Midgut of the diplopod Urostreptus atrobrunneus: structure, function, and redefinition of hepatic cells / Intestino médio do diplópodo Urostreptus atrobrunneus: estrutura, função e redefinição das células hepáticas

Abstract Diplopods are considered important macroarthropods the soil as part of its maintenance and balance. These animals usually do not occur in high densities, but population explosions caused by environmental disturbances, climate changes, and use of pesticides that eliminate possible competitors, have been reported. The millipede Urostreptus atrobrunneus Pierozzi and Fontanetti, 2006 have become a nuisance to humans in infestation sites in urban centers of the state of Sao Paulo, Brazil. As a contribution to the understanding of this potential pest, this study describes the histology, histochemistry, and ultrastructure of the U. atrobrunneus midgut, and presents the redefinition of hepatic cells somewhat controversial in the literature. The region of the midgut is characterized by the absence of a cuticular intima, and composed of a pseudostratified epithelium on a thick basal membrane, followed by a muscle layer, a layer of hepatic cells, lined by an external membrane. The morphology observed in U. atrobrunneus is similar to that reported for other species of diplopods. The hepatic cells have been previously described as randomly without forming a layer, however, the present results clearly demonstrate that these cells form a continuous layer over the whole midgut.

Resumo Diplópodos são considerados importantes macro-artrópodes do solo, uma vez que participam de sua manutenção e equilíbrio. Comumente estes animais não apresentam população numerosa, porém há relatos de explosões populacionais ocasionadas por desequilíbrios ambientais, mudanças climáticas e utilização de pesticidas que eliminam possíveis competidores. O milípede Urostreptus atrobrunneus Pierozzi e Fontanetti, 2006, têm apresentado pontos de infestação em centros urbanos, do estado de São Paulo, Brasil, causando muitos transtornos à população humana. Com objetivo de contribuir para o conhecimento desta potencial praga, este trabalho apresenta a descrição histológica, histoquímica e ultra-estrutural do intestino médio do milípede U. atrobrunneus, bem como apresenta a redefinição das células hepáticas, um tanto controversa na literatura pertinente. A região do intestino médio é caracterizada pela ausência da íntima cuticular, sendo formado por um epitélio pseudoestratificado, apoiado por uma membrana basal espessa, seguido de uma camada muscular, uma camada de células hepáticas, revestido por uma membrana externa. A morfologia observada neste trabalho assemelha-se bastante com as descrições de outras espécies de diplópodos estudadas até o momento. As células hepáticas foram previamente descritas como dispostas aleatoriamente sem a formação de uma camada, contudo, os presentes resultados demonstram claramente que estas células formam uma camada contínua ao longo de todo o intestino médio.

Histopathological analysis of tilapia gills (Oreochromis niloticus Linnaeus, 1758) exposed to sugarcane vinasse.

Sugarcane vinasse is one of the main residues generated by the transformation of cane into ethanol. Because of the high organic content (COD), high biochemical oxygen demand (BOD), low pH, the large amount that this residue is generated (15l for every liter of ethanol produced) and their use as fertilizer on the sugarcane crop, this residue is potentially polluting to the soil ecossystem and by percolation to water ecossystem too. Thus, this study aimed to assess the toxicity of vinasse by analyzing Oreochromis niloticus gills exposed to different dilutions (1%, 2.5%, 5% and 10%) in two bioassays. The gills were collected, fixed and analyzed using ultra morphological, histological, and histochemical techniques. After exposure to the vinasse, a statistically significant reduction of the ridges present on the surface of pavimentous cells was observed in one of the bioassays; such structures are responsible for mucus retention, which helps to protect the tissue. In addition, an intumescence of the cells was observed in the treatments with vinasse as well as an increase in the amount of chloridric cells. Some striking tissue changes detected in the treatments were epithelial detachment and loss of integrity of secondary lamellae, causing their rupture and consequent hemorrhage. In the first bioassay, the amount of these changes was statistically significant at the 5% dilution, and the focus of hemorrhage was significant at all dilution ratios. In the second bioassay, the epithelial disorganization was statistically significant only at the 2.5% dilution of vinasse. Moreover, for both bioassays performed, a significant increase in mucous cells was observed when compared with the control. Our results demonstrate the toxic action of sugarcane vinasse, which caused histopathological changes in the exposed animals at all four dilution tested. This highlights the need for caution in the disposal of sugarcane vinasse on the soil, especially due to its capacity for being leached or percolated into water resources, which could seriously damage aquatic fauna.

Midgut of the diplopod Urostreptus atrobrunneus: structure, function, and redefinition of hepatic cells.

Diplopods are considered important macroarthropods the soil as part of its maintenance and balance. These animals usually do not occur in high densities, but population explosions caused by environmental disturbances, climate changes, and use of pesticides that eliminate possible competitors, have been reported. The millipede Urostreptus atrobrunneus Pierozzi and Fontanetti, 2006 have become a nuisance to humans in infestation sites in urban centers of the state of Sao Paulo, Brazil. As a contribution to the understanding of this potential pest, this study describes the histology, histochemistry, and ultrastructure of the U. atrobrunneus midgut, and presents the redefinition of hepatic cells somewhat controversial in the literature. The region of the midgut is characterized by the absence of a cuticular intima, and composed of a pseudostratified epithelium on a thick basal membrane, followed by a muscle layer, a layer of hepatic cells, lined by an external membrane. The morphology observed in U. atrobrunneus is similar to that reported for other species of diplopods. The hepatic cells have been previously described as randomly without forming a layer, however, the present results clearly demonstrate that these cells form a continuous layer over the whole midgut.

Nickel exposure promotes osmoregulatory disturbances in Oreochromis niloticus gills: histopathological and energy dispersive spectrometry analysis.

Water is an essential factor for maintaining the vital functions of living beings. Nickel is the 24th most abundant element on Earth; it is a heavy metal that is genotoxic and mutagenic in its chloride form. Due to industrial use, its concentration in surface sediments increased considerably. Fish develop characteristics that make them excellent experimental models for studying aquatic toxicology. They are particularly useful because they can alert of the potential danger of chemical substances or environmental pollution. Due to water quality impairment and because there are few published studies that relate nickel to tissue alteration, this study aimed to examine the consequences of nickel in an aquatic environment. For this analysis, individuals of Oreochromis niloticus were exposed for 96 h to three different concentrations of nickel dissolved in water according to the standard established by Brazilian law and compared them to a control group. After exposure, the gills were analyzed using X-ray microanalysis, ultramorphology, and histological and histochemical analysis. The results demonstrated that all the concentrations used in the experiment altered the histophysiology of the individuals exposed. In conclusion, the nickel presents a toxic potential to fish, even at the lowest concentration tested, which is equivalent to half of the concentration allowed by law. The CONAMA resolution should be revised for this parameter because of the interference of this metal in the histophysiology of the tested organism.

Comparative study of the ovarian morphology in the order Polydesmida (Diplopoda) and description of unusual structures in the female reproductive system / Estudo comparativo da morfologia ovariana na ordem Polydesmida (Diplopoda) e descrição de estruturas incomuns do sistema reprodutor feminino

Polydesmida is the largest order in the class Diplopoda in terms of the number of species, genera and families, but there are few studies of the ovarian morphology of species in this order. This study aimed to perform a comparative study of the ovarian morphology of three species from this order to increase the understanding of the morphological evolution of this system in Polydesmida. Adults females of two of these species, Poratia salvator and Myrmecodesmus hastatus, belonging to the family Pyrgodesmidae, had a unpaired ovary that formed a tubular organ containing oocytes, with P. salvator present grouping of oocytes into a structure similar to ovisacs. This condition appeared to be apomorphic and was associated with the small size of these species ( 10 mm) and their short life cycle. The third species, Telonychopus klossae (Chelodesmidae), had a large body and an ovary with paired ovisacs, which was symplesiomorphic with other Polydesmida families.

A ordem Polydesmida é a maior da classe Diplopoda em número de espécies, gêneros e famílias, entretanto existem poucos estudos sobre a morfologia ovariana de suas espécies. Esta pesquisa tem como objetivo realizar um estudo comparativo sobre a morfologia ovariana de três espécies dessa ordem a fim de contribuir para discussões sobre a evolução morfológica deste sistema no grupo. Duas dessas espécies, Poratia salvator e Myrmecodesmus hastatus, representantes da família Pyrgodesmidae, apresentam ovário como um órgão tubular único, com a primeira possuindo os ovócitos agrupados em uma estrutura semelhante a ovissacos. Esta condição parece ser uma apomorfia aparentemente associada ao tamanho diminuto dos animais destas espécies (menos de 10 mm) e suas histórias de vida curtas. A terceira espécie, Telonychopus klossae, um quelodesmídeo de tamanho corporal grande, possui um ovário pareado com presença de ovissacos, condição simplesiomórfica semelhante à de outras famílias de Polydesmida.

Comparative study of the ovarian morphology in the order Polydesmida (Diplopoda) and description of unusual structures in the female reproductive system.

Polydesmida is the largest order in the class Diplopoda in terms of the number of species, genera and families, but there are few studies of the ovarian morphology of species in this order. This study aimed to perform a comparative study of the ovarian morphology of three species from this order to increase the understanding of the morphological evolution of this system in Polydesmida. Adults females of two of these species, Poratia salvator and Myrmecodesmus hastatus, belonging to the family Pyrgodesmidae, had a unpaired ovary that formed a tubular organ containing oocytes, with P. salvator present grouping of oocytes into a structure similar to ovisacs. This condition appeared to be apomorphic and was associated with the small size of these species (< 10 mm) and their short life cycle. The third species, Telonychopus klossae (Chelodesmidae), had a large body and an ovary with paired ovisacs, which was symplesiomorphic with other Polydesmida families.

3D reconstruction and scanning electron microscopy of salivary glands of the millipede Rhinocricus padbergi (Verhoef, 1938) (Diplopoda: Spirobolida).

The three-dimensional technique applied to the salivary gland of the millipides Rhinocricus padbergi showed that it is a single structure, dorsolaterally located to the animal gut and composed of two distinct portion: an acinar and a tubular one. The last one opening onto the oral cavity. This work shows that the salivary gland in R. padbergi, although is a single body, it displays two excretory ducts (one in each side of the anterior end of the gut) which has the function of carrying out the secretion produced toward to the oral cavity, contrary to the anterior data registered by other authors who suggested that the salivary gland in diplopod animals would be a paired structures.

A millipede infestation of an urban area of the city of Campinas, Brazil and preliminary toxicity studies of insecticide Bendiocarb® to the Urostreptus atrobrunneus Pierozzi & Fontanetti, 2006 / Infestação de uma área urbana na cidade de Campinas, Brasil, e estudo preliminar da toxicidade do inseticida Bendiocarb® para Urostreptus atrobrunneus Pierozzi & Fontanetti, 2006

Millipede infestations of urban centers have received the attention of several researchers interested in understanding the causes of this phenomenon and to develop control measures. A great numbers of millipede Urostreptus atrobrunneus have invaded a district of Campinas, São Paulo, Brazil, during the spring and summer of the last three years, apparently becoming a nuisance to local residents. The toxicity of Bendiocarb® insecticide for U. atrobrunneus was tested in laboratory. High mortality was observed 24 hours after the insecticide application. However, this insecticide may cause environmental contamination (if used for millipede control).

Infestações de milípedes em centros urbanos têm atraído a atenção de diversos pesquisadores preocupados em desvendar as causas desse fenômeno e em desenvolver métodos de controle desses animais. Este trabalho relata a invasão de grande número de milípedes da espécie Urostreptus atrobrunneus, num bairro da Cidade de Campinas, São Paulo, Brasil, durante as estações de primavera e verão dos últimos três anos, causando transtornos aos moradores do local. O inseticida Bendiocarb® foi testado em condições de laboratório em relação a sua toxicidade para U. atrobrunneu. Após 24h da aplicação ocorreu alta mortalidade dos diplópodos. Entretanto, esse inseticida pode causar contaminação ambiental (se utilizado para o controle de milípedes).

The Allium cepa bioassay to evaluate landfarming soil, before and after the addition of rice hulls to accelerate organic pollutants biodegradation.

Landfarming is a soil bioremediation technology practiced by oil refineries in order to reduce or eliminate hydrocarbons from petroleum sludge. The goal of the current study was to use Allium cepa bioassay to assess landfarming and landfarming with rice hulls amendment before and after hydrocarbons biodegradation assay in the laboratory. Three cytogenetic endpoints were used: mitotic and chromosome abnormalities (MCA), micronucleus (MN) and nuclear buds (NB). Landfarming presented 13.5 g/kg of total petroleum hydrocarbons (TPHs) and caused strong clastogenic and mutagenic effects (p<0.05) in A. cepa. After 108 days of biodegradation, the landfarming reached the rate of 26.30 mmol of CO(2) released, the concentration of TPHs decreased by 27% and there was significant reduction in MCA, MN and NB. Landfarming treated with rice hulls had the highest release of CO(2), 110.9 mmol, associated with a remarkable reduction in TPHs concentration, 59%, and had the highest decrease in MCA, MN and NB (p>0.05). Our findings showed that the use of rice hulls accelerated the biodegradation efficacy of landfarming and reduced their clastogenicity, indicating that supplementary treatments are important to improve the efficiency of bioremediation processes.

Morphometric analysis of a population of diplopods of the genus Rhinocricus Karsch, 1881.

Diplopods belonging to the subclass Helminthomorpha may present one or both leg pairs of the seventh diplosegment modified into structures that aid copulation, called gonopods. These structures are used as a taxonomic trait for the description of most species. In the genus Rhinocricus these structures are closely similar, so that it is difficult to distinguish species only on the basis of this trait. Two species, R. padbergi and R. varians, are found in the same habitat and present gonopods practically identical in shape; together they present a broad colour gradient, ranging from dark brown to light beige. Morphometric data for individuals of the experimental group were submitted to ANOVA and MANOVA, using Hotelling-Lawley Trace and generalized Mahalanobis distances (D2) tests. The results demonstrated a relationship between size and colour, with darker individuals being larger. On the basis of this preliminary analysis, we may suggest that the two species are distinct since dark individuals are distant from medium- and light-coloured individuals according to the D2 values. This seems to indicate a possible polymorphism of individuals belonging to R. padbergi which present close proximity in the values obtained. In all analyses, we observed that the main variables were diameter, length and telson size.

Composition of constitutive heterochromatin of Pseudonannolene strinatii Mauriès, 1974 (Diplopoda, Spirostreptida) analyzed by AT/CG specific fluorochromes

Triple staining with fluorochromes (DA/DAPI/CMA) and C-banding were used to characterize the composition of Pseudonannolene strinatii heterochromatin. C-banding showed C+ bands of different labeling intensity on chromosomes 1 and 2 in some cells. Fluorochrome staining revealed DAPI+ regions corresponding to the C-banding pattern, indicating that the heterochromatin of this species is abundant in AT-rich sequences.

Ultrastructural features of the fat body and oenocytes of Rhinocricus padbergi Verhoeff (Diplopoda, Spirobolida).

The fat body of the diplopod Rhinocricus padbergi is located in two preferential areas of its body: a) immediately below the tegument, denominated parietal, and b) filling the body cavity, close to the viscera, mainly the ovaries and alimentary canal, denominated perivisceral. Ultrastructurally, its cells, the adipocytes, presented varied morphology and contained organelles indicating that they are cells that mainly produce and store lipids and proteins. The presence of cells similar to the oenocytes found in insects was observed for the first time in diplopods, associated to the fat body of R. padbergi. Our observations suggest that this tissue probably maintains activity cycles, since the presence of cells undergoing apoptosis was detected.

Ultrastructural features of the fat body and oenocytes of Rhinocricus padbergi Verhoeff (Diplopoda, Spirobolida).

The fat body of the diplopod Rhinocricus padbergi is located in two preferential areas of its body: a) immediately below the tegument, denominated parietal, and b) filling the body cavity, close to the viscera, mainly the ovaries and alimentary canal, denominated perivisceral. Ultrastructurally, its cells, the adipocytes, presented varied morphology and contained organelles indicating that they are cells that mainly produce and store lipids and proteins. The presence of cells similar to the oenocytes found in insects was observed for the first time in diplopods, associated to the fat body of R. padbergi. Our observations suggest that this tissue probably maintains activity cycles, since the presence of cells undergoing apoptosis was detected.