Molecular and phenotypic characterization of Agrobacterium species from vineyards allows identification of typical Agrobacterium vitis and atypical biovar 1 strains.

AIM: To molecularly and phenotypically characterize a selection of Agrobacterium-like isolates from grapevine canes, crowns, soil and tumours in plants grown under cold conditions. METHODS AND RESULTS: Most of the strains were biovar 3 (Agrobacterium vitis), and the remaining were atypical biovar 1 (Agrobacterium tumefaciens). All of them were tumourigenic on grapevine plants but differences in other hosts were observed. Chromosomal and plasmid-borne traits were analysed by gene amplification with four primer sets. Detection of the pectin enzyme hydrolase gene clearly distinguished A. vitis from the atypical A. tumefaciens. Regarding the virulence sensor gene, limited host range tumour-inducing plasmids were found in the atypical isolates. About opine utilization, most A. vitis and some A. tumefaciens contained octopine/cucumopine plasmids, but the nopaline-type was only detected in one A. tumefaciens. CONCLUSIONS: The A. vitis strains were molecularly and phenotypically more homogeneous than those of A. tumefaciens, the latter displaying some typical A. vitis characteristics, suggesting an adaptation to life in grapevine. SIGNIFICANCE AND IMPACT OF THE STUDY: The findings of this work will help to improve detection procedures of the pathogen, and demonstrate the pathogen diversity in cold vineyards, laying the groundwork for epidemiological studies and development of control strategies of the crown and cane gall disease.

Detection and characterization of Pseudomonas syringae pv. actinidifoliorum in kiwifruit in Spain.

AIMS: Bacterial canker of kiwifruit caused by Pseudomonas syringae pv. actinidiae (Psa) is currently the major threat to its commercial production worldwide. In 2011, the most virulent type (Psa3) was detected for the first time in Northwest-Spain, in the province of Pontevedra. In 2013 surveys, leaves and flower buds with mild symptoms were observed in Actinidia deliciosa 'Hayward' vines in an orchard at the province of A Coruña, suggesting the presence of P. syringae pv. actinidifoliorum (Psaf). METHODS AND RESULTS: Isolates obtained from such orchard were characterized by morphological, biochemical and physiological tests, fatty acids (FA) profile and molecular tests (PCR, BOX-PCR, duplex PCR, multiplex PCR, real-time PCR, PCR-C, phytotoxins, housekeeping and effector genes). Pathogenicity tests were also carried out on plants and fruits of A. deliciosa 'Hayward' and on different cultivated plants and fruits. Results demonstrated the presence of P. syringae pv. actinidifoliorum in Spain. CONCLUSIONS: The work provides new information on the pathovar P. syringae pv. actinidifoliorum, which has only been found previously in New Zealand, Australia and France. SIGNIFICANCE AND IMPACT OF STUDY: The results are relevant for taxonomy of isolates of P. syringae from kiwifruit, especially those of low virulence not belonging to pathovar actinidiae.

Intensity of Interprofessional Collaboration and related factors in Intensive Care Units. A descriptive cross-sectional study with an analytical approach.

OBJECTIVE: To determine the Intensity of Collaboration between the intensive care professionals of a third level hospital. METHOD: Descriptive cross-sectional study with an analytical approach. SETTING: 6 intensive care units of a third level hospital. SAMPLE: nurses and doctors. Consecutive type non-probabilistic sampling. DATA COLLECTION: sociodemographic, economic, motivation and professional satisfaction variables, and the intensity of collaboration using the "Scale of Intensity of Interprofessional Collaboration in Health." RESULTS: A total of 102 health professionals (91 nurses and 11 doctors) were included. The mean overall Intensity of Collaboration (IoC) was moderate. Men showed higher scores in all factors (p<.05). The IoC global score was higher in the group of professionals with ≤10 years of experience (p=.043) and those who were highly satisfied with the profession (p=.037). Physicians presented higher scores in the global IdC (p=.037) and in the Collaboration mean (p=.020) independently in the multivariate models. A negative linear relationship (rho: -0,202, p=.042) was observed between age and the overall IoC score. Professionals aged ≤30years reported a higher perception of Shared Activities (p=.031). Negative linear relationships were observed between years of experience and total IoC score (rho: -0,202, p=.042) and patients' Perception score (rho: -0.241, p=0.015). The research activity also showed to be a variable related to a greater degree of Collaboration at a global level and in some of the factors (p<.05). The scale of IoC obtained a Cronbach's α of 0,9. CONCLUSIONS: The intensity of interprofessional collaboration in ICUs is moderate. Professionals with experience of ≤10 years, a higher level of satisfaction and participation in research activities show a greater intensity of collaboration. Doctors perceive collaboration more intensely than nurses. All factors contribute equally to the internal consistency of the questionnaire.

First Report of Bacterial Canker of Kiwifruit Caused by Pseudomonas syringae pv. actinidiae in Spain.

Bacterial canker of kiwifruit caused by Pseudomonas syringae pv. actinidiae was first described in Japan and Korea and is currently an emerging disease that causes major losses in China, Italy, New Zealand, France, Portugal, and Chile. Gold kiwifruit (Actinidia chinensis), especially cvs. Jin Tao and Hort 16A, seem to be more susceptible than green kiwifruit (Actinidia deliciosa) cvs. Hayward and Summer. The bacterium affects male and female woody vines equally, with young vines being more susceptible. The most characteristic symptoms that appear in early spring are reddish orange or white exudates associated with cankers and wounds in branches and/or trunk, as well as brown leaf spots. Buds and fruits were also affected (1). In Spain, 1,132 ha of kiwifruit orchards yielded 25,285 t of fruit in 2009 (2). Most Spanish kiwifruit is cultivated in Galicia (northwest Spain), where the main cultivar is Hayward. In 2010, the first plantation of cv. Jin Tao and one plantation of cv. Summer were established in this area close to Hayward woody vine. In early spring 2011, 80% of the vines in one orchard had twigs with reddish exudates and branches and trunks as well as leaves with angular spots surrounded by yellow haloes. Isolations from both Actinidia spp. were conducted on nutrient agar with sucrose. One hundred and twelve isolates were obtained and seventy-seven were aerobic, gram negative and nonfluorescent on King's B medium. Biochemical tests performed were levan, oxidase, potato rot, arginine didhydrolase, hypersensitivity in tobacco, and utilization of 49 carbohydrates by the API 50 CH system (BioMérieux, Marcy l'Etoile, France). Three PCR protocols were used: two with pathovar-specific primers (PSAF1/PSAR2 and PSAF3/PSAR4) and one with nonspecific primers (PsITSF1/PsITSR2) (3). The results of all biochemical and molecular tests were in agreement with those expected for P. syringae pv. actinidiae. The 16S-23S region of strain EFA 37 isolated from A. deliciosa cv. Summer was sequenced (GenBank Accession No. JF815537) and had 100% sequence identity with P. syringae pv. actinidiae (GenBank Accession Nos. AY342165 and D86357). Pathogenicity tests were performed on 15 plants of A. deliciosa cv. Hayward (five plants per isolate) with the Spanish representative strain EFA 37 and compared with two reference strains isolated from both Actinidia species in Italy and five plants of an untreated control. Three buds per healthy vine were wounded with a sterile needle, inoculated with 30 to 50 µl of each bacterial suspension (108 CFU/ml), sealed, and then covered with plastic. Five leaves per healthy vine were also pierced with a sterile needle and then atomized with the same suspension. Symptoms began to appear after 5 days on inoculated vines, but not on untreated control vines. The bacterium, P. syringae pv. actinidiae, was reisolated from symptomatic plants. The kiwifruit orchard with affected plants was eradicated (25 ha). To our knowledge, this is the first report of P. syringae pv. actinidiae in Spain. References: (1) EPPO Alert List. Online publication. Retrieved from http://www.eppo.org/QUARATINE/Alert_List , June, 2011. (2) Ministerio de Medio Ambiente y Medio Rural y Marino (MARM). Anuario de Estadística, Online Publication. Retrieved from http://www.marm.es/estadistica/pags/anuario/2010 , June 2011. (3) J. Rees-George et al. Plant Pathol. 59:453, 2010.

Wavelength-compensated time-sequential multiplexed color joint transform correlator.

We report a wavelength-compensated three-channel (RGB) joint transform correlator (JTC) for color pattern recognition using a ferroelectric liquid-crystal spatial light modulator (SLM) operating in binary pure phase modulation. We apply a previously reported time-multiplexing technique useful in creating wavelength-compensated diffraction patterns, based on the synchronization of properly scaled diffraction masks with the input wavelength selection obtained by applying a rotating RGB color-filter wheel to an Ar-Kr laser. The application of this technique to a JTC architecture permits real-time color object detection. In order to achieve a high light efficiency for the correlation process, we combine the design of zero-order joint power spectra in all color channels with the selection of a certain polarization configuration of the SLM, producing a broadband phase-only modulation. Excellent experimental results demonstrating color-object detection are provided.

First Report on Almond in Europe of Bacterial Spot Disease of Stone Fruits Caused by Xanthomonas arboricola pv. pruni.

Symptoms characteristic of bacterial spot disease of Prunus spp. (4) were observed on almond trees (Prunus dulcis (Mill.) Webb) in 14 localities of Comunidad Valenciana (eastern Spain) and Aragón (northeastern Spain) between 2006 and 2009. Symptoms were first noted in the spring and were observed until leaf fall. Initial infections began on leaves as small, angular, water-soaked spots, which mainly developed toward the tip and along the leaf margins. These water-soaking lesions were surrounded by chlorotic tissue, although chlorosis did not extend more than a few millimeters. Subsequently, the lesions turned light brown, necrotic, and sometimes the necrotic spots fell out. When the lesions coalesced, they produced large necrotic areas. Sometimes premature leaf drop of infected leaves was observed in severely affected trees. Infected fruits initially displayed sunken, corky lesions that oozed gum, which later became raised when the mesocarp dehydrated. Infected fruits either dropped prematurely or remained on trees after harvest. Cankers typical of bacterial spot disease of stone fruit trees were observed on branches and shoots. Isolations from diseased leaves and fruits yielded Xanthomonas-like colonies on YPGA medium (yeast extract, peptone, and glucose agar), which were subsequently purified and characterized. All strains were gram-negative rods, oxidase negative, and strictly aerobic and showed typical biochemical characteristics of the Xanthomonas genus (3). A collection of 70 strains were further identified by PCR with primers Y17CoF/Y17CoR (1) as Xanthomonas arboricola pv. pruni by comparison with reference strains ISPaVe B4 and ISPaVe B6 isolated from Prunus salicina in Italy. A selection of 46 strains were also analyzed by immunofluorescence (IF) and ELISA using commercial polyclonal antibodies from NEOGEN Europe Ltd. (Ayrshire, Scotland, UK) and SEDIAG S.A.S. (Longvic, France), respectively), although ELISA antibodies proved to be not specific for X. arboricola pv. pruni. Pathogenicity was confirmed by inoculation of 70 almond strains and the reference strains on leaves of potted almond trees and/or on detached leaves (2) with bacterial suspensions (107 CFU per ml). One leaf was inoculated at 8 to 10 sites per strain. Characteristic bacterial spot disease symptoms (4) appeared on all inoculated leaves after 1 week of incubation at 25°C and high humidity, but not on the negative controls infiltrated with sterile distilled water. The original pathogen was reisolated from lesions of inoculated leaves and confirmed by biochemical tests, IF and PCR. As observed in Spain, the disease produces serious damage on the most susceptible almond cultivars like Antoñeta, Guara, Marta, Mas Bovera, and Vayro and can be very harmful, with severity of infection depending upon the relative cultivar susceptibility and environmental conditions. Appropriate eradication measures were taken after the causal agent was confirmed as X. arboricola pv. pruni. This pathogen was previously reported on almond in Japan and New Zealand (4). To our knowledge, this is not only the first report on almond in Spain but also in Europe. References: (1) M. C. Pagani. Ph. D. thesis, North Carolina State University, Raleigh, 2004. (2) P. S. Randhawa and E. L. Civerolo. Phytopathology 75:1060, 1985. (3) L. Vauterin et al. Int. J. Syst. Bacteriol. 45:472, 1995. (4) J. M. Young. N. Z. J. Agric. Res. 20:105, 1977.

Randomized, double-blind, controlled trial, phase III, to evaluate the use of platelet-rich plasma versus hyaluronic acid in hip coxarthrosis. / Ensayo clínico fase III para evaluar la eficacia y seguridad del uso de plasma rico en plaquetas frente a ácido hialurónico en coxartrosis.

AIMS OF THE STUDY: To compare efficacy and safety of a home-made platelet-rich plasma (PRP) solution versus hyaluronic acid in patients with hip osteoarthritis not responding to conservative treatment and to correlate cellular composition of PRP to clinical outcomes. MATERIAL AND METHODS: This is a phase III clinical trial, double-blinded, controlled and randomised into two treatment groups (PRP and hyaluronic acid). Patients received one hip ultrasound-guided injection. Follow up was 12 months. Pain was assessed using VAS score, HHS and WOMAC were used as functional scores, analgesia, adverse events, cellular components (PRP group) in peripheral blood and in PRP were recorded. Clinical response was assessed using OARSI criteria. RESULTS: Seventy-four patients were included. Both groups improved in VAS, WOMAC and HHS score and reduced the amount of analgesia (p<.05). Significant differences were seen at 1 year post-treatment in HHS score (PRP 70.9 [3.7-58], hyaluronic acid 60.2[43-74.2] p<.05). No adverse events were observed in none of the groups. Platelet concentration was different between responders and non-responders (at 1 month, non-responders 449[438-578] x103 platelets/µl versus responders 565 [481-666] x103 platelets/µl, p<.044). There was a correlation between leukocytes concentration and clinical scores (VAS at six months, r=0.748, p<.013, WOMAC at 6 months r=0.748, p <.013). Patients with early stage hip OA showed higher response rate to PRP compared with late stage (11.51 OR, 95%CI 2.34-50.65, p<.03). CONCLUSIONS: Platelet-rich plasma injection improved hip function, reduced pain and the use of analgesia. It is important to bear in mind the cellular composition in order to achieve a better clinical response.

Survival of Erwinia amylovora in mature apple fruit calyces through the viable but nonculturable (VBNC) state.

AIMS: Survival of Erwinia amylovora, causal agent of fire blight in pome fruits and other rosaceous plants, was monitored inside mature apples calyces under some storage conditions utilized in fruit. METHODS AND RESULTS: Apple fruit calyces inoculated with two E. amylovora strains and their respective GFP-marked strains were maintained at 26 degrees and 5 degrees C, and the effect of copper treatment was assayed at 0.01 and 0.1 mmol l(-1) CuSO4. In nontreated apples at 26 degrees C, part of the population of E. amylovora survived in the 'viable but nonculturable' (VBNC) state, whereas at 5 degrees C the majority of the population retained culturability. In copper-treated apples, the whole population adopted the VBNC state irrespective of temperature. Regardless of temperature, copper and inoculum dose, VBNC cells recovered culturability and pathogenicity in King's B broth or by host plant passage. CONCLUSIONS: Erwinia amylovora survived for at least 35 days in mature apple calyces. Besides, the ability of the pathogen in the VBNC state to regain culturability and pathogenicity suggests that the apple fruit could be a potential carrier of E. amylovora contributing to the spreading of fire blight disease. SIGNIFICANCE AND IMPACT OF THE STUDY: The risk of E. amylovora dissemination through mature fruit transport, although low, has been demonstrated, and should be considered in pest risk assessments.

Evaluation of the efficacy of immunomagnetic separation for the detection of Clavibacter michiganensis subsp. michiganensis in tomato seeds.

AIMS: To evaluate the effectiveness of the optimized immunomagnetic separation (IMS)-plating protocol in relation to other culture, serological and molecular techniques currently used for Clavibacter michiganensis subsp. michiganensis in seed-testing laboratories. METHODS AND RESULTS: Bacterial suspensions, tomato seed extracts spiked with the pathogen and naturally infected seeds were IMS-plated for the detection of C. m. subsp. michiganensis. These results were compared with plating on general (YPGA) and semiselective (mSCM) media, double-antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA), immunofluorescent assay (IF) or polymerase chain reaction (PCR). Different seed lots and pathogen strains were also tested. IMS-plating allowed the detection of less than 10 CFU ml(-1) of pathogen in all assayed samples. The mSCM medium provided positive results for 10 CFU ml(-1) in naturally infected seeds, but up to 14 days was necessary for the typical colonies of the target to be come visible. By serological techniques, 10(3) and up to 10(4) CFU ml(-1) were detected by IF and ELISA, respectively. DNA extraction was required to obtain positive results by PCR in seed extracts containing 10(3) CFU ml(-1) or more. CONCLUSIONS: Among the evaluated methods, IMS-plating provided the best results regarding sensitivity and specificity for C. m. subsp. michiganensis detection, allowing the recovery of viable bacteria from seed extracts. SIGNIFICANCE AND IMPACT OF THE STUDY: IMS-plating increases isolation rates of C. m. subsp. michiganensis and could improve standard protocols currently used for routine analysis.

Integrated approach for detection of nonculturable cells of Ralstonia solanacearum in asymptomatic Pelargonium spp. cuttings.

Ralstonia solanacearum (biovar 2, race 3) is a soil and water-borne pathogen that causes serious diseases in several solanaceous hosts. It can also infect geranium plants, posing an important threat to their culture when latently infected cuttings are imported from countries where the pathogen is endemic. R. solanacearum can be present in very low numbers in asymptomatic geranium cuttings, and/or in a particular stressed physiological state that escapes direct isolation on the solid media usually employed. Consequently, an integrated protocol has been developed to analyze asymptomatic geranium cuttings routinely. The first screening tests include isolation and co-operational-polymerase chain reaction (Co-PCR), based on the simultaneous and co-operational action of three primers from 16S rRNA of R. solanacearum. This method was selected as the most sensitive one, able to detect only 1 cell/ml including nonculturable cells. When isolation is negative but Co-PCR is positive, the bioassay in tomato plants is proposed, since stressed bacterial cells or those present in low numbers that do not grow on solid media can be recovered from inoculated tomato plants and retain pathogenicity. This methodology has been demonstrated to be useful and has allowed us to assess the relevance of the physiological status of bacterial cells and its implications in detection. It also reveals the risk of introducing R. solanacearum through asymptomatic geranium material when relying only on bacterial isolation.

Comparative behavior of Ralstonia solanacearum biovar 2 in diverse plant species.

Ralstonia solanacearum causes bacterial wilt in numerous plant species worldwide. Although biovar 2 mostly affects solanaceous crops, identification of new hosts remains a matter of concern since there is still no clear-cut distinction between host and nonhost plants. In this work we provide data based on histological studies on the status of 20 plant species, most of them of potential interest in crop rotation. Plants were watered with a beta-glucuronidase-expressing derivative of R. solanacearum biovar 2, and after a month of incubation, sections of roots and stems were analyzed to localize the pathogen on surface, in cortex and/or xylem. Depending on whether the xylem was colonized or not, plants were classified as hosts or nonhosts, respectively. Hosts generally affected in a few xylem vessels or occasionally in all xylem bundles were classified as tolerant. These included some cabbage, kidney bean, and rutabaga cultivars, and the weed bittersweet nightshade (Solanum dulcamara). Nonhosts were the cultivars tested of alfalfa, barley, black radish, carrot, celery, colocynth, fennel, fiber flax, field bean, field pea, horseradish, maize, and zucchini. However, barley and maize, though nonhosts, may act as reservoirs for the pathogen. The present work constitutes a basis for further studies on cropping systems in fields where R. solanacearum has been detected.

Plant-pathogenic bacteria as biological weapons - real threats?

At present, much attention is being given to the potential of plant pathogens, including plant-pathogenic bacteria, as biological weapons/bioterror weapons. These two terms are sometimes used interchangeably and there is need for care in their application. It has been claimed that clandestine introduction of certain plant-pathogenic bacteria could cause such crop losses as to impact so significantly on a national economy and thus constitute a threat to national security. As a separate outcome, it is suggested that they could cause serious public alarm, perhaps constituting a source of terror. Legislation is now in place to regulate selected plant-pathogenic bacteria as potential weapons. However, we consider it highly doubtful that any plant-pathogenic bacterium has the requisite capabilities to justify such a classification. Even if they were so capable, the differentiation of pathogens into a special category with regulations that are even more restrictive than those currently applied in quarantine legislation of most jurisdictions offers no obvious benefit. Moreover, we believe that such regulations are disadvantageous insofar as they limit research on precisely those pathogens most in need of study. Whereas some human and animal pathogens may have potential as biological or bioterror weapons, we conclude that it is unlikely that any plant-pathogenic bacterium realistically falls into this category.

Mineralized alginate hydrogels using marine carbonates for bone tissue engineering applications.

The search for an ideal bone tissue replacement has led to the development of new composite materials designed to simulate the complex inorganic/organic structure of bone. The present work is focused on the development of mineralized calcium alginate hydrogels by the addition of marine derived calcium carbonate biomineral particles. Following a novel approach, we were able to obtain calcium carbonate particles of high purity and complex micro and nanostructure dependent on the source material. Three different types of alginates were selected to develop inorganic/organic scaffolds in order to correlate alginate composition with scaffold properties and cell behavior. The incorporation of calcium carbonates into alginate networks was able to promote extracellular matrix mineralization and osteoblastic differentiation of mesenchymal stem cells when added at 7 mg/ml. We demonstrated that the selection of the alginate type and calcium carbonate origin is crucial to obtain adequate systems for bone tissue engineering as they modulate the mechanical properties and cell differentiation.

Desmineralización de la Dentina Humana Producida por Irrigantes Endodónticos Combinados

El objetivo de este trabajo fue evaluar el efecto de soluciones de irrigación endodónticas solas y combinadas sobre iones calcio y fosfato de la dentina radicular ex vivo. Se emplearon 56 discos de dentina obtenidos del tercio medio radicular de premolares inferiores unirradiculares extraídos por razones ortodóncicas. Los discos se dividieron al azar en 8 grupos (n=7). Grupo I: agua destilada (AD), Grupo II: hipoclorito de sodio (NaClO) 1 %, Grupo III: EDTA 17 %, Grupo IV: ácido maleico (AM) 5 %, Grupo V: ácido acético (AA) 5 %, Grupo VI: EDTA 17 % + NaClO 1 %, Grupo VII: AM 5 % + NaClO 1 %, Grupo VIII: AA 5 % + NaClO 1 %. Los segmentos de dentina permanecieron en contacto a 37° C durante 5 min y 2,5 minutos en cada solución cuando se usaron en forma sucesiva. Se determinó la concentración de iones calcio de las soluciones mediante espectrometría de absorción atómica y la concentración de iones fosfatos mediante colorimetría (Wienner Lab.). Los resultados se expresaron en mg/ml/gr de tejido. Para el análisis estadístico se utilizó ANOVA y Test de Tukey. AA 5 % y EDTA 17 % se comportaron de manera similar utilizados solos durante 5 minutos, NaClO 1 % no mostró diferencias con el AD. AM 5 % eliminó significativamente más calcio y fosfato que todos los grupos. Todas las soluciones desmineralizaron la dentina, pero AM 5 % durante 5 min fue la solución que más afectó el componente inorgánico de la dentina.

The aim of the present study was to evaluate ex vivo irrigating solutions effect under calcium and phosphates dentin ions, using them alone and combined. In this study 56 dentin discs where used. They were obtained from middle third of mandibular single-root premolars extracted for orthodontics reasons. Discs were randomly divided into 8 groups (n:7). Group I: Distilled water (DW), Group II: 1 % sodium hypochlorite (NaOCl), Group III: 17 % EDTA, Group IV: 5 % maleic acid (MA), Group V: 5 % acetic acid (AA), Group VI: 17 % EDTA + 1 % NaOCl, Group VII: 5 % MA + 1 % NaOCl, Group VIII: 5 % AA + 1 % NaOCl. Dentin segments were kept in contact with irrigating solutions at 37°C for 5 minutes, when used alone, or for 2.5 minutes when used combined. After that, calcium ions (using absorption atomic spectrometry) and phosphorus ions (by colorimetry Wienner Lab.) were determined. Results were expressed in mg/ml/g tissue. Statistical analysis was performed by ANOVA and Tukey test. 5 % AA and 17 % EDTA eliminated similar concentrations of calcium and phosphates ions from dentin at 5 minutes exposure time, while 1 % NaOCl did not present statistical differences with control. 5 % MA eliminated significantly more calcium and phosphates ions than the rest of analyzed groups. Every tested solutions demineralized human dentin, but 5 % MA used for 5 minutes did it the most.

[Henoch-Schönlein purpura. Study of factors associated with the development and course of the disease]. / Púrpura de Schönlein-Henoch. Estudio de factores asociados con el desarrollo y evolución de la enfermedad.

INTRODUCTION: Henoch-Schönlein purpura (HSP) is the most common form of pediatric vasculitis. The objective of this study was to determine the factors associated with the development and course of this disease. PATIENTS AND METHODS: A case-control study was performed. The case group included patients with HSP followed-up at the pediatric rheumatology and nephrology units of a tertiary university hospital over a 2-year period. The control group included children followed-up at the pediatric rheumatology unit for mechanical or non-inflammatory conditions. A medical history including data on infectious conditions and previous medication was taken. A throat culture was performed and antistreptolysin 0 levels were quantified. The seroprevalence of different viruses was investigated. Subsequently, the patients were prospectively followed-up and disease manifestations were compared with reported epidemiological factors. RESULTS: Seventy patients and 58 controls were studied. A history of a recent upper respiratory infection (URI) and antibiotic intake were independently associated with development of HSP. Palpable purpura was present in 100 % of the patients. Gastrointestinal manifestations were recorded in 63 %, articular in 50 %, and renal in 18.6 %. Arthralgias were more frequent in girls and purpura duration was longer when disease onset occurred in spring or summer. Other factors studied were not associated with disease development or with a worse clinical course. CONCLUSIONS: Factors associated with the development of HSP were a history of URI and antibiotic administration. Other epidemiological factors studied were not associated with either the development or the course of the disease.

Factors Affecting Pseudomonas savastanoi pv. savastanoi Plant Inoculations and Their Use for Evaluation of Olive Cultivar Susceptibility.

ABSTRACT Pseudomonas savastanoi pv. savastanoi causes olive knot disease, which is present in most countries where olive trees are grown. Although the use of cultivars with low susceptibility may be one of the most appropriate methods of disease control, little information is available from inoculation assays, and cultivar susceptibility assessments have been limited to few cultivars. We have evaluated the effects of pathogen virulence, plant age, the dose/response relationship, and the induction of secondary tumors in olive inoculation assays. Most P. savastanoi pv. savastanoi strains evaluated were highly virulent to olive plants, but interactions between cultivars and strains were found. The severity of the disease in a given cultivar was strongly dependent of the pathogen dose applied at the wound sites. Secondary tumors developed in noninoculated wounds following inoculation at another position on the stem, suggesting the migration of the pathogen within olive plants. Proportion and weight of primary knots and the presence of secondary knots were evaluated in 29 olive cultivars inoculated with two pathogen strains at two inoculum doses, allowing us to rate most of the cultivars as having either high, medium, or low susceptibility to olive knot disease. None of the cultivars were immune to the disease.

Pregnancy, bovine somatotropin, and dietary n-3 fatty acids in lactating dairy cows: I. Ovarian, conceptus, and growth hormone-insulin-like growth factor system responses.

The objective was to examine effects of bovine somatotropin (bST), pregnancy, and dietary fatty acids on reproductive responses in lactating dairy cows. Beginning at approximately 17 d in milk (DIM), a comparison was made of isoenergetic diets comprising supplementary lipids of whole cottonseed vs. calcium salts of fish oil enriched lipid (FO). Ovulation was synchronized in cows with a presynchronization plus Ovsynch protocol, and cows were inseminated artificially by appointment or not inseminated (d 0 = time of synchronized ovulation; 77 +/- 12 DIM). On d 0 and 11, cows received bST (500 mg) or no bST. All cows were slaughtered on d 17. Number of cows in each group was as follows: control diet had 5 bST-treated cyclic (bST-C), 5 non-bST-treated cyclic (no bST-C), 4 bST-treated pregnant (bST-P), and 5 non-bST-treated pregnant (no bST-P) cows; and cyclic cows fed FO diet had 4 bST-treated (bST-FO) and 5 non-bST-treated cyclic (no bST-FO-C) cows. Feeding FO increased milk production, number of class 1 follicles (2 to 5 mm), and decreased insulin during the period before d 0 compared with control-fed cows. The bST increased milk production, pregnancy rate [83% (5/6) vs. 40% (4/10)], conceptus length (45 vs. 34 cm), and interferon-tau in the uterine luminal flushings (9.4 vs. 5.3 microg) with no effect on interferon-tau mRNA concentration in the conceptus. Treatment with bST increased plasma growth hormone (GH) and insulin-like growth factor (IGF)-I. Among control-fed cows (cyclic and pregnant), bST decreased progesterone concentration in plasma. Cows fed FO had less plasma insulin than control-fed cyclic cows, and FO altered the plasma GH (bST-FO > bST-C) and IGF-I (bST-C > bST-FO-C) responses to bST injections. Endometrial IGF-I mRNA was reduced in pregnant cows and tended to decrease in those fed FO. The IGF-II mRNA was increased in the endometrium of pregnant and bST-treated cows fed the control diet. Cows fed FO had increased concentrations of IGF-II mRNA, when bST was not injected. The insulin-like growth factor binding protein-2 (IGFBP-2) mRNA was increased in bST-P cows, whereas bST decreased the IGFBP-2 mRNA in all cyclic cows. In summary, bST and FO seemed to modulate reproductive responses that may be beneficial to the developing conceptus and pregnancy rate.

[Assessment of protein quality in foods by calculating the amino acids score corrected by digestibility]. / Evaluación de la calidad de las proteínas en los alimentos calculando el escore de aminoácidos corregido por digestibilidad.

The protein score reflects its amino acids (AA) content in comparison with the ideal protein. However, when there is a need to know the use of AA by the organism it is necessary to do a correction of the score value by protein digestibility (PDCAAS). Since this information is not available for usually consumed foods, the present work aimed at calculating the PDCAAS values of these foods. The score was calculated the limiting AA of 70 foods, taking as reference protein the AA pattern for children > 1 year old and adults proposed by the U.S. National Academy of Sciences for the year 2002. The PDCAAS value was obtained in each case by multiplying the score value by the digestibility index. For vegetable foods the obtained score values and PDCAAS were, respectively: vegetables 88.5% / 73.4%, tubercles 89.44% / 74.24%, fresh fruits 75.6% / 64.3%, dried fruits 65.6% / 48.1%, legumes in general 89.2% / 69.58%, chickpea and soybean 100% / 78%, cereals and derivatives 68.8% / 58.5%. Creation of table that contents the score values, digestibility values, and PDCAAS of foods is a useful tool when food selection for a dietary plan based on its protein quality is desirable.

Major cold shock proteins, CspA from Escherichia coli and CspB from Bacillus subtilis, interact differently with single-stranded DNA templates.

The family of bacterial major cold shock proteins is characterized by a conserved sequence of 65-75 amino acid residues long which form a three-dimensional structure consisting of five beta-sheets arranged into a beta-barrel topology. CspA from Escherichia coli and CspB from Bacillus subtilis are typical representative members of this class of proteins. The exact biological role of these proteins is still unclear; however, they have been implicated to possess ssDNA-binding activity. In this paper, we report the results of a comparative quantitative analysis of ssDNA-binding activity of CspA and CspB. We show that in spite of high homology on the level of primary structure and very similar three-dimensional structures, CspA and CspB have different ssDNA-binding properties. Both proteins preferentially bind polypyrimidine ssDNA templates, but CspB binds to the T-based templates with one order of magnitude higher affinity than to U- or C-based ssDNA, whereas CspA binds T-, U- or C-based ssDNA with comparable affinity. They also show similarities and differences in their binding to ssDNA at high ionic strength. The results of these findings are related to the chemical structure of DNA bases.

Cloning, overexpression, purification, and spectroscopic characterization of human S100P.

The calcium-binding protein S100P has been found to be associated with human prostate cancer. We have overexpressed S100P in Escherichia coli using a T7 expression system. A rapid two-step procedure for the isolation of overexpressed S100P leads to a preparation of >95% pure protein with a yield of approximately 150 mg per liter of culture. The structural integrity of recombinant S100P was analyzed using CD and fluorescence spectroscopic techniques. The far-UV CD shows that secondary structure of recombinant S100P consists predominantly of a-helical structure. Both near-UV CD and tyrosine fluorescence spectra show that aromatic residues are involved in the formation of a specific, well packed structure, indicating that the recombinant S100P protein adopts a compact folded conformation. Ca2+ has a profound effect on S100P structure. Near-UV CD and fluorescence intensity of both internal (tyrosine) and external (ANS) probes suggest significant structural rearrangements in the tertiary structure of the molecule. The similarity of far-UV CD spectrum of S100P in the presence and in the absence of Ca2+ suggests that Ca2+ binding has only minor effects on secondary structure.