Daniel Silva Fernandes

Antigenic cross-reactivity and immunogenicity of Bothrops venoms from snakes of the Amazon region

Snakebites are still a critical public health problem in developing countries or isolated areas. In Brazil, the North Region has a high distribution coefficient worsened by the significant number of eventually unreported cases, due to difficulties in access to health services, to the natural geographic barriers and the vast territory. In the Rio Negro area, the species Bothrops atrox, Bothrops brazili, Lachesis muta muta and Bothriopsis taeniata are thought to be the major species responsible for snakebites. The aim of this study was to qualitatively and quantitatively determine the antigenic cross-reactivity and expression of toxins and the immunogenicity of Bothrops venom species of the Amazon and to evaluate the general efficacy of the therapeutic sera. The in vivo assays demonstrated that the defibrinating activity of B. taeniata venom was absent but that the lethal and hemorrhagic properties were more intense than in the B. atrox venom. The results evidence venom variability among the two B. atrox populations from two distinct Amazonian regions, which may reveal a subjacent speciation process. The results point to new aspects that may guide the improvement of anti-Bothropic therapeutic serum.

Axonal and extracellular matrix responses to experimental chronic nerve entrapment

We have analyzed the ultrastructural and histopathological changes that occur during experimental chronic nerve entrapment, as well as the immunohistochemical expression of chondroitin sulfate proteoglycan (CSPG). Adult hamsters (n = 30) were anesthetized and received a cuff around the right sciatic nerve. Animals survived for varying times (5 to 15 weeks) being thereafter perfused transcardially with fixative solutions either for immunohistochemical or electron microscopic procedures. Experimental nerves were dissected based upon the site of compression (proximal, entrapment and distal). CSPG overexpression was detected in the compressed nerve segment and associated with an increase in perineurial and endoneurial cells. Ultrastructural changes and data from semithin sections were analyzed both in control and compressed nerves. We have observed endoneurial edema, perineurial and endoneurial thickening, and whorled cell-sparse pathological structures (Renaut bodies) in the compressed nerves. Morphometrical analyses of myelinated axons at the compression sites revealed: (a) a reduction both in axon sectional area (up to 30%) and in myelin sectional area (up to 80%); (b) an increase in number of small axons (up to 60%) comparatively to the control group. Distal segment of compressed nerves presented: (a) a reduction in axon sectional area (up to 60%) and in myelin sectional area (up to 90%); (b) a decrease in axon number (up to 40%) comparatively to the control data. In conclusion, we have shown that nerve entrapment is associated with a local intraneural increase in CSPG expression, segmental demyelination, perineurial and endoneurial fibrosis, and other histopathological findings.

Experimental chronic entrapment of the sciatic nerve in adult hamsters: an ultrastructural and morphometric study

Entrapment neuropathy is a group of clinical disorders involving compression of a peripheral nerve and interference with nerve function mostly through traction injury. We have investigated the chronic compression of peripheral nerves as an experimental procedure for detecting changes in ultrastructural nerve morphology. Adult hamsters (Mesocricetus auratus, N = 30) were anesthetized with a 25% pentobarbital solution and received a cuff around the right sciatic nerve. Left sciatic nerves were not operated (control group). Animals survived for varying times (up to 15 weeks), after which they were sacrificed and both sciatic nerves were immediately fixed with a paraformaldehyde solution. Experimental nerves were divided into segments based upon their distance from the site of compression (proximal, entrapment and distal). Semithin and ultrathin sections were obtained and examined by light and electron microscopy. Ultrastructural changes were qualitatively described and data from semithin sections were morphometrically analyzed both in control and in compressed nerves. We observed endoneurial edema along with both perineurial and endoneurial thickening and also the existence of whorled cell-sparse structures (Renaut bodies) in the subperineurial space of compressed sciatic nerves. Morphometric analyses of myelinated axons at the compression sites displayed a remarkable increase in the number of small axons (up to 60%) in comparison with the control axonal number. The distal segment of compressed nerves presented a distinct decrease in axon number (up to 40%) comparatively to the control group. The present experimental model of nerve entrapment in adult hamsters was shown to promote consistent histopathologic alterations analogous to those found in chronic compressive neuropathies.

The skull of Hydrodynastes gigas (Duméril, Bibron & Duméril, 1854) (Serpentes: Dipsadidae) as a model of snake ontogenetic allometry inferred by geometric morphometrics

Allometric growth is one of the dominant factors of morphological variation, although this issue is still poorly explored. Using geometric morphometrics, we evaluated and described the ontogenetic allometry in the skull of the snake Hydrodynastes gigas. Furthermore, we described the skull morphology of the species and compared it with its congener, H. bicinctus. We found that 46.91xa0% of shape variation was explained by allometry: smaller centroid sizes were mostly correlated to short postorbital process, medial region of frontals elongated, shortened nasals and elongated parietal, and shortened/oblique supratemporals in relation to the anteroposterior axis of the skull; larger centroid sizes point mainly to postorbital process elongated, elongated nasals and shortened parietals, posterior region of parietal strongly tapered, and elongated/parallel supratemporals in relation to the anteroposterior axis of the skull. In general aspect, the skull of H. gigas differs from the skull of H. bicinctus by being less dorsoventrally compressed. Comparisons between our results and the results of other studies which considered allometry in specific bones of snakes’ skull showed some common patterns, although we found a different correlation between parietal and skull length. These results reinforce the importance of further studies to evaluate the existence of recurrent patterns of allometric growth in the skull of other representatives of this group of snakes. Moreover, the analyses presented herein revealed a significant ontogenetic allometry in the skull of H. gigas and represent the first approach of geometric morphometrics with this goal for snakes.

On the identity of Chironius flavolineatus (Serpentes: Colubridae).

The snake genus Chironius Fitzinger, 1826 is endemic to the Neotropical region, occurring from Honduras to Uruguay and northeastern Argentina. Some species of the genus have taxonomic and/or nomenclatural problems, such as C. flavolineatus which lacks agreement in the literature about its authorship and type locality. Some researchers have been suggesting Jan (1863) as the author of the species since he first described C. flavolineatus based on two specimens. However, other researchers report that Jans description is so incomplete that it is not possible to ascertain what snake he had in mind and therefore suggest Boettger (1885) as the author, since he was the first to provide a detailed description of the species. In the present study one of the syntypes of C. flavolineatus, supposedly destroyed in Second World War, was found. Thus, the taxonomic identity of C. flavolineatus was redefined, its lectotype was designated and the authorship of the taxa is attributed to Jan (1863).

Reproductive Biology and Notes on Natural History of the Side-necked Turtle Acanthochelys radiolata (Mikan, 1820) in Captivity (Testudines: Chelidae)

Abstract Sixteen adult specimens (six males and ten females) of Acantochelys radiolata from different regions of the state of Rio de Janeiro, Brazil, were monitored in captivity from 1996 to 2000 in order to obtain data on species reproductive biology including breeding season, egg laying, egg incubation, and hatchlings. Mating behaviour was observed between November and March and showed a regular pattern constituted by three conspicuous phases: approximation, persecution, and copulation. Sixteen clutches were found between March and July and twelve eggs were found in natural conditions. Hatches occurred between September and November. Hatchlings showed gular region, anterior and posterior members, tail, and edges of carapace and plastron with reddish blotches resting on a black background. This pattern was modified to an orange tonality in the third month of life.

Taxonomic revision of Chironius flavolineatus (Jan, 1863) with description of a new species (Serpentes: Colubridae).

We conducted a taxonomic review of Chironius flavolineatus on the basis of continuous and discrete morphological characters. We recognize a new species which is distinguished from all currently recognized congeners by the following unique combination of characters: first third of body black or dark gray; vertebral stripe yellowish or creamish white distinct from dorsals of nape and extending throughout almost whole body length; head dorsum tan to brown, distinct from background color of first third of body; posterior temporal scales ranging one to four; cloacal shield frequently divided; two to four rows of keeled dorsal scales at midbody; venter ground color gradually darkening towards cloaca; region of medial constriction of hemipenis slightly covered with spinules separating calyces of apex from spines below region of constriction; in lateral view, sulcus spermaticus positioned on convex face of hemipenis; ascending process of premaxilla oblique anteroposteriorly to longitudinal axis of skull; optic fenestrae not exceeding frontoparietal suture; posterior border of supratemporal exceeding braincase; dorsoventral axis of quadrate oblique mesolaterally, moving away from longitudinal axis of skull. Furthermore, we provide data on morphological variation, distribution, and an emended diagnosis for C. flavolineatus.

A new species of Chironius Fitzinger, 1826 from the state of Bahia, Northeastern Brazil (Serpentes: Colubridae).

We describe a new species of Chironius Fitzinger, 1826 from the highlands of Chapada Diamantina, state of Bahia, Brazil. The new species is distinguished from all currently recognized congeners by a unique combination of states of characters on coloration, scale counts, scale ornamentation, and hemipenis. The new species closely resembles Chironius flavolineatus (Jan, 1863) in color pattern, but differs from the later taxon by the presence of two to four posterior temporal scales; cloacal shield entire; six to ten rows of keeled dorsal scales at midbody; ventral scales with posterior dark edges forming conspicuous transverse bars along almost the entire venter; conspicuous dark longitudinal stripes (in zigzag) in the midventral portion of subcaudals; region of medial constriction of hemipenis slightly covered with spinules separating calyces of apex from spines below region of constriction; and sulcus spermaticus situated on convex face of hemipenis in lateral view. The new species is apparently restricted to Chapada Diamantina, corroborating the biological importance of this region from a conservational perspective.

Taxonomic status and morphological variation of Hydrodynastes bicinctus (Hermann, 1804) (Serpentes: Dipsadidae)

Hydrodynastes bicinctus was described with no type material or locality and it has two subspecies currently recognized that are not taxonomically well defined. We tested the validity of the two subspecies through meristic, morphometric, and color pattern characters. Two apparently distinct color patterns of H. bicinctus were noticed, one from the Cerrado open formations and the other from the Amazon rainforest. These aforementioned patterns, however, exhibited a high degree of geographic overlap and many specimens showed a blended pattern. Based on these results we propose synonymizing H. bicinctus schultzi with the nominal taxon. Furthermore, we designate a neotype for the species, present data on geographic distribution, and provide morphological descriptions of the hemipenis, cephalic glands, and skull.

Morphological variation in the Brazilian Radiated Swamp Turtle Acanthochelys radiolata (Mikan, 1820) (Testudines: Chelidae)

The freshwater turtle Acanthochelys radiolata (Mikan, 1820) is endemic to the Atlantic Forest domain in Brazil and few studies have been done on the morphology, geographic variation and taxonomy of this species. In this paper we record the morphological variation, as well as sexual dimorphism and ontogenetic changes in A. radiolata throughout its distribution range. We analyzed 118 morphological characters from 41 specimens, both quantitative and qualitative, and performed statistical analyses to evaluate size and shape variation within our sample. Morphological analysis revealed that A. radiolata is a polymorphic species, especially regarding color and shape. Two color patterns were recognized for the carapace and three for the plastron. Diagnostic characteristics of the species, such as the rounded tubercles on the neck and the shallow dorsal sulcus between the 2nd and 4th vertebral scutes, varied considerably. Acanthochelys radiolata also showed a high level of ontogenetic variation characterized by a change on the color pattern of plastron and limbs starting from the 4th month of life. Sexual size dimorphism was observed for the first time on nine morphometric variables and females showed larger sizes than males. Based on these results we conclude that A. radiolata represents one single polymorphic species distributed in the lowlands of the Brazilian Atlantic Forest from the state of Alagoas to the state of Rio de Janeiro and the south of Minas Gerais state.