RESUMO
Gladiolus (Iridaceae) is a popular bulbous plant grown worldwide as an ornamental garden plant or cut flower due to its attractive color, size, and flower shape. In April 2012, leaf spots were observed on plants of Gladiolus grandiflorus varieties T-704 and Amsterdam growing in a production area of cut flowers located in the city of Viçosa, Minas Gerais. The oval to round leaf spots were brown with a dark border surrounded by a halo of yellow tissue. Infected leaf samples were deposited in the herbarium at the Universidade Federal de Viçosa (VIC31897). A fungus was isolated from the leaf spots and a single-spore pure culture was initiated and grown on corn meal carrot agar (CCA) medium in petri dishes incubated at 25°C under a 12-h photoperiod for 4 weeks. A sporulating single-spore culture was deposited at the Coleção de Culturas de fungos fitopatogênicos "Prof. Maria Menezes" (UFRPE, Brazil) code CMM 4055. On CCA medium, the fungal isolate initially appeared white, becoming dark after 14 days. Thirty conidia and conidiophores were measured for identification to species. The septate, smooth to pale brown conidiophores were present singly or in groups. The simple, straight or flexuous conidiophores were 42.5 to 82.5 × 3.5 to 7.5 µm and some had a geniculate growth pattern. The majority of conidia were curved at the third (central) cell from the base, which was usually enlarged compared to the end cells. The cells at each end of the 3-distoseptate conidia were pale brown, the intermediate cell brown or dark brown, and the third (central) cell was often the darkest. The basal cell had a protuberant hilum. Conidia were smooth and 20.0 to 33.5 × 10 to 17.5 µm. These characteristics matched well with the description of Curvularia gladioli (1). To confirm this identification, DNA was extracted using a Wizard Genomic DNA Purification Kit and the internal transcribed spacer region (ITS) of rDNA was amplified using ITS1 and ITS4 primers and the partial 28S rDNA region using primers LR0R and LR5. The sequences were deposited in GenBank as accession nos. JX995106 and JX995107, respectively. The ITS sequence matched sequence AF071337, C. gladioli, with 100% identity. This pathogen was first identified as C. lunata, but based on the characteristic of the hilum, spore size, and pathogenicity testing, the fungus was renamed C. trifolii f. sp. gladioli (3). Due to the explicit curvature of the conidia at the third cell and molecular data, the fungus was reclassified as C. gladioli (1,2). To confirm Koch's postulates, 1-month-old healthy plants of G. grandiflorus var. T-704 and Amsterdam (five plants each) were inoculated with a conidial suspension (2 × 104 conidia mL-1) by spraying the foliage and then placed on a growth chamber at 25°C. The control plants were sprayed with distilled water. Symptoms were consistent with those initially observed and all plants developed leaf spots by 4 days post-inoculation. C. gladioli was consistently recovered from the symptomatic tissue and control plants remained symptomless. To our knowledge, this is the first report of C. gladioli causing leaf spot on G. grandiflorus in Brazil. Due to a lack of chemical fungicides for management of this pathogen, further studies to evaluate the susceptibility of the main varieties of gladiolus grown in Brazil to C. gladioli may be necessary. References: (1) G. H. Boerema and M. E. C. Hamers. Neth. J. Plant Pathol. 95:1, 1989. (2) D. S. Manamgoda et al. Fungal Divers. 56:131, 2012. (3) J. A. Parmelee. Mycologia 48:558, 1956.
RESUMO
A very simple and rapid method for the determination of total mercury in fish samples using the Direct Mercury Analyser DMA-80 was developed. In this system, a previously weighted portion of fresh fish is combusted and the released mercury is selectively trapped in a gold amalgamator. Upon heating, mercury is desorbed from the amalgamator, an atomic absorption measurement is performed and the mercury concentration is calculated. Some experimental parameters have been studied and optimised. In this study the sample mass was about 100.0 mg. The relative standard deviation was lower than 8.0% for all measurements of solid samples. Two calibration curves against aqueous standard solutions were prepared through the low linear range from 2.5 to 20.0 ng of Hg, and the high linear range from 25.0 to 200.0 ng of Hg, for which a correlation coefficient better than 0.997 was achieved, as well as a normal distribution of the residuals. Mercury reference solutions were prepared in 5.0% v/v nitric acid medium. Lyophilised fish tissues were also analysed; however, the additional procedure had no advantage over the direct analysis of the fresh fish, and additionally increased the total analytical process time. A fish tissue reference material, IAEA-407, was analysed and the mercury concentration was in agreement with the certified value, according to the t-test at a 95% confidence level. The limit of quantification (LOQ), based on a mercury-free sample, was 3.0 µg kg(-1). This LOQ is in accordance with performance criteria required by the Commission Regulation No. 333/2007. Simplicity and high efficiency, without the need for any sample preparation procedure, are some of the qualities of the proposed method.