Tegumentary papillae of Echinostoma caproni cercariae (Trematoda: Echinostomatidae)

Argentophilic staining and scanning electron microscopy were used to study the tegumentary papillae of Echinostoma caproni cercariae. The most abundant tegumentary papillae were uniciliate, but multiciliate papillae were also found, mainly on the ventral aspect of the oral collar. The distribution pattern of the papillae on the body and tail was in general similar to that seen in the cercariae of other 37-collar-spined Echinostoma species. Some differences were noted between E. caproni and the allopatric species, E. trivolvis. E. caproni has a greater number of papillae associated with the collar spines than does E. trivolvis. E. caproni has uniciliate papillae on the acetabulum, whereas E. trivolvis does not. Chaetotaxy is useful to distinguish subtle morphological differences in cercarial species in the 37-collar-spined Echinostoma complex.     

Cercarial chaetotaxy of Echinostoma revolutum (Froelich, 1802) and E. echinatum (Zeder, 1803) (Trematoda, Echinostomatidae)

The life-cycles of Echinostoma revolutum and E. echinatum were carried out by using cercariae emitted by naturally infected Molluscs. E. revolutum strain was issued from Lymnaea auricularia, E. echinatum from Lymnaea truncatula and Planorbis planorbis, Planorbarius corneus produced an Echinostome which may possibly be E. echinatum or another species E. sp. Metacercarial stages and adults were obtained from laboratory experimental hosts. None of these adult Echinostomes displayed well-defined morphological differences: nevertheless their respective larval stages exhibited discrepancies used for species diagnosis. Cercarial chaetotaxy is given for every batch and compared with that of other species described as E. audyi, E. lindoense and E. caproni; discriminating features are discussed.     

Biochemical profile of Biomphalaria glabrata (Mollusca: Gastropoda) after infection by Echinostoma paraensei (Trematoda: Echinostomatidae)

The effect of infection by Echinostoma paraensei on the activity of the enzymes alanine aminotransferase (ALT) and aspartate aminotransferase (AST) and the concentration of total proteins, uric acid and urea in the hemolymph of Biomphalaria glabrata were investigated after exposure to five or 50 miracidia. The biochemical concentrations were measured weekly until the end of the fourth week after exposure. There was a significant decrease in the concentrations of total proteins in the snails exposed both to five and 50 miracidia, as well as an increase in the nitrogenous products of excretion, ALT and AST activities. The higher ALT activity in the hemolymph of the snails after infection with 50 miracidia suggests highest energetic requirement in these snails in relation to snails exposed to five miracidia. The results also suggest an increase in the use of total proteins, since there was increased formation of nitrogenous catabolites, in conformity with an increase in the aminotransferase activities, frequently associated with tissue damages. This can be explained by damage due to penetration by the miracidia and subsequent development of intramolluscan sporocysts and rediae.     

The comparative development of Echinostoma caproni (Trematoda: Echinostomatidae) adults in experimentally infected hamsters and rats

The aim of the present study was to compare the development of Echinostoma caproni (Trematoda: Echinostomatidae) adults in two host species displaying different degrees of compatibility with this parasite. For this purpose, the variability in the worm recovery, egg output, and morphology of E. caproni adults during the course of experimental infections in hamsters and rats was analyzed. Students t-tests and two-factor ANOVA analysis with the time post-infection and the host species as independent variables, and Bonferroni t-tests as post hoc analysis were used for the study. Worm recovery and egg output were host species dependent. The values were significantly higher in the worms established in hamsters than those in rats. The oral sucker area, pre-pharynx length, and pharynx area were the most conservative features, and no significant variability related to the host species was detected. In contrast, body area, collar width, esophagus length, cirrus sac area, ventral sucker area, ovarian area and anterior and posterior testicular areas were significantly higher in those worms collected from hamsters. Moreover, significant worm age-host species interactions were found for body area, ovarian area, ventral sucker area, and anterior and posterior testicular areas.     

Molecular characterization of Gastrodiscoides hominis (Platyhelminthes: Trematoda: Digenea) inferred from ITS rDNA sequence analysis

Gastrodiscoides hominis (Digenea: Paramphistomata: Gastrodiscidae) is an amphistomid intestinal fluke of pigs causing gastrodiscoidiosis. With the use of molecular tools assisting the conventional diagnostic procedures, we aimed at finding out molecular characterization of G. hominis using PCR amplifications of rDNA ITS (1, 2) sequences. The sequences obtained (GenBank accession numbers EF027096, EF027097, EF027098, EU887294, and EU887295) were compared with available sequences of other digenean parasites, particularly those having a zoonotic potential in the northeastern region of India. The BLAST search revealed a close similarity with members of the family Paramphistomidae, showing maximum similarity with the amphistome, Homalogaster paloniae (subfamily Paramphistominae). Based on various tree construction methods, phylogeny of G. hominis is discussed.     

Ultrastructure of the Miracidium of Echinostoma paraensei Lie and Basch, 1967 (Trematoda, Echinostomatidae)

The morphology of Echinostoma paraensei was studied using transmission electron microscopy. The terebratorium region has many electrondense secretory granules and many folds on the surface. The epidermal cells that cover the larval body have unique nuclear morphology, many mitochondria and vesicles being attached to the interepidermal ridges by a septate junction. The cilia present the organization 9+2 and a typical structure with a shaft, axosome, basal body and rootlet. Below the epidermal cells there is a layer of circular muscle and, adjacent to it, a layer of longitudinal muscle fibers. The excretory system has two flame cells, with internal and external ribs and leptotriches at the barrel region, an excretory vesicle and an excretory pore.     

Comparative ultrastructure of eggs in Echinostoma paraensei, E. caproni, and E. trivolvis (Trematoda: Echinostomatidae)

The 37-collar-spined echinostomes Echinostoma paraensei, E. caproni, and E. trivolvis are digeneans that live in the intestine of small mammals and birds. Comparative studies of the eggs of these species were done using light microscopy (LM), scanning electron microscopy (SEM), and transmission electron microscopy (TEM). The egg of E. caproni was the largest of the three species studied, whereas the egg of E. trivolvis was the smallest in both length and width. The SEM study showed differences in the aboperculum region of the eggs in all three species. The TEM study showed that the eggshell of all three species consisted of three layers, but no difference in eggshell structure was observed in any species. Received: 10 September 1999 / Accepted: 9 November 1999     

Taxonomy of 37-collar spined Echinostoma (Trematoda: Echinostomatidae) in studies on the population regulation in experimental rodent hosts

With reference to a recent taxonomic revision the species of the echinostomes used in key studies on the population regulation in infections with 37-collar-spined Echinostoma species in experimental rodent hosts were reconsidered. This was considered essential to prevent taxonomic problems blocking further fruitful progress within this field of experimental parasitology.     

Mayaro virus: complete nucleotide sequence and phylogenetic relationships with other alphaviruses

Mayaro (MAY) virus is a member of the genus Alphavirus in the family Togaviridae. Alphaviruses are distributed throughout the world and cause a wide range of diseases in humans and animals. Here, we determined the complete nucleotide sequence of MAY from a viral strain isolated from a French Guianese patient. The deduced MAY genome was 11,429 nucleotides in length, excluding the 5' cap nucleotide and 3' poly(A) tail. Nucleotide and amino acid homologies, as well as phylogenetic analyses of the obtained sequence confirmed that MAY is not a recombinant virus and belongs to the Semliki Forest complex according to the antigenic complex classification. Furthermore, analyses based on the E1 region revealed that MAY is closely related to Una virus, the only other South American virus clustering with the Old World viruses. On the basis of our results and of the alphaviruses diversity and pathogenicity, we suggest that alphaviruses may have an Old World origin.     

Biological aspects of a new isolate of Echinostoma paraensei (Trematoda: Echinostomatidae): susceptibility of sympatric snails and the natural vertebrate host

A new isolate of Echinostoma paraensei (Lie & Basch, 1967) was obtained from a natural vertebrate host, the water rat Nectomys squamipes. Relationships with sympatric snails, Biomphalaria glabrata; Physa marmorata and Lymnaea columella, as well as allopatric snails and the definitive hosts N. squamipes, Mus musculus, Rattus norvegicus, Mesocricetus auratus and birds were investigated. E. paraensei developed in all sympatric snail species following exposure to one or five miracidia. P. marmorata showed the highest infection rate (34% and 36%) followed by L. columella (18% and 24%) and B. glabrata (8% and 28%), respectively. Mortality was elevated in the patent period, with L. columella having the highest values. B. glabrata, P. marmorata, and L. columella did not differ in relation to cercarial encystment, and metacercariae from each species were able to infect N. squamipes with similar worm burdens. Recovered adult worms were found in clusters from the first to third sections of the small intestine. Miracidia of E. paraensei hatched after 10 days incubation in darkness in dechlorinated water at 28+/-1 micro C. Miracidia began to hatch 1 h after exposure to incandescent light.     

Comparative ultrastructure study of lamellar gastrodermal projections in Echinostoma paraensei, E. caproni, and E. trivolvis (Trematoda: Echinostomatidae)

A comparative ultrastructure study using transmission electron microscopy (TEM) of ultrathin sections, freeze-fracture replication, and scanning electron microscopy (SEM) was conducted on the gastrodermal lamellar microvilli in adults of Echinostoma paraensei, E. caproni, and E. trivolvis. Lamellar projections observed by SEM were mainly planiform and rhomboidal or paddle-shaped, and they were commonly seen in the three species. The distal margins of these lamellae were mainly smooth in E. paraensei, whereas filiform or digitiform extensions in the margins occurred occasionally in E. caproni and frequently in E. trivolvis. Freeze-fracture TEM elucidated significant interspecific differences in the numbers of intramembranous particles (IMP) per square micrometer; the numbers of IMP were significantly lower in E. trivolvis than in either E. caproni or E. paraensei. Thus, ultrastructural differences in the gastrodermal lamellae of adult echinostomes will be a taxonomically useful criterion for distinguishing species of Echinostoma. Received: 8 March 1999 / Accepted: 25 March 1999     

Molecular characterization and phylogenetic relationships among microsporidian isolates infecting silkworm, Bombyx mori using small subunit rRNA (SSU-rRNA) gene sequence analysis

The life cycle, spore morphology, pathogenicity, tissue specificity, mode of transmission and small subunit rRNA (SSU-rRNA) gene sequence analysis of the five new microsporidian isolates viz., NIWB-11bp, NIWB-12n, NIWB-13md, NIWB-14b and NIWB-15mb identified from the silkworm, Bombyx mori have been studied along with type species, NIK-1s_mys. The life cycle of the microsporidians identified exhibited the sequential developmental cycles that are similar to the general developmental cycle of the genus, Nosema. The spores showed considerable variations in their shape, length and width. The pathogenicity observed was dose-dependent and differed from each of the microsporidian isolates; the NIWB-15mb was found to be more virulent than other isolates. All of the microsporidians were found to infect most of the tissues examined and showed gonadal infection and transovarial transmission in the infected silkworms. SSU-rRNA sequence based phylogenetic tree placed NIWB-14b, NIWB-12n and NIWB-11bp in a separate branch along with other Nosema species and Nosema bombycis; while NIWB-15mb and NIWB-13md together formed another cluster along with other Nosema species. NIK-1s_mys revealed a signature sequence similar to standard type species, N. bombycis, indicating that NIK-1s_mys is similar to N. bombycis. Based on phylogenetic relationships, branch length information based on genetic distance and nucleotide differences, we conclude that the microsporidian isolates identified are distinctly different from the other known species and belonging to the genus, Nosema. This SSU-rRNA gene sequence analysis method is found to be more useful approach in detecting different and closely related microsporidians of this economically important domestic insect.     

Effect of Echinostoma friedi (Trematoda: Echinostomatidae) experimental infection on longevity, growth and fecundity of juvenile Radix peregra (Gastropoda: Lymnaeidae) and Biomphalaria glabrata (Gastropoda: Planorbidae) snails

The effect of Echinostoma friedi experimental infection on longevity, growth and fecundity of two susceptible first intermediate host snails, Radix peregra and Biomphalaria glabrata, was studied to contrast the level of compatibility. 120 R. peregra and 150 B. glabrata snails were used exposed to one, three or five miracidia and divided in three categories: INF (snails exposed and infected); ENI (exposed but not infected) and C (control or not miracidial-exposed snails). R. peregra INF snails’ death process starts sooner, but in a prolonged extension, while B. glabrata INF snails have a much shorter life span. The infection and the miracidial exposure are able to reduce R. peregra normal development (stunting). B. glabrata INF snails’ growth exceeds that of C snails (gigantism). E. friedi produces a total parasitic castration of R. peregra and B. glabrata INF snails. R. peregra would be considered as the required snail host, while B. glabrata only as an adequate snail host.     

Genetic differentiation of Artyfechinostomum malayanum and A. sufrartyfex (Trematoda: Echinostomatidae) based on internal transcribed spacer sequences

Genetic differentiation between two synonymous echinostomes species, Artyfechinostomum malayanum and Artyfechinostomum sufrartyfex was determined by using the first and second internal transcribed spacers (ITS1 and ITS2), the non-coding region of rDNA as genetic makers. Of the 699 bp of combined ITS1 and ITS2 sequences examined, 18 variable nucleotide positions (2.58 %) were observed. Of these, 17 positions could be used as diagnostic position between these two sibling species, whereas the other one variation was intraspecific variation of A. malayanum. A clade of A. malayanum was closely aligned with A. sufrartyfex and clearly distance from the cluster of other echinostomes. Our results may sufficiently suggest that the current synonymy of these species is not valid.     

The redial population of paryphostomum segregatum (Trematoda: Echinostomatidae) in the snail Biomphalaria glabrata

Summary Redial populations of Paryphostomum segregatum were examined in Biomphalaria glabrata. In mature infections the number of rediae was proportional to the size of the snail host. The maximum size of the redial population in snails of similar size remained approximately the same whether the snails were exposed to 1, 10 or 100 miracidia each.

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Zusammenfassung Redienpopulationen von Paryphostomum segregatum in Biomphalaria glabrata wurden untersucht. Lagen reife Redien vor, war deren Anzahl proportional der Größe der Schnecken. In den Schnecken ähnlicher Größe blieb die Größe der Redienpopulation annähernd gleich, ohne Rücksicht darauf, ob die Schnecken jeweils 1, 10 oder 100 Miracidien ausgesetzt waren.

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This study was supported by the University of California International Center for Medical Research and Training (UC ICMRT, Hooper Foundation, San Francisco School of Medicine) with Research Grants TW 00144 from the Office of International Research and AI 07054 from the Institute of Allergy and Infectious Diseases, NIH, U.S. Public Health Service; and by Rockefeller Foundation Grant GA-MNS-6654.

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Hooper Foundation and graduate student, Department of Pathology, University of California, San Francisco.     

Egg laying in vitro of Echinostoma caproni (Trematoda) in nutritive and nonnutritive media

 Egg laying in vitro was studied in Echinostoma caproni adults placed in 10 ml of nutritive or nonnutritive media for 48 h in petri-dish cultures maintained at 37°C in an atmosphere containing 7.6% CO₂. Maximal egg laying occurred within 24 h in the defined medium RPMI 1640. Egg laying was significantly greater in this medium than in McCoy’s or Locke’s solution. Eggs released into the RPMI medium were capable of producing miracidia that were infective to Biomphalaria glabrata snails. Received: 16 October 1995 / Accepted: 4 January 1996     

Isolation of bluetongue virus serotype 1 (BTV-1) from goats and its phylogenetic relationship to other BTV-1 isolates worldwide based on full-length sequence of genome segment-2

Eight bluetongue viruses (BTV) were isolated in BHK-21 cell culture from blood of goats suffering from peste des petits ruminants. These viruses were identified as BTV serotype 1 (BTV-1) by RT-PCR using VP2-gene-based primers coupled with sequencing of the PCR products. All of the isolates showed similar genome migration profile in 8% polyacrylamide gel electrophoresis. The genome segment-2 (seg-2) of one isolate (MKD18/India/2008) was amplified piecemeal by overlapping PCR, and the products were sequenced to obtain full-length seg-2. Phylogenetic analysis based on the seg-2 sequence revealed that MKD18 is closely related to Australian BTV-1 isolates, with 86.3–86.8% nucleotide identity. Phylogenetic analysis based on the partial sequence of seg-2 (541 bp, nucleotides 1,304–1,844) showed that the Indian BTV-1 isolates, namely, MKD18, Avikanagar, Sirsa-3 and Chennai, are very closely related to each other, with more than 99.6% nucleotide identity. Although a high degree of similarity exists, the Indian BTV-1 isolates collected over the past 25 years should be studied to demonstrate the co-existence of different VP2 antigenic profiles.