Tobacco: A New Natural Host of Tomato chlorosis virus in Spain.

In March 2013, symptoms of mild leaf curling, mosaic, and interveinal yellowing were observed in tobacco (Nicotiana tabacum) plants grown in a row surrounding the exterior of a greenhouse containing a tomato crop in Guía de Isora, Tenerife (Canary Islands, Spain). The tobacco plants were found lightly infested by the whitefly (Hemiptera: Aleyrodidae) Bemisia tabaci. The greenhouses in this area are devoted to the commercial production of tomato. The farmers grow some tobacco plants inside and outside of them as a reservoir of parasitoids and depredators of B. tabaci. This insect is the natural vector of the main viruses severely affecting tomato in the Canary Islands, the begomovirus Tomato yellow leaf curl virus and the crinivirus Tomato chlorosis virus (ToCV). ToCV was detected in Spain in 1997 (2) and has become established in most of the coastal provinces of eastern and southern continental Spain and in the Canary Islands. Approximately 50% of the tomato plants grown inside the greenhouse close to the tobacco plants showed typical ToCV symptoms, and infection by this virus was confirmed in the seven plants tested by reverse transcription (RT)-PCR using specific coat protein gene (CP) primers (see below). Total RNA was extracted with TRIzol Reagent (Invitrogen) from leaves of five tobacco plants showing the symptoms mentioned above and analyzed by dot-blot hybridization using digoxigenin-labeled RNA probes to the CP gene of ToCV. Positive signal was obtained for all five plants. RT-PCR reactions were performed with specific primers for the detection of ToCV, MA380(+) (5'-GTGAGACCCCGATGACAGAT-3') and MA381(-) (5'-TACAGTTCCTTGCCCTCGTT-3'), specific to the CP gene (ToCV RNA 2) (3), and MA396(+) (5'-TGGTCGAACAGTTTGAGAGC-3') and MA397(-) (5'-TGAACTCGAATTGGGACAGA-3'), specific to the RNA-dependent RNA polymerase (RdRp) gene (ToCV RNA 1) (1). DNA fragments of the expected size (436 and 763 bp, respectively) were obtained, thus supporting the presence of ToCV in the symptomatic samples. The amplified product of the RdRp gene fragment from one sample was directly sequenced (Macrogen Inc., South Korea) and resulted closely related to ToCV isolates from Sudan (GenBank Accession No. JN411686, 99.6% nt identity) and Spain (DQ983480, 99.4% nt identity), thereby confirming the infection by this virus. Partial sequence of the ToCV isolate from tobacco was deposited in GenBank under accession no. KJ175084. In addition, all five plants resulted positive when analyzed by ELISA for Tomato spotted wilt virus and Potato virus Y and by PCR for Tomato yellow leaf curl virus (data not shown), all three viruses reported to infect naturally tobacco. Although tobacco has been reported as an experimental host of ToCV (4), to our knowledge, this is the first report of this species as a natural host of this virus. The finding of ToCV infecting tobacco raises the question of whether this virus could emerge as a pathogen of this crop and questions the use that farmers make of tobacco as reservoirs of natural enemies for whitefly control in tomato. References: (1) G. Lozano et al. J. Virol. 83:12973, 2009. (2) J. Navas-Castillo et al. Plant Dis. 84:835, 2000. (3) H. P. Trenado et al. Eur. J. Plant Pathol. 118:193, 2007 (4) W. M. Wintermantel and G. C. Wisler. Plant Dis. 90:814, 2006.

Copy number variation and missense mutations of the agouti signaling protein (ASIP) gene in goat breeds with different coat colors.

In goats, classical genetic studies reported a large number of alleles at the Agouti locus with effects on coat color and pattern distribution. From these early studies, the dominant A(Wt) (white/tan) allele was suggested to cause the white color of the Saanen breed. Here, we sequenced the coding region of the goat ASIP gene in 6 goat breeds (Girgentana, Maltese, Derivata di Siria, Murciano-Granadina, Camosciata delle Alpi, and Saanen), with different coat colors and patterns. Five single nucleotide polymorphisms (SNPs) were identified, 3 of which caused missense mutations in conserved positions of the cysteine-rich carboxy-terminal domain of the protein (p.Ala96Gly, p.Cys126Gly, and p.Val128Gly). Allele and genotype frequencies suggested that these mutations are not associated or not completely associated with coat color in the investigated goat breeds. Moreover, genotyping and sequencing results, deviation from Hardy-Weinberg equilibrium, as well as allele copy number evaluation from semiquantitative fluorescent multiplex PCR, indicated the presence of copy number variation (CNV) in all investigated breeds. To confirm the presence of CNV and evaluate its extension, we applied a bovine-goat cross-species array comparative genome hybridization (aCGH) experiment using a custom tiling array based on bovine chromosome 13. aCGH results obtained for 8 goat DNA samples confirmed the presence of CNV affecting a region of less that 100 kb including the ASIP and AHCY genes. In Girgentana and Saanen breeds, this CNV might cause the A(Wt) allele, as already suggested for a similar structural mutation in sheep affecting the ASIP and AHCY genes, providing evidence for a recurrent interspecies CNV. However, other mechanisms may also be involved in determining coat color in these 2 breeds.

IL6, IL10 and TGFB1 gene polymorphisms in coeliac disease: differences between DQ2 positive and negative patients.

UNLABELLED: Predisposition to coeliac disease (CD) might be partially due to an individual pattern of hyper-inflammatory biased immune response. One of these patterns of intense response may be linked to the haplotype carrying HLA-DQ2 alleles and TNF -308A allele. However, 10 % of CD patients do not express the DQ2 heterodimer and these do not usually carry the TNF -308A allele. A similar response might be achieved by genes codifying other cytokines. OBJECTIVES: To study biallelic polymorphisms in genes codifying for TNFalpha, IL10, IL6 and TGFbeta1 in DQ2 negative CD patients and to compare the results with DQ2 positive patients and healthy controls, in order to establish whether any of these polymorphisms have a role in CD susceptibility. METHODS: TNF -308 (G > A), IL-6 -174 (G > C) and TGFB1 codon 10 (+ 869, T > C) and codon 25 (+ 915, G > C) polymorphisms and IL-10 haplotype of polymorphisms in positions -1082 (G > A), -819 (C > T) and -592 (C > A) were typed by a SSP-PCR technique. RESULTS: The distribution of allele frequencies for TNF -308 is different between DQ2 positive CD patients and controls and the same occurs for haplotype frequencies of the IL10 promoter (-1082, -819, -592): The frequencies of the TNF -308A allele (p = 0.027), TNF -308A carriers (p = 0.031) and of IL10GCC haplotype are increased (p = 0.013) in DQ2 positive CD patients. However, the IL6 -174 allele G is more frequent in DQ2 negative patients than in healthy controls (p = 0.018), DQ2 negative controls (p = 0,018), and DQ2 positive patients (p = 0.008). CONCLUSIONS: DQ2 negative CD patients show an increased frequency of genotypes associated to IL6 high production. These were mainly allele G homozygous for the IL6 gene (-174) polymorphism. The IL6 -174GG genotype (homozygous) may be an additional risk marker for CD in DQ2 negative patients, representing an alternative susceptibility factor for CD when TNF -308A is negative.

IL6, IL10 and TGFB1 gene polymorphisms in coeliac disease: differences between DQ2 positive and negative patients

Predisposition to coeliac disease (CD) might be partially due to an individual pattern of hyper-inflammatory biased immune response. One of these patterns of intense response may be linked to the haplotype carrying HLA-DQ2 alleles and TNF-­308A allele. However, 10 % of CD patients do not express the DQ2 heterodimer and these do not usually carry the TNF-­308A allele. A similar response might be achieved by genes codifying other cytokines. Objectives: To study biallelic polymorphisms in genes codifying for TNFá, IL10, IL6 and TGFâ1 in DQ2 negative CD patients and to compare the results with DQ2 positive patients and healthy controls, in order to establish whether any of these polymorphisms have a role in CD susceptibility. Methods: TNF-­308 (G > A), IL-6 ­174 (G > C) and TGFB1 codon 10 (+ 869, T > C) and codon 25 (+ 915, G > C) polymorphisms and IL-10 haplotype of polymorphisms in positions ­1082 (G > A), ­819 (C > T) and ­592 (C > A) were typed by a SSP-PCR technique. Results: The distribution of allele frequencies for TNF ­308 is different between DQ2 positive CD patients and controls and the same occurs for haplotype frequencies of the IL10 promoter (­1082, ­819, ­592): The frequencies of the TNF-­308A allele (p = 0.027), TNF ­308A carriers (p = 0.031) and of IL10GCC haplotype are increased (p = 0.013) in DQ2 positive CD patients. However, the IL6 ­174 allele G is more frequent in DQ2 negative patients than in healthy controls (p = 0.018), DQ2 negative controls (p = 0,018), and DQ2 positive patients (p = 0.008). Conclusions: DQ2 negative CD patients show an increased frequency of genotypes associated to IL6 high production. These were mainly allele G homozygous for the IL6 gene (­174) polymorphism. The IL6 ­174GG genotype (homozygous) may be an additional risk marker for CD in DQ2 negative patients, representing an alternative susceptibility factor for CD when TNF -308A is negative

No disponible