A new species of Rineloricaria (Siluriformes: Loricariidae: Loricariinae) from rio Daraá, rio Negro basin, Amazon, Brazil

Rineloricaria daraha, new species, is described from the rio Daraá, tributary of rio Negro, northwestern Amazonas State, Brazil. The new species is diagnosed by having seven branched pectoral-fin rays, finger-like papillae on the lower lip, a large multi-angular preanal plate, and at least four quadrangular plates of variable size surrounding the preanal plate. The new species is known only from rio Daraá and its waterfalls.

Revision of the Neotropical trahiras of the Hoplias lacerdae species-group (Ostariophysi: Characiformes: Erythrinidae) with descriptions of two new species

Hoplias lacerdae was originally described from the rio Ribeira de Iguape, Iporanga, São Paulo State. The Hoplias lacerdae group is defined as containing generally large trahiras with the medial margins of dentaries running parallel to each other and lacking teeth on the basihyal compared to the H. malabaricus group in which the medial margins of the dentaries converge towards the mandibular symphysis and which have teeth on the basihyal. A taxonomic revision of the group based on meristic and morphometric data identified five distinct species: H. lacerdae distributed in the rio Ribeira de Iguape and rio Uruguai; H. intermedius from the rio São Francisco, upper rio Paraná basin, and rio Doce; H. brasiliensis from rivers of the Atlantic Coastal drainage from the rio Paraguaçu to the rio Jequitinhonha; H. australis new species, endemic to the rio Uruguai; and H. curupira new species present in northern South America, including the rios Negro, Trombetas, Tapajós, Xingu, Tocantins and Capim in the Amazon basin, upper rio Orinoco near the rio Casiquiare (Venezuela), and coastal rivers of Guyana and Suriname. A lectotype for Hoplias intermedius and a neotype for H. brasiliensis are designated.

Morfologia dos órgãos genitais masculinos do Jurará Kinosternon scorpioides (Chelonia: Kinosternidae)

Kinosternon scorpioides é uma pequena tartaruga semi-aquática, típica de água doce, de distribuição geográfica bastante diversificada, encontrada no estado do Maranhão, onde é denominada de jurará ou muçuã. Sua carne é uma excelente fonte de proteína e a despeito da legislação vigente, é comercializado nas praias e feiras da cidade de São Luís e consumido nos restaurantes sob a forma de farofa servida em casquinha. Os órgãos genitais do macho foram estudados visando fornecer dados morfológicos da própria espécie, que poderão ser utilizados na biologia reprodutiva voltada para ações de preservação em cativeiro. Compõe-se a amostra de 10 machos adultos, obtidos mediante apreensões do IBAMA-MA (Proc. nº 020.12.002400/99-31, licença nº 002/01), os quais foram eutanaziados conforme normas do Comitê de Ética do Curso de Medicina Veterinária, Universidade Estadual do Maranhão. A cavidade celomática foi aberta e os órgãos fixados em solução aquosa de formaldeído 10%, e posteriormente dissecados. Os testículos possuem formato ovóide e coloração amarelo-ouro. Os epidídimos convolutos estavam aderidos dorsalmente à superfície medial dos testículos, terminando em um pequeno ducto deferente. Os ductos deferentes não forma-ram nenhuma ampola distinta, abrindo-se na cloaca. O pênis sulcado, localizado no assoalho da cloaca, estendeu-se até a cauda, composto de raíz, corpo e glande. A morfologia dos órgãos reprodutivos destes animais assemelha-se aos de outras tartarugas, sugerindo uma morfologia conservada entre as tartarugas.

Flow cytometry as a tool in the evaluation of blood leukocyte function in Chelonia mydas (Linnaeus, 1758) (Testudines, Cheloniidae)

Chelonia mydas is a sea turtle that feeds and nests on the Brazilian coast and a disease called fibropapillomatosis is a threat to this species. Because of this, it is extremely necessary to determine a methodology that would enable the analysis of blood leukocyte function in these sea turtles. In order to achieve this aim, blood samples were collected from C. mydas with or without fibropapillomas captured on the São Paulo north coast. Blood samples were placed in tubes containing sodium heparin and were transported under refrigeration to the laboratory in sterile RPMI 1640 cell culture medium. Leukocytes were separated by density gradient using Ficoll-PaqueTM Plus, Amershan Biociences®. The following stimuli were applied in the assessment of leukocyte function: Phorbol Miristate-Acetate (PMA) for oxidative burst activity evaluation and Zymosan A (Saccharomyces cerevisiae) Bio Particles®, Alexa Fluor® 594 conjugate for phagocytosis evaluation. Three cell populations were identified: heterophils, monocytes and lymphocytes. Monocytes were the cells responsible for phagocytosis and oxidative burst.

Hematological profile of Chelonia mydas (Testudines, Cheloniidae) according to the severity of fibropapillomatosis or its absence

The green turtle Chelonia mydas feeds and nests in the Brazilian coastal area and is considered an endangered species by the World Conservation Union (IUCN 2009) and threatened by the Red List of Brazilian Fauna (Ministério do Meio Ambiente 2009). Fibropapillomatosis is a disease characterized by benign skin tumors (fibropapillomas), and it is one of the main threats to the survival of this species. Studies suggest the involvement of viruses as infectious agents associated with environmental and genetic factors. Blood samples were collected from 45 turtles captured in the coastal area of the state of Sao Paulo, Brazil. From these, 27 were affected by fibropapillomas and 18 were tumor free. Biometrical data on the turtles, size, location and quantity of tumors were recorded. The area occupied by fibropapillomas per animal was calculated and four groups were determined according to severity of the disease or its absence. The objective of the study was to compare hemogram results of the sea turtles classified in these four groups. The lowest hematocrit value was observed in severely affected animals. In the hemoglobin assay, the highest value was observed in the group of tumor free turtles and the lowest, in animals severely affected. Lymphocyte counts and curved carapace length were on the verge of statistical significance.

Detection of metallo-'beta'-lactamases-encoding genes in environmental isolates of Aeromonas hydrophila and Aeromonas jandaei

Aims: To determine the prevalence and expression of metallo-beta-lactamases (MBL)-encoding genes in Aeromonas species recovered from natural water reservoirs in southeastern Brazil. Methods and Results: Eighty-seven Aeromonas isolates belonging to Aeromonas hydrophila (n = 41) and Aer. jandaei (n = 46) species were tested for MBL production by the combined disk test using imipenem and meropenem disks as substrates and EDTA or thioglycolic acid as inhibitors. The presence of MBL genes was investigated by PCR and sequencing using new consensus primer pairs designed in this study. The cphA gene was found in 97.6% and 100% of Aer. hydrophila and Aer. jandaei isolates, respectively, whereas the acquired MBL genes bla(IMP), bla(VIM) and bla(SPM-1) were not detected. On the other hand, production of MBL activity was detectable in 87.8% and 10.9% of the cphA-positive Aer. hydrophila and Aer. jandaei isolates respectively. Conclusions: Our results indicate that cphA seems to be intrinsic in the environmental isolates of Aer. hydrophila and Aer. jandaei in southeastern Brazil, although, based on the combined disk test, not all of them are apparently able to express the enzymatic activity. Significance and Impact of the Study: These data confirm the presence of MBL-producing Aeromonas species in natural water reservoirs. Risk of water-borne diseases owing to domestic and industrial uses of freshwater should be re-examined from the increase of bacterial resistance point of view

Phylogenetic relationships within the speciose family Characidae (Teleostei: Ostariophysi: Characiformes) based on multilocus analysis and extensive ingroup sampling

Background: With nearly 1,100 species, the fish family Characidae represents more than half of the species of Characiformes, and is a key component of Neotropical freshwater ecosystems. The composition, phylogeny, and classification of Characidae is currently uncertain, despite significant efforts based on analysis of morphological and molecular data. No consensus about the monophyly of this group or its position within the order Characiformes has been reached, challenged by the fact that many key studies to date have non-overlapping taxonomic representation and focus only on subsets of this diversity. Results: In the present study we propose a new definition of the family Characidae and a hypothesis of relationships for the Characiformes based on phylogenetic analysis of DNA sequences of two mitochondrial and three nuclear genes (4,680 base pairs). The sequences were obtained from 211 samples representing 166 genera distributed among all 18 recognized families in the order Characiformes, all 14 recognized subfamilies in the Characidae, plus 56 of the genera so far considered incertae sedis in the Characidae. The phylogeny obtained is robust, with most lineages significantly supported by posterior probabilities in Bayesian analysis, and high bootstrap values from maximum likelihood and parsimony analyses. Conclusion: A monophyletic assemblage strongly supported in all our phylogenetic analysis is herein defined as the Characidae and includes the characiform species lacking a supraorbital bone and with a derived position of the emergence of the hyoid artery from the anterior ceratohyal. To recognize this and several other monophyletic groups within characiforms we propose changes in the limits of several families to facilitate future studies in the Characiformes and particularly the Characidae. This work presents a new phylogenetic framework for a speciose and morphologically diverse group of freshwater fishes of significant ecological and evolutionary importance across the Neotropics and portions of Africa.

DESCRIPTION OF THE MALE REPRODUCTIVE SYSTEM OF THE HERMIT CRAB CALCINUS TIBICEN (DECAPODA: ANOMURA: DIOGENIDAE)

We describe the male reproductive system of the intertidal hermit crab Calcinus tihicen, with emphasis on the sexual apparatus, spermatophore, and spermatozoa. The crabs were collected on the rocky shore of Praia Grande Beach, Ubatuba, southeastern Brazil. The morphological analysis, based on 30 specimens, was made with the use of a stereomicroscope, an optical microscope, and scanning and transmission electron microscopes. The male reproductive system is composed of a pair of juxtaposed testes, located dorsally in the pleon. From each testis emerges a vas deferens that links it to the exterior by the gonopores. located on the base of the fifth pair of pereiopods. The vas deferens has three macroscopically distinct regions that contain spermatophores in different stages of maturation. The spermatophore morphology is similar to that of other members of Paguroidea, having a distal, nearly spherical ampulla containing spermatozoa; an approximately cylindrical peduncle and a proximal foot connecting the spermatophores. We describe, for the first time, the variability in the spermatophore morphology and size in the three regions of the vas deferens using the type species of the genus Calcinus. The spermatozoa have three main regions (the acrosomal vesicle, the nucleus, and the cytoplasm). The morphological similarity of the male reproductive system of C. tihicen with previously studied species of Diogenidae is an indicative of complex phylogenetic relationships among the members of the genus.

Patterns of distribution of the hermit crab Loxopagurus loxochelis (Moreira, 1901) (Decapoda, Diogenidae) in two coastal areas of southern Brazil

The present study determined the distribution pattern of the hermit crab Loxopagurus loxochelis by a comparison of catch, depth and environmental factors at two separate bays (Caraguatatuba and Ubatuba) of Sao Paulo State, Brazil. The influence of these parameters on the distribution of males, non- ovigerous females and ovigerous females was also evaluated. Crabs were collected monthly, over a period of one year (from July/2002 to June/2003), in seven depths, from 5 to 35 m. Abiotic factors were monitored as follows: superficial and bottom salinity (psu), superficial and bottom temperature (C), organic matter content (%) and sediment composition (%). In total, 366 hermit crabs were sampled in Caraguatatuba and 126 in Ubatuba. The highest frequency of occurrence was verified at 20 m during winter (July) in Caraguatatuba and 25 m during summer (January) in Ubatuba. The highest occurrences were recorded in the regions with bottom salinities ranging from 34 to 36 psu, bottom temperatures from 18 to 24 C and, low percentages of organic matter, gravel and mud; and large proportion of sand in the substrate. There was no significant correlation between the total frequency of organisms and the environmental factors analyzed in both regions. This evidence suggests that other variables as biotic interactions can influence the pattern of distribution of L. loxochelis in the analyzed region, which is considered the limit of the northern distribution of this species.

Genetic variability of the Amazon River prawn Macrobrachium amazonicum (Decapoda, Caridea, Palaemonidae)

The freshwater prawn Macrobrachium amazonicum is widely distributed in South America, and occupies habitats with a wide range of salinities. Several investigations have revealed the existence of wide intraspecific variability among different populations, although the understanding of this variability is still fragmentary and incomplete. We compared and characterized inland and coastal populations of M. amazonicum from Brazil, using molecular data (16S and COI mtDNA) to describe the degree of variability, structure, and relationships among them. Genetic divergence rates among populations showed variability at the intraspecific level. All the analyses evidenced significant genetic divergence among populations, structuring them in three groups: I-inland waters of the Amazonian Hydrographic Region (HR); II-Parana/Paraguay HR; and III-coastal systems of northern and northeastern Brazil. Phylogenetic reconstructions revealed that the populations form a single monophyletic clade, which supports their characterization as a single species. Clade I was a sister clade of that formed by clades II and III, which were themselves sister clades. Populations from Sertaozinho/Miguelopolis and Avare, introduced into the state of Sao Paulo, may have originated from natural populations in the states of Mato Grosso do Sul and Para, respectively. Geographical isolation probably contributed to the observed variation, and if this isolation continues. M. amazonicum may undergo speciation within its broad geographical distribution. The sequences obtained here can be used as name-tags for population identification, and the DNA barcodes are useful to identify the origin of specimens used in different freshwater-prawn cultures or introduced populations of unknown origin.

INTERPOPULATION COMPARISON OF REPRODUCTION OF THE ATLANTIC SHRIMP HIPPOLYTE OBLIQUIMANUS (CARIDEA: HIPPOLYTIDAE)

The genus Hippolyte is represented by typically small shrimps with intriguing mechanisms of reproduction. In order to study possible variability in reproductive aspects among different populations, we conducted an exhaustive comparative study of H. obliquimanus from South (Brazil) and Central American (Costa Rica) waters. The study focuses on fecundity and reproductive output. Mean size of ovigerous females was significantly larger, and both mean reproductive output and mean fecundity were significantly higher in specimens from Costa Rica then in those collected in Brazil. Embryo volume was significantly smaller in the Costa Rican population, and in both populations embryos doubled their volume during embryogenesis. We discuss and compare our findings with the information available regarding H. obliquimanus and other hippolytid shrimp. The reproductive traits of both populations of H. obliquimanus show some important differences which may reflect adaptations to local environmental conditions, demonstrating a high plasticity of reproductive features of the species in Brazilian and Costa Rican waters.

LIFE CYCLE AND POPULATION STRUCTURE OF AEGLA PAULENSIS (DECAPODA: ANOMURA: AEGLIDAE)

We describe growth, longevity, sex ratio, reproductive period, and recruitment of Aegla paulensis from Jaragua Stale Park, Sao Paulo, Brazil (23 degrees 27'27.9 '' S; 46 degrees 45'32.3 '' W). The population was sampled monthly (September 2007 through August 2009) with the aid of traps. Over five thousand individuals were captured, sexed, measured (carapace length = CL) and inspected for reproductive traits (females only), and then released back to the sampling site. The pattern of the reproductive cycle was strongly seasonal (austral mid autumn through late winter), with a single recruitment pulse per year. The obtained von Bertalanffy growth equations were CL = 21.25[1-e(-0.041(t + 1.250))] and CL = 16.52[1-e(-0.049(t + 1.823))] for males and females, respectively. Males (mean CL +/- SD = 11.86 +/- 2.79 mm) attain larger sizes than females (mean CL +/- SD = 10.84 +/- 2.36 mm). Aegla paulensis reproduces twice during an estimated life span of 40.2 months for females and 33.9 months for males. Temporal variation of sex ratio showed a distinctive pattern characterized by a sequence of three distinct periods that repeated from one year to another, and which suggested that a behavioral component influence the proportion of sex in adult specimens sampled with traps during reproductive and non-reproductive periods.

REPRODUCTIVE BIOLOGY IN FEMALES OF AEGLA STRINATII (DECAPODA: ANOMURA: AEGLIDAE)

Females of Aegla strinatii (n = 466) were sampled monthly (September 2003 to September 2005) by means of sieves and traps from Rio das Ostras (24 degrees 38'16.2 '' S; 48 degrees 24'05.2 '' W), at Jacupiranga State Park, South of Sao Paulo State, Brazil. The reproductive period was markedly seasonal (from May to September) encompassing the Austral late autumn through late winter. This is in accordance to the pattern of reproductive period variations in relation to the latitudinal climate variability verified in species of Aegla. The proportion of adult females exhibiting the ovigerous condition was higher in young/small specimens as compared to old/large ones, and suggests the occurrence of senescence in the latter group. Average size at the onset of functional maturity in females was estimated as 16.66 mm of carapace length (rostrum excluded). The number of eggs per ovigerous females ranged from 1 to 325. Eggs are slightly elliptical and average size varied according to embryonic stage. Mean (+/- standard deviation) carapace length of juveniles (n = 118) was 1.50 +/- 0.05 mm (range: 1.40-1.65mm).

REPRODUCTIVE BIOLOGY AND FUNCTIONAL MATURITY IN FEMALES OF AEGLA FRANCA (DECAPODA: ANOMURA: AEGLIDAE)

We describe the reproductive period. fecundity, and average size at the onset of functional maturity of female Aegla franca, the northernmost distributed aeglid species. The reproductive period is markedly seasonal and takes place front May (austral mid-autumn) to August (late winter). Ovigerous females appear quite abruptly in the population by May, and this condition is observed in all adult females sampled regardless of their size. The average size at the onset of functional maturity in females, at which 50% of the females sampled during the reproductive period were considered adults, was 12.75 mm CL. The smallest post-ovigerous female measured 12.06 mm carapace length (CL). Mean fecundity (+/- S.D.) from 41 females bearing early and intermediate eggs was 129.1 +/- 32.2 and corresponded to a mean female CL of 14.11 mm. The elliptical-shaped eggs exhibited significant increase in size along the development stages. The third pair of pleopods bore higher number of eggs than the others. Compiled information regarding the reproductive period reported for aeglids revealed all increase in the breeding period length with latitude. The reproductive period tends to be shorter in localities under larger rainfall variation and smaller temperature variability than in sites with opposite climate conditions. Eggs tend to be fewer in number and larger in size towards lower latitudes. We present an hypothesis that stream water velocity might act as a major selective pressure during the early life history of fluvial aeglids with direct effect on the reproductive pattern.