Conyza bonariensis as an alternative host for Colletotrichum species in Argentina.

AIMS: This study investigated the diversity of Colletotrichum isolates recovered from Conyza bonariensis leaves through the use of morphological characteristics, growth rate, carbon sources utilization and phylogenetic analysis. METHODS AND RESULTS: In all, 30 Colletotrichum isolates recovered from C. bonariensis leaves showing symptoms of disease were included in the present study. Based on the analysis of morphology and sequences, the isolates were distributed into six Colletotrichum species complexes. The concatenated alignment of GAPDH and ITS sequences showed that 20 out of 30 isolates were included in four species complexes which comprise the most important pathogens causing anthracnose in soybean or anthracnose and stalk rot in maize: C. truncatum, C. orchidearum, C. gloeosporioides and C. graminicola. The remaining 10 isolates were included in the C. boninense and C. destructivum species complexes or could not be assigned to any complex with the available information. CONCLUSION: Weeds belonging to genus Conyza are host to soybean and maize potential pathogenic species of Colletotrichum and could have a role as inoculum reservoir for cross contamination in the agroecosystem. SIGNIFICANCE AND IMPACT OF THE STUDY: The combined use of morphological, kinetics and physiological parameters of growth and phylogenetic analysis in Colletotrichum isolates from Conyza leaves allowed the detection of species complexes previously not identified in Argentina.

Influence of crop residues, matric potential and temperature on growth of Exserohilum turcicum an emerging maize pathogen in Argentina.

Northern corn leaf blight (NCLB) caused by Exserohilum turcicum is an endemic foliar disease in the Argentinean maize production area. Before applying a control method, it is essential to focus on factors that lead to the survival and conservation of E. turcicum inoculum. However, there is little information about the life cycle of this emerging pathogen in our country. The objective of the present work was to analyse the growth ability of three E. turcicum isolates under different type and conditions of maize, wheat and soybean residues, matric potential, temperature and their interactions. Statistical analysis demonstrated a significant effect of three factors on growth rate: residues-based media, matric potential and temperature. Among them the major effect was produced by matric potential followed by temperature. Although there were no significant differences in the growth rate among different residues types, the range of conditions in which growth occurred was different. According to these results, soybean residues would allow a better survival of E. turcicum inoculum under no-tillage system and providing an inoculum source for maize infections in subsequent year. SIGNIFICANCE AND IMPACT OF THE STUDY: Exserohilum turcicum is an emerging pathogen in Argentina that has caused significant economic losses in different maize growing areas. However, at present there is a lack of information about the life cycle of this fungal pathogen. The present study shows the influence of abiotic factors such as temperature and water potential on the growth of E. turcicum on different crop residues used in rotations with maize under no-till system cultivation. According to our results, soybean residues allowed the pathogen growth in a wider range of conditions compared to wheat and maize residue, providing an inoculum source for maize infections in subsequent season.

Impact of two bacterial biocontrol agents on bacterial and fungal culturable groups associated with the roots of field-grown maize.

AIMS: To assess the impact of Bacillus amyloliquefaciens and Microbacterium oleovorans on bacterial and fungal groups associated to the roots of field-grown maize. METHODS AND RESULTS: Identification and count of bacterial and fungal culturable populations associated to the roots of maize seedlings, changes in culturable community structure according to the richness and diversity indexes concept and shifts in microbial activity through analysis of cellulolytic, ammonification and nitrification potentials were determined, in relation to kernel treatment with biological control agents. Following the treatment of maize kernels with B. amyloliquefaciens at 10(7) CFU ml(-1), an increase in bacterial diversity was observed at the rhizoplane of resultant seedlings. Bacterial richness was significantly increased at the root inner tissues of seedlings treated with Mic. oleovorans. Fusarium, Aspergillus, Penicillium and Trichoderma were the main fungal genera isolated and there population sizes were unequally affected by the addition of biocontrol agents. CONCLUSIONS: Numbers and types of isolated bacteria and fungi changed in response to the addition of biocontrol agents, while microbial activity remained unchanged with respect to control. SIGNIFICANCE AND IMPACT OF THE STUDY: This study provides an insight of the effects of proven biocontrol agents on micro-organisms naturally associated to the target crop.

Screening of Argentine plant extracts: impact on growth parameters and aflatoxin B1 accumulation by Aspergillus section Flavi.

The effect of essential oils, ethanolic and aqueous extract of 41 vegetable species on Aspergillus section Flavi growth was evaluated. The in vitro screen was a two-stage process. A wide-spectrum initial screen which identified promising antifungal plant extracts was carried out first. After that, identified extracts were studied in more detail by in vitro assays. A total of 96 plant extracts were screened. Essential oils were found to be the most effective extract controlling aflatoxigenic strains. Clove, mountain thyme, poleo and eucalyptus essential oils were selected to study their antifungal effect. Studies on percentage of germination, germ-tube elongation rate, growth rate, and aflatoxin B1 accumulation were carried out. Clove, mountain thyme and poleo essential oils showed the most antifungal effect under all growth parameters analyzed as well as aflatoxin B1 accumulation. Our results suggest that mountain thyme and poleo, which are native vegetal species of Argentina, and clove essential oils could be used alone or in conjunction with other substances to control the presence of aflatoxigenic fungi in stored maize.

Control of Aspergillus section Flavi growth and aflatoxin accumulation by plant essential oils.

AIMS: The antifungal effect of Pimpinella anisum (anise), Pëumus boldus (boldus), Mentha piperita (peppermint), Origanum vulgare (oregano) and Minthosthachys verticillata (peperina) essential oils against Aspergillus section Flavi (two isolates of Aspergillus parasiticus and two isolates of Aspergillus flavus) was evaluated in maize meal extract agar at 0.982 and 0.955 water activities, at 25 degrees C. METHODS AND RESULTS: The percentage of germination, germ-tube elongation rate, growth rate and aflatoxin B(1) (AFB(1)) accumulation at different essential oils concentrations were evaluated. Anise and boldus essential oils were the most inhibitory at 500 mg kg(-1) to all growth parameters of the fungus. These essential oils inhibited the percentage of germination, germ-tube elongation rate and fungal growth. AFB(1) accumulation was completely inhibited by anise, boldus and oregano essential oils. Peperina and peppermint essential oils inhibited AFB(1) production by 85-90% in all concentrations assayed. CONCLUSIONS: Anise and boldus essential oils could be considered as effective fungitoxicans for Aspergillus section flavi. SIGNIFICANCE AND IMPACT OF THE STUDY: Our results suggest that these phytochemical compounds could be used alone or in conjunction with other substances to control the presence of aflatoxigenic fungi in stored maize.

Biocontrol of Bacillus subtilis against Fusarium verticillioides in vitro and at the maize root level.

Bacillus species as a group offer several advantages over other bacteria for protection against root pathogens because of their ability to form endospores, and because of the broad-spectrum activity of their antibiotics. The objectives of this work were to determine the ability of strains of Bacillus to inhibit Fusarium verticillioides growth and fumonisin B(1) accumulation in vitro, and to evaluate the ability of the best bacterium for preventing rhizosphere and endorhizosphere colonization by F. verticillioides. Bacterial populations from the maize rhizoplane were obtained, and the capacity of ten Bacillus strains to inhibit fungal growth and fumonisin B(1) accumulation in vitro was assayed. According to these results, B. subtilis CE1 was selected as the best antagonist for testing maize root colonization of F. verticillioides. Bacillus subtilis CE1 at 10(8) and 10(7) CFU ml(-1) inocula was able to reduce rhizoplane and endorhizosphere colonization of F. verticillioides in greenhouse trials. The strain B. subtilis CE1 could be a potential biological control agent against F. verticillioides at the root level.

Regulation of epidermal growth factor receptor activity by crotoxin, a snake venom phospholipase A2 toxin. A novel growth inhibitory mechanism.

Crotoxin (CT), a phospholipase A2 (PLA2) derived from the venom of Crotalus durissus terrificus, is a heterodimeric protein composed of subunit B with enzymatic activity and a binding regulatory subunit (A) without enzyme activity. Although the PLA2 activity of CT may be important in its anti-proliferative activity, its cytostatic mechanism is unknown. In this study, we examined the cytostatic effect of PLA2-associated CT activity on squamous carcinoma cells expressing distinct levels of epidermal growth factor receptor (EGFr). CT was most effective in suppressing growth on cells expressing high intrinsic levels of EGFr. Cardiotoxin, another membrane active toxin with no intrinsic PLA2 activity, had no differential anti-proliferative activity on cells expressing high EGFr levels, suggesting a correlation between EGFr expression and CT-directed anti-proliferative activity. Both chemically modified CT (MCT) devoid of PLA2 activity and covalently cross-linked CT (CCT), which is functionally unable to utilize cellular membranes as PLA2 substrate, were also without growth inhibitory activity. No evidence for direct binding of CT to EGFr was found, although pretreatment with EGF was able to partially suppress the anti-proliferative activity of CT. Tyrosine phosphorylation of EGFr, however, was stimulated by CT in intact A431 cells. Tyrosine phosphorylation of EGFr was concentration-dependently stimulated (3- to 8-fold) in cellular membranes of A431 cells treated in vitro with CT but not with anti-proliferatively inactive MCT or CCT. The data provide evidence for transmembrane receptors involved in growth signaling (namely EGFr) as cellular targets and potential effectors of PLA2-mediated anti-proliferative activity of snake venom.

Toxicity of the novel animal-derived anticancer agent, VRCTC-310: acute and subchronic studies in beagle dogs.

Acute and subchronic toxicities of VRCTC-310, a combination product of crotoxin (CT) and cardiotoxin (CD), which has shown antitumor activity in vivo, have been studied in Beagle dogs. Single i.m. doses of 0.25, 0.5 and 1.0 mg/kg resulted in dose-dependent local muscular toxicity consisting of myofiber atrophy, interstitial edema and macrophage infiltration. Also, AST, ALT and LDH levels increased on day 2, returning to normal values on days 6-8. Local lesions were absent after recovery on day 45. At 2.0 mg/kg, signs of neurotoxicity (ataxia) appeared, in addition to vomitus, salivation, hematuria and myotoxicity in tongue and diaphragm on day 8. Local lesions healed with fibrosis at the site of injection on day 45. Administration of fixed (0.025 and 0.05 mg/kg) or escalating (0.025-0.1 mg/kg) daily doses for 30 days also produced local muscular damage, which was absent at day 75. The increases in AST, ALT and LDH serum activities on days 2-4 were independent of dosing schedule and sharply decreased on day 8, despite continuation of treatment. An escalating dose schedule of 0.025-2.0 mg/kg showed local muscle damage at the site of injection on day 31, however, there were no lesions of myotoxicity in the tongue or diaphragm and no clinical signs of neurotoxicity were observed. Animals tolerated the subchronic treatment better than the acute. The resolution of serum enzymes to normal values during treatment may be attributed to a decrease of sensitivity to VRCTC-310-mediated myotoxic effects.

Studies on damage to sunflower seeds: water activity, germination, acidity index and aflatoxin B1 presence.

The significance of fungal contamination during the storage of sunflower seeds has been investigated. Samples were taken during 7 months at 45 day-intervals. Water activity, seed germination, presence of aflatoxin B1 and free fatty acids were monitored. It was demonstrated that water activity increased during storage, germinability decreased, the content of free fatty acid increased and so did the content of aflatoxin B1.

Neoadjuvant chemotherapy with cisplatin, ifosfamide and 5-fluorouracil in the treatment of locally advanced cervical cancer.

The objective of this study was to evaluate clinical and histological response, resectability, and survival in patients with cervical epidermoid carcinoma stage IB2 to IIIB with the use of neoadjuvant chemotherapy followed by radical surgery and/or radiation therapy. Between September 1989 and February 1996, 53 patients were admitted to this study. They were given three cycles of cisplatin 30 mg/m2/day, 5-fluorouracil 500 mg/m2/day, ifosfamide 2000 mg/m2/day i.v., and mesna 400 mg/m2/day i.v. at hour 0 and 400 mg/m2 at hours 4 and 8 during three days every 21-28 days. We evaluated 47 patients. Global clinical response obtained was 85% {95% (CI), 75-97%, CR in 14 patients (30%) and PR in 26 patients (55%)}. Twenty-three patients underwent surgery. Six patients (13%) had a complete histological response. Median follow-up was 42 months (5-96). In resected patients, with a median follow-up of 57 months (5-96), the estimated five-year disease-free survival was 78%. Global survival estimated to 60 months was 83% for stage IB2, 70% for IIB, and 20% for IIIB. This mode of therapy offers a new option to improve survival in locally advanced cervical cancer. Randomized trials are required in order to establish a definitive role for this therapeutic strategy.

Fumonisin production on irradiated corn kernels: effect of inoculum size.

Production of fumonisins B1, B2, and B3 by Fusarium moniliforme was evaluated on irradiated corn kernels inoculated with different spore concentrations (10, 10(2), 10(3), 10(5), and 10(6)), a water activity of 0.97, and a temperature of 25 degrees C. There was a direct relationship between the level of toxin produced and inoculum size. The highest levels of total fumonisin produced after 35 days of incubation were 5,028 and 9,063 ng/g at 10(5) and 10(6) spores per ml, respectively. The pattern of fumonisin production (FB1 > FB2 > FB3) in cultures growing from different inocula was not affected during the 35 days of incubation. The ratio between FB2 and FB1 varied from 0.15 to 0.42, whereas the ratio between FB3 and FB1 varied from 0.34 to 0.87.

Emission probabilities of the KX-rays following the decay of 237Np in equilibrium with 233Pa.

Following international EUROMET Project No. 416 and our recent measurement of the emission probabilities of the main gamma-rays of 237Np in equilibrium with 233Pa, a complementary work has been undertaken to determine the photon emission probabilities of the KX-rays associated with the decay of these nuclides. Two different analysis methods have been used: the KX-rays region was analyzed by fitting both Voigt and full response functions. Photon emission probabilities were determined with a relative uncertainty of about 2%.

Standardization and decay data of 237Np.

This paper reports contributions from participants in the EUROMET project (No. 416) which was entitled "237Np research into problems relating to purification, characterization and standardization". Primary standardizations were made by the defined low solid angle, coincidence, 4pi alpha, 2pi alpha and liquid scintillation counting methods. Secondary standardizations were made with calibrated gamma-ray spectrometers. Absolute X-ray, gamma-ray and alpha-particle emission probabilities were also determined. The results for the successful conclusion of both primary and secondary standardization are presented together with the values for alpha-particle and gamma-ray emission probabilities determined in this exercise. Several significant inconsistencies remain with the gamma-ray emission probabilities, and these are highlighted.

Reconstrucción vulvar con colgajo fasciocutáneo del surco glúteo / Vulvar reconstruction with fasciocutaneous fl ap of the gluteal fold

Introducción. La reconstrucción inmediata de la región vulvar posterior a vulvectomía radical por patología oncológica representa un desafío para el cirujano plástico dado el tamaño del defecto que genera dicha cirugía, en relación con la complejidad que posee la región anatómica a tratar. Material y métodos. Se presenta el caso de una paciente de sexo femenino de 74 años de edad con antecedentes de resección local de tumor vulvar asociado a radioterapia adyuvante en otra Institución, 5 años previos a la consulta. Actualmente presenta recidiva de su enfermedad. Se aborda de forma multidisciplinaria y se realiza vulvectomía radical más reconstrucción inmediata del defecto con dos colgajos del surco glúteo. Resultado. Una vez realizada la cirugía, la paciente fue dada de alta de la Institución al séptimo día posoperatorio y fue instruida para evitar sentarse durante 7 días más. Al 12 día posoperatorio presentó una dehiscencia parcial de la herida en región subglútea derecha, la cual se resolvió suturándola nuevamente. Al cumplirse 1 mes de la cirugía, la paciente se encontró satisfecha con el resultado estético y en condiciones de volver a su vida normal desde el punto de vista funcional y estético. Conclusión. El presente trabajo confirma que la utilización del colgajo fasciocutáneo del surco glúteo es una buena elección para la reconstrucción de la región vulvar, tanto estética como funciona

Background. The immediate reconstruction of the vulvar region after radical vulvectomy due to oncological pathology represents a challenge for the plastic surgeon do to the size of the defect generated by said surgery, in relation to the complexity of the anatomical region to be treated. Material and Methods. We present the patient 74-years-old woman with a history of local resection of vulvar tumor associated with adjuvant radiotherapy in another Institution and currently she has recurrence of the disease. Is addressed by a multidisciplinary team and radical vulvectomy is performed with immediate reconstruction of the defect with two gluteal fold fl aps. Results. Once the surgery was performed, the patient was discharged from the Institution on the seventh postoperative day and was instructed to avoid sitting for 7 days. On the 12th postoperative day, she presented a partial dehiscence of the wound, which was resolved by suturing it. One month after surgery, the patient was satisfi ed with the aesthetic result and able to return to her normal life from a functional point of view. Conclusion. The present work confi rms that the use of the fasciocutaneous fl ap of the gluteal fold is a very good option for the vulvar regionreconstruction, not only functionally as aesthetically

Enhanced retention strategies and willingness to participate among hard-to-reach female sex workers in Barcelona for HIV prevention and vaccine trials.

The potential for implementation of HIV vaccine trials in hard-to-reach female sex workers in an inner city area of Barcelona, Spain was assessed via a study of HIV risk, willingness to participate and the success of retention strategies. In 130 women, serological HIV status, behavioral risk exposures and willingness to participate in future HIV vaccine trials were recorded every six months using a confidential questionnaire. An enhanced retention (ER) strategy was compared with a control retention (CR) strategy comprising the recording of data on appointment cards. HIV seroincidence and retention rates were estimated. Retention rates after 6 and 12 mo of follow-up in the ER group were 76% and 69% respectively compared with 16% and 13% in the CR group. Among the ER group 97% were willing to participate in HIV vaccine trials at baseline and, after 12 mo of follow-up. Willingness was significantly associated with higher HIV risk exposure, and higher education level. Successfully retaining these cohorts over time in settings with a high HIV seroincidence rate is an ongoing challenge that will need to be addressed to ensure participation in future trials. Furthermore, as we have demonstrated, the fact that retaining hard-to-reach populations is difficult should not exclude this target population for HIV vaccine and prevention trials.

Production of Fusarium verticillioides biocontrol agents, Bacillus amyloliquefaciens and Microbacterium oleovorans, using different growth media: evaluation of biomass and viability after freeze-drying.

The aims of this study were to compare the viability and biomass production of B. amyloliquefaciens and M. oleovorans in different growth media, and the efficiency of a freeze-drying method as a possible formulation process. B. amyloliquefaciens and M. oleovorans were grown in 100 ml of four different media. Media water activity was modified at 0.99, 0.98, 0.97 and 0.96. Nutrient yeast dextrose broth (NYDB) and molasses soy powder (MSB) media were selected and survival levels of cells were determined before and after the freeze-drying process. B. amyloliquefaciens showed the highest survival after freeze-drying when grown in NYDB medium at 0.99 a(w), whereas, at 0.98, 0.97 and 0.96 a(w), the highest survival was obtained in MSB medium. M. oleovorans showed the highest survival in MSB medium at 0.99 a(w). MSB medium was select for biomass production due to high growth and survival after freeze-drying.

HIV vaccine trial willingness among injection and non-injection drug users in two urban centres, Barcelona and San Francisco.

Being able to recruit high-risk volunteers who are also willing to consider future participation in vaccine trials are critical features of vaccine preparedness studies. We described data from two cohorts of injection- and non-injection drug users in Barcelona, Spain [Red Cross centre] and in San Francisco, USA, [UFO-VAX study] at high risk of HIV/HCV infection to assess behaviour risk exposure and willingness to participate in future preventive HIV vaccine trials. We successfully identified drug-using populations that would be eligible for future HIV vaccine efficacy trials, based on reported levels of risk during screening and high levels of willingness to participate. In both groups, Red Cross and UFO-VAX respectively, HCV infection was highly prevalent at baseline (41% and 34%), HIV baseline seroprevalence was 4.2% and 1.5%, and high levels of willingness were seen (83% and 78%).