First Report of Tomato yellow leaf curl virus in Louisiana.

Tomato yellow leaf curl virus (TYLCV) is a begomovirus (Geminiviridae) that causes a serious disease of tomato throughout the world. In 1997, the strain from Israel of TYLCV (TYLCV-IS) was found infecting tomatoes in Florida for the first time in the United States (1). During late spring of 2000, approximately 90% of the tomato plants (Lycopersicon esculentum) in a farm near New Orleans exhibited severe stunting, leaf cupping, and chlorosis. Symptoms were similar to those caused by TYLCV. Whiteflies (Bemisia tabaci biotype B) were present in the field but in relatively low numbers. The effect on yield reduction varied from negligible (late infections) to 100% (early infections). Six selected plants showing symptoms were assayed by polymerase chain reaction (PCR) using begomovirus-specific primers. Capsicum frutescens infected with an isolate of Texas pepper virus from Costa Rica was used as positive control. DNA was extracted using Plant DNAzol Reagent (GIBCO BRL). PCR was conducted using degenerate primers AV494/AC1048 that amplify the core coat protein region of most begomoviruses (2). PCR yielded a DNA fragment of approximately 550 bp, suggesting that a begomovirus was associated with the disease. The amplified DNA of one field isolate was cloned and the nucleotide (nt) sequence determined. Sequence comparisons with other begomoviruses in the GenBank Database indicated that the Louisiana isolate shared 100% nt identity with TYLCV-IS (GenBank Accession X76319). Successful transmission (100%) to Bonny Best tomato were obtained with four groups of 10 whiteflies each (B. tabaci biotype B) that fed on TYLCV-IS infected tomato plants. Acquisition and transmission feedings were for 2 days. In all cases, the virus was diagnosed by the ability to reproduce typical TYLCV-like symptoms in tomato and PCR. The virus was also successfully graft-transmitted to tomato cv. Bonny Best, Nicotiana benthamiana, and tomatillo (Physalis ixocarpa) using scions from tomato plants infected with a whitefly transmitted virus isolate. This is the first report of TYLCV-IS in Louisiana. References: (1) J. E. Polston et al. Plant Dis. 83:984-988, 1999. (2) S. D. Wyatt and J. K. Brown. Phytopathology 86:1288-1293, 1996.

Stress, anxiety and peripheral benzodiazepine receptor mRNA levels in human lymphocytes.

Peripheral benzodiazepine receptor (PBR) mRNA levels were measured in lymphocytes obtained from a cohort of university students and clinically diagnosed anxious patients. The average level of PBR mRNA was decreased in anxious patients compared to a control group. This data confirms previously published results, but it also indicates that PBR mRNA levels cannot be used as a sole diagnostic measure of anxiety because the range of the individual PBR mRNA levels of the anxious group overlapped the range of the PBR mRNA levels of the control group. PBR mRNA levels in students following academic examinations were increased in some individuals and decreased in others. In the same cohort of students individual levels of cortisol and prolactin were predominantly increased and decreased respectively. There was no correlation between the individual changes in the hormone levels or PBR mRNA, which suggests that each of these parameters is affected by different environmental and physiological factors. Lymphocyte PBR mRNA measurement is a useful additional methodology for studying human stress responses however, its use in clinical studies would require the elucidation of PBR's physiological role.

Biological and Molecular Properties of a Begomovirus from Dicliptera sexangularis.

ABSTRACT Sixangle foldwing, Dicliptera sexangularis (Acanthaceae), showing severe yellow mottle and leaf distortion symptoms was collected from the shoreline of Calusa Island (Lee County, FL). The putative virus was transmitted from infected D. sexangularis to healthy seedlings by mechanical, whitefly (Bemisia tabaci biotype B), and graft-inoculations. Different forms of geminivirus-like DNAs were detected in total DNA extracted from infected plants by Southern blot hybridization analyses using DNA-A and -B of Bean golden mosaic virus (BGMV) from Guatemala as probes. Preliminary polymerase chain reaction experiments and sequence comparisons indicated that the virus was a distinct bipartite begomovirus. The virus was designated Dicliptera yellow mottle virus (DiYMV). Replicative dsDNAs of DiYMV were extracted, digested with selected restriction enzymes, and cloned into a plasmid vector. Both DNA-A and -B were sequenced and compared with those of other begomoviruses. Phylogenetic analyses using AV1, AC1, and BV1 nucleotide sequences indicated that DiYMV has a close relationship with the New World begomoviruses, especially those distributed in the nearby geographic areas of the Florida coast and the Caribbean Basin. However, different percent nucleotide sequence identities and phylogenetic relationships were detected when different open reading frames (ORFs) of DiYMV were compared with their counterparts from begomoviruses from the Caribbean Basin. Based on phylogenetic analyses of the AC1 and BV1 ORFs, DiYMV was closely related to BGMV type II isolates, whereas sequence comparisons of the common region and the AC4-derived amino acid sequences indicated its close relationship with Potato yellow mosaic virus from Venezuela.

Detection of a Geminivirus Infecting Sweet Potato in the United States.

In 1994, a sweet potato sample showing leaf curl symptoms was collected from the field in Louisiana. When graft-inoculated, Ipomoea nil cv. Scarlett O'Hara reacted with severe leaf distortion and chlorosis symptoms. I. aquatica reacted with a bright yellow mottle. The virus isolated was designated the United States isolate of sweet potato leaf curl virus (SPLCV-US). It was transmitted to I. nil by the sweet potato whitefly, Bemisia tabaci biotype B. DNA probes prepared with component A of pepper Huasteco geminivirus, with an isolate of bean golden mosaic geminivirus from Guatemala, with an isolate of tomato mottle geminivirus from Florida, and with an isolate of tomato yellow leaf curl geminivirus from the Dominican Republic (TYLCV-DR) hybridized with a 2.6-kb DNA band present in DNA extracts from plants infected with SPLCV-US. Probes prepared with the B component of these geminiviruses did not hybridize with these DNA extracts. We were unable to amplify SPLCV-US DNA products by polymerase chain reaction (PCR) in quantities that could be visualized by ethidium bromide staining. However, Southern blots from amplifications with primers AV494/AC1048 revealed PCR products of approximately 600 bp and 550 bp when hybridized with the TYLCV-DR probe. These results were consistently obtained from infected I. cordatotriloba and less consistently from I. aquatica or I. setosa. Fibrillar inclusions were occasionally seen, and granular aggregates of viruslike particles were observed in the nucleus of infected I. cordatotriloba. These results suggest that the virus isolated from sweet potato with leaf curl symptoms belongs to the geminivirus group.

First Report of Erwinia carotovora subsp. carotovora Causing Bacterial Root Rot of Sweetpotato (Ipomoea batatas) in Louisiana.

Bacterial root and stem rot of sweetpotato (Ipomoea batatas (L.) Lam.) was first fully characterized in the U.S. in 1977 (2). It was thought to be caused exclusively by Erwinia chrysanthemi. Although a previous report described sweetpotato as a host for E. carotovora subsp. carotovora, based on artificial inoculations, others have reported that neither E. carotovora subsp. carotovora nor E. carotovora subsp. atroseptica decay sweetpotato storage roots (1). In October 1995, storage roots of sweetpotato cv. Beauregard were received from St. Landry Parish, LA, that displayed typical bacterial root rot. Isolations from these roots yielded bacteria that showed a similarity of 0.945 to E. carotovora subsp. carotovora with the Biolog GN Bacterial Identification System (version 3.50). This isolate (Ecc-LH) also differed from isolates of E. chrysanthemi (Ech) from sweetpotato and other hosts in that it was insensitive to erythromycin, did not produce phosphatase or lecithinase, and did not produce gas from glucose. Ecc-LH differed from known strains of E. carotovora subsp. atroseptica in that it did not produce reducing substances from sucrose or acid from palatinose. When Beauregard storage roots were inoculated by inserting micropipette tips containing 50 µl of 1.0 × 108 CFU/ml, both Ecc-LH and Ech-48 produced typical bacterial root rot symptoms. However, when they were compared by infectivity titrations at 28 to 32°C, Ecc-LH was less virulent than Ech-48. Ecc-LH had an ED50 of approximately 1.0 × 106 CFU/ml and did not cause appreciable disease below inoculum concentrations of 1.0 × 105, whereas Ech-48 had an ED50 of approximately 1.0 × 108 and caused soft rot at the lowest concentration tested, 1.0 × 103. Similar disease incidence was observed in infectivity titrations at 22 to 24°C, but Ech-48 caused less severe soft rot. E. carotovora subsp. carotovora was reisolated from inoculated storage roots and its identity was reconfirmed by Biolog. When terminal vine cuttings of Beauregard were dipped in 1.0 × 108 CFU/ml and planted in a greenhouse, bacterial stem rot symptoms developed on plants inoculated with Ech-48 at about 4 weeks postinoculation, or when new growth began. However, no symptoms developed on plants inoculated with Ecc-LH. This is the first report of natural occurrence of E. carotovora subsp. carotovora causing bacterial root rot of sweetpotato in Louisiana. E. chrysanthemi remains the most important pathogen causing bacterial soft rot in sweetpotato since it is widely associated with sweetpotato, is more virulent on storage roots and also causes a stem rot. E. carotovora subsp. carotovora can cause root rot, but has been isolated in only one location to date, is less virulent on storage roots, and apparently does not cause stem rot on the predominant cultivar in U.S. sweetpotato production, Beauregard. References: (1) C. A. Clark and J. W. Moyer. 1988. Compendium of Sweet Potato Diseases. American Phytopathological Society, St. Paul, MN. (2) N. W. Schaad and D. Brenner. Phytopathology 67:302, 1977.

Purification and characterization of lipase from Aeromonas sobria LP004.

Lipase from Aeromonas sobria LP004, isolated from raw milk, was purified and characterized. The lipase was purified 10.29 fold to a homogeneous state by ultrafiltration and column chromatography on phenyl sepharose. The molecular weight of the lipase determined by SDS-PAGE was 97 kDa. Purified A. sobria LP004 lipase exhibited the maximum activity at pH 6.0 and 45 degrees C and was stable under alkaline conditions (pH 6.5-10.0) and at temperatures lower than 40 degrees C. This lipase could be classified as a 1,3-position specific enzyme and its catalytic activity was calcium dependent. PMSF, a serine enzyme inhibitor and 2-mercaptoethanol, a reducing agent, did not affect the enzyme activity.