La organización y función del genoma del virus del mosaico dorado del frijol

Programa Cooperativo Regional de Frijol para Centroamérica, México y el Caribe (PROFRIJOL)Cooperación Suiza para el Desarrollo (COSUDE)Centro Internacional de Agricultura Tropical (CIAT)UCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias Agroalimentarias::Estación Experimental Agrícola Fabio Baudrit Moreno (EEAFBM

Identificaci?n de un nuevo begomovirus en mel?n (Cucumis melo L.) en Lara, Venezuela

3 ilus. 2 tab. 35 ref.Se recolectaron muestras de ?pices foliares de plantas de mel?n (Cucumis melo L.) que mostraban s?ntomas de mosaico clor?tico en campos cultivados del Estado de Lara, Venezuela. Con el objetivo de identificar geminivirus, se utilizaron iniciadores generales y la t?cnica de reacci?n en cadena de la polimerasa (PCR) para amplificar la parte superior del componente A y la regi?n hipervariable del componente B del genoma viral. en cuatro muestras representativas, se obtuvieron fragmentos de 1400 y 400 pares de bases, respectivamente. Dichos fragmentos fueron clonados y secuenciados. Las secuencias obtenidas se compararon con otras secuencias de geminivirus en la base de datos GenBank, determin?ndose una similitud m?xima (82 por ciento) con el virus de enrollamiento de la hoja del ayote (squash leaf curl virus, SLCV). Estos an?lisis de secuencia indican que se identific? un nuevo geminivirus en mel?n en el estado Lara, Venezuela, para el cual se propuso el nombre de melon chlorotic mosaic virus (MCMV). Se report? la secuencia obtenida al GenBank (n?mero de acceso AF453447, 11 de diciembre del 2001). adem?s, se gener? una sonda molecular espec?fica para el MCMV, que se utilizar? para estudios epidemiol?gicos futuros Foliar apex samples were collected from melon (Cucumis melo L.) plants that showed chlorotic mosaic symptoms in cultivated fields of Lara, Venezuela. In order to identify geminivirus, universal primers and the polimerase chain reaction (PCR) technology were used for amplifying the top part of the A component and the hypervariable region of the B component of the viral genome. Fragments of 1400 and 400 base pairs in four representative samples were obtained. The above mentioned fragments were cloned and sequenced. The sequences obtained were compared with other geminivirus sequences from the GenBank database. A maximum similarity (82 per cent) with the squash leaf curl virus (SLCV) was observed. These sequence anlyses indicate that a new geminivirus was identified in melon cultivars from Lara, Venezuela. we propose the name of melon chlorotic mosaic virus (MCMV) for the new geminivirus and the obtained sequence was submitted to the GenBank database (accession number AF453447, December 11, 2001). A molecular specific probe was generated to detect MCMV and it will be useful for future epidemiological studies

Comparative Analysis of Three Different Total Nucleic Acid Extraction Protocols for the Diagnosis of Geminiviruses in Squash (Cucurbita moschata)

Squash (Cucurbita moschata) is one of the most important crops in tropical countries. Geminiviruses are an important group of plant pathogens. In 2002 a new begomovirus was reported to naturally infect squash and some other crops in Costa Rica. Our objective was to compare, using molecular techniques, the extraction and further purification of DNA from squash by different extraction protocols and storage methods. A single infected sample was collected, half of the material was stored frozen at ) -70C, and the remainder was stored dehydrated in silica gel (SG). Total nucleic acids (TNAs) were extracted by three different protocols and were quantified by fluorometry, and the quality was analysed by electrophoresis in agarose gels, polymerase chain reaction (PCR) of the virus genome, dot blot and Southern blot hybridization. Even though the tissue stored in SG yielded a higher amount of TNAs, the genetic material exhibited lower integrity and this made it useful exclusively for the detection of geminiviral DNA by PCR amplification of short viral sequences and by hybridization with short viral probes. The Dellaporta method proved to be the most effective for the detection of geminiviral DNA in infected squash tissue. Although the cetyltrimethylammonium bromide method showed similar results, the procedure is more time-consuming. Surprisingly, the citrate method showed either similar or worse results than the other methods.Universidad de Costa Rica/[801-A2-525]/UCR/Costa RicaInternational Centre for Genetic Engineering and Biotechnology/[CRP⁄COS-02⁄02]/ICGEB/IndiaUCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias Básicas::Centro de Investigación en Biología Celular y Molecular (CIBCM)UCR::Vicerrectoría de Docencia::Salud::Facultad de Microbiologí

Molecular characterization of the S1 gene in GI-17 and GI-13 type isolates of avian infectious bronchitis virus (IBV) in Costa Rica, from 2016 to 2019

Avian infectious bronchitis is one of the most important respiratory diseases affecting poultry production worldwide. The etiological agent of this disease is the avian infectious bronchitis virus (IBV). We analyzed 14 isolates of IBV obtained from poultry farms in Costa Rica, from 2016 through 2019. We sequenced the S1 region of the genome and the sequences obtained were submitted to GenBank. Phylogenetic analyses showed that the isolates obtained during 2016–2017 belong to the GI-17 lineage and are related to the Georgia 13-type Ga-13/14255/14 and CK/CR/1160/16 variants, with a 96.90–100% nucleotide sequence identity and a 92.25–100% amino acid sequence identity. The main differences were detected in the RBD and HVR-3 regions, where a series of mutations eliminate an N-glycosylation site in 10 out of 11 isolates. The isolates obtained during 2018–2019 belong to the GI-13 lineage and are closely related to the 4/91 vaccine variant, with over 98% sequence identity at the nucleotide and amino acids levels. Variations were detected in the RBD and HVR regions, with a possible N-glycosylation site detected in isolate CK/CR/0632/19. These results indicate that a GA13-like pathogenic variant circulated during the 2016–2017 period and that the 4/91 variant was detected after the introduction of the vaccine. The variations shown in both the GA13-like and 4/91 isolates examined, reveal the need for continuous surveillance of IBV in Costa Rica, to detect new variants that may be introduced to the country or develop during outbreaks. This information is highly relevant for vaccination planning and disease management programs.UCR::Vicerrectoría de Docencia::Ciencias Básicas::Facultad de Ciencias::Escuela de BiologíaUCR::Vicerrectoría de Docencia::Ciencias Agroalimentarias::Facultad de Ciencias Agroalimentarias::Escuela de Zootecni

Molecular characterization of an avian GA13-like infectious bronchitis virus full-length genome from Costa Rica

We describe the first whole-genome sequence of a GA13-like isolate of avian infectious bronchitis virus CK/CR/1160/16 (MN757859), obtained in 2016 in the province of Alajuela, Costa Rica. This virus caused an outbreak with great economic impact to the local poultry industry. The genome sequence is 27 696 bp in length, with the following genome organization 5′-UTR-Pol-S-3a-3b-E-4b-4c-M-5a-5b-N-6b-3′-UTR. The complete genome sequence has the highest sequence identity (94.03%) with DMV/1639/GA9977/2019 (MK878536) from Georgia, USA, and the lowest identity (86.03%) with ck/CH/LHLJ/08-6 (KX252788), from China. Analysis of the S1 subunit indicates that the Costa Rican isolate belongs to genotype I, lineage 17 (GI-17) and displays 96.89% identity with the S1 subunit of Ga-13/14255/14 (KM087780) (USA). Possible recombination events in genes S, E, M, 4b y 4c were detected, with Massachusetts, Connecticut, Arkansas and MA5 as potential parental types. This study highlights the importance of the epidemiological and molecular surveillance of avian infectious bronchitis.Fundación para el Fomento y Promoción de la Investigación y Transferencia de Tecnología Agropecuaria/[3-006-115123]/FITACORI/Costa RicaUniversidad de Costa Rica//UCR/Costa RicaUCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias Agroalimentarias::Centro de Investigación en Nutrición Animal (CINA