Intraspecies genetic variability of Ornithobacterium rhinotracheale in commercial birds in Peru

Strains of the bacterium Ornithobacterium rhinotracheale (ORT), a causal agent of respiratory diseases in birds, were microbiologically isolated, identified, and molecularly characterized. Blood-enriched culture media and biochemistry tests were used for microbiologic identification. Polymerase chain reaction (PCR) and repetitive extragenic palindromic PCR (rep-PCR) techniques were used for molecular identification and characterization, respectively, of the microorganism. ORT strains were isolated in enriched media from the trachea and air sacs of broilers, breeders, and layers from several geographic zones of Peru. Of the original 75 strains isolated from 75 clinical samples from which ORT was recovered during 1998-2000, 25 were selected for further study based on ORT as the primary pathogenic isolate (no other pathogens were detected). Selected isolates were molecularly identified and characterized by PCR using specific primers designed from the conserved zones of the 16S ribosomal genes. Primers used for the identification of ORT produced a specific fragment of 784 base pair (bp), which did not appear in Haemophilus paragallinarum or Pasteurella multocida, microorganisms with similar morphologic and biochemical characteristics that produce dinical signs identical to those of ORT. All 25 strains of ORT tested with rep-PCR had a genetic profile similar to that of ORT American Type Culture Collection 51463, indicating the presence of only one genotype in the ORT strains studied.     

Trichinella: differing effects of antigens from encapsulated and non-encapsulated species on in vitro nitric oxide production

Trichinellosis is a cosmopolitan zoonotic disease affecting a wide variety of animals, including man. Non-encapsulated and encapsulated species diverge with respect to their developmental strategies. Little is known at the molecular level about parasite-derived mediators responsible for host muscle cell transformation occurring during trichinellosis. In this context, host-parasite relationships in Trichinella-infected animals could be related to different host-immune and cell mediators, e.g. nitric oxide (NO). Here, we investigate the stimulatory/inhibitory role of L1 antigens from four encapsulated (T. spiralis, T. britovi, T. nelsoni and T. nativa) and one non-encapsulated (T. pseudospiralis) Trichinella species on NO production from rat macrophages in vitro. Our results demonstrate that encapsulated and non-encapsulated Trichinella species differ in their capacity to stimulate the secretion of NO from host macrophages. Biological significance of these differences should be further assessed in the available experimental models.     

Proximal tibial epiphysiodesis to reduce tibial plateau slope in young dogs with cranial cruciate ligament deficient stifle

The purpose of this study was to evaluate the ability of proximal tibial epiphysiodesis to reduce the tibial plateau slope in young dogs with cranial cruciate ligament (CCL) deficient stifles. Of the 14 treated dogs, eight had a bilateral injury, for a total of 22 joints. After physical and radiographical examination and measurement of tibial plateau slope, all of the dogs underwent surgery. Insertion of the screw was placed in the most proximal part of the tibial plateau, in its medio-lateral centre, aiming to the tibial shaft and using a K wire predriven as a guide; correct position of the screw was confirmed with intraoperative fluoroscopy or radiography. In all of the dogs the tibial slope was decreased at the time of physis fusion and the degree of change depended on the age and the breed of the dog at the time of surgery. The minimum change was 4 degrees and the maximum was 24 degrees. There was a statistically significant difference between tibial slope measured before surgery compared to tibial slope measured at the last follow-up visit after surgery. This study shows that the partial proximal tibial fusion in dogs with ACL injuries was effective in reducing the tibial slope during the residual growing time to such an extent to stabilize the joint, provided that the surgery had been carried out when there was still residual growing potential. The technique appeared to be mini-invasive and malalignment complications could be avoided by correct and precise insertion of the screw.     

The growing prevalence of Nosema ceranae in honey bees in Spain, an emerging problem for the last decade

Microsporidiosis caused by infection with Nosema apis or Nosema ceranae has become one of the most widespread diseases of honey bees and can cause important economic losses for beekeepers. Honey can be contaminated by spores of both species and it has been reported as a suitable matrix to study the field prevalence of other honey bee sporulated pathogens. Historical honey sample collections from the CAR laboratory (Centro Apícola Regional) were analyzed by PCR to identify the earliest instance of emergence, and to determine whether the presence of Nosema spp. in honey was linked to the spread of these microsporidia in honey bee apiaries. A total of 240 frozen honey samples were analyzed by PCR and the results compared with rates of Nosema spp. infection in worker bee samples from different years and geographical areas. The presence of Nosema spp. in hive-stored honey from naturally infected honey bee colonies (from an experimental apiary) was also monitored, and although collected honey bees resulted in a more suitable sample to study the presence of microsporidian parasites in the colonies, a high probability of finding Nosema spp. in their hive-stored honey was observed. The first honey sample in which N. ceranae was detected dates back to the year 2000. In subsequent years, the number of samples containing N. ceranae tended to increase, as did the detection of Nosema spp. in adult worker bees. The presence of N. ceranae as early as 2000, long before generalized bee depopulation and colony losses in 2004 may be consistent with a long incubation period for nosemosis type C or related with other unknown factors. The current prevalence of nosemosis, primarily due to N. ceranae, has reached epidemic levels in Spain as confirmed by the analysis of worker honey bees and commercial honey.     

Occurrence, Distribution and Epidemiology of Grapevine Yellows in Spain

The main viticultural production areas in Spain were surveyed in 1994, 1995 and 1996 for the occurrence and incidence of Grapevine Yellows diseases associated to phytoplasmas. Samples from 300 plants showing symptoms of phytoplasma infection were collected from grapevine fields in the Spanish regions of Aragón, Catalonia and Navarra and analysed by PCR with specific primers for a non-ribosomal DNA of stolbur/Bois Noir (BN) and of Flavescence dorée (FD) phytoplasma. Nested PCR with universal primers P1/P7 and fU5/rU3 was also used. In the survey conducted in 1994 and 1995 only BN/stolbur phytoplasma was detected. The incidence of symptomatic plants was low in five plots of Catalonia from 3% to 18% in 1994 and 1995, respectively, and high in two plots of Navarra, from 60% to 80%. In the survey conducted in 1996 the incidence of symptomatic plants in Catalonia increased (6–80%) due to the presence of FD in five plots in the Northeastern Catalonia. An epidemiological study was carried out in two BN-affected plots of two regions from 1994 to 1997, with the evaluation of potential vectors and of host plants. The stolbur phytoplasma was found in individuals from different insect species belonging to the families Cicadellidae and Delphacidae. Some wild plants naturally infected with stolbur phytoplasma around the infected grapevines were: Convolvulus arvensis, Lavandula officinalis, Polygonum convolvulus, Solanum nigrum, and Thymus officinalis. The incidence of the disease in one BN-infected grapevine plot increased from 3.4% in 1994 to 18.40% in 1997.     

Diversity of the calabash tree (<Emphasis Type="Italic">Crescentia cujete</Emphasis> L.) in Colombia

Germplasm of the calabash tree (Crescentia cujete L.) was collected in five major regions of Colombia, i.e. the Andes, Caribbean, Amazon, Orinoco, and Pacific regions. Collecting this multipurpose tree was guided by the indigenous knowledge of farmers and artisans in each region. Large variation in fruit shapes and sizes was found, of which some forms were typical for certain regions. Overall 56 accessions were collected and roughly classified into 22 types by eight fruit shapes and eight sizes. Molecular markers (Amplified fragment length polymorphisms) were applied to leaf tip tissue originating from vegetatively propagated plants in order to assess the diversity available in the germplasm collected as well as to detect patterns of geographical or morphological similarity. One accession each of C. alata H.B.&K. and C. amazonica Ducke were used as outgroups. Overall, genetic diversity was high (mean Nei and Li’s coefficient of 0.43). No relations could be established between either geographical provenance or fruit morphology and patterns of genetic diversity. Concerning the outgroups, the C. amazonica accession appeared to be a distinct species. The C. alata accession, however, did not seem to be sufficiently distinct from C. cujete to merit species status. The latter material may in fact be a hybrid or serve to challenge the validity of interspecific organization of the genus Crescentia.

In vitro regeneration of Salix nigra from adventitious shoots

Black willow (Salix nigra Marsh.) is the largest and only commercially important willow species in North America. It is a candidate for phytoremediation of polluted soils because it is fast-growing and thrives on floodplains throughout eastern USA. Our objective was to develop a protocol for the in vitro regeneration of black willow plants that could serve as target material for gene transformation. Unexpanded inflorescence explants were excised from dormant buds collected from three source trees and cultured on woody plant medium (WPM) supplemented with one of: (1) 0.1 mg l(-1) thidiazuron (TDZ); (2) 0.5 mg l(-1) 6-benzoaminopurine (BAP); or (3) 1 mg l(-1) BAP. All plant growth regulator (PGR) treatments induced direct adventitious bud formation from the genotypes. The percentage of explants producing buds ranged from 20 to 92%, depending on genotype and treatment. Although most of the TDZ-treated inflorescences produced buds, these buds failed to elongate into shoots. Buds on explants treated with BAP elongated into shoots that were easily rooted in vitro and further established in potting mix in high humidity. The PGR treatments significantly affected shoot regeneration frequency (P < 0.01). The highest shoot regeneration frequency (36%) was achieved with Genotype 3 cultured on 0.5 mg l(-1) BAP. Mean number of shoots per explant varied from one to five. The ability of black willow inflorescences to produce adventitious shoots makes them potential targets for Agrobacterium-mediated transformation with heavy-metal-resistant genes for phytoremediation.     

Fluorescence, browning index, and color in infant formulas during storage

Free and total fluorescent compounds, browning index, and color formation were measured in milk-based powdered infant formulas (IF) during 2 years of storage at 20 and 37 degrees C. The excitation spectra from 415 nm emission show three peaks (ex lambda1 = 270 nm, lambda2 = 325/315 nm, lambda3 = 350 nm) and from 347 nm excitation two emission peaks (415 and 520 nm), and no wavelength shifts were observed. Temperature and time of storage exert in general no significant effect on the development of fluorescence emission intensity and browning index. However, an important increase in pentodilysine was recorded-probably because of the iron and ascorbic acid contents of the samples-as well as in browning index in adapted IF. In both IF a color increase (deltaE) throughout storage was observed, this increase being greater in samples stored at 37 degrees C than in those stored at 20 degrees C. The increase in color with time fitted a linear regression model. Color appeared to be an indicator of sufficient sensitivity to measure the effect of temperature or storage time.     

Transformation of organoarsenical species by the microflora of freshwater crayfish

A study of the transformation of arsenic species by the microflora of the freshwater crayfish Procambarus clarkii was carried out. The study of the degradation of AB (arsenobetaine) was performed in aerobic conditions in two culture media (tryptic soy broth and saline medium) at two temperatures (30 and 8 degrees C). The microflora transformed AB into TMAO (trimethylarsine oxide), DMA (dimethylarsinate), MA (methylarsonate), and an unidentified compound (U1). The quickest transformations were carried out by microflora from hepatopancreas incubated in saline medium at 30 degrees C. The individualized study of other arsenic species [AC (arsenocholine), TETRA (tetramethylarsonium ion), TMAO, DMA, and MA] was also performed in saline medium. The only transformation observed was of AC into AB. The bacteria possibly responsible for AB degradation were isolated, identified by phenotypic and genotypic methods, and individually assayed for AB transformation. Only isolates allocated to the species Pseudomonas putida were able to metabolize AB.