First Report of Stewart's Wilt of Maize in Argentina Caused by Pantoea stewartii.

Stewart's wilt is a serious disease of corn (Zea mays L.) caused by the bacterium Pantoea stewartii subsp. stewartii (Pss). Typical symptoms of infected fields and dent corn are longitudinal streaks with irregular or wavy margins, which are parallel to the veins and may extend the length of the leaf. These pale to green yellow lesions become dry and brown as the disease progresses producing a leaf blight (4). During the growing seasons 2010 to 2011 and 2011 to 2012, symptoms of bacterial leaf blight of corn were observed in central Argentina maize fields, with an incidence of 54% in Córdoba province. To identify the pathogen, leaves from 10 symptomatic maize plants per field were collected from 15 fields covering a representative geographical area. High populations of morphologically uniform bacteria were isolated from leaf tissues by conventional methods using King's medium B agar (2). Ten representative facultatively anaerobic gram-negative, non-fluorescing, non-motile, catalase positive and oxidase negative rod-shaped and yellow-pigmented bacterial isolates were evaluated further. The biochemical profile obtained was: fermentative metabolism, negative indol, acetoin and hydrogen sulfide production, negative gelatin hydrolysis (22°C), positive acid production from D-glucose and lactose, negative gas production from D-glucose, and negative nitrate reduction (1). All the isolates produced a 300-bp band with PCR using the species specific primer pair PST3581/PST3909c (3). The Pss ATCC 8199 and Pseudomonas fluorescens ATCC 13525 strains were used as positive and negative controls for the PCR assays, respectively. The pathogenicity test was performed by stem inoculation of five to ten P2069 YR maize plants (one to two leaf growth stage) grown in growth chamber. Plants were inoculated by syringe with a 107 to 108 cell/ml bacterial suspension and kept in a humid chamber at 25 to 27°C. Plants inoculated with Pss ATCC 8199 or with sterile water were used as positive and negative control treatments, respectively. The development of symptoms similar to those originally found in the field was observed on all the plants inoculated with the different isolates at 7 to 10 days post inoculation. In addition, symptoms on inoculated plants were similar to those observed for the positive control treatment. No symptoms were found on negative controls. Koch's postulates were fulfilled since bacteria isolated from symptomatic tissue had identical characteristics to isolates used to inoculate plants and to the reference Pss strain for biochemical tests and PCR reaction mentioned above. To our knowledge, this is the first report of P. stewartii subsp. stewartii isolated from diseased maize in Argentina. References: (1) J. G. Holt et al. Page 179 in: Bergey's manual of determinative bacteriology. Williams and Wilkins, Baltimore, MD, 1994. (2) OEPP/EPPO. Bulletin OEPP/EPPO Bulletin, 36: 111, 2006. Pantoea stewartii subsp. stewartii diagnostic. (3) A. Wensing et al. Appl. Environ. Microbiol. 76:6248, 2010. (4) D. G. White Page 4 in: Compendium of corn disease. The American Phytopathology Society, 1999.

First Report in Argentina of Plum pox virus Causing Sharka Disease in Prunus.

Sharka disease, caused by Plum pox virus (PPV), is probably the most important disease of stone fruits crops worldwide because of tremendous yield losses from infected trees (1). During November 2004, symptoms resembling sharka disease were observed in a plum and apricot orchard consisting of 5,000 trees in Pocito, San Juan Province, Argentina. Apricot leaves showed chlorotic spots while plum leaves showed chlorotic rings, spots, and irregular edges. Plum fruits were deformed and much smaller than those from symptomless trees. Samples collected from 70 symptomatic trees were tested using double-antibody sandwich enzyme-linked immunosorbent assays with a polyclonal antiserum anti-PPV from BIOREBA (Reinach BL1, Switzerland), and immunosorbent electron microscopy with a polyclonal antiserum from our laboratory made against a recombinant PPV capsid protein (CP). The samples were also tested using double-antibody sandwich indirect enzyme-linked immunosorbent assay using the REAL kit (Durviz, Valencia, Spain) with two different monoclonal antibodies including Mab 5b that recognizes all strains of PPV and Mab 4DG5 that is specific for PPV strain D. Samples were positive with both antibodies in 80% of the cases. Leaf extracts from symptomatic plum samples were also analyzed by immuno-capture reverse-transcription polymerase chain reaction. A 1,209-bp fragment was amplified with specific primers that anneal at the 5' end of the coat protein coding region and the viral 3' end poly A tail. The amplified fragment was cloned and the nucleotide sequence was determined for two of the resulting clones (Gen-Bank Accession Nos. DQ299537 and DQ299538). The sequences were 98% identical with the PPV-strain D from the United States (GenBank Accession No. AF360579) and Germany (GenBank Accession No. X81081). The restriction sites for AluI and RsaI, previously described (2) as typical for the PPV-D strain, were present in the expected positions. To our knowledge, this is the first report of PPV-D in Argentina. Reference: (1) M. Németh. Virus, Mycoplasma, and Rickettsia Disease of Fruit Trees. Martinus Nijhoff Publishers, Dordrecht, the Netherlands, 1986. (2) T. Wetzel et al. J. Virol. Methods 33:355, 1991.

Immunocapture reverse transcription-polymerase chain reaction combined with nested PCR greatly increases the detection of Prunus necrotic ring spot virus in the peach.

A detection system based on nested PCR after IC-RT-PCR (IC-RT-PCR-Nested PCR) was developed to improve indexing of Prunus necrotic ringspot virus in peach trees. Inhibitory effects and inconsistencies of the standard IC-RT-PCR were overcome by this approach. IC-RT-PCR-Nested PCR improved detection by three orders of magnitude compared with DAS-ELISA for the detection of PNRSV in leaves. Several different tissues were evaluated and equally consistent results were observed. The main advantages of the method are its consistency, high sensitivity and easy application in quarantine programs.

A triply avulsed tooth.

Canada has a rich hockey tradition. Accordingly, Canadian dentists are well acquainted with tooth avulsion, although the emergence as the mouth guard as an essential component of protective sports equipment has reduced its incidence. Practitioners occasionally examine patients who have lost teeth as a result of accidents. In this unusual case, a patient's tooth was removed 3 times in 12 hours.

Sequencing and characterization of the coat protein and 3' non-coding region of a new sweet potato potyvirus.

A cDNA library was constructed from viral genomic RNA purified from sweet potato plants affected by "Sweet Potato Chlorotic Dwarf disease" in an attempt to clarify the etiology of this viral complex in Argentina. By sequence analysis, some of the obtained clones were found to belong to sweet potato feathery mottle potyvirus (SPFMV), to a closterovirus and to a new potyvirus. A cDNA clone of 1,103 bp representing the coat protein cistron and 3' non-coding region of the newly identified potyvirus was further characterized. The sequence contained an ORF of 855 nucleotides with a coding capacity of 285 amino acids, followed by a 3' untranslated tail of 248 nucleotides. The core and C-terminal regions have sequences well conserved among potyviruses. Furthermore, amino acid sequence comparisons of the capsid protein with those of other described potyviruses showed 63% homology with SPFMV, 68 to 70% with two different isolates of sweet potato latent potyvirus (SPLV), 57% with sweet potato G potyvirus (SPGV) and 73% with potato virus Y (PVY). These data allowed us to propose the inclusion of this virus as a new member of the family Potyviridae, genus Potyvirus with the designation sweet potato mild speckling potyvirus (SPMSV).

Gene I mutants of peanut chlorotic streak virus, a caulimovirus, replicate in plants but do not move from cell to cell.

Gene I of peanut chlorotic streak virus (PCISV), a caulimovirus, is homologous to gene I of other caulimoviruses and may encode a protein for virus movement. To evaluate the function of gene I, several mutations were created in this gene of an infectious, partially redundant clone of PCISV. Constructs with an in-frame deletion and a single amino acid substitution in gene I were not infectious. To test for replication of these mutants in primarily infected cells, an immunosorbent PCR technique was devised. Virus particles formed by mutants in plants were recovered by binding to antivirus antibodies on a solid matrix and DNase treated to discriminate against residual inoculum, and DNA of trapped virions was subjected to PCR amplification. Gene I mutants were shown to direct formation of encapsidated DNA as revealed by a PCR product. Control gene V mutants (reverse transcriptase essential for replication) did not yield a PCR product. Quantitative PCR allowed estimation of the proportion of cells initially infected by gene I mutants and the amount of extractable virus per cell. It is concluded that PCISV gene I encodes a movement protein and that the immunoselection-PCR technique is useful in studying subliminal virus infection in plants.