[Nanotechnology applied to the transport of antibiotics in orthopedics and traumatology]. / Nanotecnología aplicada a la transportación de antibióticos en ortopedia y traumatología.

Bone infection and implants are a real problem in orthopedics. The formation of biofilm as well as multi-existing pathogens to antibiotics, make fighting them a difficult challenge with the tools we have today. With the aim of knowing the current state of nanotechnology applied to the transport of antibiotics in traumatology and orthopedics, and their projection in the future. We conducted a bibliographic review in June 2019. While much development of the topic and work on humans is lacking, experimental studies show that nanotechnology applied to antibiotic transport promises to be an important weapon in the treatment of bone infections in the future.

La infección ósea y de los implantes son un verdadero problema en traumatología y ortopedia. La formación de biofilm, así como patógenos multirresistentes a antibióticos hacen que combatirlas sea un difícil reto con las herramientas que hoy tenemos. Con el objetivo de conocer el estado actual de la nanotecnología, aplicada a la transportación de antibióticos en traumatología y ortopedia y su proyección a futuro, realizamos una revisión bibliográfica en Junio de 2019. Si bien falta mucho desarrollo del tema y trabajos sobre humanos, los estudios experimentales muestran que la nanotecnología aplicada a la transportación de antibióticos promete ser un arma importante en el tratamiento de las infecciones óseas a futuro.

Técnicas «Lean» para la mejora del flujo de los pacientes críticos de una región sanitaria con epicentro en el servicio de medicina intensiva de un hospital de referencia / Lean techniques to improve the flow of critically ill patients in a health region with its epicenter in the intensive care unit of a reference hospital

OBJETIVO: Analizar si la aplicación de técnicas «Lean» mejora el flujo de pacientes críticos de una región sanitaria, tomando como epicentro el servicio de medicina intensiva (UCI) del hospital de referencia. DISEÑO: Estudio observacional con análisis pre y postintervención. ÁMBITO: UCI del hospital de referencia. PACIENTES: Diseñamos proyectos y un mapa de flujo y comparamos características pre y postintervención. INTERVENCIONES: Registramos datos demográficos, de traslados de pacientes por el SEM por falta de camas y los tiempos de demora en la hora de alta de la UCI a planta de hospitalización. Realizamos reuniones multidisciplinarias y panel visual diario, con priorización de altas de UCI. Promovimos la reubicación temporal de pacientes críticos en otras áreas especiales del hospital. Cuestionario de satisfacción profesional con valoración pre y postintervención. Análisis estadístico de las comparaciones pre y postintervención. RESULTADOS: Se planificó durante 2013 y se implementó de forma progresiva en 2014. Las medidas principales fueron: 1) análisis de la entrada de pacientes al flujo del proceso de críticos, evaluando los pacientes que deben trasladarse por falta de camas, centrados en un diagnóstico y un área: 10/22 pre vs. 3/21 post (p = 0,045); 2) análisis del tiempo de demora en la hora de alta de UCI a planta de hospitalización: 360,8 ± 163,9 min en el primer periodo vs. 276,7 ± 149,5 en el segundo (p = 0,036); y 3) cuestionario de satisfacción profesional personal, con 6,6 ± 1,5 puntos pre vs. 7,5 ± 1,1 en post (p = 0,001). Análisis de los indicadores de UCI, como son las infecciones adquiridas, los días de estancia, la tasa de reingresos y la mortalidad, sin diferencias significativas entre ambos periodos. CONCLUSIONES: La aplicación de técnicas ‘Lean’ en el proceso de críticos tuvo un impacto positivo en la mejora del flujo de pacientes dentro de la región sanitaria, observando una disminución de los traslados fuera de la región por falta de camas, una reducción en la demora del alta de UCI a hospitalización convencional y un aumento de la satisfacción de los profesionales de la UCI de referencia

OBJECTIVE: To analyze whether the application of Lean techniques to improve the flow of critically ill patients in a health region with its epicenter in the intensive care unit (ICU) of a reference hospital. DESIGN: Observational study with pre and post intervention analysis. SETTING: ICU of a reference hospital. PATIENTS: We design projects and a value stream map of flow and compared pre and post intervention. INTERVENTIONS: We recorded demographic data, patient transfers by EMS for lack of beds and delay times in the discharge from ICU to ward. Multidisciplinary meetings and perform daily visual panel, with high priority ICU discharge. We promote temporary relocation of critically ill patients in other special areas of the hospital. We performed a professional satisfaction questionnaire with pre and post implementation of process. We make a statistical analysis of pre and post-intervention comparisons. Results: We planned for 2013 and progressively implemented in 2014. Analysis of patients entering the critical process flow 1) evaluate patients who must transfer for lack of beds, focusing on a diagnosis: pre 10/22 vs. 3/21 post (P = .045); 2) analysis of time delay in the discharge from the ICU to ward: 360.8 ± 163.9 minutes in the first period vs. 276.7 ± 149.5 in the second (P = .036); and 3) personal professional satisfaction questionnaire, with 6.6 ± 1.5 points pre vs. 7.5 ± 1.1 in post (P = .001). Analysis of indicators such as the ICU acquired infections, length of ICU stay, the rate of re-admissions and mortality, with no significant differences between the two periods. Conclusions: The application of Lean techniques in the critically ill process had a positive impact on improving patient flow within the health region, noting a decrease of transfers outside the region due to lack of beds, reduced delayed discharge from ICU to conventional ward and increased satisfaction of ICU professionals

Guanylin and uroguanylin stimulate lipolysis in human visceral adipocytes.

BACKGROUND/OBJECTIVES: Uroguanylin and guanylin are secreted by intestinal epithelial cells as prohormones postprandially and act on the hypothalamus to induce satiety. The impact of obesity and obesity-associated type 2 diabetes (T2D) on proguanylin and prouroguanylin expression/secretion as well as the potential role of guanylin and uroguanylin in the control of lipolysis in humans was evaluated. SUBJECTS/METHODS: Circulating and gastrointestinal expression of proguanylin (GUCA2A) and prouroguanylin (GUCA2B) were measured in 134 subjects. In addition, plasma proguanylin and prouroguanylin were measured before and after weight loss achieved either by Roux-en-Y gastric bypass (RYGB) (n=24) or after a conventional diet (n=15). The effect of guanylin and uroguanylin (1-100 nmol l(-1)) on lipolysis was determined in vitro in omental adipocytes. RESULTS: Circulating concentrations of prouroguanylin, but not proguanylin, were decreased in obesity in relation to adiposity. Weight loss achieved by RYGB increased plasma proguanylin and prouroguanylin. Obese T2D individuals showed higher expression of intestinal GUCA2A as well as of the receptors of the guanylin system, GUCY2C and GUCY2D, in omental adipocytes. The incubation with guanylin and uroguanylin significantly stimulated lipolysis in differentiated omental adipocytes, as evidenced by hormone-sensitive lipase phosphorylation at Ser563, an increase in fatty acids and glycerol release together with an upregulation of several lipolysis-related genes, including AQP3, AQP7, FATP1 or CD36. CONCLUSIONS: Both guanylin and uroguanylin trigger lipolysis in human visceral adipocytes. Given the lipolytic action of the guanylin system on visceral adipocytes, the herein reported decrease of circulating prouroguanylin concentrations in obese patients may have a role in excessive fat accumulation in obesity.

Lean techniques to improve the flow of critically ill patients in a health region with its epicenter in the intensive care unit of a reference hospital. / Técnicas «Lean¼ para la mejora del flujo de los pacientes críticos de una región sanitaria con epicentro en el servicio de medicina intensiva de un hospital de referencia.

OBJECTIVE: To analyze whether the application of Lean techniques to improve the flow of critically ill patients in a health region with its epicenter in the intensive care unit (ICU) of a reference hospital. DESIGN: Observational study with pre and post intervention analysis. SETTING: ICU of a reference hospital. PATIENTS: We design projects and a value stream map of flow and compared pre and post intervention. INTERVENTIONS: We recorded demographic data, patient transfers by EMS for lack of beds and delay times in the discharge from ICU to ward. Multidisciplinary meetings and perform daily visual panel, with high priority ICU discharge. We promote temporary relocation of critically ill patients in other special areas of the hospital. We performed a professional satisfaction questionnaire with pre and post implementation of process. We make a statistical analysis of pre and post-intervention comparisons. RESULTS: We planned for 2013 and progressively implemented in 2014. Analysis of patients entering the critical process flow 1) evaluate patients who must transfer for lack of beds, focusing on a diagnosis: pre 10/22 vs. 3/21 post (P=.045); 2) analysis of time delay in the discharge from the ICU to ward: 360.8±163.9minutes in the first period vs. 276.7±149.5 in the second (P=.036); and 3) personal professional satisfaction questionnaire, with 6.6±1.5 points pre vs. 7.5±1.1 in post (P=.001). Analysis of indicators such as the ICU acquired infections, length of ICU stay, the rate of re-admissions and mortality, with no significant differences between the two periods. CONCLUSIONS: The application of Lean techniques in the critically ill process had a positive impact on improving patient flow within the health region, noting a decrease of transfers outside the region due to lack of beds, reduced delayed discharge from ICU to conventional ward and increased satisfaction of ICU professionals.

Citrus Viroids: Symptom Expression and Performance of Washington Navel Sweet Orange Trees Grafted on Carrizo Citrange.

Citrus are natural hosts of several viroid species. Citrus exocortis viroid (CEVd) and Hop stunt viroid (HSVd) are the causal agents of two well-known diseases of citrus, exocortis and cachexia. Other viroids have been found to induce specific symptoms and different degrees of stunting in trees grafted on trifoliate orange and trifoliate orange hybrids. A field assay was initiated in 1989 to establish the effect of CEVd, HSVd, Citrus bent leaf viroid (CBLVd), Citrus dwarfing viroid (CDVd), and Citrus bark cracking viroid (CBCVd) on Washington navel sweet orange trees grafted on Carrizo citrange rootstock. Here we report the effect of viroid infection on symptom expression, tree size, fruit production and quality evaluated from 2004 to 2007. Vegetative growth was affected by viroid infection with height and canopy volume being reduced. No bark scaling symptoms were observed in CEVd-infected trees albeit they presented lesions and blisters in the roots. Bark cracking symptoms were consistently observed in CBCVd-infected trees that were smaller with enhanced productivity and fruit size. No major effects were found as a result of infection with CBLVd, HSVd, or CDVd. The quality of the fruits was not affected by viroid infection, except for the low diameter of the fruits harvested from HSVd-infected trees. An interesting effect was identified in terms of tree productivity increase (yield/canopy volume) as a result of infection with CEVd, CDVd, and especially CBCVd.

Body mass index classification misses subjects with increased cardiometabolic risk factors related to elevated adiposity.

CONTEXT: Body mass index (BMI) is widely used as a measure of overweight and obesity, but underestimates the prevalence of both conditions, defined as an excess of body fat. OBJECTIVE: We assessed the degree of misclassification on the diagnosis of obesity using BMI as compared with direct body fat percentage (BF%) determination and compared the cardiovascular and metabolic risk of non-obese and obese BMI-classified subjects with similar BF%. DESIGN: We performed a cross-sectional study. SUBJECTS: A total of 6123 (924 lean, 1637 overweight and 3562 obese classified according to BMI) Caucasian subjects (69% females), aged 18-80 years. METHODS: BMI, BF% determined by air displacement plethysmography and well-established blood markers of insulin sensitivity, lipid profile and cardiovascular risk were measured. RESULTS: We found that 29% of subjects classified as lean and 80% of individuals classified as overweight according to BMI had a BF% within the obesity range. Importantly, the levels of cardiometabolic risk factors, such as C-reactive protein, were higher in lean and overweight BMI-classified subjects with BF% within the obesity range (men 4.3 ± 9.2, women 4.9 ± 19.5 mg l(-1)) as well as in obese BMI-classified individuals (men 4.2 ± 5.5, women 5.1 ± 13.2 mg l(-1)) compared with lean volunteers with normal body fat amounts (men 0.9 ± 0.5, women 2.1 ± 2.6 mg l(-1); P<0.001 for both genders). CONCLUSION: Given the elevated concentrations of cardiometabolic risk factors reported herein in non-obese individuals according to BMI but obese based on body fat, the inclusion of body composition measurements together with morbidity evaluation in the routine medical practice both for the diagnosis and the decision-making for instauration of the most appropriate treatment of obesity is desirable.

Bovine insulin-phosphatidylcholine mixed Langmuir monolayers: behavior at the air-water interface.

The behavior of the binary mixed Langmuir monolayers of bovine insulin (INS) and phosphatidylcholine (PC) spread at the air-water interface was investigated under various subphase conditions. Pure and mixed monolayers were spread on water, on NaOH and phosphate-buffered solutions of pH 7.4, and on Zn(2+)-containing solutions. Miscibility and interactions between the components were studied on the basis of the analysis of the surface pressure (π)-mean molecular area (A) isotherms, surface compression modulus (C(s)(-1))-π curves, and plots of A versus mole fraction of INS (X(INS)). Our results indicate that intermolecular interactions between INS and PC depend on both the monolayer state and the structural characteristics of INS at the interface, which are strongly influenced by the subphase pH and salt content. Brewster angle microscopy (BAM) was applied to investigate the peptide aggregation pattern at the air-water interface in the presence of the studied lipid under any experimental condition investigated. The influence of the lipid on the INS behavior at the interface strongly depends on the subphase conditions.

First Report of Citrus viroid V in Moro Blood Sweet Orange in Iran.

Viroids are nonencapsidated, small, circular, single-stranded RNAs that replicate autonomously when inoculated in their host plants in which they may elicit diseases (sensitive hosts) or replicate as latent infections (tolerant hosts). Citrus viroid V (CVd-V) was initially identified in Spain (1) and later found to be present in the United States, Nepal, and the Sultanate of Oman (2). CVd-V is a member of the Apscaviroid genus within the Pospiviroidae family. Like other members of this genus, CVd-V has a restricted host range but it is able to infect a wide range of citrus and citrus related species (1,2). Within the framework of a comprehensive survey of the sanitary status of the citrus industry in Iran, a sample from a private orchard of symptomless Moro blood sweet orange (Citrus sinensis) trees grafted on Mexican lime (C. aurantifolia) located at Javanan in the southern inland region was found to be infected with CVd-V. Briefly, RNAs of nucleic acid preparations from bark tissues were separated by 5% polyacrylamide gel electrophoresis (PAGE), electrotransferred to positively charged nylon membranes, immobilized by UV cross-linking, and hybridized with a full length CVd-V specific digoxigenin (DIG)-labeled DNA probe (2). A positive identification of CVd-V was made in these extracts. This positive detection of CVd-V was confirmed by reverse transcription-PCR using CVd-V specific primers of opposite polarity (5'-GACGAAGGCCGGTGAGCAGTAAGCC-3') and (5'-GACGACGACAGGTGAGTACTTTC-3') corresponding to CVd-V positions 90 to 114 and 69 to 89, respectively. Analysis of the sequence of the 293-bp amplicon (Genbank Accession No. GQ466068) revealed 99% identity with the reference sequence (Genbank Accession No. NC010165) of CVd-V. The rod-like predicted minimum free energy secondary structure of this new variant has 68.3% paired nucleotides. The changes with respect to the reference CVd-V variant are: (i) a deletion (48→-U) located in a loop of the V domain; (ii) a substitution (155A→C) located in a loop of the TR domain of the viroid secondary structure; and (iii) two compensatory substitutions located in the upper (46A→G) and lower (244U→C) strands of the viroid secondary structure. As shown earlier, the genome of CVd-V allows little variation with a large loop located in the segment I of the secondary structure (2) being the most amenable for mutations/changes. Among the viroids that have been found naturally infecting citrus, the members of the genus Apscaviroid are not associated with specific diseases but they cause a reduction of tree size and fruit harvest (3), an effect that is enhanced when several viroids coinfect the same plant (4). Therefore, the presence of CVd-V should be considered in further indexing tests aimed at the production and distribution of pathogen-free plants in Iran. References: (1) P. Serra et al. Virology 370:102, 2008. (2) P. Serra et al. Phytopathology 98:1199, 2008. (3) C. Vernière et al. Plant Dis. 88:1189, 2004. (4) C. Vernière et al. Phytopathology 96:356, 2006.

Sirolimus for treatment of steroid-refractory acute graft-versus-host disease.

Acute GVHD (aGVHD) is a major cause of morbidity and mortality in hematopoietic allograft recipients. The best therapy for patients failing to respond, or not tolerating, systemic glucocorticoids remains undefined. We evaluated the efficacy of sirolimus in 34 patients, median age of 49 (23-67) years, with steroid-refractory (n=31) or steroid-intolerant (n=3) aGVHD. aGVHD was diagnosed at a median of 34 (7-1042) days post allografting, and confirmed by biopsy in all cases. Initial aGVHD treatment consisted of prednisone up to 2 mg/kg. Sirolimus was initiated at a median of 9 (1-255) days after glucocorticoid initiation. A sirolimus loading dose was administered to 19 (56%) of 34 patients, median 6 (3-8) mg, followed by maintenance of 1-2 mg/day to target therapeutic trough levels between 4 and 12 ng/ml. Overall response rate was 76%. Fifteen (44%) of 34 patients achieved CR, defined as complete resolution of aGVHD sustained for at least 1 month, after sirolimus initiation without additional immunosuppressive agents. CR was achieved in 11 (42%) of 31 steroid-refractory and 2 (67%) of 3 steroid-intolerant patients. Median OS after initiation of sirolimus was 5.6 months, and 1-year OS was 44% (95% CI: 27-60%). Sirolimus is effective in controlling steroid-refractory aGVHD.

[Therapeutic use and profile of toxicity of the FOLFOX4 regimen]. / Uso terapéutico y perfil de toxicidad del esquema FOLFOX4.

INTRODUCTION: Since the publication of the MOSAIC test results in 2004, the FOLFOX4 regimen has been established as an adjuvant treatment which is recommended in stage III colorectal cancer. The aim of this study is to assess the use of this regimen in our field and to describe its toxicity. METHODS: Descriptive study of treatments with FOLFOX4 prescribed between April 2005 and March 2007. The data was obtained from the Farhos Oncología programme and clinical records. The following data was collected: age, gender, diagnosis, stage of the illness (TNM classification) and adverse reactions, expressing severity according to Common Toxicity Criteria 2.0. RESULTS: The FOLFOX4 regimen was prescribed for 39 patients (24 men and 15 women) with an average age of 59. The diagnoses were: 28 colon cancer (4 stage II, 17 stage III, and 7 stage IV), 10 rectal cancer (1 stage II, 4 stage III, and 5 stage IV) and 1 stage IV gastric cancer. The most frequent adverse reactions were peripheral neuropathy (82 %), neutropenia (56.4 %) and diarrhoea (53.9 %.) When the study was completed, 9 patients continued active treatment with the regimen (average 6.8 cycles.) Of the 30 remaining patients only 16 people completed the 12 planned cycles. 14 patients stopped their treatment (an average of 8.1 cycles) due to toxicity in 10 cases, clinical progression in 3 cases and one patient died. Of the total 368 cycles administered, 68 suffered administration delays and 22 had the dosage reduced. CONCLUSION: The use of the FOLFOX4 regimen has been adjusted to uses with some solid scientific evidence, but its toxicity has limited its use and has made administering the planned dosage levels difficult.

La actividad física determinada mediante acelerometría es menor en los pacientes obesos / Obese subjects exhibit a reduced physical activity assessed by accelerometry

Objetivos: Un descenso de actividad física (AF) puede ser un factor de obesidad. El objetivo del presente estudio fue evaluar la utilidad del acelerómetro Actical® para medir de manera objetiva la AF y establecer diferencias entre sujetos normoponderales, con sobrepeso u obesidad. Material y métodos: Se midió la AF con el Actical® y se comparó con la información obtenida con el SenseWear®Arm- Band. Se midió la AF de 46 sujetos normoponderales (NP), 17 con sobrepeso (SP) y 31 obesos (OB). También se midieron diferentes variables antropométricas y bioquímicas. Resultados: La AF registrada fue similar independientemente del lugar de colocación del dispositivo y comparable con la obtenida con el ArmBand. Los pacientes obesos mostraron un nivel reducido de AF evidenciado por la cantidad de actividad registrada por el acelerómetro (NP: 357 ± 16, SP: 338 ± 23, OB: 298 ± 14, cuentas/min P<0,05). La cantidad de AF se correlacionó, entre otras variables, con el IMC (r=-0,38, P<0,05), la circunferencia de la cintura (r=-0.42, P<0.05) y la concentración de ácidos grasos libres (r=-0,44, P<0,01). Conclusiones: El acelerómetro Actical® proporciona una medida objetiva del grado de AF de un individuo permitiendo establecer diferencias entre los individuos normoponderales y los pacientes obesos. Un bajo grado de AF se asocia con un peor perfil lipídico y mayor resistencia a la insulina, sobre todo en los sujetos obesos, aumentado su riesgo cardiometabólico (AU)

Aim: Reduced physical activity (PA) play a role in obesity. The amount and intensity of PA in free-living conditions is difficult to measure. The aim was to evaluate the usefulness of the accelerometer Actical® to measure PA and to discriminate potential differences between lean and obese subjects. Material and Methods: We studied information obtained with the Actical® and compared it with the SenseWear®Arm- Band. PA of 46 lean (LE), 17 overweight (OW) and 31 obese (OB) volunteers. Anthropometric variables and blood biochemistry were analysed. Results: Data regarding PA obtained with the Actical® was similar independently of the location and comparable to the one obtained with the ArmBand. Obese subjects exhibited a reduced PA as evidenced by the amount of movement recorded by the accelerometer (LE: 357 ± 16, OW: 338 ± 23, OB: 298 ± 14, counts/min P<0.05). The amount of PA showed a significant correlation with BMI (r=-0.38, P<0.05), waist circumference (r=-0.42, P<0.05), and circulating free fatty acids concentrations (r=-0.44, P<0.01). Conclusions: The accelerometer Actical® worn on the wrist represents a useful tool in the clinical setting allowing to discriminate differences between lean and obese subjects. A low PA level is accompanied by a worst lípid profile and higher insulin resistance, particullarly in obese subjects, increasing its cardiometabolic risk (AU)

Uso terapéutico y perfil de toxicidad del esquema FOLFOX4* / Therapeutic use and profile of toxicity of the FOLFOX4 regimen

Introducción: Desde la publicación de los resultados del estudio MOSAIC en 2004, el esquema FOLFOX4 se ha establecido como un tratamiento adyuvante recomendado en los cánceres colorrectales estadio III. El objetivo de este estudio es valorar la utilización de este esquema en nuestro ámbito y describir su toxicidad. Métodos: Estudio descriptivo de los tratamientos con FOLFOX4 prescritos desde abril de 2005 a marzo de 2007. Los datos se obtuvieron del programa Farhos Oncología® y las historias clínicas. Se recogieron las variables siguientes: edad, sexo, diagnóstico, estadio de la enfermedad (clasificación TNM) y reacciones adversas, expresando su gravedad según los Common Toxicity Criteria 2.0. Resultados: El esquema FOLFOX4 ha sido prescrito a 39 pacientes (24 varones y 15 mujeres), con una mediana de edad de 59 años. Los diagnósticos fueron: 28 cáncer de colon (4 estadio II, 17 III y 7 IV), 10 cáncer de recto (1 estadio II, 4 III y 5 IV) y 1 cáncer gástrico estadio IV. Las reacciones adversas más frecuentes fueron neuropatía periférica (82 %), neutropenia (56,4 %) y diarrea (53,9 %). Al finalizar el estudio 9 pacientes seguían en tratamiento activo con este esquema (media, 6,8 ciclos). De los 30 restantes, sólo 16 completaron los 12 ciclos previstos. En 14 pacientes se suspendió el tratamiento (media, 8,1 ciclos), siendo los motivos: toxicidad en 10 casos, progresión clínica en 3 y fallecimiento en 1. Del total de los 368 ciclos administrados, 68 tuvieron retrasos en la administración y en 22 se redujo la dosis. Conclusión: La utilización del esquema FOLFOX4 se ha ajustado a usos con unas evidencias científicas sólidas, pero su toxicidad ha limitado el uso y dificultado la administración de la intensidad de dosis prevista (AU)

Introduction: Since the publication of the MOSAIC test results in 2004, the FOLFOX4 regimen has been established as an adjuvant treatment which is recommended in stage III colorectal cancer. The aim of this study is to assess the use of this regimen in our field and to describe its toxicity. Methods: Descriptive study of treatments with FOLFOX4 prescribed between April 2005 and March 2007. The data was obtained from the Farhos Oncología® programme and clinical records. The following data was collected: age, gender, diagnosis, stage of the illness (TNM classification) and adverse reactions, expressing severity according to Common Toxicity Criteria 2.0. Results: The FOLFOX4 regimen was prescribed for 39 patients (24 men and 15 women) with an average age of 59. The diagnoses were: 28 colon cancer (4 stage II, 17 stage III, and 7 stage IV), 10 rectal cancer (1 stage II, 4 stage III, and 5 stage IV) and 1 stage IV gastric cancer. The most frequent adverse reactions were peripheral neuropathy (82 %), neutropenia (56.4 %) and diarrhoea (53.9 %.) When the study was completed, 9 patients continued active treatment with the regimen (average 6.8 cycles.) Of the 30 remaining patients only 16 people completed the 12 planned cycles. 14 patients stopped their treatment (an average of 8.1 cycles) due to toxicity in 10 cases, clinical progression in 3 cases and one patient died. Of the total 368 cycles administered, 68 suffered administration delays and 22 had the dosage reduced. Conclusion: The use of the FOLFOX4 regimen has been adjusted to uses with some solid scientific evidence, but its toxicity has limited its use and has made administering the planned dosage levels difficult (AU)

A novel hybridization approach for detection of citrus viroids.

Citrus plants are natural hosts of several viroid species all belonging to the family Pospiviroidae. Previous attempts to detect viroids from field-grown species and cultivars yielded erratic results unless analyses were performed using Etrog citron a secondary bio-amplification host. To overcome the use of Etrog citron a number of RT-PCR approaches have been proposed with different degrees of success. Here we report the suitability of an easy to handle northern hybridization protocol for viroid detection of samples collected from field-grown citrus species and cultivars. The protocol involves: (i) Nucleic acid preparations from bark tissue samples collected from field-grown trees regardless of the growing season and storage conditions; (ii) Separation in 5% PAGE or 1% agarose, blotting to membrane and fixing; (iii) Hybridization with viroid-specific DIG-labelled probes and detection with anti-DIG-alkaline phosphatase conjugate and autoradiography with the CSPD substrate. The method has been tested with viroid-infected trees of sweet orange, lemon, mandarin, grapefruit, sour orange, Swingle citrumello, Tahiti lime and Mexican lime. This novel hybridization approach is extremely sensitive, easy to handle and shortens the time needed for reliable viroid indexing tests. The suitability of PCR generated DIG-labelled probes and the sensitivity achieved when the samples are separated and blotted from non-denaturing gels are discussed.

Effect of a Field-Source Mixture of Citrus Viroids on the Performance of 'Nules' Clementine and 'Navelina' Sweet Orange Trees Grafted on Carrizo Citrange.

A field-source mixture of citrus viroids was characterized and shown to contain Citrus exocortis viroid (CEVd), Hop stunt viroid (HSVd), Citrus bent leaf viroid (CBLVd), and Citrus dwarfing viroid (CDVd). Sequencing results showed that: (i) CEVd contained the PL and PR characteristic of class A variants; (ii) HSVd was a noncachexia variant; (iii) CBLVd was related to CVd-Ia variants; (iv) CDVd was a mixture of two types (CVd-IIIa and CVd-IIIb) of variants. The presence of the same type of variants in inoculated clementine (Citrus clementina 'Nules') and sweet orange (C. sinensis 'Navelina') trees on Carrizo citrange (Poncirus trifoliata × C. sinensis) rootstocks was confirmed. The effect of infection was determined by assessing the performance of infected and noninfected trees growing in the field. Infection resulted in small trees with reduced canopy, yielding a reduced crop. Fruit characteristics were also affected: (i) clementine and sweet orange fruits from infected trees were larger than those from noninfected trees; (ii) clementine fruits from infected trees differed in shape from those of noninfected trees; (iii) sweet orange fruits from infected trees had maturity indexes and juice contents higher than those from noninfected trees; (iv) in both species, the density of the juice, the amount of soluble solids, and the acidity of the fruits from infected trees were lower than those of fruits from noninfected trees. Infected trees had a poorly developed root system with fibrous roots containing fewer amyloplasts than noninfected trees. The results of an in vitro assay on the induction and development of roots in cultured explants are discussed.

A set of novel RNAs transcribed from the chloroplast genome accumulates in date palm leaflets affected by brittle leaf disease.

Brittle leaf disease or maladie des feuilles cassantes (MFC) is a lethal disorder of date palms that has assumed epidemic proportions in the oases of southern Tunisia. After a prolonged period during which palms are declining, the disease ends with the death of the palms. Whereas no pathogen could ever be associated with the disease, leaflets of affected palms have been previously shown to be deficient in manganese. Analysis of RNA preparations from leaflets of MFC-affected palms revealed the presence of a set of novel RNAs (MFC-RNAs) of sense and antisense polarities, which are homologous to various regions of the date palm chloroplast genome, such as the regions containing genes rrn5S-trnR(ACG) and trnM(CAU)-atpE. In the RNA preparations obtained from leaflets of affected palms, some of these RNAs are present as double-stranded species (MFC-dsRNAs), as witnessed by results from cellulose chromatography, end labeling, RNase digestion, and northern hybridization with strand specific probes. These MFC-RNAs represent a novel type of host-derived RNAs, and their presence in MFC-affected date palms is of diagnostic value.

Citrus viroid V: occurrence, host range, diagnosis, and identification of new variants.

The recently described Citrus viroid V (CVd-V) has been proposed as a new species of the genus Apscaviroid within the family Pospiviroidae. Analysis of 64 samples from different citrus-growing areas has shown that CVd-V is present in the United States, Spain, Nepal, and the Sultanate of Oman. CVd-V found in six sweet orange sources from the Sultanate of Oman was identical to the reference CVd-V variant, whereas three new variants with sequence identities of 98.6% (CVd-VCA), 97.3% (CVd-VST), and 94.9% (CVd-VNE) were identified in sources from California, Spain, and Nepal, respectively. These results suggest that this viroid has not emerged recently and that it is relatively widespread. Transmission assays to sweet orange, mandarin, and mandarin hybrids, clementine, satsuma, lemon, sour orange, Tahiti lime, Palestine sweet lime, calamondin, bergamot, and kumquat have shown that all these citrus species and citrus relatives are hosts for CVd-V. Several indexing approaches, including slot blot, northern blot hybridization, and reverse transcription-polymerase chain reaction, have been evaluated for detecting CVd-V, either using Etrog citron as an amplification host or directly from commercial species and cultivars.

A single nucleotide change in Hop stunt viroid modulates citrus cachexia symptoms.

Cachexia disease of citrus is caused by Hop stunt viroid (HSVd). In citrus, pathogenic and non-pathogenic strains differ by a "cachexia expression motif" of five to six nucleotides located in the variable domain of the proposed rod-like secondary structure. Here, site-directed mutants were generated to investigate if all these nucleotides were required for infectivity and/or symptom expression. Specifically an artificial cachexia inducing mutant M0 was generated by introducing the six nucleotides changes of the "cachexia expression motif" into a non-pathogenic sequence variant and M0 was used as a template to systematically restore some of the introduced changes. The resulting mutants in which specific changes introduced to generate M0, were restored presented a variety of responses: (i) M1, obtained by introducing two insertions forming a base-pair, was infectious but non-pathogenic; (ii) M2, obtained by introducing an insertion and restoring a substitution, presented low infectivity and the resulting progeny reverted to M0; (iii) M3, obtained by restoring a single substitution in the lower strand of the viroid secondary structure, was infectious but induced only mild cachexia symptoms; (iv) M4, obtained by restoring a single substitution in the upper strand of the viroid secondary structure, was non-infectious. These results confirm that the "cachexia expression motif" plays a major role in inciting cachexia symptoms, and that subtle changes within this motif affect symptom severity and may even suppress symptom expression.

Citrus viroid V: molecular characterization and synergistic interactions with other members of the genus Apscaviroid.

Studies on Atalantia citroides, a citrus relative, revealed the existence of a viroid not described previously. The new viroid has a GC-rich genome of 293-294 nucleotides and contains the central conserved region characteristic of members of the genus Apscaviroid, and the terminal conserved region present in this and other genera of the family Pospiviroidae. The secondary structure of minimum free energy predicted for the new viroid is a rod-like conformation with 68.7% paired nucleotides and showing sequence identities with other viroids always lower than 90%, the conventional limit that separates different species within a given genus. Infectivity assays showed that the new viroid induces mild but characteristic symptoms on the indicator Etrog citron. Co-inoculation of CVd-V with either Citrus bent leaf viroid or Citrus viroid III, two other members of the genus Apscaviroid infecting citrus, disclosed synergistic interactions manifested in enhanced leaf symptoms and very pronounced dwarfing. Viroid titers, however, remained unaltered in co-infected plants. Possible mechanisms underlying the observed synergistic effects are discussed. According to its molecular and biological properties and its unusual ability to replicate in A. citroides, the new viroid, tentatively named Citrus viroid V (CVd-V), should be considered a new species of the genus Apscaviroid.

Molecular characterization of CEVd strains that induce different phenotypes in Gynura aurantiaca: structure-pathogenicity relationships.

Two Citrus exocortis viroid isolates (CEVd-s and CEVd-129) that induce severe and mild symptoms in Gynura aurantiaca, respectively, have been characterized. They present nucleotide sequences in the pathogenicity motifs P(L), C and P(R) similar to those of "Class A" and "Class B". Infectivity and symptom expression in G. aurantica and tomato were evaluated with a selection of sequence variants recovered from both isolates. As expected, the two variants selected from CEVd-s induced severe symptoms. The variants selected from CEVd-129 induced mild symptoms, except one of them, named MJ, that presented an unusual genotype and induced severe symptoms in G. aurantiaca. The biological properties of MJ show that the two nucleotide changes of the C domain normally associated with the P(L) and P(R) motifs of "Class B" strains are not implicated in symptom expression. The relationship between "Class A" and "Class B" strains with the symptoms induced in clementine trees grafted on trifoliate orange is discussed.