Size-assortative mating in the earthworm Eisenia fetida (Oligochaeta, Lumbricidae)

In many species of simultaneous hermaphrodites, body size correlates with fecundity, and larger partners are preferred to small ones. Since sperm exchange is usually reciprocal, small individuals may be rejected by larger partners resulting in size-assortative mating. We studied the mating patterns in a natural population of the simultaneous hermaphroditic earthworm Eisenia fetida (Oligochaeta, Lumbricidae). We found that size-assortative mating processes existed, with variance in body weight within pairs lower than between pairs in mating earthworms. This non-random mating pattern probably reveals the existence of mate selection in this species, which lives at elevated densities with high availability of potential mates.     

Seasonal population dynamics of Eisenia fetida (Savigny, 1826) (Oligochaeta, Lumbricidae) in the field

In order to assess the response of epigeic earthworms to seasonal changes we monitored the population dynamics of Eisenia fetida (Oligochaeta, Lumbricidae) in a manure heap in the field during a year. Earthworms were hand-sorted from five 0.25 x 0.25 x 0.20 m blocks around the heap in November (autumn) 1999 and in January (winter), April (spring) and August (summer) 2000 to determine earthworm population dynamics. Earthworms of each block were classified into different age classes: mature, preclitellate, juvenile, hatchling and cocoon, and afterwards counted and weighed. Seasonality had a strong effect on the density, biomass and reproductive activity of the population. The population of E. fetida was characterized by a high density of individuals and the predominance of mature individuals throughout the year. Maximum density, mating activity and size of cocoons were achieved in spring, but there were not changes in the number of cocoons per mature earthworm throughout the year. Unexpectedly, the smallest cocoons were produced in winter by the largest individuals. These results suggest that E. fetida is able to allocate resources to growth and/or reproduction in response to environmental fluctuations.     

Eisenia fetida (Oligochaeta, Lumbricidae) Activates Fungal Growth, Triggering Cellulose Decomposition During Vermicomposting

Cellulose is the most abundant polymer in nature and constitutes a large pool of carbon for microorganisms, the main agents responsible for soil organic matter decomposition. Cellulolysis occurs as the result of the combined action of fungi and bacteria with different requirements. Earthworms influence decomposition indirectly by affecting microbial population structure and dynamics and also directly because the guts of some species possess cellulolytic activity. Here we assess whether the earthworm Eisenia fetida (Savigny 1826) digests cellulose directly (i.e., with its associated gut microbiota) and also whether the effects of E. fetida on microbial biomass and activity lead to a change in the equilibrium between fungi and bacteria. By enhancing fungal communities, E. fetida would presumably trigger more efficient cellulose decomposition. To evaluate the role of E. fetida in cellulose decomposition, we carried out an experiment in which pig slurry, a microbial-rich substrate, was treated in small-scale vermireactors with and without earthworms. The presence of earthworms in vermireactors significantly increased the rate of cellulose decomposition (0.43 and 0.26% cellulose loss day⁻¹, with and without earthworms, respectively). However, the direct contribution of E. fetida to degradation of cellulose was not significant, although its presence increased microbial biomass (C_mic) and enzyme activity (cellulase and β-glucosidase). Surprisingly, as fungi may be part of the diet of earthworms, the activity of E. fetida triggered fungal growth during vermicomposting. We suggest that this activation is a key step leading to more intense and efficient cellulolysis during vermicomposting of organic wastes.     

Ranges and cold hardiness of two earthworm subspecies (Eisenia nordenskioldi, Lumbricidae, Oligochaeta)

The ranges of two earthworm subspecies, Eisenia nordenskioldi nordenskioldi (Eisen 1879) and E. n. pallida Malevi 1956, differ in area and partially overlap. E. n. nordenskioldi populates the entire Asian Russia and eastern regions of the Russian Plain, from the lower reaches of the Volga and Don rivers to the Arctic Ocean coasts, while E. n. pallida has not expanded to the Asian permafrost zone and does not occur in European Russia and in the Urals. These subspecies “hold the record” in cold hardiness: the worms and cocoons of the nominotypical subspecies withstand temperatures down to ?34 and ?40°C, and those of E. n. pallida, to ?28 and ?23°C, respectively. Hence, their distribution is independent of subzero temperatures, and their ability to overwinter at any phase of the life cycle makes them also independent of heat supply during the summer period. Differences in geographic range may also be due to biological features of the subspecies. The nominotypical subspecies feeds belongs to the epiendogeic morphoecological type (feeding on the ground surface), whereas E. n. pallida is a true endogeic earthworm. Both subspecies have similar requirements for soil acidity; however, conditions in coarse-humus organomineral horizons of frozen soils appear to be unfavorable for E. n. pallida, which accounts for the absence of this subspecies in the permafrost zone.     

Glycosylated trypsin-like proteases from earthworm Eisenia fetida

Although groups of earthworm proteases have been found by several laboratories, it is still unclear how many of the isolated trypsin-like fibrinolytic enzymes are in glycosylated form. Here, eight glycosylated fibrinolytic proteases (EfP-0-1, EfP-0-2, EfP-I-1, EfP-I-2, EfP-II-1, EfP-II-2, EfP-III-1 and EfP-III-2) were isolated from an earthworm species (Eisenia fetida) through a stepwise-purification procedure: ammonium sulfate precipitation, affinity chromatography on a Sepharose-4B column coupled with soybean trypsin inhibitor (SBTI), and ionic chromatography with a DEAE-Cellulose-52 column. Among the eight purified trypsin-like glyco-proteases, EfP-0-2 and EfP-II-2 were newly isolated isozymes. Glycoprotein staining of the proteases on native-PAGE with a Schiff's reagent (sodium meta-periodate) revealed that the eight proteases were glycoproteins. Measurements of the glycan content with sodium meta-periodate and glycoprotein-test reagent showed that these proteases had different carbohydrate contents. Dot-blotting assay with ConA suggested the oligosaccharides were composed of mannose residues.     

Transmission of nephridial bacteria of the earthworm Eisenia fetida

The lumbricid earthworms (annelid family Lumbricidae) harbor gram-negative bacteria in their excretory organs, the nephridia. Comparative 16S rRNA gene sequencing of bacteria associated with the nephridia of several earthworm species has shown that each species of worm harbors a distinct bacterial species and that the bacteria from different species form a monophyletic cluster within the genus Acidovorax, suggesting that there is a specific association resulting from radiation from a common bacterial ancestor. Previous microscopy and culture studies revealed the presence of bacteria within the egg capsules and on the surface of embryos but did not demonstrate that the bacteria within the egg capsule were the same bacteria that colonized the nephridia. We present evidence, based on curing experiments, in situ hybridizations with Acidovorax-specific probes, and 16S rRNA gene sequence analysis, that the egg capsules contain high numbers of the bacterial symbiont and that juveniles are colonized during development within the egg capsule. Studies exposing aposymbiotic hatchlings to colonized adults and their bedding material suggested that juvenile earthworms do not readily acquire bacteria from the soil after hatching but must be colonized during development by bacteria deposited in the egg capsule. Whether this is due to the developmental stage of the host or the physiological state of the symbiont remains to be investigated.     

Effects of chronic gamma irradiation on reproduction in the earthworm Eisenia fetida (Oligochaeta)

Eisenia fetida were exposed continuously to (60)Co gamma radiation during two generations (F(0) and F(1)). Adult F(0) reproduction capacity (i.e., number of cocoons produced, hatchability and number of F(1) hatchlings) in controls and at five dose rates (0.18, 1.7, 4, 11 and 43 mGy/h) was measured over a 13-week exposure period. Survival, growth and sexual maturation of F(1) hatchlings were observed for 11 weeks. F(1) adults were exposed for a further 13 weeks to determine their reproduction capacity. There was no radiation-induced effect on the cocoon production rate in either F(0) or F(1). For F(0), hatchability of cocoons produced during the first 4 weeks was reduced to 60% at 43 mGy/h (98% in controls), and none of the cocoons produced at 5-13 weeks hatched. At 11 mGy/h the cocoon hatchability was reduced to 25% at 9-13 weeks. In addition, the number of hatchlings per hatched cocoon was reduced at 11 and 43 mGy/h. Correspondingly, at these dose rates, the total number of F(1) hatchlings per adult F(0) was significantly lower than in the control. This number was also reduced at 4 mGy/h, but the effect was of borderline significance. For adult F(1), the hatchability of cocoons at 11 mGy/h was reduced to 45-69% during the 13-week exposure period. The number of hatchlings (F(2)) per cocoon and the total number of F(2) individuals produced was also reduced. However, and in contrast to the results observed for F(0), hatchability increased with time, suggesting a possible acclimatization or adaptation of the F(1) individuals. In conclusion, chronic irradiation reduced the reproduction capacity of E. fetida, but extensive exposure periods (13 weeks) were needed for these effects to be expressed. The lowest dose rates at which an effect was observed were 4 mGy/h in F(0) and 11 mGy/h in F(1).     

Glial cells in the central nervous system of earthworm, Eisenia fetida

Glial elements in the central nervous system of Eisenia fetida were studied at light- and electron microscopic level. Cells were characterized with the aid of toluidine blue, Glial Fibrillary Acidic Protein (GFAP), S100 staining. We identified neurilemmal-, subneurilemmal-, supporting-nutrifying- and myelinsheath forming glial cells. Both neuronal and non-neuronal elements are S100-immunoreactive in the CNS. Among glial cells neurilemmal and subneurilemmal cells are S100-immunopositive. With the antibody against the S100 protein one band is visible at 15 kDa. GFA P-immunopositive supporting-nutrifying glial cells are localized around neurons and they often appear as cells with many vacuoles. GFA P-positive cell bodies of elongated neurilemmal glial cells are also visible. Western blot analysis shows a single 57 kDa GFA P immunoreactive band in the Eisenia sample. At ultrastructural level contacts between neuronal and glial cells are recognizable. Glial cell bodies and their filopodia contain a granular and vesicular system. Close contacts between neuronal cell membranes and glial filopodia create a special environment for material transport. Vesicles budding off glial cell granules move towards the cell membranes, probably emptying their content with kiss and run exocytosis. The secreted compounds in return may help neuronal survival, provide nutrition, and filopodia may also support neuronal terminals.     

Reorganization of the GABAergic system following brain extirpation in the earthworm (Eisenia fetida,Annelida, Oligochaeta)

The reorganization of the GABAergic system was studied by means of immunohistochemistry after the symmetrical and asymmetrical (unilateral) extirpation of the brain of the annelid Eisenia fetida. GABA-immunoreactive neurons were first observed in the wound tissue on the 3rd postoperative day. Thereafter the number of labelled cells gradually increased, and by postoperative days 76-80 all GABA-immunoreactive cells (approx. 140 neurons) could be found in their final positions in the symmetrically regenerated brain. After asymmetrical brain extirpation, nearly all cells (70-75) could be detected in the regenerating hemisphere by postoperative days 50-56. In the early stages of the asymmetrical regeneration of the brain, more GABAergic cells were concentrated dorsally and laterally in the preganglion than during the symmetrical type of regeneration. In both types of regeneration, the immunoreactive neurons in the regenerated brain originated in part from undifferentiated neuroblasts situated in different parts of the body, and in part from dividing neurons localized mainly in the pharyngeal nerve plexus. Both exogenous GABA and picrotoxin, applied during the early stages (days 10- 12) of brain regeneration, inhibited the development of the wound tissue and the migration of the neuroblasts and the enteric neurons. At the same time, exogenous GABA application accelerated the proliferation of the pharyngeal neurons. No effect on the process of regeneration could be demonstrated when exogenous GABA and picrotoxin were given together.     

Multiple mating increases cocoon hatching success in the earthworm Eisenia andrei (Oligochaeta: Lumbricidae)

In simultaneous hermaphrodites with reciprocal mating, multiple mating may be a male strategy that conflicts with female interests, and therefore an intra‐individual sexual conflict regarding the number of matings may be expected. The evolutionary outcome of this sexual conflict will depend on the costs and benefits that extra mating entails for each sexual function. In the present study, we investigated the costs and benefits of multiple mating on cocoon number, cocoon mass, and cocoon hatching success in the redworm Eisenia andrei, a simultaneous hermaphrodite with reciprocal insemination, by manipulating the number of matings with different partners. We did not detect any reduction in the female reproductive output (number and mass of cocoons) with increasing number of mating partners. However, we found that multiple mating showed benefits for female reproduction that increased the hatching success of the cocoons. This effect may be a result of increased quantity and/or diversity of sperm in the spermathecae of multiple mated earthworms. Further studies are required to clarify the mechanism underlying the increased cocoon hatching success when redworms engage in multiple matings. © 2012 The Linnean Society of London, Biological Journal of the Linnean Society, 2012, ?? , ??–??.     

Reorganization of monoaminergic systems in the earthworm,Eisenia fetida,following brain extirpation

The present study describes the major aspects of how monoaminergic (serotonin, dopamine) systems change in the course of regeneration of the brain in the earthworm (Eisenia fetida), investigated by immunocytochemistry, HPLC assay, and ligand binding. Following brain extirpation, the total regeneration time is about 80 days at 10 degrees C. On the 3rd postoperative day serotonin, and on the 11th postoperative day tyrosine hydroxylase-immunoreactive neurons can be observed in the wound tissue. Thereafter the number of the immunoreactive cells increases gradually, and by the 76th-80th postoperative days all serotonin- and tyrosine hydroxylase-immunopositive neurons can be found in their final positions, similarly to those observed in the intact brain. Labeled neurons located in the dorsal part of the regenerated brain appear earlier than the cells in lateral and ventral positions. Both serotonin- and tyrosine hydroxylase-immunoreactive neurons of the newly formed brain seem to originate from undifferentiated neuroblasts situated within and around the ventral ganglia and the pleura. Dopaminergic (tyrosine hydroxylase-immunoreactive) elements may additionally derive from the proliferation of neurons localized in the subesophageal ganglion and the pharyngeal nerve plexus. Following brain extirpation, both serotonin and dopamine levels, assayed by HPLC, first increase in the subesophageal ganglion; by the 25th day of regeneration, the monoamine content decreases in it and increases in the brain. Hence it is suggested that monoamines are at least partly transported from this ganglion to the regenerating brain. At the same time, (3)H-LSD binding can be detected in the regenerating brain from the 3rd postoperative day, showing a continuous increase until the 80th postoperative day, suggesting a guiding role of postsynaptic elements in the monoaminergic reinnervation of the newly formed brain.     

阿维菌素B1a对土壤微生物和蚯蚓的影响

研究了不同阿维菌素B_1a浓度对种土壤微生物生长和呼吸强度的影响及对土壤中蚯蚓的急性毒性.结果表明,土壤中阿维菌素B_1a浓度在83.3 mg·kg^-1以上时对种土壤细菌均表现出明显的抑制作用,但对土壤真菌不表现抑制作用.高浓度阿维菌素B_1a对土壤微生物的呼吸强度有抑制作用,并且在不同土壤其作用有差异.采用滤纸接触试验和人工土壤试验测定阿维菌素对蚯蚓的急性毒性(半数致死量,LD₅₀),接触毒性LD₅₀为.63μg·cm^-2,土壤法试验测定的LD₅₀在处理后第7 d和第1 d分别为2.13和17.06 mg·kg^-1.     

8-hydroxydeoxyguanosine generated in the earthworm Eisenia fetida grown in metal-containing soil

Heavy metal pollution of soil causes biological problems, such as mutagenicity to living organisms, including human beings. However, few methods have been developed to assess metal mutagenicity in soil. To avoid metal mutagenicity, an adequate bio-monitoring method is required. In the present study, to determine if the analysis of oxidative DNA damage generated in the earthworm is a useful bio-monitoring method for soil mutagenicity, the accumulation of 8-hydroxydeoxyguanosine (8-OH-dG), a major form of oxidative DNA damage, in Eisenia fetida (Savigny, 1826) treated with cadmium chloride (CdCl2) or nickel chloride (NiCl2) was analyzed. E. fetida was treated with Cd (10 or 200 microg/g soil) or Ni (10 or 200 microg/g soil) for 1, 2, and 3 weeks or 3 months. After metal exposure, the metal concentration in E. fetida was analyzed by atomic absorption spectrometry and the 8-OH-dG accumulated in E. fetida was analyzed by HPLC analyses and immunohistochemistry. Atomic absorption spectrometry revealed that Cd, but not Ni, accumulated within E. fetida. The 8-OH-dG levels in the DNA of E. fetida treated with Cd for 3 months were significantly higher than those in control E. fetida. Moreover, immunohistochemical analyses revealed that positive signals for 8-OH-dG accumulation in seminal vesicles were detected only in E. fetida treated with 10 microg of Cd for 3 months. Although some points remain unresolved, a bio-monitoring system analyzing the DNA damage generated in the earthworm might be useful for the assessment of the mutagenicity of soil contaminated with various heavy metals, such as Cd.     

Cold-stress induced formation of calcium and phosphorous rich chloragocyte granules (chloragosomes) in the earthworm Eisenia fetida

The cytochemical and functional characteristics of chloragocytes of both 'control' and cold-stressed Eisenia fetida were examined. Flow cytometry revealed the heterogeneity of chloragocytes: the first group was characterized by low, the second one by high acid phosphatase (AcP) content. In 'control' animals the former, in cold-stressed ones the latter type were the dominant form. The elevated AcP-activity correlated with the accumulation of autophagic vacuoles (AVs) in chloragocytes. Both AVs and all small chloragosomes showed high AcP activity, while most of the large chloragosomes did not display any. Most 'control' granules (0.75-1.25 μm) contained high amounts of Ca and P, with less and variable quantities of S, Cl, K, Fe and Zn. Small chloragosomes with low Ca and P concentrations were seldom found. In cold-stressed animals the number of small granules (0.25-0.75 μm) increased up to 40% of total population. Their Ca and P contents were significantly lower; S and Fe concentrations were higher than those of large chloragosomes (1.0-1.5 μm). Our results prove that the formation and elemental composition of chloragosomes can be influenced by environmental stressors and suggest that the mature chloragosomes are tertiary lysosomes and their formation is coupled to autophagocytosis.     

Enantioselective toxicity and degradation of chiral herbicide fenoxaprop-ethyl in earthworm Eisenia fetida

The enantioselective degradation of fenoxaprop-ethyl in ecological indicator earthworm was studied and the main metabolites (fenoxaprop, 6-chloro-2,3-dihydrobenzoxazol-2-one, ethyl-2-(4-hydroxyphenoxy)propanoate, 2-(4-hydroxyphenoxy)propanoic acid) were also monitored on an enantiomeric level. The individual enantiomers of fenoxaprop-ethyl and its three chiral metabolites were prepared to study the acute toxicity to earthworm. Chiral analysis methods were set up based on HPLC–MS/MS with chiralpak IC chiral column. Fenoxaprop-ethyl was not found in earthworms, while the primary metabolite fenoxaprop was in relatively high levels indicating a quick hydrolysis degradation. Fenoxaprop was accumulated almost exclusively with R-enantiomer in earthworms and the bio-concentration factors of R-fenoxaprop and S-fenoxaprop were 1.39 and 0.17 respectively with the enantiomer fraction (EF) values about 0.99. The degradation of R-fenoxaprop in earthworms followed first-order kinetics with half-life of 1.82 day. The other metabolites could not be detected in earthworms. The calculated LC₅₀ values showed ecological indicator earthworm was more sensitive to the four metabolites than fenoxaprop-ethyl. Furthermore, earthworm was more sensitive to the R-form of the chiral metabolites than the S-form and rac-form. The results suggested metabolites and enantioselectivity should be taken into consideration to better predict the exposure concentration and apply ecological indicators in toxicological studies.     

Distribution and ecology of the earthworm Eisenia nordenskioldi pallida (Oligochaeta,Lumbricidae) dominant in southern Siberia and the Russian Far East

A literature review and the results of re-examination of collections of the earthworm Eisenia nordenskioldi are presented. Peculiarities of the geographic and habitat distribution, as well the ecology of the diploid subspecies E. n. pallida, originally described as a non-pigmented form of E. nordenskioldi, are analyzed. E. n. pallida inhabits mainly the south of Siberia and the Russian Far East, penetrating as far northwards as the forest-tundra zone along the valleys of large rivers. It is the most abundant in the low-mountain belt in dark coniferous forests, being the dominant form of Lumbricidae there, but it is quite rare above the timberline. The range of E. n. nordenskioldi, which can be found together with E. n. pallida in the south of the territories studied, is much wider, extending further to the north and including some Arctic islands. In addition, E. n. nordenskioldi is known from the eastern parts of European Russia. Both subspecies show high levels of resistance to unfavorable abiotic factors. Differences in the latitudinal and altitudinal distribution patterns of both subspecies of E. nordenskioldi are determined by their different morpho-ecological forms.     

Characterization of two endoglucanases for the classification of the earthworm,Eisenia fetida Waki

Eisenia fetida and Eisenia andrei are vermicomposting species that are used as model animals for testing chemical material toxicology. Eisenia spp. are grown commercially in various fields in Japan. However, these two species have not been classified because it is difficult to distinguish them morphologically; thus, all bred earthworms are called E. fetida. However, it has been proposed that these two species have different expression regulation mechanisms. Here, we classified a sample of earthworms purchased from several farms, confirming that both E. fetida and E. andrei are present in Japanese earthworm breeding programs. We also characterized two highly active endoglucanases (EfEG1 and EfEG2) from the E. fetida Waki strain, which contained strong fibrinolytic enzymes for improving human health. We confirmed that EfEG1 is 1371 bp long and belongs to GHF9. Thus, E. fetida Waki may have commercial application for biomass utilization and as a dietary health supplement.     

Phoresy of the entomopathogenic nematode Steinernema feltiae by the earthworm Eisenia fetida

The free-living stage of entomopathogenic nematodes occurs in soil, and is an environmental-friendly alternative for biological control. However, their dispersal capability is limited. Earthworms improve soil characteristics, changing soil structure and influencing many edaphic organisms. Thus, earthworms could be used as vectors to introduce/disperse beneficial organisms. Nevertheless this interaction has not been studied in detail. This study presents the infectivity results of Steinernema feltiae after passing through the Eisenia fetida gut. Although entomopathogenic nematodes have no deleterious effects on earthworms, their passage through E. fetida gut seriously affected their mobility and virulence.     

土壤低剂量芘污染对蚯蚓若干生化指标的影响

通过人工污染土壤的方法,设计芘的暴露浓度为0、60、120、240、480、960μg.kg-1.暴露实验进行1、3、7和14d后,分别检测蚯蚓内脏中细胞色素P450含量、谷胱甘肽转移酶(GST)、超氧化物歧化酶(SOD)、过氧化物酶(POD)、过氧化氢酶(CAT)活性和丙二醛(MDA)含量.结果表明,在供试浓度范围内,蚯蚓内脏中各生化指标对污染物暴露指示的敏感性存在差异:其中P450含量、GST和SOD活性最为敏感;POD和CAT活性次之;而MDA含量未对低剂量的芘暴露起到明显的指示作用.研究同时发现,低剂量污染物暴露的时间效应要强于剂量效应的影响.因而,在进行生态毒性诊断时,采用多指标和多时段的检测对增强指示的灵敏性和有效性尤为重要.     

Distribution and action of some putative neurotransmitters in the stomatogastric nervous system of the earthworm,Eisenia fetida (Oligochaeta,Annelida)

Brain Cell Biology - The chemical neuroanatomy of the stomatogastric nervous system of the earthworm, Eisenia fetida, has been investigated, using antibodies raised against serotonin, tyrosine...