Virginia Agricultural Experiment Station

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https://hdl.handle.net/10919/24235
The Virginia Agricultural Experiment Station conducts research on food and fiber systems, their impact on the environment, and their relation to the future needs of Virginia, the nation, and the world.

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Browsing Virginia Agricultural Experiment Station by Subject "0605 Microbiology"

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    Age related differences in phylogenetic diversity, prevalence of Shiga toxins, Intimin, Hemolysin genes and select serogroups of Escherichia coli from pastured meat goats detected in a longitudinal cohort study
    Ndegwa, Eunice; Alahmde, Aber; Kim, Chyer; Kaseloo, Paul; O'Brien, Dahlia (BMC, 2020-07-30)
    Background: Little is known on significance, diversity and characteristics of gut E. coli in goats despite their importance as food animals globally. We characterized the temporal dynamics in diversity of E. coli in fecal samples from a cohort of goat kids and adult meat goats on pasture over a one-year period. Isolates were characterized based on phylogenetic grouping, virulence genes; shiga toxins 1 and 2 (Stx1&Stx2) (STEC), intimin (eaeA), hemolysin (hly) and select important sero-groups (026, 045, 0103, 0126 and 0146) using molecular methods. Results: A total of 516 E. coli isolates were screened. Prevalence of virulence genes and STEC was 65 and 56% respectively. Prevalence of virulence genes and STEC was significantly higher in goat kids less than six months (76% /66%) than adults (48% /28%). Isolates with virulence profiles of two or more genes were also higher in young goat kids (50%) than adults (20%). Entero-pathogenic E. coli (EPEC-eaeA gene only) were mostly from pre-weaned goat kids while hly gene only isolates were significantly higher in adults. The stx1, stx2 and hly genes peaked around weaning (60, 63 and 52%) respectively. Goats kids were mostly hosts to group D (59%) while adults older than one year had B1 (75%) isolates. Group D isolates were most abundant at weaning (64%) and diarrhea samples (74%). Group B2 isolates overall (6%) were mostly detected around weaning (63%) while A isolates were 4% overall. Twenty-four isolates belonged to sero-groups 026, 0103 and 0146 with 70% of the isolates detected around weaning. Nineteen of these isolates were STEC with most harboring the stx1/stx2/hly/eae (25%) profile. Most belonged to O26 sero-group (75%) and phylogroup D (75%). Conclusion: To our knowledge this is the first study to highlight longitudinal age related differences in E. coli phylogenetic diversity, abundance of virulence genes and select important sero-groups in goats. Differences detected suggest a possible role of age and weaning stress in influencing E. coli diversity in the gut of goats. The findings are relevant to both animal and public health to advise on further studies on caprine E. coli isolates as animal and human pathogens.
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    Conidial production and viability of Calonectria pseudonaviculata on infected boxwood leaves as affected by temperature, wetness, and dryness periods
    Avenot, Herve F.; Baudoin, Antonius B.; Hong, Chuanxue (Wiley, 2021-10-31)
    Calonectria pseudonaviculata causes lesions on boxwood leaves and twigs. Controlled-environment experiments were conducted to determine the effects of temperature and leaf wetness period on C. pseudonaviculata sporulation on diseased (cv. Suffruticosa) leaves and of dryness periods and high temperature on conidial survival. Infected leaves were incubated in moist chambers and subjected to six temperatures (9, 13, 17, 21, 25, and 29°C) and six leaf wetness periods (0, 12, 24, 40, 48, and 72 h). Spore production was influenced significantly by wetness period, temperature, and their interaction. Increasing duration of leaf wetness and increasing temperature generally increased sporulation, with no sporulation occurring at 29°C or 9 and 13°C, except at 72 h of wetness exposure, while it was optimal at 21°C. Detached leaves with profuse conidia were subjected to a range of drying (relative humidity at 65%) times (0, 2, 4, 6, and 8 h) at two temperatures of 21 and 29°C. Conidia were then harvested and plated on water agar. Germinating conidia were counted to measure the spore viability. Spore mortality increased with increasing dryness duration at both temperatures but occurred more quickly and severely at 29 than 21°C. Overall, this study extended biological knowledge of conditions required for crucial stages of the C. pseudonaviculata disease cycle and the obtained results will be vital for developing boxwood blight forecasting and management tools.
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    Diversity and Dynamics of Salmonella enterica in Water Sources, Poultry Litters, and Field Soils Amended With Poultry Litter in a Major Agricultural Area of Virginia
    Gu, Ganyu; Strawn, Laura K.; Zheng, Jie; Reed, Elizabeth A.; Rideout, Steven L. (Frontiers, 2019-12-17)
    The Eastern Shore of Virginia (ESV) is a major agricultural region in Virginia and in the past has been linked to some tomato-associated outbreaks of salmonellosis. In this study, water samples were collected weekly from irrigation ponds and wells in four representative vegetable farms (Farms A–D, each farm paired with one pond and one well) and a creek as well. In addition, water samples from two sites in the Chesapeake Bay on the ESV were collected monthly. Poultry litter was sampled monthly from three commercial broiler farms. Soil samples were collected monthly after fertilization with poultry litter from 10 farms in 2014 and another 14 farms in 2015. A most probable number method was used to detect Salmonella enterica presence and concentration in collected samples. Presumptive Salmonella colonies were confirmed by the cross-streaking method. Molecular serotyping was carried out to determine the Salmonella serovars. The average prevalence of Salmonella in pond, well, creek, and bay water samples was 19.3, 3.3, 24.2, and 29.2%, respectively. There were significant spatial and temporal differences for Salmonella incidence in various water sources. The prevalence of S. enterica in four tested ponds from farms A, B, C, and D were 16, 12, 22, and 27%, respectively. While the prevalence of S. enterica in irrigation wells was significantly lower, some well water samples tested positive during the study. Salmonella Newport was found to be the predominant serovar isolated from water samples. All poultry houses of the three tested broiler farms were Salmonella-positive at certain sampling points during the study with prevalence ranging from 14.3 to 35.4%. Salmonella was found to be able to survive up to 4 months in poultry litter amended soils from the tested farms in 2014, and up to 6 months in 2015. This research examined the dynamics of S. enterica in relationship to water source, poultry litter, and amended soil in a major agricultural area, and provides useful information for food safety risk assessments.
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    Exploring Rain as Source of Biological Control Agents for Fire Blight on Apple
    Llontop, Marco E. Mechan; Hurley, Kelly; Tian, Long; Galeano, Vivian A. Bernal; Wildschutte, Hans K.; Marine, Sasha C.; Yoder, Keith S.; Vinatzer, Boris A. (2020-02-14)
    Poor survival on plants can limit the efficacy of Biological Control Agents (BCAs) in the field. Yet bacteria survive in the atmosphere, despite their exposure to high solar radiation and extreme temperatures. If conditions in the atmosphere are similar to, or more extreme than, the environmental conditions on the plant surface, then precipitation may serve as a reservoir of robust BCAs. To test this hypothesis, two hundred and fifty-four rain-borne isolates were screened for in vitro inhibition of Erwinia amylovora, the causal agent of fire blight, as well as of other plant pathogenic bacteria, fungi and oomycetes. Two isolates showed strong activity against E. amylovora and other plant pathogenic bacteria, while other isolates showed activity against fungal and oomycete pathogens. Survival assays suggested that the two isolates that inhibited E. amylovora were able to survive on apple blossoms and branches similarly to E. amylovora. Pathogen population size and associated fire blight symptoms were significantly reduced when detached apple blossoms were treated with the two isolates before pathogen inoculation, however, disease reduction on attached blossoms within an orchard was inconsistent. Using whole genome sequencing, the isolates were identified as Pantoea agglomerans and P. ananatis, respectively. A UV-mutagenesis screen pointed to a phenazine antibiotic D-alanylgriseoluteic acid synthesis gene cluster as being at the base of the antimicrobial activity of the P. agglomerans isolate. Our work reveals the potential of precipitation as an under-explored source of BCAs, whole genome sequencing as an effective approach to precisely identify BCAs, and UV-mutagenesis as a technically simple screen to investigate the genetic basis of BCAs. More field trials are needed to determine the efficacy of the identified BCAs in fire blight control.
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    Proof of Concept for Shoot Blight and Fire Blight Canker Management with Postinfection Spray Applications of Prohexadione-Calcium and Acibenzolar-S-Methyl in Apple
    Aćimović, Srđan G.; Meredith, Christopher L.; Santander, Ricardo Delgado; Khodadadi, Fatemeh (Scientific Societies, 2021-12-03)
    To reduce the severity of shoot blight and prevent the resulting development of cankers on perennial apple wood, we evaluated eight fire blight postinfection spray programs of prohexadione-calcium (PCA) alone or with acibenzolar-S-methyl (ASM) over 2 years. On mature trees of cultivar Royal Court, a single application of the high PCA rate (247 mg/liter) at 2 to 3 days after inoculation resulted in 89.5 and 69.5% reduction of shoot blight severity after inoculation. Two applications of PCA 247 mg/liter 12 or 14 days apart, with the first one applied 2 to 3 days after inoculation, resulted in 78.8 and 74.5% reduction of shoot blight severity in both years. A 100% control of canker incidence on perennial wood from infected shoots in both years was achieved with a single application of PCA (247 mg/liter) applied at 2 or 3 days after the inoculation, and three applications of PCA (125 mg/liter) + ASM (25 mg/liter) 12 to 16 days apart reduced canker incidence by 83.5 and 69% in the 2 years. The other programs with lower PCA rates and frequencies of application reduced shoot blight severity 50.8 and 51.8% (PCA) and 62.6 to 72% and 59.3% (PCA + ASM) over 2 years, respectively. Reduction of canker incidence on wood by the other programs was 66.5% and 69 to 90.4% in the two years, respectively. As fire blight cankers lead to death of dwarf apple trees and serve as primary sources of inoculum, our effective PCA and PCA + ASM programs could serve as viable postinfection management options. These treatments can reduce or prevent canker development and thus significantly abate tree losses in high-density apple orchards after fire blight epidemics occur.
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    Strain-level identification of bacterial tomato pathogens directly from metagenomic sequences
    Mechan Llontop, Marco Enrique; Sharma, Parul; Aguilera Flores, Marcela; Yang, Shu; Pollock, Jill; Tian, Long; Huang, Chengjie; Rideout, Steven L.; Heath, Lenwood S.; Li, Song; Vinatzer, Boris A. (Scientific Societies, 2019-12-12)
    Routine strain-level identification of plant pathogens directly from symptomatic tissue could significantly improve plant disease control and prevention. Here we tested the Oxford Nanopore Technologies (ONT) MinIONTM sequencer for metagenomic sequencing of tomato plants either artificially inoculated with a known strain of the bacterial speck pathogen Pseudomonas syringae pv. tomato (Pto), or collected in the field and showing bacterial spot symptoms caused by either one of four Xanthomonas species. After species-level identification using ONT's WIMP software and the third party tools Sourmash and MetaMaps, we used Sourmash and MetaMaps with a custom database of representative genomes of bacterial tomato pathogens to attempt strain-level identification. In parallel, each metagenome was assembled and the longest contigs were used as query with the genome-based microbial identification Web service LINbase. Both the read-based and assembly-based approaches correctly identified Pto strain T1 in the artificially inoculated samples. The pathogen strain in most field samples was identified as a member of Xanthomonas perforans group 2. This result was confirmed by whole genome sequencing of colonies isolated from one of the samples. Although in our case, metagenome-based pathogen identification at the strain-level was achieved, caution still needs to be exerted when interpreting strain-level results because of the challenges inherent to assigning reads to specific strains and the error rate of nanopore sequencing.