Evolutionary relationships within 'pygmaeus' group microphallids using genetic analysis and scanning electron microscopy

There are four species of 'pygmaeus' microphallids, namely Microphallus pygmaeus, M. piriformes, M. pseudopygmaeus and M. triangulatus (Trematoda: Microphallidae) which are parasites of marine birds and their sporocysts give rise to transmissible metacercariae inside littoral gastropods (mostly littorines). Universally primed polymerase chain reaction (UP-PCR) showed no apparent pattern between genetic diversity of the metacercariae as estimated by genomic banding profiles and their geographic region or molluscan host species. At the same time UP-PCR product cross-hybridization showed that M. pseudopygmaeus and M. triangulatus are genetically very similar, indicating that these taxa represent one species complex. In contrast, M. pygmaeus and M. piriformes are genetically well separated from each other and also from the pseudopygmaeus-triangulatus complex. Scanning electron microscopy of ventral spines, and analyses of spine angles and the number of teeth per spine, showed that all species differed significantly from one another. It was concluded that M. piriformes represents the original western member of the 'pygmaeus' group. Microphallus pygmaeus probably diverged from M. piriformes as it progressively specialized for sea duck final hosts. Microphallus pseudopygmaeus and M. triangulatus diverged from each other and the piriformes-pygmaeus ancestral line relatively recently. Microphallus pseudopygmaeus specialized for adoption of a wide range of gastropod host species and M. triangulatus developed morpho-functional specialization associated with final host exploitation.     

Larval helminths in intermediate hosts: does competition early in life determine the fitness of adult parasites?

Density-dependent effects on parasite fitness have been documented from adult helminths in their definitive hosts. There have, however, been no studies on the cost of sharing an intermediate host with other parasites in terms of reduced adult parasite fecundity. Even if larval parasites suffer a reduction in size, caused by crowding, virtually nothing is known about longer-lasting effects after transmission to the definitive host. This study is the first to use in vitro cultivation with feeding of adult trematodes to investigate how numbers of parasites in the intermediate host affect the size and fecundity of adult parasites. For this purpose, we examined two different infracommunities of parasites in crustacean hosts. Firstly, we used experimental infections of Maritrema novaezealandensis in the amphipod, Paracalliope novizealandiae, to investigate potential density-dependent effects in single-species infections. Secondly, we used the crab, Macrophthalmus hirtipes (Ocypodidae), naturally infected by the trematodes, M. novaezealandensis and Levinseniella sp., the acanthocephalan, Profilicollis spp., and an acuariid nematode. These four helminths all develop and grow in their crustacean host before transmission to their bird definitive host by predation. In experimental infections, we found an intensity-dependent establishment success, with a decrease in the success rate of cercariae developing into infective metacercariae with an increasing dose of cercariae applied to each amphipod. In natural infections, we found that M. novaezealandensis-metacercariae achieved a smaller volume, on average, when infrapopulations of this parasite were large. Small metacercariae produced small in vitro-adult worms, which in turn produced fewer eggs. Crowding effects in the intermediate host thus were expressed at the adult stage in spite of the worms being cultured in a nutrient-rich medium. Furthermore, excystment success and egg-production in M. novaezealandensis in naturally infected crabs were influenced by the number of co-occurring Profilicollis cystacanths, indicating interspecific interactions between the two species. Our results thus indicate that the infracommunity of larval helminths in their intermediate host is interactive and that any density-dependent effect in the intermediate host may have lasting effects on individual parasite fitness.     

Intra- and interspecific competition among helminth parasites: effects on Coitocaecum parvum life history strategy, size and fecundity

Larval helminths often share intermediate hosts with other individuals of the same or different species. Competition for resources and/or conflicts over transmission routes are likely to influence both the association patterns between species and the life history strategies of each individual. Parasites sharing common intermediate hosts may have evolved ways to avoid or associate with other species depending on their definitive host. If not, individual parasites could develop alternative life history strategies in response to association with particular species. Three sympatric species of helminths exploit the amphipod Paracalliope fluviatilis as an intermediate host in New Zealand: the acanthocephalan Acanthocephalus galaxii, the trematode Microphallus sp. and the progenetic trematode Coitocaecum parvum. Adult A. galaxii and C. parvum are both fish parasites whereas Microphallus sp. infects birds. We found no association, either positive or negative, among the three parasite species. The effects of intra- and interspecific interactions were also measured in the trematode C. parvum. Both intra- and interspecific competition seemed to affect both the life history strategy and the size and fecundity of C. parvum. Firstly, the proportion of progenesis was higher in metacercariae sharing their host with Microphallus sp., the bird parasite, than in any other situation. Second, the intensity of intraspecific competition apparently constrained the ability of metacercariae to adopt progenesis and limited both the growth and egg production of progenetic individuals. These results show that the life history strategy adopted by a parasite may be influenced by other parasites sharing the same host.     

Host partitioning by parasites in an intertidal crustacean community

Patterns of host use by parasites throughout a guild community of intermediate hosts can depend on several biological and ecological factors, including physiology, morphology, immunology, and behavior. We looked at parasite transmission in the intertidal crustacean community of Lower Portobello Bay, Dunedin, New Zealand, with the intent of: (1) mapping the flow of parasites throughout the major crustacean species, (2) identifying hosts that play the most important transmission role for each parasite, and (3) assessing the impact of parasitism on host populations. The most prevalent parasites found in 14 species of crustaceans (635 specimens) examined were the trematodes Maritrema novaezealandensis and Microphallus sp., the acanthocephalans Profilicollis spp., the nematode Ascarophis sp., and an acuariid nematode. Decapods were compatible hosts for M. novaezealandensis, while other crustaceans demonstrated lower host suitability as shown by high levels of melanized and immature parasite stages. Carapace thickness, gill morphology, and breathing style may contribute to the differential infection success of M. novaezealandensis and Microphallus sp. in the decapod species. Parasite-induced host mortality appears likely with M. novaezealandensis in the crabs Austrohelice crassa, Halicarcinus varius, Hemigrapsus sexdentatus, and Macrophthalmus hirtipes, and also with Microphallus sp. in A. crassa. Overall, the different parasite species make different use of available crustacean intermediate hosts and possibly contribute to intertidal community structure.     

Hitch-hiking parasite: a dark horse may be the real rider

Many parasites engaged in complex life cycles manipulate their hosts in a way that facilitates transmission between hosts. Recently, a new category of parasites (hitch-hikers) has been identified that seem to exploit the manipulating effort of other parasites with similar life cycle by preferentially infecting hosts already manipulated. Thomas et al. (Evolution 51 (1997) 1316) showed that the digenean trematodes Microphallus papillorobustus (the manipulator) and Maritrema subdolum (the hitch-hiker) were positively associated in field samples of gammarid amphipods (the intermediate host), and that the behaviour of Maritrema subdolum rendered it more likely to infect manipulated amphipods than those uninfected by M. papillorobustus. Here I provide experimental evidence demonstrating that M. subdolum is unlikely to be a hitch-hiker in the mentioned system, whereas the lucky candidate rather is the closely related but little known species, Microphallidae sp. no. 15 (Parassitologia 22 (1980) 1). As opposed to the latter species, Maritrema subdolum does not express the appropriate cercarial behaviour for hitch-hiking.     

Redescription and life cycle of the monorchiid Postmonorcheides maclovini Szidat, 1950 (Digenea) from the Southwestern Atlantic Ocean: Morphological and molecular data

The adult monorchiid, Postmonorcheides maclovini Szidat, 1950, digenean parasite of the Patagonian blennie Eleginops maclovinus (Cuvier) (Eleginopidae) from Puerto Deseado (47° 45′ S, 65° 55′ W), Argentina, was characterized and its life cycle elucidated. P. maclovinus is the only species of the genus Postmonorcheides, proposed by Szidat (1950) from Tierra del Fuego province (~ 54° S), Argentina. This digenean uses the Patagonian blennie as definitive host, and the intertidal bivalve Lasaea adansoni (Gmelin) (Lasaeidae) as both first and second intermediate hosts (metacercariae encyst inside sporocysts), being the first record of this clam as intermediate host of trematode parasites. The cercaria may, in addition to encysting in the sporocyst, emerge and presumably infect other intermediate hosts. This is the second report of a monorchiid species with metacercariae encysting inside the sporocyst. Adults were found parasitizing the fish stomach, pyloric caeca and intestine with a prevalence of 100%; sporocysts with cercariae and/or metacercariae were found parasitizing the gonad of the bivalve with a prevalence of 2.78%. The cercariae possess a well-developed tail and eye-spots are absent. The ITS1 sequence from the adult digeneans found in the Patagonian blennie, identified as P. maclovini, was found to be identical to the ITS1 sequences obtained both from sporocysts containing cercariae and encysted metacercariae found in L. adansoni.     

滨鹬马蹄吸虫(复殖目:微茎科)的尾蚴和囊蚴期的季节动态

对科威特湾微茎科滨鹬马蹄吸虫幼虫期的中间宿主双带盾桑椹螺(Clypeomorus bifasciata)及小相手蟹(Nanosesarma minutum)的季节动态进行了研究。调查期超过一年,在检查的1 600只螺和415只蟹中, 11.8 %的螺感染了8种马蹄属线虫中的一种,且以滨鹬马蹄吸虫的感染占优势(9.9 %螺感染) ; 80 %的蟹感染滨鹬马蹄吸虫囊蚴。虽然一年四季两种宿主都会感染,但吸虫的流行和尾蚴(指成熟期感染)在夏季呈现高蜂。从螺体排出的尾蚴具有明显季节性,在此海湾必须要超过最低温度20℃。总的感染率在较大(较老)的螺里有所下降,显示吸虫影响宿主生存并随之影响宿主群体结构。囊蚴的感染丰度与蟹的个体大小有明显相关性;较大的蟹感染较多的囊蚴,显示宿主能耐受更多的吸虫。调查显示,囊蚴的感染率与蟹的大小或性别无相关性。囊蚴体外脱囊以及产卵吸虫的释放证明,成熟虫体终年存在于所有大小和性别不同的蟹里,显示从蟹到鸟的持续感染是可能的。总的来说,滨鹬马蹄吸虫在海湾的传播动态是由这两种无脊椎动物宿主来协调,并似乎是被一系列依赖于温度的活动控制,这些活动影响易感宿主种群及感染性幼虫期尾蚴和囊蚴的存在。     

A review of the currently recognised opecoelid cercariae,including the identification and emergence ecology of Cercaria capricornia XII (Digenea: Opecoelidae) from Nassarius olivaceus (Gastropoda: Nassariidae) in Central Queensland,Australia

A new opecoelid cercaria, Cercaria capricornia XII, is reported from Nassarius olivaceus in Capricornia, Central Queensland, Australia. Combined molecular and morphological data indicate that this cercaria is a member of the subfamily Opecoelinae. Cercaria capricornia XII is the first known opecoeline cercaria reported from a nassariid gastropod. Cercaria capricornia XII can be distinguished from other opecoelid cercariae by the combination of the presence of a 2-pointed stylet, body length and width, and the size of the tail. The emergence pattern for C. capricornia XII in captivity was erratic; rapid emergences of thousands of cercariae were interspersed by periods that sometimes exceeded a month in which no emergence occurred. There was no detectable pattern to or stimulus of the emergence. The molluscan host range of opecoelids is analysed in detail. Gastropods from the Buccinoidea, Cerithioidea and Rissooidea are hosts to both opecoeline and plagioporine cercariae, but the dominant subfamily infecting the Cerithioidea and Rissooidea is the Plagioporinae. The dominant marine host gastropod superfamily for opecoeline cercariae is the Buccinoidea; the family Nassariidae is contained in the Buccinoidea. The range of gastropod superfamilies known as hosts of plagioporines is much broader than that for opecoelines, which may be explained by the relative size of the two opecoelid families and perhaps by indications that the Plagioporinae is polyphyletic.     

Life cycle and description of a new species of brachylaimid (Trematoda: Digenea) in Spain

The life cycle of Brachylaima llobregatensis n. sp. (Trematoda: Brachylaimidae) is elucidated. Embryonated, operculated, asymmetric eggs (30.9 x 18.2 microm) are eliminated with feces of natural hosts wood mouse Mus spretus; white-toothed shrew, Crocidura russula; and an experimental host, domestic mouse, Mus musculus var. domesticus. The eggs are ingested by the helicid gastropod Helix (Cornu) aspersa, the only natural and experimental first intermediate host. The miracidium hatches from the egg, infects the snails, and develops into a branched sporocyst in the digestive gland. Microcaudate cercariae emerge from this snail and develop into unencysted metacercariae in the kidney of second intermediate host snails H. (C.) aspersa and Otala punctata (natural hosts) and Theba pisana (experimental host). Ingestion of infected snails leads to the infection of definitive hosts, with the adults inhabiting the middle part of the small intestine. There is a chaetotaxic pattern specific on the acetabular (S(II) 5-6 papillae) and body (papillae absent on P(II)) levels. Three types of cercaria papillae were observed by scanning electron microscopy: argentophilic papillae with fingerlike processes (cephalic, body, and acetabular levels); argentophilic papillae with opening (2 papillae in the M body level); and nonargentophilic dome-shaped papillae (on the cephalic C(II) level, alternating with argentophilic S(II) papillae on the ventral sucker). Suckers are subequal, with the acetabulum located in the posterior part of the anterior third of body. Vitellaria extend from anterior margin of acetabulum to between middle level and anterior margin of anterior testis.     

Genetic heterogeneity in natural populations of Microphallus piriformes and M. pygmaeus parthenites (Trematoda: Microphallidae)

The random amplified polymorphic DNA (RAPD) technique was applied for the studies of genetic heterogeny between several natural populations of trematodes belonging to the Microphallida family. Initially, the metacercariae from the daughter sporocysts infestating Littorina saxatilis and Littorina littorea periwinkles were used. Comparison of the banding patterns obtained for the different metacercariae within one sporocyst gave an unpredicted results. For two of three studied species (M. pygmaeus and M. pseudopygmaeus), the considerable differences in RAPD patterning was detected. According to the classical point of view, the process of cercariae (metacercariae in case of the "pygmaeus" group of microphallides) formation does not include DNA recombination. Because of that, all metacercariae within one single sporocyst should be genetically identical. However, data obtained clearly shows that at least in some cases it is not so. We can hardly believe that such result could be a methodological artifact, for not single difference in a RAPD patterns was recorded between the metacercariae within sporocysts of M. piriformes. Moreover, even the 100 fold dilution of the total DNA used for PCR amplification does not change the banding patterns. Hence, our results can not be explained by slight fluctuations in the DNA concentrations between the samples. The most evident conclusion is that we came across yet strange, but real phenomenon--some degree of genetic difference within the progeny of each of the sporocysts--metacercariae. However, the detailed study is needed to understand and interpret these observations correctly. Amplification of the total DNA extracted from the whole sporocysts (containing metacercariae) never showed any differences in RAPD patterns between the parasites been derived from one infestated snail (local parasite hemipopulation). That allowed us to compare different parasite populations referring the RAPD pattern of one sporocyst from a snail to as a representative of one local hemipopulation. Analysis of the RAPD-loci frequencies showed a considerable genetic differences between the subpopulations of M. piriformes, infestating different paraxenic intermediate hosts--L. saxatilis and L. obtusata. This phenomenon was statistically significant for 2 localities of 3 studied. No heterogeny within populations was recorded for M. pygmaeus. Both M. piriformes and M. pygmaeus are characterized by the genetic differentiation in the microgeographic scale (within the Chupa bay of the White sea, the longest distance between the analyzed localities is 20 km). According to the frequencies of the RAPD-loci, parasites from the sheltered locality differ significantly from the parasites of other two localities exposed to the open sea. For both species the degree of genetic similarity between the populations correlates positively with the distance between the localities. We can suppose that the population structure of microphallids depended mainly upon the population structures of their intermediate hosts, definitive hosts and geographical structure of the areal. However, taking into consideration the low motility of snails, we believe that the distribution, migration and species composition of the definitive hosts play the key role in the genetic structuring of M. pygmaeus and M. piriformes hemipopulations. As an addition, the RAPD analysis of the parasite populations from the Barents Sea (East cost of Murman peninsula) and North Sea (Western cost of Sweden) revealed no significant genetic differences between the worms from those places and from the White Sea. However in case of this macrogeographic comparison, insufficient number of samples does not allow us to draw any final conclusions.     

Spatial covariation between infection levels and intermediate host densities in two trematode species

Both theoretical arguments and empirical evidence suggest that parasite transmission depends on host density. In helminths with complex life cycles, however, it is not clear which host, if any, is the most important. Here, the relationships between the abundance of metacercariae in second intermediate hosts, and the local density of both the first and second intermediate hosts of two trematode species, are investigated. Samples of the snail Potamopyrgus antipodarum, the amphipod Paracalliope fluviatilis and the isopod Austridotea annectens were collected from ten stations in a New Zealand lake. In the trematode Coitocaecum parvum, neither the density of the snail first intermediate host nor that of the amphipod second intermediate host correlated with infection levels in amphipods. In contrast, in the trematode Microphallus, infection levels in isopod second intermediate hosts were positively associated with isopod density and negatively associated with the density of snail first intermediate hosts. These relationships are explained by a negative correlation between snail and isopod densities, mediated in part by their different use of macrophyte beds in the lake. Overall, the results suggest that, at least for Microphallus, local infection levels depend on local intermediate host densities.     

Infection by Paragonimus miyazakii cercariae of their crab hosts, Geothelphusa dehaani, by percutaneous penetration

Identification of the transmission routes of the trematode parasite Paragonimus miyazakii into different intermediate hosts would help to explain the natural distribution of the parasite. The behavior of P. miyazakii cercariae released from snails into water and in the presence of a living host or a whole crab leg was observed by stereoscopic or light microscopy at various times after exposure started. On encountering a crab leg or cheliped, the cercariae became entangled with the host via mucoid strands arising from the cercariae. Within 3 hr, most cercariae were attached to the host; cuticular penetration took between 5 and 6 hr, after which cercariae were found in the cavity of the leg. Crabs examined 102-149 days after exposure to the cercariae contained fully developed metacercariae. The metacercariae were fed to 2 rats, and the rats were killed 83 or 111 days later. Some of the metacercariae had reached maturity in the rats. That the cercariae were not ingested by the crabs but penetrated the crabs percutaneously (through hard as well as soft tissue) means that transmission can occur even in areas in which crabs and the host snails do not coexist, as they would if the usual route were oral (when the crabs ate infected snails).     

Effects of ultraviolet radiation on the transmission process of an intertidal trematode parasite

The transmission of parasites takes place under exposure to a range of fluctuating environmental factors, one being the changing levels of solar ultraviolet radiation (UVR). Here, we investigated the effects of ecologically relevant levels of UVR on the transmission of the intertidal trematode Maritrema novaezealandensis from its first intermediate snail host (Zeacumantus subcarinatus) to its second intermediate amphipod host (Paracalliope novizealandiae). We assessed the output of parasite transmission stages (cercariae) from infected snail hosts, the survival and infectivity of cercariae, the susceptibility of amphipod hosts to infection (laboratory experiments) and the survival of infected and uninfected amphipod hosts (outdoor experiment) when exposed to photo-synthetically active radiation only (PAR, 400-700 nm; no UV), PAR+UVA (320-700 nm) or PAR+UVA+UVB (280-700 nm). Survival of cercariae and susceptibility of amphipods to infection were the only two steps significantly affected by UVR. Survival of cercariae decreased strongly in a dose-dependent manner, while susceptibility of amphipods increased after exposure to UVR for a prolonged period. Exposure to UVR thus negatively affects both the parasite and its amphipod host, and should therefore be considered an influential component in parasite transmission and host-parasite interactions in intertidal ecosystems.     

Molecular identification of developmental stages in Opecoelidae (Digenea)

Nuclear ribosomal DNA sequences represent a useful tool for distinction of poorly differentiated developmental stages, such as trematode cercariae or metacercariae. Here, the complete internal transcribed spacer region of the ribosomal DNA (ITS 1 + 5.8S + ITS 2) was sequenced for 29 specimens of the digenean family Opecoelidae, including 16 adult specimens and 13 undescribed larval stages (nine cercariae and four metacercariae) occurring in various marine host organisms. Six cercariae and three metacercariae were found to match their corresponding adult form. This work also revealed that cercariae of the same species are able to infect more than one gastropod host species, suggesting that the specificity for the first intermediate host within the Digenea may be lower than previously thought.     

In vivo and in vitro encystment of Echinochasmus liliputanus cercariae and biological activity of the metacercariae

In vivo and in vitro encystment of the cercariae of Echinochasmus liliputanus and biological activity of the metacercariae were studied. In vivo encystment of cercariae occurred in the gills of goldfish, the second intermediate host. However, the cercariae also encysted in vitro in Locke solution (0.6x to 1.2x strength), 0.7-1.2% NaCI, artificial gastric juice, and human gastric juice. Locke or NaCI solutions were shown to be appropriate for in vitro encystment to occur within 24 hr; however, full-strength Locke solution was shown to be optimal. The 1-day-old metacercariae formed in vivo and treated in 0.1% sodium deoxycholate excystation medium at 37 C for 1 hr showed 88.5% excystation. The metacercariae formed in vitro, however, showed 88.6% and 85.0% excystation for normal and abnormal ones, respectively. Abnormal cysts at room temperature usually die within 10 days. About 70% of the normal cysts, both in vivo and in vitro, can still excyst after being stored in Locke 0.5x solution at 4 C for 3 mo. Cysts formed in vivo and in vitro were equally infective. The encystment of the cercariae in vitro could be inhibited when the cercariae were treated with 1 micromol silver nitrate. Because silver nitrate binds to the papillae, especially to the ciliated papillae, on the cercaria surface, it is suggested that papillary chemoreceptors may be involved in encystment of the cercariae. The finding of E. liliputanus cercariae encysting in vitro, especially in human gastric juice, might be helpful in elucidating mechanisms of the definitive hosts that are directly infected by the cercariae.     

Effects of the herbicide atrazine's metabolites on host snail mortality and production of trematode cercariae

Environmental stressors have the potential to greatly impact the transmission of parasites with complex, multi-host life cycles such as those of trematodes. The commonly used herbicide atrazine has been shown to affect the susceptibility of second intermediate hosts (such as larval amphibians) to trematode infection, as well as the longevity and infectivity of the free-swimming cercariae, but not eggs or the free-swimming miracidia that infect the gastropod first intermediate hosts. However, we do not know if this pesticide influences the survival of infected snails or whether it affects cercariae production within, or emergence from, these hosts. In addition, previous studies of host-parasite dynamics have only examined the parent atrazine compound, not any of the long-lasting metabolites commonly present in water bodies. Here, we report that a concentration of 0.33 μg/L of an atrazine metabolite, desethyl atrazine, increased the mortality of freshwater gastropods ( Stagnicola elodes ) infected with a gymnocephalus type of cercaria but not that of uninfected snails or those harboring a mature or dormant infection of Echinoparyphium sp. In contrast, 2 wk of exposure to desethyl atrazine did not affect the emergence of gymnocephalus cercariae from snails, although a trend for a decrease in the emergence of Echinoparyphium sp. cercariae was observed. We suggest that simultaneous trematode infection and exposure to contaminants may represent a significant combined stress to gastropods, but this is likely parasite species-specific as well as dependent on whether cercariae are being actively produced.     

Acanthoparyphium tyosenense (Digenea: Echinostomatidae): experimental confirmation of the cercaria and its complete life history in Korea

Cercaria yamagutii Ito, 1957, was found in the marine mesogastropods Lunatia fortuni and Glossaulax didyma from the tidelands of Simpo located at the estuary of the Mankyoung River, which runs to the western coast of Korea. Metacercariae were found in a marine bivalve Mactra veneriformis after being infected with C. yamagutii experimentally. When a sea gull, Larus crassiostris, was fed with the metacercariae collected from the infected M. veneriformis, adult worms were recovered 10 days later. It was confirmed that the parasites collected from L. crassiostris were Acanthoparyphium tyosenense Yamaguti, 1939. From the results of this life cycle study, it was determined that the first intermediate hosts of A. tyosenense are L. fortuni and G. didyma. The second intermediate and final hosts are M. veneriformis and L. crassiostris, respectively. Mactra veneriformis was experimentally infected with C. yamagutii isolated from L. fortuni and G. didyma by maintaining them in a water tank for 30 min at about 20 C. The cercariae entered M. veneriformis through their incurrent siphons. Five hours after infection, the cercariae tails began to separate from the bodies, and the cercariae formed cysts. Mature cysts were formed 340 hr (14 days) after infection and identified as the metacercariae of A. tyosenense. The prevalence of A. tyosenense metacercariae was 99.5% in naturally infected M. veneriformis. This is the first report of C. yamagutii as the cercaria of A. tyosenense, and the complete life cycle of A. tyosenense was established in Korea.     

Diversity of trematode genetic clones within amphipods and the timing of same-clone infections

The genetic diversity of trematodes within second intermediate hosts has important implications for the evolution of trematode populations as these hosts are utilized after the parasites reproduce asexually within first intermediate hosts and before sexual reproduction within definitive hosts. We characterised the genetic clonal diversity of the marine trematode Maritrema novaezealandensis within amphipod (Paracalliope novizealandiae) second intermediate hosts using four to six microsatellite loci to determine if multiple copies of identical trematode clones existed within naturally infected amphipods. To determine the relative timing of infections by identical clones within hosts, trematode metacercariae were assigned to six developmental stages and the stages of identical clones were compared. The genotypes of 306 trematodes were determined from 44 amphipods each containing more than one trematode. Six pairs of identical trematode clones were recovered in total (representing five amphipods: 11% of amphipods with greater than one trematode) and all pairs of clones belonged to the same developmental stage. This suggests that identical clone infections are effectively synchronous. A general decrease in the number of metacercariae recovered, prevalence, and mean intensity of infection for each subsequent developmental stage coupled with large numbers of metacercariae (>9) only being recovered from recent infections, supports the occurrence of post-infection amphipod mortality and/or within-host trematode mortality. Taken together, our results indicate that natural infections are characterised by high genetic diversity, but that amphipods also periodically encounter "batches" of genetically identical clones, potentially setting the stage for interactions within and between clonal groups inside the host.     

Discovery of Maritrema jebuensis n. sp. (Digenea: Microphallidae) from the Asian Shore Crab, Hemigrapsus sanguineus, in Korea

Maritrema spp. (Digenea: Microphallidae) are parasites of birds, but have not been found in the Republic of Korea. In this study, metacercariae of Maritrema sp. were discovered in the Asian shore crab, Hemigrapsus sanguineus, caught in the mud-flats of Jebu-do, Hwasung-gun, Gyeonggi-do, and the adult flukes were confirmed by experimental infection into mice. Based on the symmetric ribbon-like vitellarium, adult flukes of Maritrema sp. were identified, but did not belong to previously described species in terms of the following morphologic characteristics: ceca reaching to the lateral wall at the anterior border of the ovary; ventral sucker larger than oral sucker; a prominent metraterm; and vitellarium forming a complete ring. Hence, we named this microphallid M. jebuensis n. sp. after the island where the second intermediate hosts were collected. From this study, it has been shown that Maritrema sp. is distributed in Korea and transmitted by the Asian shore crab, H. sanguineus.