Ovary micromorphology in hormogastrid earthworms with a particular emphasis on the organization of the germline cysts.

There is a gap in our knowledge of microorganization and the functioning of ovaries in earthworms (Crassiclitellata) and allied taxa. Recent analyses of ovaries in microdriles and leech-like taxa revealed that they are composed of syncytial germline cysts accompanied by somatic cells. Although the pattern of cyst organization is conserved across Clitellata - each cell is connected via one intercellular bridge (ring canal) to the central and anuclear cytoplasmic mass termed the cytophore - this system shows high evolutionary plasticity. In Crassiclitellata, only the gross morphology of ovaries and their segmental localization is well known, whereas ultrastructural data are limited to lumbricids like Dendrobaena veneta. Here we present the first report about ovarian histology and ultrastructure in Hormogastridae, a small family of earthworms inhabiting the western parts of the Mediterranean sea basin. We analyzed three species from three different genera and showed that the pattern of ovary organization is the same within this taxon. Ovaries are cone-like, with a broad part connected to the septum and a narrow distal end forming an egg string. Ovaries are composed of numerous cysts uniting a small number of cells, eight in Carpetania matritensis. There is a gradient of cysts development along the long ovary axis, and three zones can be distinguished. In zone I, cysts develop in complete synchrony and unite oogonia and early meiotic cells (till diplotene). Then (zone II), the synchrony is lost, and one cell (prospective oocyte) grows faster than the rest (prospective nurse cells). In zone III, oocytes pass the growth phase and gather nutrients; at this time, their contact with the cytophore is lost. Nurse cells grow slightly, eventually die via apoptosis, and are removed by coelomocytes. The most characteristic feature of hormogastrid germ cysts is the inconspicuous cytophore in the form of thread-like thin cytoplasmic strands (reticular cytophore). We found that the ovary organization in studied hormogastrids is very similar to that described for D. veneta and propose the term "Dendrobaena" type of ovaries. We expect the same microorganization of ovaries will be found in other hormogastrids and lumbricids.

Macrofauna del suelo y condiciones edafoclimáticas en un gradiente altitudinal de zonas cafeteras, Huila, Colombia / Soil macrofauna and edaphoclimatic conditions in an altitude gradient of coffee growing regions, Huila, Colombia

Introducción: La macrofauna del suelo cumple un rol clave en los procesos del suelo, por lo tanto, regula la oferta se servicios ecosistémicos; sin embargo, su nivel de actividad depende en gran parte de las condiciones edafoclimáticas. Objetivo: El presente trabajo evaluó el efecto del gradiente de altitud sobre comunidades de macrofauna del suelo y propiedades edafoclimáticas en zonas cafeteras del Norte del Huila. Métodos: El estudio incluyó 12 lotes de café separados en dos gradientes de altitud: bajo (1 300-1 600 m.s.n.m) y alto (1 600-1 900 m.s.n.m), se recolectó la macrofauna del suelo mediante monolitos (25 x 25 cm a 30 cm de profundidad) y se estudiaron las condiciones edafoclimáticas. Resultados: En total se registró 9 520 individuos m-2 y una riqueza específica de 14 táxones. Las condiciones edafoclimáticas con mayor diferencia estadística entre los gradientes de altitud, fue la temperatura ambiente y humedad relativa con diferencias de 4.9 °C y 10.4 %, respectivamente. Conclusiones: La mayor abundancia de Coleoptera, Hemiptera, Isoptera y Lepidoptera se presentó en sitios más cálidos, es decir a un gradiente de altitud entre 1 300-1 600 m.s.n.m, mientras que Chilopoda y Diplopoda se adaptaron mejor a sitios más fríos en el gradiente altitudinal entre 1 600-1 900 m.s.n.m.

Introduction: Soil macrofauna plays a key role in soil processes and therefore regulates the supply of ecosystem services; however, its level of activity depends largely of the edaphoclimatic conditions. Objective: This work evaluated the effect of altitude gradient on soil macrofauna communities and edafoclimatic properties in coffee zones of North Huila. Methods: The study included 12 coffee lots separated in two altitude gradients: low (1 300-1 600 masl) and high (1 600-1 900 masl). Soil macrofauna was collected by means of monoliths (25 x 25 cm with 30 cm depth) and edaphoclimatic conditions were studied. Results: A total of 9 520 individuals m-2 and a specific richness of 14 taxa were registered. The edaphoclimatic conditions with the greatest statistical difference between altitude gradients were the environmental temperature and relative humidity with differences of 4.9 °C and 10.4 %, respectively. Conclusions: The highest abundance of Coleoptera, Hemiptera, Isoptera, Lepidoptera was presented in warmer sites, that is, altitude gradient 1 300-1 600 masl, while Chilopoda and Diplopoda adapted better to colder sites in altitude gradient 1 600-1 900 m.a.s.l.

Asian pheretimoid earthworms in North America north of Mexico: An illustrated key to the genera Amynthas, Metaphire, Pithemera, and Polypheretima (Clitellata: Megascolecidae).

The invasion of the pheretimoid earthworms in North America, especially the genera Amynthas and Metaphire, has raised increasing concerns among ecologists and land managers, in turn increasing the need for proper identification. However, the commonly used keys to this group are more than 30 years old with outdated taxonomic information and are based primarily on internal morphology. The requirement of significant amount of taxonomic expertise and dissection, even from the first entry of the key, has prevented broader use of these keys. As a result, many publications in the United States have used Amynthas spp. to represent the group without identifying the species. We present here a new key and diagnoses for the 16 pheretimoid earthworm species recorded in North America north of Mexico, including four genera: Amynthas (10), Metaphire (4), Pithemera (1), and Polypheretima (1). The descriptions were based on published records with modifications following inspection of specimens archived at the National Museum of Natural History, Smithsonian Institution, Washington, D.C. Photos of external and internal characters, including male pores, spermathecal pores, genital markings, spermathecae, prostate glands, and intestinal caeca, are presented to help identification. A summary of current knowledge about the ecology and historical context is provided for each species. We also highlight the previously overlooked and potentially common and widespread co-occurrence of three species-A. agrestis, A. tokioensis, and M. hilgendorfi-and point out that many recent claims of invasion of A. agrestis need to be re-evaluated for potential misidentification.

New species of Dichogaster Beddard, 1888 (Clitellata: Benhamiidae) with additional records of earthworms from Guadeloupe (French West Indies).

A survey of the earthworms of Guadeloupe was undertaken to improve knowledge of the diversity of native and introduced earthworms of the island and to evaluate native species for use in vermicomposting. Twenty-eight species were found, 8 introduced, 4 uncertain, and 16 presumed native. The introduced species are Pontoscolex corethrurus, Pontoscolex spiralis, Amynthas rodericensis, Perionyx excavatus, Polypheretima elongata, Dichogaster affinis, Eudrilus eugeniae, and Drawida sp. Juveniles of the genera Righiodrilus, Glossodrilus and Periscolex were found, but one cannot say if they are introduced or not without a confirmed identification. Omodeoscolex divergens was collected in several locations, with variation in color, size and male sexual function suggestive of long residence on the island, but we are counting this as of uncertain status and tentatively placing all in the same species. Late juveniles of a large endogeic species of a heretofore unknown genus of the Rhinodrilidae were found only in the most remote location sampled and are presumed to be native. Seven previously reported native Dichogaster were found. Eight non-recorded species of Dichogaster were found, five represented by single specimens or juveniles, for which we give only a brief precis. The three described here are Dichogaster spolstoni n. sp., Dichogaster caraibensis n. sp.,.and Dichogaster etaspiqui n. sp. These are compared to and differentiated from previously known Dichogaster species of the world and of Guadeloupe.

The Ovary of Tubifex tubifex (Clitellata, Naididae, Tubificinae) Is Composed of One, Huge Germ-Line Cyst that Is Enriched with Cytoskeletal Components.

Recent studies on the ovary organization and oogenesis in Tubificinae have revealed that their ovaries are small polarized structures that are composed of germ cells in subsequent stages of oogenesis that are associated with somatic cells. In syncytial cysts, as a rule, each germ cell is connected to the central cytoplasmic mass, the cytophore, via only one stable intercellular bridge (ring canal). In this paper we present detailed data about the composition of germ-line cysts in Tubifex tubifex with special emphasis on the occurrence and distribution of the cytoskeletal elements. Using fixed material and live cell imaging techniques, we found that the entire ovary of T. tubifex is composed of only one, huge multicellular germ-line cyst, which may contain up to 2,600 cells. Its architecture is broadly similar to the cysts that are found in other clitellate annelids, i.e. a common, anuclear cytoplasmic mass in the center of the cyst and germ cells that are connected to it via intercellular bridges. The cytophore in the T. tubifex cyst extends along the long axis of the ovary in the form of elongated and branched cytoplasmic strands. Rhodamine-coupled phalloidin staining revealed that the prominent strands of actin filaments occur inside the cytophore. Similar to the cytophore, F-actin strands are branched and they are especially well developed in the middle and outermost parts of the ovary. Microfilaments are also present in the ring canals that connect the germ cells with the cytophore in the narrow end of the ovary. Using TubulinTracker, we found that the microtubules form a prominent network of loosely and evenly distributed tubules inside the cytophore as well as in every germ cell. The well-developed cytoskeletal elements in T. tubifex ovary seem to ensure the integrity of such a huge germ-line cyst of complex (germ cells-ring canals-cytophore) organization. A comparison between the cysts that are described here and other well-known female germ-line cysts is also made.

Immune system participates in brain regeneration and restoration of reproduction in the earthworm Dendrobaena veneta.

Earthworm decerebration causes temporary inhibition of reproduction which is mediated by certain brain-derived neurohormones; thus, cocoon production is an apposite supravital marker of neurosecretory center functional recovery during brain regeneration. The core aim of the present study was to investigate aspects of the interactions of nervous and immune systems during brain regeneration in adult Dendrobaena veneta (Annelida; Oligochaeta). Surgical brain extirpation was combined, either with (i) maintenance of immune-competent coelomic cells (coelomocytes) achieved by surgery on prilocaine-anesthetized worms or (ii) prior extrusion of fluid-suspended coelomocytes by electrostimulation. Both brain renewal and cocoon output recovery were significantly faster in earthworms with relatively undisturbed coelomocyte counts compared with individuals where coelomocyte counts had been experimentally depleted. These observations provide empirical evidence that coelomocytes and/or coelomocyte-derived factors (e.g. riboflavin) participate in brain regeneration and, by implication, that there is close functional synergy between earthworm neural and immune systems.

Three new earthworm species of the genus Polypheretima Michaelsen, 1934 (Oligochaeta: Megascolecidae) from Vietnam.

The paper provides descriptions of three new species of the earthworm genus Polypheretima Michaelsen, 1934 from Dong Nai Province, South Vietnam. They are named Po. cattienensis sp. nov., Po. militium sp. nov., and Po. cordata sp. nov.. All three species are characterized by spermathecal pores in 5/6/7 and the absence of genital markings. Po. cattienensis sp. nov. is distinguished by paired spermathecal pores and seven spermathecae per porus. Po. militium sp. nov. is diagnosed by paired spermathecal pores and a variable number of spermathecae, 21-40 altogether, with 7-17 in 5/6 and 11-23 in 6/7. Po. cordata sp. nov. is recognized by one pair of spermathecal pores in 5/6 and two in 6/7, by only one spermatheca per porus, and by a heart-shaped spermathecal ampulla. 

Earthworms of the 'acaecate' Pheretima group in Vietnam (Oligochaeta: Megascolecidae), with description of a new species from the Mekong delta.

The 'acaecate' Pheretima group from Vietnam is revised, with description of a new species, Polypheretima mekongmontis sp. nov. A total of 18 species of three genera, Metapheretima Michaelsen, 1928, Planapheretima Michaelsen, 1934, and Polypheretima Michaelsen, 1934 has been recorded from Vietnam. Caecate species of Planapheretima are included here as well. Planapheretima arboricola (Rosa, 1890), Pl. lacertina (Chen, 1946), Pl. tenebrica (Chen, 1946), Polypheretima elongata (Perrier, 1872), and Po. taprobanae (Beddard, 1892) are originally recorded from other countries; all remaining species are native to Vietnam. The species Polypheretima tani (Thai, 1996) and Po. tiencanhensis (Pham, 1995) are transferred to the genus Metapheretima. A key to species and distribution maps are also presented.

First record of Dero (Aulophorus) bimagnasetus Harman (Oligochaeta) from Brazil and habitat characteristics / Primeiro registro de Dero (Aulophorus) bimagnasetus Harman (Oligochaeta) do Brasil e notas ecológicas

Annelid worms represent a significant part of freshwater benthic communities worldwide and Oligochaeta is a particularly species-rich group. Dero (A) bimagnasetus (Naididae) previously found and described from a small marsh in Surinam in 1974, has now been found for the first time in Barra Lake, MG, Brazil. Due to the scarce biological data and absence of ecological information in the literature regarding this species we are presenting morphological information on the specimens obtained and the physical and chemical characteristics of the habitat they were found. This species occurred only in the littoral zone of Barra Lake, in muddy, low oxygen, low conductivity and low organic matter sediment. The four individuals collected ranged 3.17-4.15 mm total length; 0.25 - 0.26 mm body width and 0.16-0.21 mm3 total volume. Considering the present anthropic pressures on freshwater biota and fast biodiversity losses worldwide it is now recognized that attention must be paid to low abundance species and the urgency for preservation of their habitats.

Vermes anelídeos representam uma parte significativa das comunidades de água doce em todo o mundo e Oligochaeta é um grupo particularmente rico em espécies. Dero (A) bimagnasetus (Naididae) havia sido encontrado e descrito em um pequeno pântano no Suriname em 1974, e agora também coletado na lagoa Barra, MG, Brasil. Devido ao baixo número de espécimes coletados e à ausência de informações ecológicas sobre esta espécie, estamos apresentando informações sobre a morfometria dos espécimes obtidos e as características físicas e químicas de seu habitat. Esta espécie ocorreu apenas na região litorânea da lagoa da Barra, em sedimento lamacento e com pouca matéria orgânica, baixa concentração de oxigênio e baixa condutividade elétrica na água. Os quatro indivíduos encontrados tiveram 3,17-4,15 mm de comprimento total; 0,25-0,26 mm de largura e 0,16-0,21 mm3 de biovolume. Considerando as atuais pressões antrópicas sobre a biota de água doce e as rápidas perdas de biodiversidade em todo o mundo, é agora reconhecido que muita atenção deve ser dada às espécies pouco abundantes e que é urgentemente necessário preservar seus habitats.

Life history, distribution and abundance of the giant earthworm Rhinodrilus alatus RIGHI 1971: conservation and management implications / História de vida, distribuição e abundância do minhocuçu Rhinodrilus alatus RIGHI 1971: implicações para sua conservação e manejo

Rhinodrilus alatus is an endemic giant earthworm of the Brazilian Cerrado hotspot used as live bait for about 80 years. The goal of this study was to gather ecological data about this species, which will support the establishment of management strategies. The life history, distribution and abundance of R. alatus were investigated in Cerrado, pastures and Eucalyptus plantation areas following the harvesting activities of the local extractors of this species. We found that this earthworm is abundant in all of the sampled areas, showing its resilience to land-use conversion. The Capture Per Unit Effort was 4.4 ± 5 individuals per 100 metres of transect and 5.6 ± 3 individuals per hour. The earthworm's annual cycle is markedly seasonal, with an aestivation period throughout the driest and coldest season of the year. Significant differences in the length and diameter of the body and in the diameter and depth of the aestivation chambers were found between the juveniles and adults. The distribution range of the species was expanded from two to 17 counties. The life history, abundance, distribution and resilience of R. alatus to certain perturbations are key elements to be considered in conservation and management strategies for this species.

Rhinodrilus alatus é um minhocuçu endêmico do hotspot Cerrado, utilizado como isca viva por cerca de 80 anos. O objetivo deste estudo foi obter informações ecológicas sobre essa espécie, as quais servirão para o estabelecimento de estratégias para seu manejo. A história de vida, a distribuição e a abundância de R. alatus foram investigados em áreas de Cerrado, pastagens e plantações de Eucalyptus, acompanhando sua extração pela comunidade local. Rhinodrilus alatus foi abundante em todas as áreas amostradas, mostrando-se resiliente à conversão do Cerrado em outros usos. A Captura por Unidade de Esforço foi de 4.4 ± 5 indivíduos por 100 metros de transecto e 5.6 ± 3 indivíduos por hora. O ciclo anual do minhocuçu é marcadamente sazonal, apresentando estivação no período mais frio e seco do ano. Jovens e adultos diferiram significativamente no comprimento e diâmetro do corpo e no diâmetro e profundidade da câmara de estivação. A distribuição da espécie foi expandida de dois para 17 municípios. A história de vida, a abundância, a distribuição e resiliência de R. alatus por certos distúrbios são elementos chave a serem considerados nas estratégias de conservação e manejo dessa espécie.

Ovary organization and oogenesis in two species of Lumbriculida (Annelida, Clitellata).

The aim of the present study is to describe the organization of the ovary and mode of oogenesis at the ultrastructural level in two representatives of Lumbriculida -Lumbriculus variegatus and Stylodrilus heringianus. In both species studied, the ovaries are small and conically shaped structures that are attached to the intersegmental septum via a thin ligament. The ovaries are composed of germline cysts formed by germ cells interconnected by stable cytoplasmic bridges. As a rule, the cyst center is occupied by a poorly developed anuclear cytoplasmic mass, termed a cytophore, whereas the germ cells are located at the periphery of the cyst. Germline cysts are enveloped by somatic cells. The ovaries of the species studied are polarized, i.e., along the long axis of the ovary there is an evident gradient of germ cell development. The data obtained suggest ovary meroism, i.e., two categories of germ cells were found: oocytes, which continue meiosis, gather nutrients, grow and protrude into the body cavity, and nurse cells, which do not grow and are supposed to supply oocytes with cell organelles and macromolecules via the cytophore. The ovary structure and mode of oogenesis in the species studied were compared with those of other clitellate annelids. As a rule, in all clitellates studied to date, the ovaries are composed of germline cysts equipped with a cytophore and associated with somatic cells; however, the ovary morphology differs between taxa regarding several quantitative and qualitative features. The ovary organization and mode of oogenesis in L. variegatus and S. heringianus strongly resemble those found in Tubificinae and Branchiobdellida studied to date. Our results also support a sister-group relationship between Lumbriculida and a clade comprising ectoparasitic clitellates (i.e., Branchiobdellida, Acanthobdellida and Hirudinida) with Branchiobdellida as a plesiomorphic sister group to Acanthobdellida and Hirudinida.

Características bio-ecológicas en poblaciones de lombrices (Oligochaeta: Glossoscolecidae) de una sabana natural y una protegida en los llanos centrales de Venezuela / Bioecological characteristics of earthworm populations (Oligochaeta: Glossoscolecidae) in a natural and a protected savanna in the central Llanos of Venezuela

In tropical savannas, the earthworm communities have a predominant role since they regulate the soil structure and dynamics of the organic matter. To study the effect on earthworm populations in two differently managed savannas, we compared the general aspects of the biology and ecology of earthworm populations from a 40 years protected savanna (SP) with no fire or cattle raising at the Estación Biológica de los Llanos, Venezuela (EBLL), and a natural savanna (SNI), under normal burning and cattle raising management conditions. Sampling was carried out at the end of the dry season (April), and at the peak of the wet season (July-August). The main physical properties of soils per system were estimated. In each system, in plots of 90x90m, five fixed sampling units were selected at random; and at each sampling point one soil monolith of 25x25x30cm was collected per unit. Earthworms were extracted using the hand sorting extraction method; and the flotation method was used to estimate the density of cocoons. The earthworms were classified in different ecological categories considering their pigmentation, size and depth profile distribution. As a result of the savanna protection, physical parameters were modified in relation to SNI. The SP soils had higher soil moisture when compared to SNI. Soil moisture varied with depth during the dry season since, after the start of the rainy season, the soil was saturated. Field capacity in the SP was greater than that in the SNI. The surface apparent bulk density of soil was lower in the SP respect SNI, reflecting a lower soil compaction. Total average for the density and biomass of earthworms differed greatly, showing higher values in the SP. The earthworm density average in SP ranged between 25.6-85 individuals/m2 and the average biomass between 6.92-23.23g/m2. While in SNI, earthworms were only found in August, with a mean density of 22.40individuals/m2 and a mean biomass of 5.17g/m2. The vertical distribution pattern was only analyzed for the SP, and as in the SNI, no earthworms were found during the dry season; in the SP, a migration of earthworms to deeper layers during the dry season was observed; while in the rainy season they moved to upper soil layers to feed. The systems were characterized by abundance in juvenile Glossoscolecidae family earthworms which might be included in the endo-anecic ecological category. Results suggest that savanna agricultural management can modify an important fraction of pedofauna, particularly their earthworm communities, by modification of soil properties.

Las lombrices de tierra tienen un papel importante en el ciclado de nutrientes y el mantenimiento de la fertilidad de los suelos. En este estudio se empleo el método de extracción manual para determinar la densidad y la biomasa de las lombrices de tierra en diferentes profundidades y en diferentes estaciones, seleccionando al azar cinco monolitos de 25cm2x30cm, ubicados en un área de 90x90m en una Sabana Protegida (SP) y una Sabana Natural Intervenida (SNI). Adicionalmente, se estimó la densidad de los capullos, se analizaron las clases de etapas de desarrollo y se clasificaron los individuos en categorías ecológicas. Se estimó la humedad del suelo, la capacidad de campo, la temperatura, la densidad aparente y la textura del suelo. La SP presentó mejores condiciones físicas en el suelo que la SNI, mientras que la actividad de las lombrices de tierra estuvo limitada a los meses de lluvia y se observó un patrón de migración estacional en el perfil del suelo. En la SNI sólo se observó presencia de individuos en agosto y no se encontró diferencia significativa en sus biomasas con la SP. Los sistemas estuvieron dominados por individuos jóvenes con una categoría ecológica de tipo endo-anécica pertenecientes a la familia.

Tres nuevas especies de Righiodrilus Zicsi 1995 (Annelida, Oligochaeta: Glossoscolecidae) de la Amazonía colombiana / Três novas espécies de Righiodrilus Zicsi 1995 (Annelida, Oligochaeta: Glossoscolecidae) da Amazônia colombiana / Three new species of RighiodrilusZicsi, 1995

Se describieron tres nuevas especies de Righiodrilus Zicsi, 1995 para la Amazonía: R. andake sp. n., R. inga sp. n. y R. muinanei. Con las nuevas adiciones el género suma 26 especies. Se presenta una clave para las especies y algunos comentarios relacionados con las características y presencia en usos del terreno de Righiodrilus.

Three new species of RighiodrilusZicsi, 1995 from Amazonia were described: R. andake sp. n., R. inga sp. n. y R. muinanei sp. n. With these new additions the genus now contains 26 species. A key for species is provided and some remarks related to characteristics and presence of Righiodrilusland uses are furnished.

Expression of PACAP-like compounds during the caudal regeneration of the earthworm Eisenia fetida.

The regeneration of the ventral nerve cord ganglion and peripheral tissues was investigated by radioimmunoassay and immunohistochemistry in the model animal, Eisenia fetida (Annelida, Oligochaeta). It is now well-established that pituitary adenylate cyclase-activating polypeptide (PACAP) is a neurotrophic factor, playing important roles in the development of the nervous system in vertebrate animals. Based on the apparent evolutionary conservation of PACAP and on the several common mechanisms of vertebrate and invertebrate nervous regeneration, the question was raised whether PACAP has any role in the regeneration of the earthworm nervous system. As a first step, we studied the distribution, concentration, and time-course of PACAP-like immunoreactivity during caudal regeneration of both lost segments and the ventral nerve cord ganglia in E. fetida. A strong upregulation of PACAP-like immunoreactivity was observed in most tissues following injury as determined by radioimmunoassay and immunohistochemistry. Significant increases in the concentration of PACAP-like compounds were found in the body wall, alimentary canal, and in coelomocytes. The most characteristic morphological feature was the accumulation of immunolabeled neoblasts in the injured tissues, especially in the ventral nerve cord ganglion that initiates and mediates regeneration processes. Our present results show that PACAP/PACAP-like peptides accumulate in the regenerating tissues of the earthworm, suggesting trophic functions of these compounds in earthworm tissues similarly to vertebrate species.

Changes in the expression of PACAP-like compounds during the embryonic development of the earthworm Eisenia fetida.

Pituitary adenylate cyclase-activating polypeptide (PACAP) is expressed at very early stages in the vertebrate nervous system, and its functions in the embryonic development have been shown by various studies. PACAP is an extremely conserved molecule in phylogeny; however, little is known about its presence and functions in invertebrates. Our previous studies have shown the occurrence of PACAP-like immunoreactivity in the invertebrate nervous system. The aim of this study was to investigate the presence and localization of PACAP-like compounds during the embryonic development of earthworms from cocoon deposition to hatching using immunological methods (radioimmunoassay, dot blot, immunohistochemistry). PACAP-like immunoreactive compounds were detected at very early stages of the embryonic development of the earthworm Eisenia fetida. No significant changes were observed during the early stages in the developing embryo, but a marked increase occurred before hatching. In contrast, during the embryonic development, the level of PACAP-like compounds gradually decreased in cocoon fluids. Immunohistochemistry revealed the presence of PACAP-like immunoreactive cell bodies and processes in the developing body wall, prostomium, pharyngeal wall, and central nervous system. Cells located in the body wall correspond to putative progenitor cells of primary sensory cells. In the present study, we also showed that the clitellum (reproductive organ) of sexually mature worms contained significantly higher levels of PACAP-like immunoreactivity than other regions of the same animals or the clitellar region of a non-reproducing animal. In summary, these observations provide a morphological basis and suggest a role of PACAP(-like peptides) in the reproductive and developmental functions of invertebrates.

Comparative anatomy of PACAP-immunoreactive structures in the ventral nerve cord ganglia of lumbricid oligochaetes.

By means of a whole mount immunocytochemical approach, the distribution patterns of pituitary adenylate cyclase-activating polypeptide (PACAP)-27 and PACAP-38 were identified in the ventral nerve cord (VNC) ganglia of the earthworms Eisenia fetida and Lumbricus terrestris. Each PACAP form appears to occur in a distinct neuron population. Positions of these populations, as well as numbers and sizes of the constituting neurons do not essentially differ between the two species. The data suggest that in Lumbricid Oligochaetes, PACAP-27 and PACAP-38 neuron populations may mediate distinct physiological processes.

Assessing soil ecotoxicity of methyl tert-butyl ether using earthworm bioassay; closed soil microcosm test for volatile organic compounds.

An earthworm bioassay was conducted to assess ecotoxicity in methyl tert-butyl ether (MTBE)-amended soils. Ecotoxicity of MTBE to earthworms was evaluated by a paper contact method, natural field soil test, and an OECD artificial soil test. All tests were conducted in closed systems to prevent volatilization of MTBE out of test units. Test earthworm species were Perionyx excavatus and Eisenia andrei. Mortality and abnormal morphology of earthworms exposed to different concentrations of MTBE were examined. MTBE was toxic to both earthworm species and the severity of response increased with increasing MTBE concentrations. Perionyx excavatus was more sensitive to MTBE than Eisenia andrei in filter papers and two different types of soils. MTBE toxicity was more severe in OECD artificial soils than in field soils, possibly due to the burrowing behavior of earthworms into artificial soils. The present study demonstrated that ecotoxicity of volatile organic compounds such as MTBE can be assessed using an earthworm bioassay in closed soil microcosm with short-term exposure duration.

Sublethal doses of malathion alter male reproductive parameters of Eisenia foetida / Malathion en dosis subletales altera los parámetros reproductivos masculinos de Eisenia foetida

Malathion es un insecticida organofosforado cuyo efecto agudo principal es la inhibición de la enzima acetilcolinesterasa. Este pesticida es usado ampliamente y se ha informado como dañino para la espermatogénesis de ratones. El objetivo de este trabajo fue evaluar el riesgo tóxico de malathion. Gusano de tierra Eisenia foetida (Annelida, Oligochaeta) se usó como modelo biológico y se analizaron testículos y espermatozoides de animales controles y tratados. Se usaron gusanos de tierra con clitelo desarrollado, los cuales fueron aspersados con dosis sub letales de 0, 80, 150, 300 y 600 mg de malathion por kilogramo de tierra (LD50 = 880 mg/kg de tierra seca). A los grupos controles se les agregó solo agua destilada. Cada grupo se analizó a los 1, 5, 15 y 30 días post tratamiento. Los resultados observados en los animales tratados indican un alza significativa en el número de espermatozoides a los 1 y 5 días después del tratamiento, seguido por una disminución espermática entre los 15 y 30 días. El uso de naranja de acridina mostró un aumento significativo de espermatozoides metacromáticos anormales en animales tratados (fluorescencia roja). La histología testicular en animales tratados mostró desorganización del tejido y vacuolización, con la presencia de núcleos pequeños e hipercromáticos, indicativo de posible apoptosis. Además, malathion causó contracción corporal caracterizada por la cola enrollada. Se concluye que malathion provoca una alteración significativa en el recuento y en la calidad espermática, en la histología testicular y en la morfología corporal de Eisenia foetida. Este modelo biológico resulta ser excelente para supervisar el daño citotóxico de pesticidas organofosaforados.

Cadmium effect on the ovarian structure in earthworm Dendrobaena veneta (Rosa).

Cadmium effects on the ovary structure and oocytes were tested in earthworms Dendrobaena veneta exposed to 10 and 50 mg Cd kg(-1) in soil after 10 and 20 days of the experiment. In both experimental doses cadmium caused damage to the structure of the ovary but the effects were different in each group. At 10 mg Cd kg(-1) concentration in soil, young stages of oocytes and trophocytes were most sensitive to cadmium deleterious effects whereas somatic cells in the ovarian stroma were only slightly affected. Cadmium. at a concentration of 50 mg Cd kg(-1) in soil caused most damage in the somatic cells leading to the occurrence of unnaturally swollen elements and desmosomes destruction. At both experimental concentrations cadmium induced degenerative changes in cell nuclei and an increase in number of cell organelles (RER and Golgi complex elements) in the cytoplasm of oocytes and trophocytes. These also proved to be more active. No ultrastructural changes were manifested in oogonia. In both experimental groups degenerative changes occurred as early as after 10 days of Cd exposure.

Cocoon production, morphology, hatching pattern and fecundity in seven tropical earthworm species - a laboratory-based investigation.

Data on the reproductive biology of seven Indian species of earthworms, viz. Perionyx excavatus Perrier, Lampito mauritii Kinberg, Polypheretima elongata (Perrier), Pontoscolex corethrurus (Muller), Eutyphoeus gammiei (Beddard), Dichogaster modiglianii (Rosa) and Drawida nepalensis Michaelsen are presented. The peregrine earthworms such as Perionyx excavatus, Pontoscolex corethrurus, Dichogaster modiglianii, and Polypheretima elongata are considered to be continuous breeders with high fecundity. Native Lampito mauritii and Drawida nepalensis are semi-continuous and Eutyphoeus gammiei discrete breeders. There is a dramatic increase in cocoon production by most earthworm species of Tripura in the summer and monsoon with a corresponding peak during April and July. Cocoon production decreased or ceased during winter. Temperature affected the incubation period of cocoons. With increase in temperature, incubation period increased in the endogeic worms, Pontoscolex corethrurus, Polypheretima elongata and Drawida nepalensis and decreased in the epigeic worms, Perionyx excavatus and Dichogaster modiglianii, within a temperature range between 28-32 degrees C under laboratory conditions. There was a significant (P < 0.05) positive correlation between number of hatchlings per cocoon and incubation period in Lampito mauritii. High rate of cocoon production, short development time with high hatching success, as well as continuous breeding strategies in the epigeic species Perionyx excavatus and Dichogaster modiglianii and the top soil endogeic species, Pontoscolex corethrurus, Drawida nepalensis and Lampito mauritii, indicate their possible usefulness in vermiculture. The giant anecic worm, Eutyphoeus gammiei, which has a very long cocoon development time, discrete breeding strategy and very low rate of cocoon production, is not a suitable species for vermiculture.