Polyclonal antibodies to the coat protein of Apple stem grooving virus expressed in Escherichia coli: production and use in immunodiagnosis

The coat protein gene of Apple stem grooving virus (ASGV) was amplified by RT-PCR, cloned, sequenced and subcloned in the expression vector pMal-c2. This plasmid was used to transform Escherichia coli BL21c+ competent cells. The ASGV coat protein (cp) was expressed as a fusion protein containing a fragment of E. coli maltose binding protein (MBP). Bacterial cells were disrupted by sonication and the ASGVcp/MBP fusion protein was purified by amylose resin affinity chromatography. Polyclonal antibodies from rabbits immunized with the fusion protein gave specific reactions to ASGV from infected apple (Malus domestica) cv. Fuji Irradiada and Chenopodium quinoa at dilutions of up to 1:1,000 and 1:2,000, respectively, in plate trapped ELISA. The ASGVcp/MBP fusion protein reacted to a commercial antiserum against ASGV in immunoblotting assay. The IgG against ASGVcp/MBP performed favorably in specificity and sensitivity to the virus. This method represents an additional tool for the efficient ASGV-indexing of apple propagative and mother stock materials, and for use in support of biological and molecular techniques.

Detection and partial characterization of a carlavirus causing stem necrosis of soybean in Brazil

Stunting and stem necrosis were noticed in soybeans (Glycine max) grown in 2000/2001 in West Central Brazil the same condition was also observed in the following year in plantations as far as 2,000 km from the initial area. Based on transmission (mechanical, graft, insect vector), purification and serology, electron microscopy and molecular studies the causal agent was determined to be a whitefly-borne carlavirus which is possibly related to Cowpea mild mottle virus (CpMMV).

Diagnóstico biológico e molecular e análise da seqüência de nucleotídeos do gene da proteína capsidial de um isolado do Apple stem pitting virus

O Apple stem pitting virus (ASPV) foi detectado por RT-PCR em amostras de cultivares de pereiras européias (Pyrus communis L.) cvs. Starkrimson e Abate Fetel, e asiáticas (P. pyrifolia var. culta) cvs. Kousui e Housui coletadas no início do outono de 2003 em pomar da Estação Experimental da Embrapa Uva e Vinho, Vacaria, RS. Utilizando várias combinações de oligonucleotídeos, foram amplificados fragmentos de DNA de 269 e 1554 pb, este último contendo o gene completo (1131 nt) da proteína capsidial do ASPV. Outro fragmento amplificado de 291 pb compreende parte do gene da polimerase viral. Estes fragmentos constituem-se em um excelente instrumento de diagnóstico do ASPV em pereiras. A comparação das seqüências de nucleotídeos do gene da proteína capsidial do ASPV com seqüências do banco de dados GenBank, revelou identidades de 89% com seqüências de um isolado alemão de macieira e de 85 a 88% com isolados poloneses, de pereiras. A indicadora herbácea Nicotiana occidentalis cv. 37B, inoculada mecanicamente com extrato foliar da cv. Housui, apresentou lesões locais necróticas, necrose foliar marginal e das nervuras. O ASPV também foi detectado por dot-ELISA nas cvs. Abate Fetel e Kousui, na cv. Starkrimson por imunoblot, e em Pyronia veitchii (Trabut) Guill. por enxertia de borbulhas da cv. Abate Fetel infetada.

Foliar anatomy and histochemistry in seven species of Eucalyptus

This work aimed to describe the foliar anatomy of seven species of Eucalyptus, emphasizing the characterization of secretory structures and the chemical nature of the compounds secreted and /or present in the leaves. Anatomical characterization and histochemical evaluation to determine the nature and localization of the secondary compounds were carried out in fully expanded leaves, according to standard methodology. Anatomical differences were verified among the species studied, especially in E. pyrocarpa. Sub-epidermal cavities were the only secretory structures found in the seven species studied, with higher density in E. pellita and lower in E. pilularis. The following compounds were histochemically detected: lipophilic compounds, specifically lipids of the essential or resin-oil type and sesquiterpene lactones found in the lumen of the cavities of the seven species; and hydrophilic compounds, of the phenolic compound type found in the mesophyll of all the species studied and on the epidermis of some of them. The results confirmed the complexity of the product secreted by the cavities, stressing the homogeneous histochemistry nature of these compounds among the species. However, the phenolic compounds results may be an indication of important variations in adaptations and ecological relations, since they show differences among the species.

Modalities for soil preparation and gypsum application in ultisol: stem productivity of sugarcane

The study was conducted in an area of expansion of sugarcane at Vale do Paraná factory in Suzanápolis city - São Paulo (SP), in Brazil, in the northwestern region of the State of São Paulo. It was used the sugarcane variety RB92-5345, 1.5m of spacing between rows, in an Ultisol. The study aimed to evaluate the productivity of sugarcane and first ratoon and some soil chemical attributes in function of soil tillage and application or not of gypsum. The experimental design was randomized blocks with six treatments, in a factorial 3x2 and six replicates, the main treatments were soil tillage with three equipments, moldboard plow, chisel plow, and heavy harrow, and two secondary treatments with application of 1 t ha-1of gypsum and no gypsum. After each harvest of cane, the soil was characterized as to its fertility indicators in layers of 0.0-0.15; 0.15-0.30 and 0.30-0.45m. Differences in values of soil chemical attributes due to the methods of preparation occurred in the sugarcane did not last until the harvest of the 1st ratoon cane, and also did not influence the crop productivity. The gypsum application resulted in higher values ​​of total recoverable sugar (TRS) and the productivity of tons of stems per hectare (TSH) to sugarcane and 1st ratoon cane, respectively, confirming the initial hypothesis.

Establishment of a protocol for obtention of neuronal stem cells lineages from the dog olfactory epithelium

A morphological and cell culture study from nasal mucosa of dogs was performed in order to establish a protocol to obtain a cell population committed to neuronal lineage, as a proposal for the treatment of traumatic and degenerative lesions in these animals, so that in the future these results could be applied to the human species. Twelve mongrel dogs of 60-day aged pregnancy were collected from urban pound dogs in São Paulo. Tissue from cribriform ethmoidal lamina of the fetuses was collected at necropsy under sterile conditions around 1h to 2h postmortem by uterine sections and sections from the fetal regions described above. Isolated cells of this tissue were added in DMEM/F-12 medium under standard conditions of incubation (5% CO², >37ºC). Cell culture based on isolated cells from biopsies of the olfactory epithelium showed rapid growth when cultured for 24 hours, showing phase-bright sphere cells found floating around the fragments, attached on culture flasks. After 20 days, a specific type of cells, predominantly ellipsoids or fusiform cells was characterized in vitro. The indirect immunofluorescence examination showed cells expressing markers of neuronal precursors (GFAP, neurofilament, oligodendrocyte, and III â-tubulin). The cell proliferation index showed Ki67 immunostaining with a trend to label cell groups throughout the apical region, while PCNA immunostaining label predominantly cell groups lying above the basal lamina. The transmission electron microscopy from the olfactory epithelium of dogs revealed cells with electron-dense cytoplasm and preserving the same distribution as those of positive cell staining for PCNA. Metabolic activity was confirmed by presence of euchromatin in the greatest part of cells. All these aspects give subsidies to support the hypothesis about resident progenitor cells among the basal cells of the olfactory epithelium, committed to renewal of these cell populations, especially neurons.

Comparative anatomy of the thigh nerves of Cebus libidinosus (Rylands et al., 2000)

Non-human primates have constituted an important group among animals subjected to various studies. Ethological, evolutionary and paleontological studies have revealed changes in anatomical structures linked to the evolution of primates, considered in studies on the comparative anatomy between Cebus libidinosus and other neotropical monkeys or those from the Old World, and the detailed knowledge on their anatomy may represent an important factor for their preservation and protection when the animals are brought to veterinary clinics after accidents or illnesses. In terms of veterinary importance, sometimes these animals arrive in the veterinary medical clinics after accidents, needing surgery or clinical treatment, but the little data available on anatomy has impaired the correct proceedings. The main justification for studies on C. libidinosus, is due to little information about the anatomy related to C. libidinosus in Brazilian and worldwide scientific literature. In this study, the distribution, enervation and path of the femoral and sciatic nerves of the pelvic limb (thigh) of C. libidinosus were studied and these results were compared with literature on the anatomy of humans, chimpanzees and baboons. In general, the enervation model of the four primates is identical, but in specific terms, the differences in enervations indicate evolution convergence closer to the branch of baboons in the evolutionary tree, and these data corroborate other comparative studies in relation to the same primates to vessels, muscles and nerves. In conclusion, the nerve organization in the thigh of C. libidinosus is identical to baboon, chimpanzee and homo, but more similar to baboon. The specific differences found indicate an ancient phylogenic origin to C. libidinosus and baboons (data corroborated by other studies).

Morphology and topographic anatomy of the spinal cord of the red-footed tortoise (Geochelone carbonaria Spix, 1824)

The aim of this study was to describe the topography of the spinal cord of the red-footed tortoise to establish a morphological basis for applied research in anesthesiology and morphology. Six tortoises from the state of Maranhão (Brazil) that had died of natural causes were used. The common carotid artery was used to perfuse the arterial system with saline solution (heated to 37ºC) and to fix the material with a 20% formaldehyde solution. The specimens were then placed in a modified decalcifying solution for 72 hours to allow dorsal opening of the carapace with a chisel and an orthopedic hammer. Dissection of the dorsal musculature and sectioning of the vertebral arches were performed to access the spinal cord. The results revealed the spinal cord of G. carbonaria to be an elongated, whitish mass that reached the articulation between the penultimate and last caudal vertebrae. The cervical intumescence (Intumescentia cervicalis) was located between vertebral segments C5 and T1, whereas the lumbosacral intumescence (Intumescentia lumbalis) was located between T6 and Ca1.

Morphology and morphometry of feline bone marrow-derived mesenchymal stem cells in culture

Mesenchymal stem cells (MSC) are increasingly being proposed as a therapeutic option for treatment of a variety of different diseases in human and veterinary medicine. Stem cells have been isolated from feline bone marrow, however, very few data exist about the morphology of these cells and no data were found about the morphometry of feline bone marrow-derived MSCs (BM-MSCs). The objectives of this study were the isolation, growth evaluation, differentiation potential and characterization of feline BM-MSCs by their morphological and morphometric characteristics. in vitro differentiation assays were conducted to confirm the multipotency of feline MSC, as assessed by their ability to differentiate into three cell lineages (osteoblasts, chondrocytes, and adipocytes). To evaluate morphological and morphometric characteristics the cells are maintained in culture. Cells were observed with light microscope, with association of dyes, and they were measured at 24, 48, 72 and 120h of culture (P1 and P3). The non-parametric ANOVA test for independent samples was performed and the means were compared by Tukey's test. On average, the number of mononuclear cells obtained was 12.29 (±6.05x10(6)) cells/mL of bone marrow. Morphologically, BM-MSCs were long and fusiforms, and squamous with abundant cytoplasm. In the morphometric study of the cells, it was observed a significant increase in average length of cells during the first passage. The cell lengths were 106.97±38.16µm and 177.91±71.61µm, respectively, at first and third passages (24 h). The cell widths were 30.79±16.75 µm and 40.18±20.46µm, respectively, at first and third passages (24 h).The nucleus length of the feline BM-MSCs at P1 increased from 16.28µm (24h) to 21.29µm (120h). However, at P3, the nucleus length was 26.35µm (24h) and 25.22µm (120h). This information could be important for future application and use of feline BM-MSCs.

Osteology and radiographic anatomy of the hind limbs in Marshdeer (Blastocerus dichotomus)

Abstract: The knowledge of anatomical structures found in wild animals is important for the practice of medical and surgical clinic. Thus, the aim of this study was to describe the osteology and radiographic anatomy of the femur, patella, tibia, fibula, tarsal, metatarsal and phalanges of the Marshdeer Blastocerus dichotomus as a reference for clinical use and species identification. Most structures were similar to those found in domestic animals, with special features of this species. Noteworthy is, for example, the absence of the third trochanter of the femur. Although a ruminant, the Marshdeer has a fibuyla similar to the one described for the horse. B. dichotomus has four fingers on each limb, formed through three phalanges, only the third and fourth finger touch the ground, and the second and fifth finger is rudimentary. It has four proximal and two distal sesamoid bones, and sesamoid bones near the gastrocnemius muscle do not exist.

Leaf blade quantitative anatomy of sugarcane cultivars and clones

The objective of this study was to make a quantitative assess of the anatomic characteristics of leaf blade of the sugarcane cultivars RB855113, SP80-1842, SP80-1816, RB867515 and clone RB957689 presenting different sensitivity to the mixture of sodium trifloxysulfuron + ametryn herbicides. Compared to the other cultivars assessed, RB855113 cultivar, considered more sensitive to the herbicide mixture, presented relevant differences such as greater proportion of bulliform cells, greater tissue proportion in the transverse section of the leaf blade, greater stomata and trichome density on both surfaces, thinner epidermis on the adaxial surface and length of stomata on both surfaces. The external paraclinal wall of the bulliform cells was thinner than in the common epidermis cells in all the genotypes on the adaxial and abaxial surfaces. Multivariate analysis of the data on the variables considered most relevant to explain the herbicide penetration singled out the sensitive RB855113 from the other materials. Such characteristics can explain the greater penetration, and consequently, greater sensitivity of this cultivar to the sodium trifloxysulfuron + ametryn mixture.

Foliar anatomy of the species Lantana camara and L. radula (Verbenaceae)

The species Lantana camara, commonly used as ornamental, has spread worldwide becoming one of the world's most important weeds. To develop new methods of control of this plant, it is essential to distinguish it from other species of the same genus, and this is usually accomplished through taxonomic studies of fertile samples. Considering the similarity between L. camara and L. radula, and the consequent difficulty in distinguishing one from the other when only sterile samples are available, this work aimed to investigate the use of the anatomical characteristics of the leaves of both species as tools for supporting correct classification. The leaves of L. camara and L. radula were anatomically examined by light microscopy and scanning electron microscopy. The major differences were observed in the petiole, which presented secretory idioblasts in L. camara. Secretory idioblasts were observed in the leaf blades of L. camara and Crystalliferou idioblasts were found in L. radula. Glandular and nonglandular trichomes as well as the abaxial surface are different in each species. Such results can support the strategies aiming at the control of L. camara without interfering with L. radula.