomass per second is of the same magnitude as for crushing, the probability of successful shelling is very low. Apparently the fish prefer prey with lowest risks.     

Population genetic structure of Biomphalaria glabrata in a schistosomiasis-endemic region in Brazil

The population genetic structure of the freshwater snail, Biomphalariaglabrata, was assessed in the municipality of Virgem das Graças(Minas Gerais, Brazil), a schistosomiasis-endemic area. Theseven microsatellite loci that were used to genotype 326 individualsfrom 17 collection sites within a 3-km radius revealed highgenetic variability. However, we found a deficit of heterozygotesrelative to Hardy–Weinberg expectations for most of theloci and in many collection sites, indicating a high level ofinbreeding. This may be the result of founder effects and sib-mating.A high level of overall population genetic differentiation amongB. glabrata collection sites suggested low gene flow at thisrelatively small spatial scale in this region. However, subsequentanalyses also indicated that these patterns may also resultfrom founder events and that an exchange of individuals betweenparticular collection sites does exist. The isolation-by-distancecorrelation between collection sites of B. glabrata was foundto be significant. However, only approximately 6% of the variancein F_ST was explained by geographic distances suggesting thatthere are other factors affecting genetic differentiation amongthese collection sites. It is likely that the current populationgenetic structure reflects patterns of both migration and founderevents. Results from this study are important for understandingthe potential for local adaptation between schistosomes andtheir snail hosts at this epidemiologically relevant scale.     

Decoupling of molecular and morphological evolution in deep lineages of a meiobenthic harpacticoid copepod

Molecular and biochemical genetic analyses have revealed that many marine invertebrate taxa, including some well-studied and presumably cosmopolitan species, are actually complexes of sibling species. When morphological differences are slight and estimated divergence times are old, data suggest either unusually high rates of sequence evolution or long-term morphological stasis. Here, five gene regions (mitochondrial cytochrome oxidase subunit I and large-subunit ribosomal 16S rDNA and nuclear ITS1, 5.8S rDNA, and ITS2) were analyzed in four geographic samples of the meiobenthic harpacticoid copepod Cletocamptus deitersi. Molecular sequences revealed four extremely differentiated molecular lineages with unalignable nuclear intergenic spacers and mitochondrial uncorrected divergences reaching 25% (cytochrome oxidase) and 36% (16S rDNA). These levels of divergence are greater than those reported previously for congeneric species in diverse invertebrate taxa, including crustaceans. The nominally intraspecific divergence matches or exceeds the corresponding divergence from a known congener (Cletocamptus helobius). A molecular clock applied to the cytochrome oxidase subunit I data suggests that these lineages split in the Miocene, consistent with the fossil record of a North American Cletocamptus from the same period. Morphological differences among the major lineages are subtle but congruent with the patterns of genetic differentiation. Our conclusion, based on concordant patterns of variation in two mitochondrial and three nuclear gene regions, as well as morphological observations, is that C. deitersi in North America is composed of at least four separate species by the genealogical concordance, phylogenetic, and morphological-species criteria. Alternative explanations for the deep phylogenetic nodes and apparent morphological stasis, including high rates of sequence evolution, balancing selection, and genetic signatures of historical events, are considered unlikely.     

Insights from the shotgun-based proteomics analysis of Biomphalaria glabrata

Biomphalaria glabrata is an important host in the transmission of human schistosomiasis in the Caribbean and South America. Therefore, it is of interest to analyse the proteome data of Biomphalaria glabrata hemolymph to identify immunity related proteins in host-pathogen relationship. We used shotgun proteomic and bioinformatic analyses of the non-depleted and depleted [0.5 and 0.75% Trifluoroacetic acid (TFA) depletion] hemolymph of B. glabrata (LE strain). Analysis showed 148 proteins from the hemolymph. 148 were obtained from the 0.5% TFA-depleted sample. 62 proteins follow this from the 0.75% TFA-depleted sample. However, only 59 were found from non-depleted hemolymph. A number of proteins were identified from the hemolymph of this schistosomiasis snail vector linked to immunity related functions. This provides insights to the understanding of schistosome-snail interaction.