Metabolomic analysis reveals altered amino acid metabolism and mechanisms underlying Eimeria infection in laying hens.

Avian coccidiosis, caused by Eimeria spp, is a devastating disease in laying hens. Previous studies have suggested that amino acids may be involved in Eimeria infection of broiler chickens. However, their metabolic features in laying hens, as well as the effect of multiple Eimeria species challenges on poultry hosts have not been elucidated yet. Here, a targeted metabolomics approach was employed to identify altered amino acid metabolism and mechanisms in laying hens with multiple Eimeria species challenges. Laying hens, Hy-Line W-36 aged 25 wk, were randomly assigned to a control group and groups inoculated with varying levels of mixed Eimeria species (E. maxima, E. tenella, and E. acervulina). Serum samples from each group were collected at 6 d and 14 d of postinoculation (6 and 14 DPI) for metabolite profiling. Metabolomic analysis revealed notable metabolic variations between control and infected groups, especially at 6 DPI stage. Varying levels of Eimeria dosages did not show a significant metabolic difference, and metabolites were sensitive to low-level infection. With statistical analysis, differentially expressed compounds (3-methylhistidine, alanine, aspartate, lysine, asparagine, methionine, ornithine, and tryptophan) were selected, and their metabolic network was identified by pathway enrichment analysis. In the network, the lysine biosynthesis pathway was upregulated, while the arginine and proline metabolic pathway was downregulated under infection. Other pathways showed complex patterns of metabolic relationships. Based on the results, biological implications of metabolic changes were elucidated and discussed. Last, the results were further confirmed with our previous study (phenotype and gene expression results) using the same set of samples. Our finding provides in-depth information on altered amino acid metabolism and mechanisms in laying hens upon multiple Eimeria species infection.

Metabolomic analysis combined with machine learning algorithms enables the evaluation of postharvest pecan color stability.

Nut kernel color is a crucial quality indicator affecting the consumers first impression of the product. While growing evidence suggests that plant phenolics and their derivatives are linked to nut kernel color, the compounds (biomarkers) responsible for kernel color stability during storage remain elusive. Here, pathway-based metabolomics with machine learning algorithms were employed to identify key metabolites of postharvest pecan color stability. Metabolites in phenylpropanoid, flavonoid, and anthocyanin biosynthetic pathways were analyzed in the testa of nine pecan cultivars using liquid chromatography-mass spectrometry. With color measurements, different machine learning models were compared to find relevant biomarkers of pecan color phenotypes. Results revealed potential marker compounds that included flavonoid precursors and anthocyanidins as well as anthocyanins (e.g., peonidin, delphinidin-3-O-glucoside). Our findings provide a foundation for future research in the area, and will help select genes/proteins for the breeding of pecans with stable and desirable kernel color.

Metabolomic analysis reveals linkage between chemical composition and sensory quality of different floral honey samples.

Honey has a distinct flavor characterized by various volatiles and non-volatiles from diverse origins. In this study, metabolomics combined with sensory analysis was performed to identify relationships between chemical profile and sensory quality of honey. Targeted metabolomic analysis was conducted to determine volatile and non-volatile profiles of seven different honey. Volatile profile was analyzed using headspace solid-phase microextraction (HS-SPME) coupled to GC - MS. LC - MS/MS, HPLC - UV, and HPLC-RI were employed to analyze flavonoids, organic acids, and sugars, respectively. Authentic standards were utilized for confirmation of metabolites. Sensory evaluation included quantitative descriptive analysis and consumer acceptance test. The results showed that sucrose (sweetness) was responsible for a positive hedonic perception, while organic acids and flavonoids (sourness, astringency, bitterness) negatively affected consumer acceptance. Volatiles with floral notes (e.g. decyl formate) were preferred, but others with off-flavors (e.g. 2-methylbenzofuran) were not preferred by consumers. Flavor familiarity was strongly correlated with the consumer acceptance of honey, indicating that the balance between volatiles and non-volatiles is significant for honey flavor quality. This work demonstrates the role of key flavor compounds in honey quality, and may be applicable to the quality control of honey.

Pea hull fiber supplementation does not modulate uremic metabolites in adults receiving hemodialysis: a randomized, double-blind, controlled trial.

Background: Fiber is a potential therapeutic to suppress microbiota-generated uremic molecules. This study aimed to determine if fiber supplementation decreased serum levels of uremic molecules through the modulation of gut microbiota in adults undergoing hemodialysis. Methods: A randomized, double-blinded, controlled crossover study was conducted. Following a 1-week baseline, participants consumed muffins with added pea hull fiber (PHF) (15 g/d) and control muffins daily, each for 4 weeks, separated by a 4-week washout. Blood and stool samples were collected per period. Serum p-cresyl sulfate (PCS), indoxyl sulfate (IS), phenylacetylglutamine (PAG), and trimethylamine N-oxide (TMAO) were quantified by LC-MS/MS, and fecal microbiota profiled by 16S rRNA gene amplicon sequencing and specific taxa of interest by qPCR. QIIME 2 sample-classifier was used to discover unique microbiota profiles due to the consumption of PHF. Results: Intake of PHF contributed an additional 9 g/d of dietary fiber to the subjects' diet due to compliance. No significant changes from baseline were observed in serum PCS, IS, PAG, or TMAO, or for the relative quantification of Akkermansia muciniphila, Faecalibacterium prausnitzii, Bifidobacterium, or Roseburia, taxa considered health-enhancing. Dietary protein intake and IS (r = -0.5, p = 0.05) and slow transit stool form and PCS (r = 0.7, p < 0.01) were significantly correlated at baseline. PHF and control periods were not differentiated; however, using machine learning, taxa most distinguishing the microbiota composition during the PHF periods compared to usual diet alone were enriched Gemmiger, Collinsella, and depleted Lactobacillus, Ruminococcus, Coprococcus, and Mogibacteriaceae. Conclusion: PHF supplementation did not mitigate serum levels of targeted microbial-generated uremic molecules. Given the high cellulose content, which may be resistant to fermentation, PHF may not exert sufficient effects on microbiota composition to modulate its activity at the dose consumed.

Relationships between sensory properties and metabolomic profiles of different apple cultivars.

Flavor is a critical factor in apple quality. To better understand apple flavor, this study aimed to identify the relationships between sensory attributes and the chemical composition (volatiles and non-volatiles) of apples using a combined metabolomic and sensory evaluation. Sensory results showed the positive (apple, fruity, pineapple, sweetness, sourness) and negative (cucumber) flavor attributes of apples. A metabolomic analysis with statistical correlations revealed significant metabolites related to the flavor attributes of apples. Volatile esters (e.g., hexyl acetate and 2-methylbutyl acetate for apple and fruity notes) and non-volatile sugars and acids (total sugars, tartaric acid, and malic acid for balanced sweet and tart flavors) were associated with the apple flavor preferred by consumers. Some aldehydes and alcohols (e.g., (E)-2-nonenal) contributed to a negative hedonic perception (cucumber). The collected information demonstrated the roles of key chemical compounds in apple flavor quality, and may be applicable to quality control.

Influence of antibiotic-resistance and exudate on peroxyacetic acid tolerance in O157 and non-O157 Shiga toxin producing E. coli.

Shiga toxin producing Escherichia coli (STEC) continues to cause foodborne outbreaks associated with beef and beef products despite consistent use of antimicrobial interventions. In this study, the influence of antibiotic resistance (ABR) in E. coli O157:H7 H1730, O157:H7 43,895, O121:H19 and O26:H11 on tolerance to peroxyacetic acid (PAA) was evaluated. Further, bactericidal concentrations of PAA in the presence of nutrient rich media (Tryptic Soy Broth, TSB and beef exudates) and nutrient deficient media (Sterile Deionized Water, SDW and Phosphate Buffered Saline, PBS) were evaluated for all bacterial strains. Antibiotic resistance to ampicillin (amp C), or ampicillin and streptomycin (amp P strep C) was generated in each bacterial strain through incremental exposure to the antibiotics or by plasmid transformation (n = 12 total strains). The mean bactericidal concentrations of PAA were higher (p ≤ 0.05) in nutrient rich media (205.55 ± 31.11 in beef exudate and 195.83 ± 25.00 ppm in TSB) than in nutrient deficient media (57.91 ± 11.97 ppm in SDW and 56.66 ± 9.56 ppm in PBS). Strain O157: H7 ampP strepC was the most tolerant to PAA (p ≤ 0.05). At 200 ppm in nutrient rich media and 60 ppm in nutrient deficient media, all bacterial strains declined in population to below the limit of detection. Analysis of the beef exudates indicated the presence of diverse amino acids that have been associated with acid tolerance. The results from this study indicate that beef exudates could contribute to acid tolerance and suggest that some STEC bacterial strains with certain ABR profiles might be more tolerant to PAA.

Increased susceptibility to Chrysanthemum Yellows phytoplasma infection in Atcals7ko plants is accompanied by enhanced expression of carbohydrate transporters.

MAIN CONCLUSION: Loss of CALS7 appears to confer increased susceptibility to phytoplasma infection in Arabidopsis, altering expression of genes involved in sugar metabolism and membrane transport. Callose deposition around sieve pores, under control of callose synthase 7 (CALS7), has been interpreted as a mechanical response to limit pathogen spread in phytoplasma-infected plants. Wild-type and Atcals7ko mutants were, therefore, employed to unveil the mode of involvement of CALS7 in the plant's response to phytoplasma infection. The fresh weights of healthy and CY-(Chrysanthemum Yellows) phytoplasma-infected Arabidopsis wild type and mutant plants indicated two superimposed effects of the absence of CALS7: a partial impairment of photo-assimilate transport and a stimulated phytoplasma proliferation as illustrated by a significantly increased phytoplasma titre in Atcal7ko mutants. Further studies solely dealt with the effects of CALS7 absence on phytoplasma growth. Phytoplasma infection affected sieve-element substructure to a larger extent in mutants than in wild-type plants, which was also true for the levels of some free carbohydrates. Moreover, infection induced a similar upregulation of gene expression of enzymes involved in sucrose cleavage (AtSUS5, AtSUS6) and transmembrane transport (AtSWEET11) in mutants and wild-type plants, but an increased gene expression of carbohydrate transmembrane transporters (AtSWEET12, AtSTP13, AtSUC3) in infected mutants only. It remains still unclear how the absence of AtCALS7 leads to gene upregulation and how an increased intercellular mobility of carbohydrates and possibly effectors contributes to a higher susceptibility. It is also unclear if modified sieve-pore structures in mutants allow a better spread of phytoplasmas giving rise to higher titre.

Critical review: Metabolomics in dairy science - Evaluation of milk and milk product quality.

Milk and milk products are nutritionally rich and consumed globally. Maintaining nutritional quality and ensuring safety of these products have become one of the major topics in dairy research. Dairy products contain metabolites including key nutritional elements, which are derived from dairy animals and elsewhere (e.g. milk processing, fermentation). Since the level and type of metabolites can vary by diverse factors from farm to table dairy, metabolites may represent the quality of milk and milk products in terms of nutritional value, authenticity, safety, and so on. In this review, we introduce metabolomics as a powerful tool to obtain a comprehensive snapshot of metabolite composition and dynamic changes, and focus on its recent progress and applications in dairy product quality. Factors (pre- and post-harvest effects, contamination, adulteration, etc.) affecting the quality and safety of products are dissected, and examples of related metabolomics works are provided. Potential metabolite indicators and metabolic mechanisms associated with the quality factors of dairy products are presented. With cases of single metabolomics approach, current trends in the integration of metabolomics with other omics techniques (so called multi-omics) in dairy science, as well as future perspectives of metabolomics in the field are also explored and discussed.

Salicylic acid mediated immune response of Citrus sinensis to varying frequencies of herbivory and pathogen inoculation.

BACKGROUND: Plant immunity against pathogens and pests is comprised of complex mechanisms orchestrated by signaling pathways regulated by plant hormones [Salicylic acid (SA) and Jasmonic acid (JA)]. Investigations of plant immune response to phytopathogens and phloem-feeders have revealed that SA plays a critical role in reprogramming of the activity and/or localization of transcriptional regulators via post-translational modifications. We explored the contributing effects of herbivory by a phytopathogen vector [Asian citrus psyllid, Diaphorina citri] and pathogen [Candidatus Liberibacter asiaticus (CaLas)] infection on response of sweet orange [Citrus sinensis (L.) Osbeck] using manipulative treatments designed to mimic the types of infestations/infections that citrus growers experience when cultivating citrus in the face of Huanglongbing (HLB) disease. RESULTS: A one-time (7 days) inoculation access period with CaLas-infected vectors caused SA-associated upregulation of PR-1, stimulating defense response after a long period of infection without herbivory (270 and 360 days). In contrast, while repeated (monthly) 'pulses' of 7 day feeding injury by psyllids stimulated immunity in CaLas-infected citrus by increasing SA in leaves initially (up to 120 days), long-term (270 and 360 days) repeated herbivory caused SA to decrease coincident with upregulation of genes associated with SA metabolism (BMST and DMR6). Similarly, transcriptional responses and metabolite (SA and its analytes) accumulation in citrus leaves exposed to a continuously reproducing population of D. citri exhibited a transitory upregulation of genes associated with SA signaling at 120 days and a posterior downregulation after long-term psyllid (adults and nymphs) feeding (270 and 360 days). CONCLUSIONS: Herbivory played an important role in regulation of SA accumulation in mature leaves of C. sinensis, whether or not those trees were coincidentally infected with CaLas. Our results indicate that prevention of feeding injury inflicted by D. citri from the tritrophic interaction may allow citrus plants to better cope with the consequences of CaLas infection, highlighting the importance of vector suppression as a component of managing this cosmopolitan disease.

Pathway-Based Metabolomics Analysis Reveals Biosynthesis of Key Flavor Compounds in Mango.

Mango is a tropical fruit with global demand as a result of its high sensory quality and nutritional attributes. Improving fruit quality at the consumer level could increase demand, but fruit quality is a complex trait requiring a deep understanding of flavor development to uncover key pathways that could become targets for improving sensory quality. Here, a pathway-based metabolomics (untargeted and targeted) approach was used to explore biosynthetic mechanisms of key flavor compounds with five core metabolic pathways (butanoate metabolism, phenylalanine biosynthesis and metabolism, terpenoid backbone biosynthesis, linoleic and linolenic acid pathway, and carbon fixation and sucrose metabolism) in three mango cultivars. The relationships between flavor precursors and flavor compounds were identified using correlation analysis. With these novel strategies, differentially regulated metabolic flux through the pathways was first elucidated, demonstrating possible mechanisms of key flavor formation and regulation in mango fruits.

Metabolomic analysis elucidates how shade conditions ameliorate the deleterious effects of greening (Huanglongbing) disease in citrus.

Under tropical and subtropical environments, citrus leaves are exposed to excess sunlight, inducing photoinhibition. Huanglongbing (HLB, citrus greening), a devastating phloem-limited disease putatively caused by Candidatus Liberibacter asiaticus, exacerbates this challenge with additional photosynthetic loss and excessive starch accumulation. A combined metabolomics and physiological approach was used to elucidate whether shade alleviates the deleterious effects of HLB in field-grown citrus trees, and to understand the underlying metabolic mechanisms related to shade-induced morpho-physiological changes in citrus. Using metabolite profiling and multinomial logistic regression, we identified pivotal metabolites altered in response to shade. A core metabolic network associated with shade conditions was identified through pathway enrichment analysis and metabolite mapping. We measured physio-biochemical responses and growth and yield characteristics. With these, the relationships between metabolic network and the variables measured above were investigated. We found that moderate-shade alleviates sink limitation by preventing excessive starch accumulation and increasing foliar sucrose levels. Increased growth and fruit yield in shaded compared with non-shaded trees were associated with increased photosystem II efficiency and leaf carbon fixation pathway metabolites. Our study also shows that, in HLB-affected trees under shade, the signaling of plant hormones (auxins and cytokinins) and nitrogen supply were downregulated with reducing new shoot production likely due to diminished needs of cell damage repair and tissue regeneration under shade. Overall, our findings provide the first glimpse of the complex dynamics between cellular metabolites and leaf physiological functions in citrus HLB pathosystem under shade, and reveal the mechanistic basis of how shade ameliorates HLB disease.

Comparative Leaf Volatile Profiles of Two Contrasting Mandarin Cultivars against Candidatus Liberibacter asiaticus Infection Illustrate Huanglongbing Tolerance Mechanisms.

Huanglongbing (HLB), presumably caused by Candidatus Liberibacter asiaticus (CaLas), is a devastating citrus disease worldwide. While all citrus are affected by HLB, some cultivars display greater tolerance; however, the underlying mechanisms are not fully understood. Here, volatile changes in HLB-tolerant LB8-9 Sugar Belle (SB) and HLB-sensitive Murcott mandarins after CaLas infection were comprehensively compared to determine if specific volatiles are associated with HLB responses and to discern the underlying tolerance mechanisms. These cultivars emitted qualitatively and quantitatively different volatiles in response to HLB induced by artificial graft or natural psyllid inoculation. Increasing amounts of total volatiles and de novo-synthesized new volatiles were two key responses to HLB of both cultivars. Markers potentially associated with HLB and host susceptibility were identified. Terpenoid biosynthetic pathway, green leaf volatile, and thymol metabolic pathways responsive to CaLas infection were dramatically altered. SB mandarin allows simultaneous defense and growth, contributing to its greater HLB tolerance.

Metabolomic Analysis Provides New Insight Into Tolerance of Huanglongbing in Citrus.

There have been efforts to develop citrus cultivars that are tolerant of Huanglongbing (HLB), a catastrophic phloem-limited disease. Previous studies demonstrated that continuous plant growth with phloem regeneration is one of the major characteristics of HLB tolerance. In this study, the metabolic mechanisms of HLB tolerance in citrus were elucidated using a multiple pathway-targeted metabolomic approach. Comparative analysis of healthy and infected HLB-tolerant and HLB-sensitive mandarin cultivars (Citrus reticulata) revealed differentially expressed metabolic responses among different groups. Pathway enrichment analysis indicated aspartate and glutamate metabolism, purine metabolism, and biosynthesis of plant hormones were upregulated in the tolerant group, except salicylic acid signaling. Catabolic pathways linked to energy-yielding metabolism were also upregulated in the tolerant group. These metabolisms and pathways were interconnected with each other, unveiling a pivotal metabolic network associated with HLB tolerance. In the network, auxins and cytokinins, the plant hormones responsible for plant growth and phloem regeneration, were accumulated. In addition, purine metabolites serving as energy carriers and nitrogen sources of plants were increased. Only salicylic acid-related metabolites for plant defense responses were decreased in the tolerant group. Our findings may evidence the strategy of HLB-tolerant cultivars that sustain plant growth and phloem formation rather than displaying direct plant defense to overcome the disease.

Mediterranean Diet Adherence in People With Parkinson's Disease Reduces Constipation Symptoms and Changes Fecal Microbiota After a 5-Week Single-Arm Pilot Study.

Introduction: Non-motor symptoms of Parkinson's disease (PD) such as gastrointestinal (GI) dysfunction are common, yet little is known about how modifying dietary intake impacts PD symptoms. The aim of this study in individuals with PD was to determine whether a Mediterranean diet intervention is feasible and affects GI function, intestinal permeability and fecal microbial communities. Methods: A single-arm, 5-week Mediterranean diet intervention study was conducted in eight people with PD. Daily and weekly questionnaires were administered to determine changes in GI symptoms. Urine and stool samples were collected at baseline and after 5 weeks to assess intestinal permeability and fecal microbial communities. Additionally, live-in partners of the participants with PD were matched as controls (n = 8) for baseline urine and stool samples. Results: Participants with PD increased intake of Mediterranean diet based on adherence scores from baseline to week 5 (4.4 ± 0.6 vs. 11.9 ± 0.7; P < 0.01 with >10 representing good adherence), which was linked with weight loss (77.4 kg vs. 74.9 kg, P = 0.01). Constipation syndrome scores decreased after 5 weeks (2.3 ± 0.5 vs. 1.5 ± 0.3; P = 0.04). Bilophila, was higher at baseline in PD (0.6 ± 0.1% vs. 0.2 ± 0.1% P = 0.02) and slightly decreased after the diet intervention (0.5 ± 0.1%; P = 0.01). Interestingly, the proportion of Roseburia was significantly lower in PD compared to controls (0.6 ± 0.2% vs. 1.6 ± 0.3%; P = 0.02) and increased at week 5 (0.9 ± 0.2%; P < 0.01). No differences were observed for markers of intestinal permeability between the control and PD groups or post-intervention. Conclusions: Short-term Mediterranean diet adherence is feasible in participants with PD; correlated with weight loss, improved constipation, and modified gut microbiota. Clinical Trial Registration: ClinicalTrials.gov, identifier: NCT03851861.

In older women, a high-protein diet including animal-sourced foods did not impact serum levels and urinary excretion of trimethylamine-N-oxide.

Diets including red meat and other animal-sourced foods may increase proteolytic fermentation and microbial-generated trimethylamine (TMA) and, subsequently, trimethylamine-N-oxide (TMAO), a metabolite associated with increased risk of cardiovascular disease and dementia. It was hypothesized that compared to usual dietary intake, a maintenance-energy high-protein diet (HPD) would increase products of proteolytic fermentation, whereas adjunctive prebiotic, probiotic, and synbiotic supplementation may mitigate these effects. An exploratory aim was to determine the association of the relative abundance of the TMA-generating taxon, Emergencia timonensis, with serum and urinary TMAO. At 5 time points (usual dietary intake, HPD diet, HPD + prebiotic, HPD + probiotic, and HPD + synbiotic), urinary (24-hour) and serum metabolites and fecal microbiota profile of healthy older women (n = 20) were measured by liquid chromatography-tandem mass spectrometry and 16S rRNA gene amplicon sequencing analyses, respectively. The HPD induced increases in serum levels of l-carnitine, indoxyl sulfate, and phenylacetylglutamine but not TMAO or p-cresyl sulfate. Urinary excretion of l-carnitine, indoxyl sulfate, phenylacetylglutamine, and TMA increased with the HPD but not TMAO or p-cresyl sulfate. Most participants had undetectable levels of E.timonensis at baseline and only 50% during the HPD interventions, suggesting other taxa are responsible for the microbial generation of TMA in these individuals. An HPD diet with or without a prebiotic, probiotic, or synbiotic elicited an increase in products of proteolytic fermentation. The urinary l-carnitine response suggests that the additional dietary l-carnitine provided was primarily bioavailable, providing little substrate for microbial conversion to TMA and subsequent TMAO formation.

Screening of Phosphodiesterase-5 Inhibitors and Their Analogs in Dietary Supplements by Liquid Chromatography-Hybrid Ion Trap-Time of Flight Mass Spectrometry.

An accurate and reliable method based on ion trap-time of flight mass spectrometry (IT-TOF MS) was developed for screening phosphodiesterase-5 inhibitors, including sildenafil, vardenafil, and tadalafil, and their analogs in dietary supplements. Various parameters affecting liquid chromatographic separation and IT-TOF detection were investigated, and the optimal conditions were determined. The separation was achieved on a reversed-phase column under gradient elution using acetonitrile and water containing 0.2% acetic acid at a flow rate of 0.2 mL/min. The chromatographic eluents were directly ionized in the IT-TOF system equipped with an electrospray ion source operating in the positive ion mode. The proposed screening method was validated by assessing its linearity, precision, and accuracy. Sequential tandem MS was conducted to obtain structural information of the references, and the fragmentation mechanism of each reference was proposed for providing spectral insight for newly synthesized analogs. Structural information, including accurate masses of both parent and fragment ions, was incorporated into the MSn spectral library. The developed method was successfully applied for screening adulterated dietary supplement samples.

Analysis of Nicotine Metabolites in Hair and Nails Using QuEChERS Method Followed by Liquid Chromatography-Tandem Mass Spectrometry.

Many studies have analyzed nicotine metabolites in blood and urine to determine the toxicity caused by smoking, and assess exposure to cigarettes. Recently, hair and nails have been used as alternative samples for the evaluation of smoking, as not only do they reflect long-term exposure but they are also stable and easy to collect. Liquid-liquid or solid-phase extraction has mainly been used to detect nicotine metabolites in biological samples; however, these have disadvantages, such as the use of toxic organic solvents and complex pretreatments. In this study, a modified QuEChERS method was proposed for the first time to prepare samples for the detection of nicotine metabolite cotinine (COT) and trans-3'-hydroxycotinine (3-HCOT) in hair and nails. High-performance liquid chromatography-tandem mass spectrometry (LC-MS/MS) was used to analyze traces of nicotine metabolites. The established method was validated for selectivity, linearity, lower limit of quantitation, accuracy, precision and recovery. In comparison with conventional liquid-liquid extraction (LLE), the proposed method was more robust, and resulted in higher recoveries with favorable analytical sensitivity. Using this method, clinical samples from 26 Korean infants were successfully analyzed. This method is expected to be applicable in the routine analysis of nicotine metabolites for environmental and biological exposure monitoring.

A novel dried blood spots analysis combined with on-spot reaction for determination of trimethylamine N-oxide and its related compounds.

The application of dried blood spots in clinical research is becoming increasingly popular owing to its convenient collection, storage, and transportation compared to that of conventional biological samples. The potential of trimethylamine N-oxide and its related compounds as biomarkers for various cardiovascular diseases, such as atherosclerosis, stroke, thrombosis, and heart failure, was recently highlighted, which was the driving force behind the development of an analytical method to identify trimethylamine N-oxide and eight related compounds in dried blood spots. In the proposed method, a novel "on-spot reaction" approach was introduced to overcome the low loading efficiency of trimethylamine in dried blood spots. Upon the addition of 50 µL of blood onto the filter paper pretreated with dilute HCl, an acid-base neutralization reaction in the blood spots transformed the volatile trimethylamine to a salt. Next, a punched disc with a diameter of 6.0 mm was eluted by agitation with 20 mM ammonium formate for 10 min and derivatized with 1.0 M ethyl bromoacetate at 80 °C for 60 min. A surrogate analyte approach was employed for quantification of these endogenous compounds in the complex matrix. Analysis was carried out using zwitterionic hydrophilic interaction liquid chromatography-high-resolution mass spectrometry. The established method was validated and applied to monitor real samples from 30 clinical cases. The proposed new methodology based on dried blood spots could greatly improve the convenience, analytical sensitivity, and selectivity of cardiovascular disease testing.

Long-term, sustained feeding by Asian citrus psyllid disrupts salicylic acid homeostasis in sweet orange.

BACKGROUND: Phloem-feeding insects are known to modulate the salicylic acid (SA) signaling pathway in various plant-insect interaction models. Diaphorina citri is a phloem feeding vector of the deadly phytopathogens, Candidatus Liberibacter americanus and Candidatus Liberibacter asiaticus, and the interactions of D. citri with its host that may modulate plant defenses are not well understood. The objectives of this study were to investigate the molecular mechanisms involved in transcriptional regulation of SA modification and activation of defense-associated responses in sweet orange (Citrus sinensis) exposed to various durations (7-, 14- and 150- days) of continuous feeding by D. citri. RESULTS: We quantified expression of genes involved in SA pathway activation and subsequent modification, as well as, associated SA metabolites (SA methyl ester, 2,3-DHBA, and SA 2-O-ß-D-glucoside). NPR1 and PR-1 expression was upregulated in plants exposed to continuous feeding by D. citri for 14 days. Expression of BSMT-like, MES1-like and DMR6-like oxygenase, as well as, accumulation of their respective SA metabolites (SA methyl ester, 2,3-DHBA) was significantly higher in plants exposed to continuous feeding by D. citri for 150 days than in those without D. citri infestation. Concomitantly, expression of UGT74F2-like was significantly downregulated and its metabolite, SA 2-ß-D-glucoside, was highly accumulated in trees exposed to 150 d of feeding compared to control trees without D. citri. CONCLUSIONS: D. citri herbivory differentially regulated transcription and SA-metabolite accumulation in citrus leaves, depending on duration of insect feeding. Our results suggest that prolonged and uninterrupted exposure (150 d) of citrus to D. citri feeding suppressed plant immunity and inhibited growth, which may highlight the importance of vector suppression as part of huanglongbing (HLB) management in citrus.

Evaluation of California-Grown Blood and Cara Cara Oranges Through Consumer Testing, Descriptive Analysis, and Targeted Chemical Profiling.

The acceptability of Moro, Tarocco, Cara Cara, Shahani, Bream Tarocco, Boukhobza, and Sanguinelli oranges from both commercial and research orchards was tested with adult (n = 152) and child (n = 72) consumers. Qualitative focus groups were also conducted to understand consumer familiarity and thoughts about the fruit. Sensory descriptive and chemical analyses were carried out to identify drivers of liking. Overall, consumers preferred the lighter colored varieties consisting of Tarocco, Cara Cara, and Boukhobza. One cluster of adults (n = 80) showed preferences towards sweet and fruity flavors and away from sourness and citric acid. The second adult cluster (n = 72) was tolerant of the sour fruit but did not like fruit high in bitterness and flavonoid content. The largest child cluster (n = 42) showed preferences for samples higher in orange and tropical flavors (Cara Cara, Tarocco, and Boukhobza varieties). The appearance of the Cara Cara was strongly liked by the consumer population in both quantitative and qualitative settings. Hunter scale a color values strongly correlated to the higher berry/dried fruit flavors, and concentrations of naringenin. Focus group participants noted that they were relatively unfamiliar with blood oranges. Growers and producers may want to invest in the lighter colored varieties, such as Cara Cara, Tarocco, Boukhobza and Shahani, as these were liked by a majority of consumers and were low in less desirable sensory characteristics, such as bitterness and sourness. PRACTICAL APPLICATION: Through consumer tests, sensory evaluation, and chemical analyses, this research uncovered which sensory properties may drive consumer acceptance of blood and Cara Cara oranges, and informed potential production and marketing strategies for increasing their consumption. This information should benefit the citrus industry as a whole and may enhance the use of specialty oranges by the food, beverage, and food service industries.