Judith K Brown

doi:10.1128/JVI.00880-12.

Abstract:

A Cotton leaf crumple virus (CLCrV)-based gene silencing vector containing a fragment of the Gossypium hirsutum Magnesium chelatase subunit I was used to establish endogenous gene silencing in cotton of varied genetic backgrounds. Biolistic inoculation resulted in systemic and persistent photo-bleaching of the leaves and bolls of the seven cultivars tested, however, the intensity of silencing was variable. CLCrV-VIGS-mediated expression of green fluorescent protein was used to monitor the in planta distribution of the vector, indicating successful phloem invasion in all cultivars tested. Acala SJ-1, one of the cotton cultivars, was identified as a particularly optimal candidate for CLCrV-VIGS-based cotton reverse-genetics. © 2010 Elsevier Ltd.

Abstract:

The ultrastructure of the endosymbionts of several populations of whitefly (Homoptera: Aleyrodidae) was examined using transmission electron microscopy. Consistent differences in morphology and relative number of endosymbionts were observed between species and biotypes of whitefly within the Bemisia taxon. Bemisia argentifolii (=B. tabaci B biotype) individuals from Hawaii, Florida, and Arizona contained two morphological types of microorganisms housed within the mycetocyte cells of immature whiteflies. In contrast, individuals from populations of B. tabaci A biotype from Arizona and Mexico, and B. tabaci Jatropha biotype from Puerto Rico, consistently contained three distinct morphological types of microorganisms within their mycetocytes. Organisms from B. tabaci A and Jatropha biotypes differed from each other in the relative frequency of each type of microorganism. These observations suggest that different whitefly biotypes may have variable combinations of micro-fauna, with some possibly unique to each group, and furthers the hypothesis that variation in whitefly endosymbionts may be associated with the development of biotypes. © 1995 Springer-Verlag.

Abstract:

Cucurbit yellow stunting disorder virus (CYSDV) (genus, Crinivirus: family, Closteroviridae) is an emerging plant pathogen, transmitted by the sweet potato whitefly Bemisia tabaci Gennadius (Hemiptera: Aleyrodidae), which infects cucurbit crops causing significant economic losses. A TaqMan^® real-time fluorescent, one-step reverse transcription (RT), polymerase chain reaction (PCR) assay for the detection of the virus has been developed and optimized. The assay is over 100-fold more sensitive than conventional RT-PCR and involves template preparation that does not require RNA purification. The assay can be accomplished either by first spotting the sap extract on a positively charged nylon membrane and elution, or by the direct addition of crude plant extract into the real-time reaction cocktail. Several factors affecting the efficiency of the tests were studied, such as the type and amount of reverse transcription (RT) enzymes and the use of different additives on the elution extract. The addition of 5 units of RT enzymes in the real-time PCR cocktail and the use of Tween 20, Triton X and Betaine in the virus release buffer resulted in improved detection efficiency. The applicability of the real-time RT-PCR assay was validated with CYSDV isolates from the USA, Mexico, the Mediterranean Basin, Jordan, and the United Arab Emirates and provides a simple, efficient and accurate detection technique, whereas the membrane preparation techniques can be used for long-term storage of samples allowing the shipment of samples from the field to remote laboratories for testing without compromising the reliability of the test. ©, 2009 Blackwell Verlag GmbH.

PMID: 18943795;Abstract:

The bipartite DNA genome of Cotton leaf crumple virus (CLCrV), a whitefly-transmitted begomovirus from the Sonoran Desert, was cloned and completely sequenced. The cloned CLCrV genome was infectious when biolistically delivered to cotton or bean seedlings and progeny virus was whitefly- transmissible. Koch's postulates were completed by the reproduction of characteristic leaf crumple symptoms in cotton seedlings infected with cloned CLCrV DNA, thereby verifying the etiology of leaf crumple disease, which has been known in the southwestern United States since the 1950s. Sequence comparisons confirmed that CLCrV has a genome organization typical of yet sufficiently divergent from all other bipartite begomoviruses to justify recognition as a distinct species. Phylogenetic analyses indicated that CLCrV has a complex evolutionary history probably involving both recombination and reassortment. The relatively low nucleotide sequence identity (77%) of the common region shared by the CLCrV DNA-A and DNA-B components and the distinct phylogenetic relationships of each component are consistent with component reassortment. Sequence analyses indicated that the CLCrV DNA-A component was likely derived by recombination among ancestors of two divergent clades (e.g., the Squash leaf curl virus [SLCV] clade and the Abutilon mosaic virus clade) of Western Hemisphere begomoviruses. The CLCrV DNA-B component also may have originated by recombination among an ancestor of the SLCV clade and another distantly related but unknown Western Hemisphere begomovirus.