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Abstract Background Aspergillus flavus is an important agricultural and food safety threat due to its production of carcinogenic aflatoxins. It has high level of genetic diversity that is adapted to various environments. Recently, we reported two reference genomes of A. flavus isolates, AF13 (MAT1-2 and highly aflatoxigenic isolate) and NRRL3357 (MAT1-1 and moderate aflatoxin producer). Where, an insertion of 310 kb in AF13 included an aflatoxin producing gene bZIP transcription factor, named atfC. Observations of significant genomic variants between these isolates of contrasting phenotypes prompted an investigation into variation among other agricultural isolates of A. flavus with the goal of discovering novel genes potentially associated with aflatoxin production regulation. Present study was designed with three main objectives: (1) collection of large number of A. flavus isolates from diverse sources including maize plants and field soils; (2) whole genome sequencing of collected isolates and development of a pangenome; and (3) pangenome-wide association study (Pan-GWAS) to identify novel secondary metabolite cluster genes. Results Pangenome analysis of 346 A. flavus isolates identified a total of 17,855 unique orthologous gene clusters, with mere 41% (7,315) core genes and 59% (10,540) accessory genes indicating accumulation of high genomic diversity during domestication. 5,994 orthologous gene clusters in accessory genome not annotated in either the A. flavus AF13 or NRRL3357 reference genomes. Pan-genome wide association analysis of the genomic variations identified 391 significant associated pan-genes associated with aflatoxin production. Interestingly, most of the significantly associated pan-genes (94%; 369 associations) belonged to accessory genome indicating that genome expansion has resulted in the incorporation of new genes associated with aflatoxin and other secondary metabolites. Conclusion In summary, this study provides complete pangenome framework for the species of Aspergillus flavus along with associated genes for pathogen survival and aflatoxin production. The large accessory genome indicated large genome diversity in the species A. flavus, however AflaPan is a closed pangenome represents optimum diversity of species A. flavus. Most importantly, the newly identified aflatoxin producing gene clusters will be a new source for seeking aflatoxin mitigation strategies and needs new attention in research.

AbstractThe series of experiments summarized here were conducted with the objective to evaluate the benefits of using biochar for coating corn seeds. Seeds coated with a slurry containing bio-based ingredients and biochar were tested for germination and vigor, and for their potential to being infected by the fungus Aspergillus flavus, using a novel single seed incubator specifically designed for these purposes. Biochar-treated seeds were also planted for two years in experimental fields in the Mississippi Delta to evaluate their effect on corn yield and aflatoxin contamination of kernels. Experiments were conducted with two types of commercial biochar; one was obtained from hardwood residues and the other from coconut shells. Application of both types of biochar for coating the seeds did not affect seed germination and vigor. However, treated seeds showed increased wettability and a more rapid water uptake. This resulted in a 8.5% shortening of germination time. Microbiological analysis using plate culturing and qPCR methods showed that biochar was not conducive to the growth of A. flavus. This was also confirmed by analyzing soil samples that were collected from experimental fields located in the Mississippi Delta. Most importantly, although aflatoxin contamination was different in the two experimental years, aflatoxin contamination of corn kernels was not affected by biochar-based formulations.

Macrophomina phaseolina (Mp) is a fungal pathogen proposed to enter host roots by releasing toxins that induce local necrosis in roots allowing entry of hyphae. Mp is reported to produce several potent phytotoxins, including (-)-botryodiplodin and phaseolinone, but isolates that do not produce these phytotoxins retain virulence. One hypothesis explaining these observations is that some Mp isolates may produce other unidentified phytotoxin(s) responsible for virulence. A previous study of Mp isolates from soybean found 14 previously unreported secondary metabolites using LC-MS/MS, including mellein, which has various reported biological activities. This study was conducted to investigate the frequency and amounts of mellein produced in culture by Mp isolates from soybean plants exhibiting symptoms of charcoal rot and to investigate the role of mellein in any observed phytotoxicity. LC-MS/MS analysis of cell-free culture filtrates (CCFs) from 89 Mp isolates revealed that 28.1% produced mellein (49–2,203 µg/L). In soybean seedlings in hydroponic culture, Mp CCFs diluted to 25% (vol/vol) in hydroponic growth medium induced phytotoxic symptoms with frequencies of 73% chlorosis, 78% necrosis, 7% wilting, and 16% death, and at 50% (vol/vol) induced phytotoxicity with frequencies of 61% chlorosis, 82% necrosis, 9% wilting, and 26% death. Commercially-available mellein (40–100 µg/mL) in hydroponic culture medium induced wilting. However, mellein concentrations in CCFs exhibited only weak, negative, insignificant correlations with phytotoxicity measures in soybean seedlings, suggesting that mellein does not contribute substantially to observed phytotoxic effects. Further investigation is needed to determine if mellein plays any role in root infection.

Kamal Hamed, Tatiana Wiktorowicz, Maziar Assadi Gehr Basilea Pharmaceutica International Ltd., Basel, SwitzerlandCorrespondence: Kamal HamedBasilea Pharmaceutica International Ltd., Grenzacherstrasse 487, Basel 4005, SwitzerlandTel +41 61 567 1588Fax +41 61 606 1216Email kamal.hamed@basilea.comPurpose: Drug-induced immune hemolytic anemia (DIIHA) is a rare but serious adverse event associated with a number of drugs, including second- and third-generation cephalosporins. A positive direct antiglobulin test (DAT) is a reliable finding in DIIHA, but positive results without evidence of hemolysis can occur, particularly in hospitalized patients. There have been no reports of hemolytic anemia in four previous Phase 3 trials or from post-marketing surveillance of the advanced-generation, broad-spectrum cephalosporin, ceftobiprole. The aim of this analysis was to review the incidence of positive DAT results and any evidence of hemolytic anemia from three recent Phase 3 trials of ceftobiprole.Patients and Methods: Patients were enrolled in three Phase 3 randomized controlled trials: 94 pediatric patients with pneumonia received ceftobiprole in the BPR-PIP-002 trial; 335 adults with acute bacterial skin and skin structure infections received ceftobiprole in the TARGET trial; and 201 adults with Staphylococcus aureus bacteremia have been randomized 1:1 to ceftobiprole or daptomycin ± aztreonam in the ongoing ERADICATE trial. In all three trials, DAT results were obtained at baseline, and follow-up tests were performed either at the test of cure (TOC) visit (BPR-PIP-002), end-of-treatment (EOT) visit (TARGET), or both EOT and post-treatment Day 70 visits (ERADICATE).Results: In the BPR-PIP-002 trial, five patients (all ceftobiprole treated) had a documented negative DAT result at baseline followed by a positive result at the TOC visit. One patient in the ongoing, blinded ERADICATE trial had a positive DAT result at both baseline and EOT. Results from other laboratory investigations showed no evidence of hemolytic anemia in these patients. No positive DAT results were reported in the TARGET trial.Conclusion: No evidence of hemolytic anemia associated with ceftobiprole was observed in either adults or children across several indications in this analysis of three large Phase 3 trials.Keywords: DIIHA, direct antiglobulin test, hemolysis, pneumonia, ABSSSIs, Staphylococcus aureus bacteremia

Mohammad Alizadeh,1,* Moludi Jalal,2 Khodaei Hamed,3 Amir Saber,2,* Sorayya Kheirouri,1 Fatemeh Pourteymour Fard Tabrizi,3 Negin Kamari2 1Nutrition Research Center, Tabriz University of Medical Sciences, Tabriz, Iran; 2Department of Nutritional Sciences, School of Nutritional Sciences and Food Technology, Kermanshah University of Medical Sciences, Kermanshah, Iran; 3Department of Biochemistry and Diet Therapy, Faculty of Nutrition, Tabriz University of Medical Sciences, Tabriz, Iran*These authors contributed equally to this workCorrespondence: Moludi JalalDepartment of Nutrition, School of Nutritional Sciences and Food Technologies, Kermanshah University of Medical Sciences, Isar Sq., Across from Farabi Hospital, Kermanshah, IranTel +98-83 37102009Fax +98-83 37102002Email jalal.moludi68@gmail.comKhodaei HamedDepartment of Biochemistry and Diet Therapy, Faculty of Nutrition, Tabriz University of Medical Sciences, Daneshgah Street, Tabriz, IranEmail hamedkhodaei@yahoo.comAbstract: The furan nucleus is found in a large number of biologically active materials. In recent years, many natural furan derivatives were isolated and their biological effects were investigated. In this review, we focused on the anti-inflammatory and antimicrobial effects of some natural furans and discussed their effects on the immune system. Our investigation revealed that furan natural derivatives have effective antioxidant activities and exert regulatory effects on various cellular activities by modifying some signaling pathways such as MAPK (mitogen-activated Protein Kinase) and PPAR-ɣ (peroxisome proliferator-activated receptor gamma). The antimicrobial activity of these natural compounds was performed through selective inhibition of microbial growth and modification of enzymes. Further studies are needed for isolation and detection of different furan derivatives from natural compounds and investigation of their precise mechanisms for revealing health beneficial effects of these compounds.Keywords: anti-inflammatory, antimicrobial, furans, benzofurans, furan natural derivatives

Charcoal rot, caused by Macrophomina phaseolina, is a major soybean disease resulting in significant yield loss and poor seed quality. Currently, no resistant soybean cultivar is available in the market and resistance mechanisms to charcoal rot are unknown, although the disease is believed to infect plants from infected soil through the roots by unknown toxin-mediated mechanisms. The objective of this research was to investigate the association between seed sugars (sucrose, raffinose, stachyose, glucose, and fructose) and their role as biomarkers in the soybean defense mechanism in the moderately resistant (MR) and susceptible (S) genotypes to charcoal rot. Seven MR and six S genotypes were grown under irrigated (IR) and non-irrigated (NIR) conditions. A two-year field experiment was conducted in 2012 and 2013 at Jackson, TN, USA. The main findings in this research were that MR genotypes generally had the ability to maintain higher seed levels of sucrose, glucose, and fructose than did S genotypes. Conversely, susceptible genotypes showed a higher level of stachyose and lower levels of sucrose, glucose, and fructose. This was observed in 6 out of 7 MR genotypes and in 4 out of 6 S genotypes in 2012; and in 5 out of 7 MR genotypes and in 5 out of 6 S genotypes in 2013. The response of S genotypes with higher levels of stachyose and lower sucrose, glucose, and fructose, compared with those of MR genotypes, may indicate the possible role of these sugars in a defense mechanism against charcoal rot. It also indicates that nutrient pathways in MR genotypes allowed for a higher influx of nutritious sugars (sucrose, glucose, and fructose) than did S genotypes, suggesting these sugars as potential biomarkers for selecting MR soybean plants after harvest. This research provides new knowledge on seed sugars and helps in understanding the impact of charcoal rot on seed sugars in moderately resistant and susceptible genotypes.

Pre-harvest aflatoxin contamination occurs in maize following kernel colonization by Aspergillus flavus. Aflatoxin contamination resistance is a highly desired trait in maize breeding programs. The identification of novel sources of resistance to pre-harvest aflatoxin contamination is a major focus in germplasm screening efforts. Here, we performed a field evaluation of 64 inbred lines over two years for pre-harvest aflatoxin contamination. Topcrosses were also performed with two testers, B73 and Mo17, to generate 128 F1 hybrids which were also evaluated over two years. Hybrid performance was used to calculate both general combining ability (GCA) of the inbreds, and observed heterosis for aflatoxin resistance. Over both years of the study, aflatoxin concentrations ranged from 80 ± 47 to 17,617 ± 8816 μg kg−1 for inbreds, and from 58 ± 39 to 2771 ± 780 μg kg−1 for hybrids with significant variation between years and lines. The inbred lines CML52, CML69, CML247, GT-603, GEMS-0005, Hi63, Hp301, and M37 W showed

An isolate of Macrophomina phaseolina from muskmelons (Cucumis melo) was reported by Dunlap and Bruton to produce red pigment(s) in melons and in culture in the presence of added glycine, alanine, leucine, or asparagine in the medium, but not with some other amino acids and nitrogen-containing compounds. We explored the generality and mechanism of this pigment production response using pathogenic M. phaseolina isolates from soybean plants expressing symptoms of charcoal rot disease. A survey of 42 M. phaseolina isolates growing on Czapek-Dox agar medium supplemented with glycine confirmed pigment production by 71% of isolates at the optimal glycine concentration (10 g/L). Studies in this laboratory have demonstrated that some pathogenic isolates of M. phaseolina produce the mycotoxin (−)-botryodiplodin, which has been reported to react with amino acids, proteins, and other amines to produce red pigments. Time course studies showed a significant positive correlation between pigment and (−)-botryodiplodin production by selected M. phaseolina isolates with maximum production at seven to eight days. Pigments produced in agar culture medium supplemented with glycine, beta-alanine, or other amines exhibited similar UV-vis adsorption spectra as did pigments produced by (±)-botryodiplodin reacting in the same agar medium. In a separate study of 39 M. phaseolina isolates, red pigment production (OD520) on 10 g/L glycine-supplemented Czapek-Dox agar medium correlated significantly with (−)-botryodiplodin production (LC/MS analysis of culture filtrates) in parallel cultures on un-supplemented medium. These results support pigment production on glycine-supplemented agar medium as a simple and inexpensive in-culture method for detecting (−)-botryodiplodin production by M. phaseolina isolates.

Anne Santerre Henriksen, Jennifer I Smart, Kamal Hamed Basilea Pharmaceutica International Ltd., Basel 4005, Switzerland Purpose: Lower respiratory tract infections (LRTIs) can cause significant morbidity and mortality and are becoming increasingly difficult to treat because of the growing prevalence of resistance to conventional antimicrobial agents. This study aimed to assess the current in vitro susceptibility of respiratory tract pathogens collected from the UK and Ireland to ceftobiprole, an advanced-generation cephalosporin, as compared with other antibiotics. Methods: Pathogens isolated from patients with LRTIs were analyzed as part of the British Society for Antimicrobial Chemotherapy Antimicrobial Resistance Surveillance Programme during 2014–2015. Antibiotic susceptibility was evaluated using European Committee on Antimicrobial Susceptibility Testing breakpoints, including the ceftobiprole pharmacokinetic/pharmacodynamic non-species-specific breakpoint when species-specific breakpoints were not available. Results: One thousand one hundred and sixty-eight isolates from community-onset LRTIs and 1,264 isolates from hospital-onset LRTIs were analyzed. The ceftobiprole susceptibility rate was 99.8% (428/429) for Streptococcus pneumoniae, 100% (502/502) for Haemophilus influenzae, and 99.6% (236/237) for Moraxella catarrhalis. All Staphylococcus aureus isolates, including methicillin-susceptible S. aureus (MSSA; N=181) and methicillin-resistant S. aureus (MRSA; N=35), were susceptible to ceftobiprole. Overall, ceftobiprole susceptibility was observed in 88.1% (215/244) of Escherichia coli isolates, 83.4% (156/187) of Klebsiella pneumoniae isolates and 86.7% (98/113) of Enterobacter spp. isolates. Conclusion: Ceftobiprole had in vitro activity against all S. aureus (both MSSA and MRSA) isolates, and almost all S. pneumoniae isolates, as well as against Gram-negative bacteria associated with community-onset or hospital-onset LRTIs. Based on this analysis, ceftobiprole is a good treatment option when broad-spectrum antibiotic coverage is needed for LRTIs. Keywords: antibiotic, BSAC, cephalosporin, MIC, resistance, RTI

Charcoal rot is a major disease of soybean (Glycine max) caused by Macrophomina phaseolina and results in significant loss in yield and seed quality. The effects of charcoal rot on seed composition (seed protein, oil, and fatty acids), a component of seed quality, is not well understood. Therefore, the objective of this research was to investigate the impact of charcoal rot on seed protein, oil, and fatty acids in different soybean genotypes differing in their charcoal rot susceptibility under irrigated and non-irrigated conditions. Two field experiments were conducted in 2012 and 2013 in Jackson, TN, USA. Thirteen genotypes differing in charcoal rot resistance (moderately resistant and susceptible) were evaluated. Under non-irrigated conditions, moderately resistant genotypes showed either no change or increased protein and oleic acid but had lower linolenic acid. Under non-irrigated conditions, most of the susceptible genotypes showed lower protein and linolenic acid but higher oleic acid. Most of the moderately resistant genotypes had higher protein than susceptible genotypes under irrigated and non-irrigated conditions but lower oil than susceptible genotypes. The different responses among genotypes for protein, oil, oleic acid, and linolenic acid observed in each year may be due to both genotype tolerance to drought and environmental conditions, especially heat differences in each year (2012 was warmer than 2013). This research showed that the increases in protein and oleic acid and the decrease in linolenic acid may be a possible physiological mechanism underlying the plant’s responses to the charcoal rot infection. This research further helps scientists understand the impact of irrigated and non-irrigated conditions on seed nutrition changes, using resistant and susceptible genotypes.

Two important mycotoxins, aflatoxin and fumonisin, are among the most potent naturally occurring carcinogens, contaminating maize (Zea mays) and affecting crop yield and quality. Resistance of maize to pre-harvest mycotoxin contamination, specifically aflatoxin produced by Aspergillus flavus and fumonisin produced by Fusarium verticillioides, is a goal in breeding programs that screen for these important traits with the aim of developing resistant commercial hybrids. We conducted two years of field evaluations on 87 inbred lines originating primarily in China and Mexico and not previously screened for resistance. The objectives of our study were to identify resistant germplasm for breeding purposes and to examine possible relationships between resistances to the two mycotoxins. Aflatoxin and fumonisin were present in samples harvested from all lines in both years. Concentrations of total aflatoxin ranged from 52.00 ± 20.00 to 1524.00 ± 396.00 μg kg−1, while those of fumonisin ranged from 0.60 ± 0.06 to 124.00 ± 19.50 mg kg−1. The inbred lines TUN15, TUN61, TUN37, CY2, and TUN49 showed the lowest aflatoxin accumulation and CN1, GT601, TUN09, TUN61, and MP717 the lowest fumonisin accumulation. TUN61 showed the lowest accumulation of both mycotoxins. This study confirmed previous observations that high levels of aflatoxin can coexist with fumonisin, with 55 maize lines showing a positive correlation coefficient between the concentrations of aflatoxin and fumonisin and 32 lines showing a negative correlation coefficient. These selected lines, particularly TUN61, may provide sources of resistance to mycotoxin contamination in breeding programs. However, the mechanism of resistance in this germplasm remains to be identified. Future research should also address factors that influence the fungus–plant interaction, such as herbivory and environmental stress.

Kamal Hamed,1 Valentino Conti,2 Hengfeng Tian,1 Emil Loefroth3 1Novartis Pharmaceuticals Corporation, East Hanover, NJ, USA; 2Novartis Global Service Center, Dublin, Ireland; 3Novartis Sverige AB, Täby, Sweden Purpose: Tobramycin inhalation powder (TIP), the first dry-powder inhaled antibiotic for pulmonary Pseudomonas aeruginosa infection, is associated with reduced treatment burden, increased patient satisfaction, and higher self-reported adherence for cystic fibrosis (CF) patients. We compared adherence in CF patients newly treated with TIP with those newly treated with the traditional tobramycin inhalation solution (TIS), using US insurance claims data.Patients and methods: From the Truven MarketScan® database, we identified CF patients chronically infected with P. aeruginosa who had been prescribed TIP between May 1, 2013 to December 31, 2014, or TIS between September 1, 2010 to April 30, 2012 with at least 12 months of continuous medical and pharmacy benefits prior to and following prescription. TIP and TIS adherence levels were assessed.Results: A total of 145 eligible patients were identified for the TIP cohort and 306 for the TIS cohort. Significant differences in age distribution (25.0 vs 21.9 years for TIP vs TIS, respectively, P=0.017), type of health plan (P=0.014), employment status (72.4% vs 63.4% of TIP vs TIS patients in full-time employment, P=0.008), and some comorbidities were observed between the two cohorts. Although a univariate analysis found no significant differences between TIP and TIS (odds ratio [OR] 1.411, 95% confidence interval [CI] 0.949–2.098), TIP was moderately associated with higher adherence levels compared with TIS in a multivariable analysis, once various demographic and clinical characteristics were adjusted for. These included geographic location (OR: 1.566, CI: 1.016–2.413) and certain comorbidities.Conclusion: This study of US patient data supports previous findings that TIP is associated with better adherence compared with TIS; however, further studies will be required to fully elucidate differences in adherence between TIP and TIS. Keywords: Adherence, convenience, inhaler, nebulizer, Pseudomonas aeruginosa, real-world evidence, tobramycin, treatment burden

A popular pre-harvest strategy to mitigate aflatoxin contamination of corn involves field application of non-aflatoxigenic strains of Aspergillus flavus. The basis of this biological control may involve multiple factors, but competitive displacement of aflatoxigenic strains by the biocontrol strains is a likely mechanism. Three biocontrol strains (NRRL 21882, 18543, and 30797) were applied annually, over a 4-year period, to the same 3.2-ha commercial corn field in the Mississippi Delta, where we monitored their post-release establishment, spread, and persistence. Within 2 months of the first biocontrol application, the percentage of soil-inhabiting aflatoxigenic A. flavus strains in some plots was reduced from 48 to 9% of the population. The frequency of aflatoxigenic A. flavus strains was also significantly reduced in the rest of field. After 4 years, neighboring plots that had never received a biocontrol treatment, and distanced from our treatment plots by at least 20 meters, had less than 20% aflatoxigenic isolates. This significant halo effect might be attributed to movement of soil through tillage operations, but the aflatoxigenicity shift could be detected in the untreated plots within 2 months of the initial applications, at a time when there was no tillage. The A. flavus populations that colonized the grain were also monitored and found to be less than 15% toxigenic in the fourth year for all treatments. Over all treatments and years, less than 2 ppb of aflatoxin was detected, which could be a consequence of the field-wide shift of the inherent A. flavus population to predominately non-aflatoxigenic strains. This study supports the efficacy of using non-aflatoxigenic A. flavus strains as pre-harvest biocontrol, and shows that most of its effectiveness occurs with the first application.

Macrophomina phaseolina (Tassi) Goid., the causal agent of charcoal rot disease of soybean, is capable of causing disease in more than 500 other commercially important plants. This fungus produces several secondary metabolites in culture, including (-)-botryodiplodin, phaseolinone and mellein. Given that independent fungal isolates may differ in mycotoxin and secondary metabolite production, we examined a collection of 89 independent M. phaseolina isolates from soybean plants with charcoal rot disease using LC-MS/MS analysis of culture filtrates. In addition to (-)-botryodiplodin and mellein, four previously unreported metabolites were observed in >19% of cultures, including kojic acid (84.3% of cultures at 0.57–79.9 µg/L), moniliformin (61.8% of cultures at 0.011–12.9 µg/L), orsellinic acid (49.4% of cultures at 5.71–1960 µg/L) and cyclo[L-proline-L-tyrosine] (19.1% of cultures at 0.012–0.082 µg/L). In addition, nine previously unreported metabolites were observed at a substantially lower frequency (M. phaseolina and on food and feed safety, if any of them contaminate the seeds of infected soybean plants.

Armando Gonzalez-Ruiz,1 R Andrew Seaton,2 Kamal Hamed3 1Darent Valley Hospital, Dartford, UK; 2Queen Elizabeth University Hospital, Glasgow, UK; 3Novartis Pharmaceuticals Corporation, East Hanover, NJ, USA Abstract: Infections caused by Gram-positive pathogens remain a major public health burden and are associated with high morbidity and mortality. Increasing rates of infection with Gram-positive bacteria and the emergence of resistance to commonly used antibiotics have led to the need for novel antibiotics. Daptomycin, a cyclic lipopeptide with rapid bactericidal activity against a wide range of Gram-positive bacteria including methicillin-resistant Staphylococcus aureus, has been shown to be effective and has a good safety profile for the approved indications of complicated skin and soft tissue infections (4 mg/kg/day), right-sided infective endocarditis caused by S. aureus, and bacteremia associated with complicated skin and soft tissue infections or right-sided infective endocarditis (6 mg/kg/day). Based on its pharmacokinetic profile and concentration-dependent bactericidal activity, high-dose (>6 mg/kg/day) daptomycin is considered an important treatment option in the management of various difficult-to-treat Gram-positive infections. Although daptomycin resistance has been documented, it remains uncommon despite the increasing use of daptomycin. To enhance activity and to minimize resistance, daptomycin in combination with other antibiotics has also been explored and found to be beneficial in certain severe infections. The availability of daptomycin via a 2-minute intravenous bolus facilitates its outpatient administration, providing an opportunity to reduce risk of health care-associated infections, improve patient satisfaction, and minimize health care costs. Daptomycin, not currently approved for use in the pediatric population, has been shown to be widely used for treating Gram-positive infections in children. Keywords: daptomycin, efficacy, Gram-positive infections, high dose, OPAT, safety

Interest in using harvest aids (defoliants or crop desiccants) such as paraquat, carfentrazone-ethyl, glyphosate, and sodium chlorate (NaClO3) have become increasingly important to assure harvest efficiency, producer profit, and to maintain seed quality. However, information on the effects of harvest aids on seed nutrition (composition) (protein, oil, fatty acids, sugars, and amino acids) in soybean is very limited. The objective of this research was to investigate the influence of harvest aids on seed protein, oil, fatty acids, sugars, and amino acids in soybean. Our hypothesis was that harvest aid may influence seed nutrition, especially at R6 as at R6 the seeds may still undergo biochemical changes. Field experiments were conducted in 2012 and 2013 under Midsouth USA environmental conditions in which harvest aids were applied at R6 (seed-fill) and R7 (yellow pods) growth stages. Harvest aids applied included an untreated control, 0.28 kg ai ha−1 of paraquat, 0.28 kg ai ha−1 of paraquat, and 1.015 kg ai ha−1 of carfentrazone-ethyl (AIM); 6.72 kg ai ha−1 sodium chlorate, 1.015 kg ai ha−1 carfentrazone-ethyl; and 2.0 kg ae ha−1 glyphosate. Results showed that the application of harvest aids at either R6 or R7 resulted in the alteration of some seed composition such as protein, oil, oleic acid, fructose, and little effects on amino acids. In addition, harvest aids affected seed composition constituents differently depending on year and growth stage. This research demonstrated the possible alteration of some nutrients by harvest aids. This research helps growers and scientists to advance the understanding and management of harvest aids and investigate possible effects of harvest aids on seed nutrition.

Information on the effects of agricultural practices such as seeding rate (S), row spacing (RS), herbicide apical treatment (T), and nitrogen application (N) on soybean seed nutrition (protein, oil, fatty acids, sugars, and amino acids) is limited. Although seed composition (nutrition) constituents are genetically controlled, agricultural practices and environmental conditions significantly influence the amount and quality of seed nutrition. Therefore, the objective of this research was to understand the responses of these seed composition constituents to these practices, the environment, and cultivar differences. Two-field experiments were conducted, in 2015 and 2016, in Milan, TN, USA. The experiments were irrigated with four replications and included: two soybean cultivars, two seeding rates, three different row spacings, two N rates, and Cobra herbicide apical treatment. The results showed significant effects of S, RS, N, and T on some seed composition constituents, including protein; oleic, linolenic, and stearic acids; sugars; and some amino acids. The current research demonstrated that single or twin row with a seeding rate of 40,000 seeds ha−1 resulted in higher protein, oleic, some sugars, and some amino acids. However, a high seeding rate of 56,000 seeds ha−1 resulted in lower protein, oleic acid, some sugars, and some amino acids due to plant competition for soil nutrients. Herbicide apical application of Cobra1X resulted in higher linolenic acid and some amino acids. Application of nitrogen resulted in higher protein, linolenic, and some amino acids. This research is beneficial to the scientific communities, including breeders and physiologists through advancing knowledge on the interactions between cultivars and environment for seed nutritional quality selection, and to soybean producers through consideration of best agricultural management to maintain high seed nutritional qualities.

Toxins have been proposed to facilitate fungal root infection by creating regions of readily-penetrated necrotic tissue when applied externally to intact roots. Isolates of the charcoal rot disease fungus, Macrophomina phaseolina, from soybean plants in Mississippi produced a phytotoxic toxin, (−)-botryodiplodin, but no detectable phaseolinone, a toxin previously proposed to play a role in the root infection mechanism. This study was undertaken to determine if (−)-botryodiplodin induces toxic responses of the types that could facilitate root infection. (±)-Botryodiplodin prepared by chemical synthesis caused phytotoxic effects identical to those observed with (−)-botryodiplodin preparations from M. phaseolina culture filtrates, consistent with fungus-induced phytotoxicity being due to (−)-botryodiplodin, not phaseolinone or other unknown impurities. Soybean leaf disc cultures of Saline cultivar were more susceptible to (±)-botryodiplodin phytotoxicity than were cultures of two charcoal rot-resistant genotypes, DS97-84-1 and DT97-4290. (±)-Botryodiplodin caused similar phytotoxicity in actively growing duckweed (Lemna pausicostata) plantlet cultures, but at much lower concentrations. In soybean seedlings growing in hydroponic culture, (±)-botryodiplodin added to culture medium inhibited lateral and tap root growth, and caused loss of root caps and normal root tip cellular structure. Thus, botryodiplodin applied externally to undisturbed soybean roots induced phytotoxic responses of types expected to facilitate fungal root infection.

Charcoal rot disease, caused by the fungus Macrophomina phaseolina, results in major economic losses in soybean production in southern USA. M. phaseolina has been proposed to use the toxin (-)-botryodiplodin in its root infection mechanism to create a necrotic zone in root tissue through which fungal hyphae can readily enter the plant. The majority (51.4%) of M. phaseolina isolates from plants with charcoal rot disease produced a wide range of (-)-botryodiplodin concentrations in a culture medium (0.14−6.11 µg/mL), 37.8% produced traces below the limit of quantification (0.01 µg/mL), and 10.8% produced no detectable (-)-botryodiplodin. Some culture media with traces or no (-)-botryodiplodin were nevertheless strongly phytotoxic in soybean leaf disc cultures, consistent with the production of another unidentified toxin(s). Widely ranging (-)-botryodiplodin levels (traces to 3.14 µg/g) were also observed in the roots, but not in the aerial parts, of soybean plants naturally infected with charcoal rot disease. This is the first report of (-)-botryodiplodin in plant tissues naturally infected with charcoal rot disease. No phaseolinone was detected in M. phaseolina culture media or naturally infected soybean tissues. These results are consistent with (-)-botryodiplodin playing a role in the pathology of some, but not all, M. phaseolina isolates from soybeans with charcoal rot disease in southern USA.

Drought stress in the field has been shown to exacerbate aflatoxin contamination of maize and peanut. Drought and heat stress also produce reactive oxygen species (ROS) in plant tissues. Given the potential correlation between ROS and exacerbated aflatoxin production under drought and heat stress, the objectives of this study were to examine the effects of hydrogen peroxide (H2O2)-induced oxidative stress on the growth of different toxigenic (+) and atoxigenic (−) isolates of Aspergillus flavus and to test whether aflatoxin production affects the H2O2 concentrations that the isolates could survive. Ten isolates were tested: NRRL3357 (+), A9 (+), AF13 (+), Tox4 (+), A1 (−), K49 (−), K54A (−), AF36 (−), and Aflaguard (−); and one A. parasiticus isolate, NRRL2999 (+). These isolates were cultured under a H2O2 gradient ranging from 0 to 50 mM in two different media, aflatoxin-conducive yeast extract-sucrose (YES) and non-conducive yeast extract-peptone (YEP). Fungal growth was inhibited at a high H2O2 concentration, but specific isolates grew well at different H2O2 concentrations. Generally the toxigenic isolates tolerated higher concentrations than did atoxigenic isolates. Increasing H2O2 concentrations in the media resulted in elevated aflatoxin production in toxigenic isolates. In YEP media, the higher concentration of peptone (15%) partially inactivated the H2O2 in the media. In the 1% peptone media, YEP did not affect the H2O2 concentrations that the isolates could survive in comparison with YES media, without aflatoxin production. It is interesting to note that the commercial biocontrol isolates, AF36 (−), and Aflaguard (−), survived at higher levels of stress than other atoxigenic isolates, suggesting that this testing method could potentially be of use in the selection of biocontrol isolates. Further studies will be needed to investigate the mechanisms behind the variability among isolates with regard to their degree of oxidative stress tolerance and the role of aflatoxin production.

Increased aflatoxin contamination in corn by the fungus Aspergillus flavus is associated with frequent periods of drought and heat stress during the reproductive stages of the plants. The objective of this study was to evaluate the relationship between aflatoxin contamination and physiological responses of corn plants under drought and heat stress. The study was conducted in Stoneville, MS, USA under irrigated and non-irrigated conditions. Five commercial hybrids, P31G70, P33F87, P32B34, P31B13 and DKC63-42 and two inbred germplasm lines, PI 639055 and PI 489361, were evaluated. The plants were inoculated with Aspergillus flavus (K-54) at mid-silk stage, and aflatoxin contamination was determined on the kernels at harvest. Several physiological measurements which are indicators of stress response were determined. The results suggested that PI 639055, PI 489361 and hybrid DKC63-42 were more sensitive to drought and high temperature stress in the non-irrigated plots and P31G70 was the most tolerant among all the genotypes. Aflatoxin contamination was the highest in DKC63-42 and PI 489361 but significantly lower in P31G70. However, PI 639055, which is an aflatoxin resistant germplasm, had the lowest aflatoxin contamination, even though it was one of the most stressed genotypes. Possible reasons for these differences are discussed. These results suggested that the physiological responses were associated with the level of aflatoxin contamination in all the genotypes, except PI 639055. These and other physiological responses related to stress may help examine differences among corn genotypes in aflatoxin contamination.

This research examined the expression patterns of 94 stress-related genes in seven maize inbred lines with differential expressions of resistance to aflatoxin contamination. The objective was to develop a set of genes/probes associated with resistance to A. flavus and/or aflatoxin contamination. Ninety four genes were selected from previous gene expression studies with abiotic stress to test the differential expression in maize lines, A638, B73, Lo964, Lo1016, Mo17, Mp313E, and Tex6, using real-time RT-PCR. Based on the relative-expression levels, the seven maize inbred lines clustered into two different groups. One group included B73, Lo1016 and Mo17, which had higher levels of aflatoxin contamination and lower levels of overall gene expression. The second group which included Tex6, Mp313E, Lo964 and A638 had lower levels of aflatoxin contamination and higher overall levels of gene expressions. A total of six “cross-talking” genes were identified between the two groups, which are highly expressed in the resistant Group 2 but down-regulated in susceptible Group 1. When further subjected to drought stress, Tex6 expressed more genes up-regulated and B73 has fewer genes up-regulated. The transcript patterns and interactions measured in these experiments indicate that the resistant mechanism is an interconnected process involving many gene products and transcriptional regulators, as well as various host interactions with environmental factors, particularly, drought and high temperature.

A polymerase chain reaction (PCR) based diagnostic assay was used to develop markers for detection of Fusarium verticillioides (=F. moniliforme), a fumonisin producing fungus in maize tissues. Species-specific primers were designed based on sequence data from the polyketide synthase (PKS) gene (FUM1- previously FUM5) responsible for fumonisin production in fungi. Four sets of oligonucleotide primers were tested for their specificity using 24 strains of F. verticillioides, 10 F. proliferatum, and 12 of other Fusarium species. In addition, 13 species of other fungal genera, from four phyla, were tested as negative controls. Among the four sets, primer set B consistently amplified a 419- bp fragment from the DNA 96% of all F. verticillioides strains and 83% of F. proliferatum. All other fungi tested were negative using primer set B. A total of 38% of the F. verticillioides strains grown on a selective liquid medium produced fumonisin and 92% formed the toxin on standard rice medium. When fumonisin formed in culture, PCR assay using primer set B detected every strain of F. verticillioides, but only amplified 80% of F. proliferatum strains that produced the toxin. PCR detection was consistent at 100 pg/μl concentration of genomic DNA from 4 F. verticillioides strains, but varied at 10 pg/μl. Two duplicate greenhouse tests using artificially inoculated maize plants, had greater levels of F. verticillioides detected after re-evaluting using primer set B than from culturing of the tissues. The molecular protocols described in this study requires only 1 day for completion compared to approximately 10 days for cultural work and morphological determination. In conclusion, conventional PCR assay using primer set B provides a sensitive and accurate detection assay that can be used as a primary or secondary confirmation method for identification and occurrence of F. verticillioides within the maize tissues. However, studies using primer set B for fumonisin production determined by strains of F. verticillioides and F. proliferatum will require further verification.

Information on the effects of management practices on soybean seed composition is scarce. Therefore, the objective of this research was to investigate the effects of planting date (PD) and seeding rate (SR) on seed composition (protein, oil, fatty acids, and sugars) and seed minerals (B, P, and Fe) in soybean grown in two row-types (RT) on the Mississippi Delta region of the Midsouth USA. Two field experiments were conducted in 2009 and 2010 on Sharkey clay and Beulah fine sandy loam soil at Stoneville, MS, under irrigated conditions. Soybean were grown in 102 cm single-rows and 25 cm twin-rows in 102 cm centers at seeding rates of 20, 30, 40, and 50 seeds m–2. The results showed that in May and June planting, protein, glucose, P, and B concentrations increased with increased SR, but at the highest seeding rates (40 and 50 seeds m–2), the concentrations remained constant or declined. Palmitic, stearic, and linoleic acid concentrations were the least responsive to SR increases. Early planting resulted in higher oil, oleic acid, sucrose, B, and P on both single and twin-rows. Late planting resulted in higher protein and linolenic acid, but lower oleic acid and oil concentrations. The changes in seed constituents could be due to changes in environmental factors (drought and temperature), and nutrient accumulation in seeds and leaves. The increase of stachyose sugar in 2010 may be due to a drier year and high temperature in 2010 compared to 2009; suggesting the possible role of stachyose as an environmental stress compound. Our research demonstrated that PD, SR, and RT altered some seed constituents, but the level of alteration in each year dependent on environmental factors such as drought and temperature. This information benefits growers and breeders for considering agronomic practices to select for soybean seed nutritional qualities under drought and high heat conditions.

The new Early Soybean Production System (ESPS), developed in the Midsouth USA, including the Mississippi delta, resulted in higher yield under irrigated and non-irrigated conditions. However, information on the effects of the agricultural practices such as row-type (RT: twin- vs. single-row), row-spacing, (RS), seeding rate (SR), soil-type (ST) on seed nutrition under the ESPS environment in the Mississippi delta is very limited. Our previous research in the Mississippi delta showed these agricultural practices altered seed nutrients in one cultivar only. However, whether these effects on seed nutrients will be exhibited by other soybean cultivars with earlier and later maturities across multiple years are not yet known. Therefore, the objective of this research was to evaluate the effects of agricultural practices and cultivar (Cv) differences on seed nutrition in clay and sandy soils under ESPS environment of high heat and drought. Two field experiments were conducted; one experiment was conducted in 2009 and 2010, and the other in 2008, 2009, and 2010 under irrigated conditions. Soybean were grown on 102 cm single-rows and on 25 cm twin-rows with 102 cm centers at seeding rates of 20, 30, 40, and 50 seeds m(-2). Two soybean cultivars (94M80 with earlier maturity; and GP 533 with later maturity) were used. Results showed that increasing seeding rate resulted in increases of protein, sucrose, glucose, raffinose, B, and P concentrations on both single- and twin-rows. However, this increase became either constant or declined at the higher rates (40 and 50 seeds m(-2)). Protein and linolenic acid concentrations were higher in GP 533 than in 94M80 on both row-types, but oil and oleic acid concentrations were in 94M80 than GP 533. Generally, cultivar GP 533 accumulated more seed constituents in seeds than 94M80. In 2010, there were no clear responses of seed nutrients to SR increase in both cultivars, perhaps due to drier year and high heat in 2010. It is concluded that RT and SR can alter seed nutrition under clay and sandy soils, especially under high heat and drought conditions as in the Mississippi delta.

A nontoxigenic Aspergillus flavus strain, K49, is currently being tested as a biological control agent in corn fields in the Mississippi Delta. However, little is known about the overall genetic diversity of A. flavus from year to year in corn fields and specifically in Mississippi. Our objective was to assess the genetic variability of A. flavus isolates from different seasons, inoculum sources, and years, from a no-till corn field. Of the 175 A. flavus isolates examined, 74 and 97 had the typical norB-cypA type I (1.5 kb) and type II (1.0 kb) deletion patterns, respectively. Variability in the sequence of the omtA gene of the majority of the field isolates (n=118) was compared to strain K49. High levels of haplotypic diversity (24 omtA haplotypes; Hd = 0.61 ± 0.04) were found. Among the 24 haplotypes, two were predominant, H1 (n=71), which consists of mostly toxigenic isolates, and H49 (n=18), which consists of mostly atoxigenic isolates including K49. Toxigenic isolates were prevalent (60%) in this natural population. Nonetheless, about 15% of the population likely shared the same ancestral origin with K49. This study provides valuable information on the diversity of A. flavus. This knowledge can be further used to develop additional biological control strains.

The effect of corn-soybean rotation on mycotoxin contamination in corn (Zea mays L.) and soybean (Glycine max L. Merrill.) grains has not been fully evaluated. Therefore, this research investigated the effect of corn-soybean rotation on aflatoxin and fumonisin contamination in respective grains. The results showed that aflatoxin levels in soybean averaged 2.3,

No abstract available. Article truncated after 150 words. A 19 year old man with acute lymphocytic leukemia was admitted to the ICU with septic shock. Due to areas of cellulitis on the right side of the chest and neck and femoral venous thrombi, a left subclavian central access was attempted. The chest x-ray obtained after line placement is shown below (Figure 1). Blood gas done from the line was consistent with venous blood and venous tranduction was seen. A CT of the chest (Figure 2) confirmed the diagnosis of persistent left superior vena cava (PLSVC). A persistent left superior vena cava is the most common congenital thoracic venous anomaly, seen in 0.3-0.5% of the population (1). Incidence is increased in patients with congenital heart disease to 5%. In most patients a right sided SVC is also present; hence the term SVC duplication has also been used. Embryologically a PLSVC is formed when the left anterior cardinal vein is ...

Four glyphosate resistant corn (Zea mays L.) hybrids, a glufosinate-ammonium resistant hybrid, and a conventional atrazine resistant hybrid gown at Stoneville, MS in 2005, 2006, and 2007 with furrow irrigation were treated with their respective herbicides and their growth, yield, and mycotoxin incidence were compared with untreated cultivated plots. Leaf area index (LAI) and dry matter accumulation (DMA) were collected on a weekly basis beginning at growth stage V3 and terminating at anthesis. Crop growth rates (CRGs) and relative growth rates (RGRs) were calculated. Plots were later harvested, yield and yield component data collected, and kernel samples analyzed for aflatoxin and fumonisin. Leaf area index, DMA, CRG, and RGR were not different among the herbicide treated plots and from those that were cultivated. Curves for LAI and DMA were similar to those previously reported. Aflatoxin and fumonisin were relatively low in all plots. Herbicide application or the lack thereof had no negative impact on the incidence of kernel contamination by these two mycotoxins. Herbicides, especially glyphosate on resistant hybrids, have no negative effects on corn yields or kernel quality in corn produced in a humid subtropical environment.

Mycotoxins that contaminate grain can cause the devaluation of agricultural products and create health risks for the consumer. Fumonisins are one such mycotoxin. Produced primarily by Fusarium verticillioides (Hypocreales: Nectriaceae) (Nirenberg, 1976) on corn, fumonisins' economic impact can be significant by causing various diseases in livestock if contaminated corn is not monitored and removed from animal feed. Finding safe alternatives to the destruction and waste of contaminated grain and restoring its economic value is needed for a sustainable future. Safe reintroduction into the farm food web may be possible through a consumable intermediary such as insects. This study demonstrates the suitability of the house cricket, Acheta domesticus L. as an alternative protein source in domestic animal feed by quantifying fumonisin B1 (FB1) levels in their subsequent insect meal and frass. Small colonies of 2nd instar A. domesticus were reared to 5th instar adults on nutrient-optimized corn-based diets treated with 4 levels of FB1 from 0 to 20 ppm. Increasing levels of FB1 had no adverse effects on the survivorship or growth of A. domesticus. Insect meals prepared from A. domesticus had significantly lower levels of FB1, at 3%–5% of their respective diets, while frass did not differ significantly from their diet. The successful rearing to adulthood of A. domesticus on fumonisin-contaminated diet paired with lower levels of FB1 in their processed insect meal supports the idea that more sustainable agricultural practices can be developed through remediation of low-value mycotoxin-contaminated grain with safer, higher-value insects as livestock feed components. [ABSTRACT FROM AUTHOR]

Abstract Background The utilization of the Internet can lead to adverse outcomes in students’ lives. Prolonged Internet usage may result in academic failure, an elevated risk of musculoskeletal problems, and the emergence of issues impacting the lives of students and their peers. This study aimed to explore the relationship between Internet addiction and musculoskeletal disorders (MSDs) among students in the field of medical sciences. Methods In this cross-sectional study, 314 medical sciences students in Semnan, Iran, were assessed using the census sampling method. The participants’ Internet addiction status and the presence of MSDs were determined through Young’s Internet Addiction questionnaire and the Nordic self-report questionnaire, respectively. Data underwent analysis employing logistic regression test to ascertain the relationship between Internet addiction and MSDs. Results Among the 314 participating students in this research, about 21% and 1.6% of the participants were lay open to and suffered from Internet addiction, respectively. Internet addiction was associated with the presence of pain, discomfort, burning, or numbness in the neck, wrist, upper back, hips, and thighs. For each unit increase in the Internet addiction score, the chance of neck and wrist pain rises by 3% compared to a 2% increase in upper back and hip/thigh pain. Conclusion The findings of this study indicate that excessive Internet use is linked to a heightened risk of MSDs in medical sciences students. Consequently, it is imperative to raise awareness about these complications and promote the adoption of proper posture when using the Internet. Further research is also warranted among diverse student populations to investigate the impact of Internet addiction.

Abstract Background Colonic diverticulosis, the most common lesion found in routine colonoscopy, affects more than 50% of individuals aged ≥ 60 years. Emerging evidence suggest that dysbiosis of gut microbiota may play an important role in the pathophysiology of diverticular disease. However, specific changes in microbial species and metabolic functions in asymptomatic diverticulosis remain unknown. Methods In a cohort of US adults undergoing screening colonoscopy, we analyzed the gut microbiota using shotgun metagenomic sequencing. Demographic factors, lifestyle, and medication use were assessed using a baseline questionnaire administered prior to colonoscopy. Taxonomic structures and metabolic pathway abundances were determined using MetaPhlAn3 and HUMAnN3. We used multivariate association with linear models to identify microbial species and metabolic pathways that were significantly different between asymptomatic diverticulosis and controls, while adjusting for confounders selected a priori including age at colonoscopy, sex, body mass index (BMI), and dietary pattern. Results Among 684 individuals undergoing a screening colonoscopy, 284 (42%) had diverticulosis. Gut microbiome composition explained 1.9% variation in the disease status of asymptomatic diverticulosis. We observed no significant differences in the overall diversity of gut microbiome between asymptomatic diverticulosis and controls. However, microbial species Bifidobacterium pseudocatenulatum and Prevotella copri were significantly enriched in controls (q value = 0.19 and 0.14, respectively), whereas Roseburia intestinalis, Dorea sp. CAG:317, and Clostridium sp. CAG: 299 were more abundant in those with diverticulosis (q values = 0.17, 0.24, and 0.10, respectively). We observed that the relationship between BMI and diverticulosis appeared to be limited to carriers of Bifidobacterium pseudocatenulatum and Roseburia intestinalis (P interaction = 0.09). Conclusions Our study provides the first large-scale evidence supporting taxonomic and functional shifts of the gut microbiome in individuals with asymptomatic diverticulosis. The suggestive interaction between gut microbiota and BMI on prevalent diverticulosis deserves future investigations.