Caudal regeneration in Tubifex tubifex (Oligochaeta, Tubificidae) following copper exposure

Abstract. The oligochaete, Tubifex tubifex (Oligochaeta, Tubificidae), is capable of the auto-tomization and regeneration of missing body parts. The posterior end of the worm is subject to autotomy (e.g., after predation or after exposure to toxic substances) and can be regenerated thereafter. Despite the ecological importance of T. tubifex , processes of autotomy and regeneration have been little studied. This work describes daily morphological and histological observations of caudal regeneration of this worm. We also compared regeneration processes when autotomy was induced by artificial transsection of the worms (control worms) and by exposure to copper ion, a frequently encountered environmental contaminant. The worms regenerated a functional posterior end (defecation possible) in 7 d as well as a prepygidial zone from which new segments could be regenerated. In T. tubifex , regeneration involved the presence of activated mesoblastocytes in the worm, increased number of neoblasts, and the increased migration of these cells. However, Cu²⁺ did not affect these processes, since no difference could be detected between regeneration of control and Cu-autotomized worms.     

Selective feeding by the aquatic oligochaete Tubifex tubifex (Tubificidae,Clitellata)

The particle size distribution of faecal pellets produced by the tubificid worm Tubifex tubifex in laboratory culture, was measured with a Coulter® Multisizer. The faecal material from worms cultured in a range of sediments was composed of particles with a mean diameter of less than 63 m, and only a few isolated larger particles were found by microscopic analysis. This suggests that this species actively selects the silt-clay fraction, avoiding larger sand particles. A more detailed analysis of faeces revealed that about 75%, by volume, was composed of particles with a mean diameter < 25m, and the mode was < 10m. T. tubifex fed selectively on the organic rich particles of the sediment, and this feeding was independent of particle size. Measurement of the organic content of faeces (measured as % loss on ignition) showed that they had a consistently higher organic content than the sediment, considered as whole sediments or the <63 m sieved fraction. On the basis of these results, we hypothesise that this species exhibits two levels of selectivity in its feeding behaviour. Thus selection is primarily based on particle size, avoiding the ingestion of sand particles and also, on the preferential selection of particles associated with organic material, within the fine (silt-clay) fraction of the sediment.     

Interactions among two strains of Tubifex tubifex (Oligochaeta: Tubificidae) and Myxobolus cerebralis (Myxozoa)

Host-parasite interactions influence host population growth, host evolution and parasite success. We examined the interactions among Myxobolus cerebralis, the parasite that causes salmonid whirling disease, and resistant and susceptible strains of the oligochaete host Tubifex tubifex. Strains of T. tubifex with diverse genotypes often coexist in nature and have variable susceptibilities to M. cerebralis infection. Further, parasite proliferation differs by several orders of magnitude among T. tubifex strains. We examined total biomass produced by individual T. tubifex, including progeny production and adult growth, parasite proliferation and prevalence of infection using 2 strains of T. tubifex at 2 myxospore doses in a response-surface experimental design. Total biomass production per individual oligochaete and progeny biomass produced by an individual adult oligochaete were density-dependent for both resistant and susceptible individuals and the effects did not change with the addition of myxospores. However, both resistant and susceptible adults had highest growth when exposed to M. cerebralis. The presence of resistant oligochaetes in mixed cultures did not reduce the infection prevalence or parasite proliferation in susceptible individuals. In natural aquatic communities, resistant strains of T. tubifex may not reduce the effects of M. cerebralis on the salmonid host, particularly if sufficient numbers of susceptible T. tubifex are present.     

Growth of Tubifex tubifex Müller (Oligochaeta,Tubificidae) under Various Trophic Conditions

Growth of Tubifex tubifex on highly caloric activated sludge and poor in organic matter natural silt was experimentally studied. The relationship between oxygen consumption rate, weight, and caloric value of T. tubifex body in culture on activated sludge was determined to calculate the growth efficiency (K₂ = P/P + R). The growth rate has been determined, and models of T. tubifex growth are given under various trophic conditions conforming to results of the experiment. Somatic growth, generative growth and life span of this species were compared on the basis of the experiments and literature. Acceleration of growth and prolongation of life under conditions of caloric food and delay of these processes in silt poor in organic matter have been established and measured. T. tubifex was found to have advantages as compared to other tubificid species in response to increasing amounts of nutrient food in waters polluted by organic matter and, therefore, to produce abundant populations under these conditions.     

Density-dependent processes in cohorts of Tubifex tubifex,with special emphasis on the control of fecundity

Laboratory cohort cultures of the tubificid Tubifex tubifex with different initial densities were carried out at 20° C with the condition of unlimited food. The main results were: 1) Intracocoon mortality was 37% of the laid eggs (observation of 689 eggs); 2) The principal bionomic parameters (generation time, r, R₀) appeared to be density dependent; 3) Recruitment was regulated through the percentage of worms that actually attained the ovigerous stage, specific fecundity, and the duration of the egg laying stage, which appeared to be inversely correlated with density.     

What is Tubifex tubifex (O. F. Müller) (Oligochaeta,Tubificidae)?

A review of the fairly muddled conditions in the early history of Tubifex tubifex (O. F. Müller, 1774) is given. The identity of Müller's (1774) Lumbricus tubifex is discussed, analytical examinations of samples from the most likely type-locality of the species are accounted for, and a neo type has been selected. For comparison, other "Tubifex" specimens, both European and American, are likewise analysed and a scrutiny of the literature concerning the species and its synonyms is undertaken. The survey results in a clearer picture of T. tubifex and a revised view of the qualifying characters on the specific level. Diagnoses of the species T. tubifex (O. F. Müller) and T. fissidens sp.n., Kernall, MS, 1914, are given.     

The effect of cohabitation of Tubifex tubifex (Oligochaeta: Tubificidae) populations on infections to Myxobolus cerebralis (Myxozoa: Myxobolidae)

The competitive interactions between susceptible and resistant Tubifex tubifex (Oligochaeta: Tubificidae) exposed to Myxobolus cerebralis (Myxozoa: Myxobolidae) infections were investigated in two laboratory trials. Competition was assessed by the total parasite production over the course of the trials in mixed and pure cultures of M. cerebralis exposed worms, and by the genetic analyses of worms from the control and experimental groups at the beginning and end of the experiments. Mixed cultures of resistant and susceptible worms showed a 70% reduction in production of parasites released when compared with pure cultures of susceptible worms. In studies with laboratory and field-collected oligochaetes the mixed cultures at the end of the cohabitation experiments were dominated by resistant Tubifex from lineage V (HB strain) this strain of Tubifex has a competitive advantage over worms from other lineages. The results of this study suggest that certain species of Tubifex may be dead-end hosts to M. cerebralis by absorbing or inactivating the parasite and may also show greater survival compared to susceptible oligochaetes in certain whirling disease enzootic habitats.     

Cuticular ultrastructure in some marine oligochaetes (Tubificidae)

Abstract. The ultrastructure of the thin, non-cellular cuticle is described for 6 marine oligochaetes, representing 3 of the subfamilies (Phallodrilinae, Limnodriloidinae, and Rhyacodrilinae) of the Tubificidae. The main components of the cuticle in these 6 species, as in most other oligochaetes examined, are: (1) a fiber zone closest to the epidermis, consisting of collagen fibers embedded in a matrix, (2) an epicuticle, which is a continuation of the matrix outside the fiber zone, and (3) epicuticular projections, which are membrane-bound bodies covering the outer surface of the epicuticle. The projections are probably formed by the microvilli that penetrate the cuticle from the epidermal cells below, but this was confirmed only in the studied limnodriloidines. Three of the species examined, Duridrilus turdus, Olavius vacuus , and Heterodrilus paucifascis , lack microvilli. The morphology of the components in the cuticle differs between the studied species. The collagen fibers may form an "orthogonal grid" (i.e., layers of parallel fibers perpendicular to the layers immediately above and below), or they may form parallel layers, or be irregularly scattered. The number of dense layers in the epicuticle, as well as the shape and internal structure of the epicuticular projections, also vary. All these characters might be useful in future phylogenetic analyses to achieve better hypotheses of relationships within oligochaetes as well as to other groups.     

Molecular phylogeny of Coriaria, with special emphasis on the disjunct distribution

Coriaria, which has the most conspicuously disjunct distribution of the flowering plants, is distributed in four separate areas of the world. The phylogenetic relationships of 12 Coriaria species collected from the representative disjunct areas were inferred by comparing 2416 bp of the combined data set of rbcL (a large subunit of ribulose 1,5-bisphosphate carboxylase/oxygenase) and matK (maturase K) genes. The phylogenetic tree shows that the Chile-Papua New Guinea-New Zealand-Pacific islands species and the Central America-northern South America species form a sister group, and the Eurasian clade is more basal to them. The divergence time between the Eurasian group and the other species was estimated as 63 or 59 million years ago using rbcL and matK molecular clocks, respectively. These results do not support previously proposed hypotheses which explain the disjunct distribution on the basis of continental drift but suggest that the distribution pattern was formed by several geographical migrations and separations in the Cenozoic.     

The development of Eustrongylides tubifex (Nematoda: Dioctophymatoidea) in oligochaetes

Egg development of Eustrongylides tubifex (Nitzsch in Rudolphi, 1819) J?gerski?ld, 1909, was studied at 5, 10, 15, 20, 25, and 30 C. Eggs developed at 20, 25, and 30 C. Development ceased at 0, 5, 10, and 15 C but resumed when temperatures were raised. Eggs contained first-stage larvae in 23-26 days at 25 C. Seven species of aquatic oligochaetes were exposed experimentally to eggs of E. tubifex containing first-stage larvae. Third-stage larvae of E. tubifex developed in the aquatic oligochaetes, Limnodrilus hoffmeisteri Claparède, 1862, and Tubifex tubifex (Müller, 1774). Larvae developed in the ventral blood vessel of oligochaetes. Second-stage larvae were seen first in oligochaetes 30 days postinfection and third-stage larvae were seen 70-109 days postinfection at 25 C. Third-stage larvae in oligochaetes retained the cuticle of the first and second molt.     

Molecular phylogeny of the Herpestidae (Mammalia, Carnivora) with a special emphasis on the Asian Herpestes

Until now, phylogenetic studies of the mongooses (Carnivora, Herpestidae) have not included an exhaustive sampling of the Asian members of this family. In this study, we used mitochondrial (Cytochrome b and ND2), nuclear (β-fibrinogen intron 7 and Transthyretin intron 1) sequences from almost all of the recognized mongoose species to produce a well-resolved phylogeny of the Herpestidae. We also performed molecular dating analyses to infer divergence dates of the different lineages within the Herpestidae. Our results confirmed the paraphyly of the Herpestes genus and other phylogenetic relationships, which previously had only been moderately supported. The Asian herpestid species were found to form a monophyletic group within the Herpestidae. Within the Asian species, a cyto-nuclear conflict was discovered between the small Indian mongoose (Herpestes auropunctatus), the Indian gray mongoose (Herpestes edwardsii) and the Javan mongoose (Herpestes javanicus), which may have occurred through interspecific hybridization. This study inferred an Early Miocene origin for the Herpestidae and a Middle Miocene origin for the Asian mongooses.     

Acute tolerance tests with the oligochaetes Nais communis (Naididae) and Ilyodrilus frantzi (Tubificidae)

96-h LC50's were determined for Nais communis Piguet and Ilyodrilus frantzi Brinkhurst in relation to selected pollutants (Hg, NaPCP) and environmental factors (pH, temperature, salinity). The naidid N. communis was consistently less tolerant than the tubificid I. frantzi in all test treatments. Comparison with previously determined sensitivities for nine tubificids and one lumbriculid shows that I. frantzi is in the mid to upper tolerance range, while N. communis is almost consistently the least tolerant of all species tested.     

On the role of tubificid oligochaetes in relation to fish disease with special reference to the myxozoa

Whirling disease is caused by the myxozoan parasite Myxobolus cerebralis, Hofer. It has been associated with a serious decline in wild rainbow trout populations in the western United States. The alternate host is the tubificid oligochaete Tubifex tubifex, and perhaps other tubificids. Tubifex is a very adaptable taxon, being found in various morphological forms. Polyploidy has been demonstrated. There is some suspicion that sibling species may be found to exist. Infection rates in tubificids are low. Related myxosporeans are associated with a variety of aquatic oligochaete worms belonging to three families, Tubificidae, Naididae and Lumbriculidae, and also with polychaetes. Myxozoans are now recognized as members of the phylum Cnidaria. Other fish parasites, such as the cestodes Archigetes and Caryophyllaeus and the nematode Eustrongylides, are transmitted by tubificids. The tubificid worm population in an ecosystem may provide an infective reservoir as the disease persists for long periods in the worm. Control of worm populations might be achieved by reducing organic inputs, lowering erosion, increasing flow and removing objects that accumulate pockets of silt. The use of pesticides is unlikely to be effective because of a lack of specific impact and consequent disruption of the whole benthic community, at least.     

Nuclear fragmentation characterises paraspermiogenesis in Tubifex tubifex (Annelida, Oligochaeta)

It is known that tubificine oligochaetes produce two types of spermatozoa: eusperm, fertilizing sperm with regular haploid DNA content; and parasperm, with a much lower DNA content, protecting and carrying the eusperm. Whereas mature spermatozoa and spermatids of the two lines are easily recognized by their morphology and DNA content, little is known about the first steps of differentiation of the two lines. This subject is addressed here in two ways: we have measured DNA content by a new method based on confocal laser microscopy and found that the total DNA content of parasperm cysts is extremely variable and equal or lower than total DNA content of eusperm cysts. Then we focused on the spermatocytes, and we found that the cells which will form paraspermatids undergo a peculiar kind of nuclear fragmentation which differ greatly from a regular cell division. During fragmentation DNA is distributed unevenly among the spermatids and this gives rise to a great and variable number of parasperm with variable DNA content. Immunocytochemical assays revealed that a proper meiotic spindle is never formed during fragmentation and that actin may play an important role in the chromatin division. Electron micrographs showed that the centrioles undergo a phenomenon of mass reproduction similar to that found in ciliated epithelia which supplies each of the numerous paraspermatids of its basal body. Mol. Reprod. Dev. 59: 442-450, 2001.     

The histochemistry of the clitellum of Tubifex tubifex (annelida: oligochaeta)

The histochemistry of the clitellar glands of Tubiflex tubiflex is reported, together with the ultrastructure of mature secretion units. The distribution and ultrastructure of mucous and granule cells are consistent with published accounts of tubificid citella, thus indicating an absence of intrageneric variation. Mucous cells contain homogenous electron lucent globules which are rich in neutral and acidic mucopolysaccharides and are non-reactive with techniques designed to reveal protein. Type I cells possess structured granules composed of tyrosine-containing basic protein and carboxylated acid mucopolysaccharides. The granules of type II cells are heterogenous and contain several 0.3--0.7 micrometer inclusions of acidic protein with identifiable tyrosine, tryptophan and cystine. The inclusions occur within a matrix composed of neutral mucopolysaccharides and carboxymucin. Type III cell granules possess 0.2--0.5 micrometer inclusions within an electron dense matrix and are rich in neutral and carboxylated acid mucopolysaccharides and basic protein containing tyrosine, tryptophane and cystine. Clitelar carboxymucins are characterized by their stability to beta-glucoronidase, hyaluronidase, neurominidase and acid hydrolysis. A pyridine-extractable lipid moiety within the clitellar secretions was not detected. The results are discussed in relation to cell function during cocoon deposition and, comparatively, with those documented for other oligochaete families.     

Molecular phylogeny of litostome ciliates (Ciliophora, Litostomatea) with emphasis on free-living haptorian genera

The monophyly of the litostomes was tested using nine newly sequenced and four previously unpublished small subunit ribosomal RNA (SSrRNA) gene sequences from free-living Haptoria as well as from endosymbiotic Trichostomatia: the vestibuliferids Balantidium coli and Isotricha prostoma, the cyclotrichiid Mesodinium pulex, and the haptorids Loxophyllum rostratum, Dileptus sp., Enchelyodon sp., Enchelys polynucleata, Epispathidium papilliferum (isolates A and B), Spathidium stammeri, Arcuospathidium muscorum, Arcuospathidium cultriforme, and the unusual Teuthophrys trisulca. Phylogenetic analyses depicted the litostomes as a monophyletic group consisting of the trichostomes (subclass Trichostomatia) and the free-living haptorians (subclass Haptoria). The cyclotrichiids Mesodinium and Myrionecta (order Cyclotrichiida) branched either basally within or outside the Litostomatea. In most analyses, the haptorians did not receive support as a monophyletic group. Instead, Dileptus branched basally to all litostome taxa, and Epispathidium papilliferum grouped with the Subclass Trichostomatia. Some subgroupings, however, of haptorian genera corresponded to suggested superfamilial taxa (e.g., orders Spathidiida and Pleurostomatida). Within the monophyletic trichostomes, we can distinguish three clades: (1) an Australian clade; (2) the order Entodiniomorphida; and (3) the order Vestibuliferida. However, Balantidium, currently classified in the Vestibuliferida, did not group with the other vestibuliferids, suggesting that this order may be paraphyletic.     

Two sperm types in the spermatozeugmata of Tubifex tubifex (annelida,oligochaeta)

The spermatozeugmata (sperm bundles lacking a distinct wall) from the spermathecae of Tubifex tubifex are composed of two different zones: an internal axial cylinder containing conventional spermatozoa and an external cortex composed of modified spermatozoa, tightly packed together. The conventional spermatozoa conform to the classical clitellate scheme: very long and thin with a complex acrosome, a filiform nucleus, small mitochondria, and a flagellum with Y links and β glycogen granules as accessory structures. The modified spermatozoa show “empty” acrosomes, degenerating nuclei, and tails which contain γ glycogen granules. The tails are helically wound around the spermatozeugma and are connected to each other by junctional complexes. The tips of the cortical tails are free and move with a metachronal wave. The presence of two sperm types in tubificids is discussed and a protective function for the modified cortical spermatozoa is proposed.     

A morphological reappraisal of Tubifex blanchardi Vejdovský, 1891 (Clitellata: Tubificidae)

Tubifex blanchardi Vejdovský, 1891 is a freshwater tubificid, often living in sympatry with Tubifex tubifex (Müller 1774). Although considered from its discovery as a species on its own, its biological status is debated. During the early seventies T. blanchardi was reduced to a mere form of T. tubifex, as a particular case of polymorphism in chaetal pattern. Using classical histological techniques, microdissections of portions of the male genital apparatus and phalloidin staining of dissected copulatory organs we investigated 163 mixed individuals of T. blanchardi and T. tubifex belonging to sympatric populations from the Lambro River (Milan, Northern Italy). The internal morphology of T. blanchardi is described for the first time. Our results show that T. tubifex and T. blanchardi differ in several characters concerning both their external and internal morphology, and in the fine organization of their copulatory organs. Several independent character sets support the separation of T. blanchardi from T. tubifex, suggesting that it is an independent species. This study also supports the idea that T. blanchardi and T. bergi (Hrabě, 1935), another species closely related to T. tubifex, are not conspecific. The observed morphological differences between allopatric populations of T. tubifex are discussed.