An in vivo study of Lactobacillus rhamnosus GG intervention in mice following dietary exposure to a deoxynivalenol/zearalenone mix

Presenter: Paul Turner                                Status: Faculty

Authors: Murphy L. Y. Wan, Hani El-Nezami Paul C Turner


Abstract: Deoxynivalenol (DON) and zearalenone (ZEA) are mycotoxins produced by Fusarium species, which naturally co-occur in foods and feeds. The gastrointestinal tract represents the first barrier met by xenobiotics. It is covered by a continuous layer of mucus gel, which serves as a medium for protection, lubrication and transport. Such mucus layer is one of the important components of innate host defense and forms the front line of defense against the entry of xenobiotics, including mycotoxins. The purpose of the present study was to investigate the ability of Lactobacillus rhamnosus GG (LGG) to improve intestinal mucosal barrier functions in Balb/c mice (6 weeks old) fed diets containing DON (12μg/g)/ZEA (0.5μg/g) mix. An exposure regimen, which stimulated human exposure experience, was designed. Two different protocols that vary the time-points of oral administration of LGG (1 x 108 CFU per day) were used to determine whether it could prevent and treat unwanted effects induced by DON/ZEA mix. Chronic ingestion of DON/ZEA mix regulated mucus secretion and supplementation of LGG before and after DON/ZEA exposure can improve mildly the mucus barrier. However, future work is still necessary to substantiate the safety and efficacy for their application for dietary intervention against intestinal exposure to mycotoxins.


Fungicide sensitivity of common Phytophthora species in Maryland nurseries

Presenter: Justine Beaulieu                         Status: Faculty

Authors: Justine Beaulieu, Will Still, Yilmaz Balci


Abstract: Phytophthora is a plant pathogen commonly found in Maryland nurseries.            In this study, a collection of Phytophthora isolates from five Maryland nurseries and forests in the Northeastern United States was screened against several fungicides used to manage the pathogen. In total, 243 isolates (77 P. cinnamomi, 23 P. citrophthora, 21 P. pini, 15 P. multivora, and 107 P. plurivora), sampled from a range of hosts and substrates, were screened for sensitivity to mefenoxam and fosetyl-Al. Insensitive isolates [≥50% growth rate relative to controls (RG)] were further tested using higher concentrations of these fungicides along with dimethomorph, dimethomorph + ametoctradin and fluoxastrobin. Mefenoxam and fosetyl-Al-insensitive isolates were detected only for P. cinnamomi, P. multivora and P. plurivora and included 4%, 13% and 12% of screened isolates, respectively, and remained insensitive at the higher concentrations. These isolates were also insensitive to fluoxastrobin, but completely sensitive (0% RG) to dimethomorph and dimethomorph + ametoctradin. Based on our previous genotyping study using AFLP it was found that, while the most common genotypes found in Maryland nurseries appear to be sensitive or intermediately sensitive (<50% RG), the least common genotypes included the insensitive isolates. Our studies suggest discrepancies in fungicide sensitivity among these five common Phytophthora species, signifying that a genus-level fungicide application plan is insufficient for the management of all species in ornamental nursery production. Insensitive isolates are present within the population and thus the inclusion of chemicals such as dimethomorph and ametoctradin into management plans is recommended.


Phyllosphere and rhizosphere microbiomes of tomato plants grown in various organic soil amendments

Presenter: Sarah Allard                                Status: Graduate Student

Authors: Sarah Allard and Shirley Micallef

Abstract: Due to the intimate association between plants and their microbial symbionts, an assessment of the influence of agricultural practices on microbial community structure and diversity will lead to a more comprehensive view of plant health and produce safety. To assess whether organic fertilizer type impacts rhizosphere- and phyllosphere microbial communities associated with tomato plants, tomato cv. ‘BHN602’ was grown in Maryland soils amended with various manure-based fertilizers, in a randomized complete block design. Culture independent DNA was extracted from washes of tomato roots, blossoms and fruit . To characterize bacterial communities, PCR amplicons of the V1-V3 region of the 16S rRNA gene were sequenced on an Illumina MiSeq. Bioinformatic analyses were performed using QIIME. Root, blossom, and fruit surfaces supported distinct bacterial communities as analyzed by principal coordinate analysis. However, there were no significant differences in taxal abundance at the family or genus level in response to fertilizer type. Soil amendment choice did not significantly influence microbial community structure of tomato roots, blossoms or fruit at the time of harvest, indicating that manure application before planting did not exert a long-term influence on the tomato microbiome. These data will contribute to an improved understanding of how agricultural practices associated with tomato production may influence plant health and food safety risk.  


Comparison of single and multi-analyte methods for determination of total deoxynivalenol in Swedish urine samples

Presenter: Allison Gost                                 Status: Graduate Student

Authors: Allison Gost, Stina Wallin, Lucia Gambacorta Michele Solfrizzo, Natalia Kotova Monica Olsen, Paul C. Turner.


Abstract: Deoxynivalenol (DON) is a frequent fungal contaminant of cereal crops in temperate world regions. Deoxynivalenol (DON) and its glucuronides (total DON) can be quantified in urine to estimate exposure. While several groups are developing methods these methods to date have not been compared. In this study, data from a single mycotoxin (SM) and a multi-mycotoxin (MM) method were compared using human urine samples (n=299 and 278, respectively, of which 256 were common for both methods)  from Riksmaten, the Swedish national survey of dietary habits. Both methods included immunoaffinity cleanup of urine pre-digested with β-glucuronidase and LC-MS/MS for identification and quantifictation of DON.;  Hhowever, the specific reagents and conditions were not identical in part because the MM method measures additional mycotoxins. Low DON concentrations were generally measured in tested samples with 84 and 86%, respectively, of detects containing less than 10 ng DON/ml, for both methods. Linear regression showed a significant albeit modest correlation between the two measures (p=0.0001, r = 0.591). Overall data are in reasonable agreement, and the differences observed may reflect subtle handling differences in DON extraction and quantitation between the methods; possibly reflecting the differing ability to detect different glucuronides of DON.


Salmonella Newport Responds to Phyllosphere Derived Nitrogenous Stress on Tomato Fruit and Leaves

Presenter: Angela Marie C. Ferelli                           Status: Graduate Student

Authors: Angela Ferelli and Shirley A. Micallef


Abstract:   Previous work in the Micallef lab has shown an upregulation of genes related to reactive nitrogen species stress response when Salmonella Typhimurium was inoculated onto sterile tomatoes. To delve deeper into this observation, we investigated whether mitigation of nitrogenous stress is important to human enteric pathogen colonization of plants.     To accomplish this, we treated a range of tomato cultivars with the nitric oxide scavenger 2-4-carboxyphenyl-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (cPTIO) and assessed growth through direct plating methods. Growth results revealed cultivars ‘Heinz’, ‘Nyagous,’ and ‘Money Maker’ fruit all exhibited higher colonization levels (in log CFU) of Salmonella Newport at 48 hours on NO-scavenged tissue relative to control (p=0.0086, p=0.0001, p=0.0078, respectively). Interestingly, ‘Heinz’ fruit showed a suppression of growth relative to other cultivars tested at 24 hours regardless of treatment. On sterile 3 week old seedlings of cv. ‘Heinz’, we measured larger populations of Salmonella Newport (p=0.0073) following treatment with cPTIO, with the rpoS partial mutant Salmonella Typhimurium LT2 unable to produce similar results.     Findings from this study suggest that there are cultivar specific plant derived mechanisms that perceive enteric bacterial colonization and release nitric oxide that is capable of restricting bacterial populations.  Furthermore, fruit response intensity to colonizing Salmonella differs among cultivars. Finally, seedling data of S. Typhimurium LT2 compared to S. Newport suggests that Salmonella perceives and interprets abiotic stress when colonizing plants, regulated in-part by rpoS.  



The Relationship between Dietary Intake and Biomarkers of Carotenoids and Physical Functioning Among U.S. Older Adults

Presenter: Olfat Sheikomar                         Status: Graduate Student

Authors: Olfat Sheikomar, MS and Proffessor Nadine Sahyoun Ph.D., R.D.


Abstract: Decline of physical function [PF] in old age might be related to oxidative damage caused by free radicals, and antioxidants may play a role in reducing the risk of physical functional limitations [PFL]. Yet little is known about the role of carotenoids in PFL. The purpose of this study was to assess the association of total and daily dietary intakes of carotenoids, fruit and vegetables [FV] and their biomarkers with PF among U.S. older adults. Data were from 2,905 men and women [≥ 60 years] in the National Health and Nutrition Examination Survey [NHANES] 2003-2006. Using logistic regression, we found that serum concentration of carotenoids was associated with limitations in PF. In the fully adjusted model, the ORs [95% CI] of having limitation in activities of daily living [ADLs], instrumental activities of daily living [IADLs] and movement difficulties [MD] were 2.03 [1.16 - 3.53], 2.34 [1.61 - 3.42], and 2.15 [1.46 - 3.18], respectively, comparing the lowest quintile of serum carotenoids to the highest. Total intake and dietary intake of carotenoids were found to be associated with limitations in IADL. However, low FV consumptions were not significantly associated with PF domains. In conclusion, elevated levels of serum carotenoids are significantly associated with better physical functional performance and may play an important role in delaying the onset of physical decline.


TRPV4 calcium channels regulate macrophage foam cell formation

Presenter: Michael Merth                            Status: Graduate Student

Authors: M. Merth1, R. Goswami1, S. Sharma1, S. O. Rahaman1*


Abstract: TRPV4 calcium channels regulate macrophage foam cell formation     M. Merth1, R. Goswami1, S. Sharma1, S. O. Rahaman1*    1University of Maryland, Department of Nutrition and Food Science, College Park, MD 20742    *Corresponding author     Cardiovascular disease is the number one cause of death in United States, and atherosclerosis, a chronic arterial disease, is the most dominant underlying pathology. Porphyromonas gingivalis (P.g), a predominant causative agent of periodontitis, has been linked to development of atherosclerosis. However, the mechanisms by which P.g infection contributes to atherogenesis remain elusive. The formation of lipid-laden macrophage foam cells is critically important to the in vivo development and progression of atherosclerosis. We have identified the transient receptor potential (TRP) channel of the vanilloid subfamily, TRPV4, a calcium-permeable channel, as a novel regulator of macrophage foam cell formation. We demonstrate that TRPV4 channels are expressed and functional in mouse macrophage cell line (RAW cells). Importantly, we show that TRPV4 activity (Ca2+ influx) is increased in response to P.g-derived LPS (pgLPS). TRPV4 antagonism blocks pgLPS-induced exacerbation of oxidized low density lipoprotein (oxLDL)-derived macrophage foam cell formation. Mechanistically, we show that TRPV4 channel activity regulates oxLDL uptake in RAW cells. Altogether, our results suggest that TRPV4 channels regulate macrophage foam cell formation by modulating uptake of oxLDL.      Funding: AHA (13SDG17310007), Startup and MAES grant from University of Maryland 


TRPV4 mechanosensing regulates normal human lung fibroblast migration

Presenter: Rishov Goswami                         Status: Graduate Student

Authors: R. Goswami, S. Sharma, K. Adlerz, M. Merth, J. H. Aranda-Espinoza, S. O. Rahaman*


Abstract: Systemic Sclerosis (SSc) is a multisystem idiopathic connective tissue disease with high morbidity and mortality. The major cause of death in SSc patients is pulmonary dysfunction resulting from lung fibrosis. Fibroblast migration and differentiation into myofibroblasts is recognized as a critical process in fibrogenesis. Although the influence of both a mechanical signal e.g., matrix stiffness, and a biochemical signal, e.g., transforming growth factor beta1 (TGFbeta1), are essential for fibroblast migration, the precise mechanisms and the identity of a plasma membrane mechanosensor by which migratory signals are transduced/progressed are unknown. Recently we discovered that the transient receptor potential (TRP) channel of the vanilloid subfamily, TRPV4, a mechanosensitive, calcium-permeable, plasma membrane channel, is required for TGFbeta1/matrix stiffness-induced fibroblast differentiation and lung fibrosis development (Rahaman et al., J Clin Invest. 2014, 124:5225-38). Here, we report that TRPV4 is required for both TGFbeta1- and matrix stiffness-induced migratory response (chemotaxis, durotaxis, scratch wound closure, and adhesion) of normal human lung fibroblasts. Furthermore, TRPV4 antagonism by a recently discovered selective small molecule antagonist abrogates calcium influx, stiffness-induced stress fiber formation, and both TGFbeta1 and matrix stiffness-induced focal adhesion kinase (FAK) activation. Mechanistically, we show that TRPV4 channel activity is required for stiffness-induced force generation in migrating fibroblasts as determined by Traction Force Microscopy. Altogether, our results suggest that TRPV4 and FAK interact to drive fibroblast migration by potentiating TGFbeta1 signaling in a manner dependent on matrix stiffness. Successful manipulations of the TRPV4 channel activity may be a novel therapeutic approach to the treatment of SSc.