Institute of Agrifood Research and Technology

autophagic responses. Here, we critically discuss current methods of assessing autophagy and the information they can, or cannot, provide. Our ultimate goal is to encourage intellectual and technical innovation in the field.

This revision of the classification of eukaryotes follows that of Adl et al., 2012 [J. Euk. Microbiol. 59(5)] and retains an emphasis on protists. Changes since have improved the resolution of many nodes in phylogenetic analyses. For some clades even families are being clearly resolved. As we had predicted, environmental sampling in the intervening years has massively increased the genetic information at hand. Consequently, we have discovered novel clades, exciting new genera and uncovered a massive species level diversity beyond the morphological species descriptions. Several clades known from environmental samples only have now found their home. Sampling soils, deeper marine waters and the deep sea will continue to fill us with surprises. The main changes in this revision are the confirmation that eukaryotes form at least two domains, the loss of monophyly in the Excavata, robust support for the Haptista and Cryptista. We provide suggested primer sets for DNA sequences from environmental samples that are effective for each clade. We have provided a guide to trophic functional guilds in an appendix, to facilitate the interpretation of environmental samples, and a standardized taxonomic guide for East Asian users.

PURPOSE: PD-1 inhibitors are established agents in the management of non-small cell lung cancer (NSCLC); however, only a subset of patients derives clinical benefit. To determine the activity of PD-1/PD-L1 inhibitors within clinically relevant molecular subgroups, we retrospectively evaluated response patterns among EGFR-mutant, anaplastic lymphoma kinase (ALK)-positive, and EGFR wild-type/ALK-negative patients. EXPERIMENTAL DESIGN: We identified 58 patients treated with PD-1/PD-L1 inhibitors. Objective response rates (ORR) were assessed using RECIST v1.1. PD-L1 expression and CD8(+) tumor-infiltrating lymphocytes (TIL) were evaluated by IHC. RESULTS: Objective responses were observed in 1 of 28 (3.6%) EGFR-mutant or ALK-positive patients versus 7 of 30 (23.3%) EGFR wild-type and ALK-negative/unknown patients (P = 0.053). The ORR among never- or light- (≤10 pack years) smokers was 4.2% versus 20.6% among heavy smokers (P = 0.123). In an independent cohort of advanced EGFR-mutant (N = 68) and ALK-positive (N = 27) patients, PD-L1 expression was observed in 24%/16%/11% and 63%/47%/26% of pre-tyrosine kinase inhibitor (TKI) biopsies using cutoffs of ≥1%, ≥5%, and ≥50% tumor cell staining, respectively. Among EGFR-mutant patients with paired, pre- and post-TKI-resistant biopsies (N = 57), PD-L1 expression levels changed after resistance in 16 (28%) patients. Concurrent PD-L1 expression (≥5%) and high levels of CD8(+) TILs (grade ≥2) were observed in only 1 pretreatment (2.1%) and 5 resistant (11.6%) EGFR-mutant specimens and was not observed in any ALK-positive, pre- or post-TKI specimens. CONCLUSIONS: NSCLCs harboring EGFR mutations or ALK rearrangements are associated with low ORRs to PD-1/PD-L1 inhibitors. Low rates of concurrent PD-L1 expression and CD8(+) TILs within the tumor microenvironment may underlie these clinical observations. Clin Cancer Res; 22(18); 4585-93. ©2016 AACRSee related commentary by Gettinger and Politi, p. 4539.

Production of biogas from different organic materials is a most interesting source of renewable energy. The biomethane potential (BMP) of these materials has to be determined to get insight in design parameters for anaerobic digesters. Although several norms and guidelines for BMP tests exist, inter-laboratory tests regularly show high variability of BMPs for the same substrate. A workshop was held in June 2015, in Leysin, Switzerland, with over 40 attendees from 30 laboratories around the world, to agree on common solutions to the conundrum of inconsistent BMP test results. This paper presents the consensus of the intense roundtable discussions and cross-comparison of methodologies used in respective laboratories. Compulsory elements for the validation of BMP results were defined. They include the minimal number of replicates, the request to carry out blank and positive control assays, a criterion for the test duration, details on BMP calculation, and last but not least criteria for rejection of the BMP tests. Finally, recommendations on items that strongly influence the outcome of BMP tests such as inoculum characteristics, substrate preparation, test setup, and data analysis are presented to increase the probability of obtaining validated and reproducible results.

The traditional role of food packaging is continuing to evolve in response to changing market needs. Current drivers such as consumer's demand for safer, "healthier," and higher-quality foods, ideally with a long shelf-life; the demand for convenient and transparent packaging, and the preference for more sustainable packaging materials, have led to the development of new packaging technologies, such as active packaging (AP). As defined in the European regulation (EC) No 450/2009, AP systems are designed to "deliberately incorporate components that would release or absorb substances into or from the packaged food or the environment surrounding the food." Active packaging materials are thereby "intended to extend the shelf-life or to maintain or improve the condition of packaged food." Although extensive research on AP technologies is being undertaken, many of these technologies have not yet been implemented successfully in commercial food packaging systems. Broad communication of their benefits in food product applications will facilitate the successful development and market introduction. In this review, an overview of AP technologies, such as antimicrobial, antioxidant or carbon dioxide-releasing systems, and systems absorbing oxygen, moisture or ethylene, is provided, and, in particular, scientific publications illustrating the benefits of such technologies for specific food products are reviewed. Furthermore, the challenges in applying such AP technologies to food systems and the anticipated direction of future developments are discussed. This review will provide food and packaging scientists with a thorough understanding of the benefits of AP technologies when applied to specific foods and hence can assist in accelerating commercial adoption.

Abstract The need to accelerate breeding for increased yield potential and better adaptation to drought and other abiotic stresses is an issue of increasing urgency. As the population continues to grow rapidly, the pressure on resources (mainly untouched land and water) is also increasing, and potential climate change poses further challenges. We discuss ways to improve the efficiency of crop breeding through a better physiological understanding by both conventional and molecular methods. Thus the review highlights the physiological basis of crop yield and its response to stresses, with special emphasis on drought. This is not just because physiology forms the basis of proper phenotyping, one of the pillars of breeding, but because a full understanding of physiology is also needed, for example, to design the traits targeted by molecular breeding approaches such as marker-assisted selection (MAS) or plant transformation or the way these traits are evaluated. Most of the information in this review deals with cereals, since they include the world's main crops, however, examples from other crops are also included. Topics covered by the review include the conceptual framework for identifying secondary traits associated with yield potential and stress adaptation, and how to measure these secondary traits in practice. The second part of the review deals with the real role of molecular breeding for complex traits from a physiological perspective. This part examines current developments in MAS and quantitative trait loci (QTL) detection as well as plant transformation. Emphasis is placed on the current limitations of these molecular approaches to improving stress adaptation and yield potential. The essay ends by presenting some ideas regarding future avenues for crop breeding given the current and possible future challenges, and on a multidisciplinary approach where physiological knowledge and proper phenotyping play a major role. Keywords: breedingcerealsdroughtyield potential ACKNOWLEDGMENTS We are very grateful to Prof. Martin AJ Parry, Head of Plant Science Centre for Crop Genetic Improvement, Rothamsted Research (Email: martin.parry@bbsrc.ac.uk) and Prof. Roberto Tuberora, head of the Biotechnology Applied to Plant Breeding Group, Department of Agroenvironmental Sciences and Technology, University of Bologna, (roberto.tuberosa@unibo.it) for their very valuable comments and corrections. This study was supported in part by the European research project OPTIWHEAT (INCO-STRIP 015460) and by the Spanish Ministry of Science and Technology projects, AGL-2006-13541-C02-1 (for J.L. Araus and M.D. Serret), AGL 2006-07814/AGR (for G.A. Slafer) and AGL-2006-09226-C02-01 (for C. Royo). Referee: Richard Richards, Program Leader, 'High Performance Crops for Australia', CSIRO Plant Industry, POB 1600 Canberra 2601 Notes a Additive effect for grain yield (q ha− 1) computed as half of the difference between the mean phenotypic values of the RILs homozygous for the Svevo and Kofa alleles [(Svevo – Kofa) / 2]. † QTLs influencing more than one trait in a range of environments.

Abstract Life cycle thinking is increasingly seen as a key concept for ensuring a transition towards more sustainable production and consumption patterns. As food production systems and consumption patterns are among the leading drivers of impacts on the environment, it is important to assess and improve food-related supply chains as much as possible. Over the years, life cycle assessment has been used extensively to assess agricultural systems and food processing and manufacturing activities, and compare alternatives "from field to fork" and through to food waste management. Notwithstanding the efforts, several methodological aspects of life cycle assessment still need further improvement in order to ensure adequate and robust support for decision making in both business and policy development contexts. This paper discusses the challenges for life cycle assessment arising from the complexity of food systems, and recommends research priorities for both scientific development and improvements in practical implementation. In summary, the intrinsic variability of food production systems requires dedicated modelling approaches, including addressing issues related to: the distinction between technosphere and ecosphere; the most appropriate functional unit; the multi-functionality of biological systems; and the modelling of the emissions and how this links with life cycle impact assessment. Also, data availability and interpretation of the results are two issues requiring further attention, including how to account for consumer behaviour.

The bioassessment of aquatic ecosystems is currently based on various biotic indices that use the occurrence and/or abundance of selected taxonomic groups to define ecological status. These conventional indices have some limitations, often related to difficulties in morphological identification of bioindicator taxa. Recent development of DNA barcoding and metabarcoding could potentially alleviate some of these limitations, by using DNA sequences instead of morphology to identify organisms and to characterize a given ecosystem. In this paper, we review the structure of conventional biotic indices, and we present the results of pilot metabarcoding studies using environmental DNA to infer biotic indices. We discuss the main advantages and pitfalls of metabarcoding approaches to assess parameters such as richness, abundance, taxonomic composition and species ecological values, to be used for calculation of biotic indices. We present some future developments to fully exploit the potential of metabarcoding data and improve the accuracy and precision of their analysis. We also propose some recommendations for the future integration of DNA metabarcoding to routine biomonitoring programs.

Recent anecdotal and scientific reports have provided evidence of a link between COVID-19 and chemosensory impairments, such as anosmia. However, these reports have downplayed or failed to distinguish potential effects on taste, ignored chemesthesis, and generally lacked quantitative measurements. Here, we report the development, implementation, and initial results of a multilingual, international questionnaire to assess self-reported quantity and quality of perception in 3 distinct chemosensory modalities (smell, taste, and chemesthesis) before and during COVID-19. In the first 11 days after questionnaire launch, 4039 participants (2913 women, 1118 men, and 8 others, aged 19-79) reported a COVID-19 diagnosis either via laboratory tests or clinical assessment. Importantly, smell, taste, and chemesthetic function were each significantly reduced compared to their status before the disease. Difference scores (maximum possible change ±100) revealed a mean reduction of smell (-79.7 ± 28.7, mean ± standard deviation), taste (-69.0 ± 32.6), and chemesthetic (-37.3 ± 36.2) function during COVID-19. Qualitative changes in olfactory ability (parosmia and phantosmia) were relatively rare and correlated with smell loss. Importantly, perceived nasal obstruction did not account for smell loss. Furthermore, chemosensory impairments were similar between participants in the laboratory test and clinical assessment groups. These results show that COVID-19-associated chemosensory impairment is not limited to smell but also affects taste and chemesthesis. The multimodal impact of COVID-19 and the lack of perceived nasal obstruction suggest that severe acute respiratory syndrome coronavirus strain 2 (SARS-CoV-2) infection may disrupt sensory-neural mechanisms.

The geographic and temporal origins of the domestic dog remain controversial, as genetic data suggest a domestication process in East Asia beginning 15,000 years ago, whereas the oldest doglike fossils are found in Europe and Siberia and date to >30,000 years ago. We analyzed the mitochondrial genomes of 18 prehistoric canids from Eurasia and the New World, along with a comprehensive panel of modern dogs and wolves. The mitochondrial genomes of all modern dogs are phylogenetically most closely related to either ancient or modern canids of Europe. Molecular dating suggests an onset of domestication there 18,800 to 32,100 years ago. These findings imply that domestic dogs are the culmination of a process that initiated with European hunter-gatherers and the canids with whom they interacted.

Abstract Deep learning (DL) constitutes a modern technique for image processing, with large potential. Having been successfully applied in various areas, it has recently also entered the domain of agriculture. In the current paper, a survey was conducted of research efforts that employ convolutional neural networks (CNN), which constitute a specific class of DL, applied to various agricultural and food production challenges. The paper examines agricultural problems under study, models employed, sources of data used and the overall precision achieved according to the performance metrics used by the authors. Convolutional neural networks are compared with other existing techniques, and the advantages and disadvantages of using CNN in agriculture are listed. Moreover, the future potential of this technique is discussed, together with the authors’ personal experiences after employing CNN to approximate a problem of identifying missing vegetation from a sugar cane plantation in Costa Rica. The overall findings indicate that CNN constitutes a promising technique with high performance in terms of precision and classification accuracy, outperforming existing commonly used image-processing techniques. However, the success of each CNN model is highly dependent on the quality of the data set used.

The quantification of the biological diversity in environmental samples using high-throughput DNA sequencing is hindered by the PCR bias caused by variable primer-template mismatches of the individual species. In some dietary studies, there is the added problem that samples are enriched with predator DNA, so often a predator-specific blocking oligonucleotide is used to alleviate the problem. However, specific blocking oligonucleotides could coblock nontarget species to some degree. Here, we accurately estimate the extent of the PCR biases induced by universal and blocking primers on a mock community prepared with DNA of twelve species of terrestrial arthropods. We also compare universal and blocking primer biases with those induced by variable annealing temperature and number of PCR cycles. The results show that reads of all species were recovered after PCR enrichment at our control conditions (no blocking oligonucleotide, 45 °C annealing temperature and 40 cycles) and high-throughput sequencing. They also show that the four factors considered biased the final proportions of the species to some degree. Among these factors, the number of primer-template mismatches of each species had a disproportionate effect (up to five orders of magnitude) on the amplification efficiency. In particular, the number of primer-template mismatches explained most of the variation (~3/4) in the amplification efficiency of the species. The effect of blocking oligonucleotide concentration on nontarget species relative abundance was also significant, but less important (below one order of magnitude). Considering the results reported here, the quantitative potential of the technique is limited, and only qualitative results (the species list) are reliable, at least when targeting the barcoding COI region.

Abstract The design details and a sample set of field test results for a line source sprinkler plot irrigation system are presented. The system produces a water application pattern which is uniform along the length of the plot and continuously, but uniformly variable across the plot. By applying a fertility variable along a plot (at right angles to the water variable) planted in some test crop, the system offers a convenient means for developing crop production function data. The system test area and water supply are both small. However, the application of the system may be limited by wind and all water application levels within a plot must be supplied at the same irrigation frequency.

Gut microbiota-related metabolites are potential clinical biomarkers for cardiovascular disease (CVD). Circulating succinate, a metabolite produced by both microbiota and the host, is increased in hypertension, ischemic heart disease, and type 2 diabetes. We aimed to analyze systemic levels of succinate in obesity, a major risk factor for CVD, and its relationship with gut microbiome. We explored the association of circulating succinate with specific metagenomic signatures in cross-sectional and prospective cohorts of Caucasian Spanish subjects. Obesity was associated with elevated levels of circulating succinate concomitant with impaired glucose metabolism. This increase was associated with specific changes in gut microbiota related to succinate metabolism: a higher relative abundance of succinate-producing Prevotellaceae (P) and Veillonellaceae (V), and a lower relative abundance of succinate-consuming Odoribacteraceae (O) and Clostridaceae (C) in obese individuals, with the (P + V/O + C) ratio being a main determinant of plasma succinate. Weight loss intervention decreased (P + V/O + C) ratio coincident with the reduction in circulating succinate. In the spontaneous evolution after good dietary advice, alterations in circulating succinate levels were linked to specific metagenomic signatures associated with carbohydrate metabolism and energy production with independence of body weight change. Our data support the importance of microbe-microbe interactions for the metabolite signature of gut microbiome and uncover succinate as a potential microbiota-derived metabolite related to CVD risk.

Adopting the concept of One Welfare could help to improve animal welfare and human wellbeing worldwide, argue Rebeca García Pinillos, Michael Appleby, Xavier Manteca, Freda Scott-Park, Charles Smith and Antonio Velarde.

Grain yield is a major goal for the improvement of durum wheat, particularly in drought-prone areas. In this study, the genetic basis of grain yield (GY), heading date (HD), and plant height (PH) was investigated in a durum wheat population of 249 recombinant inbred lines evaluated in 16 environments (10 rainfed and 6 irrigated) characterized by a broad range of water availability and GY (from 5.6 to 58.8 q ha(-1)). Among the 16 quantitative trait loci (QTL) that affected GY, two major QTL on chromosomes 2BL and 3BS showed significant effects in 8 and 7 environments, with R2 values of 21.5 and 13.8% (mean data of all 16 environments), respectively. In both cases, extensive overlap was observed between the LOD profiles of GY and PH, but not with those for HD. QTL specific for PH were identified on chromosomes 1BS, 3AL, and 7AS. Additionally, three major QTL for HD on chromosomes 2AS, 2BL, and 7BS showed limited or no effects on GY. For both PH and GY, notable epistasis between the chromosome 2BL and 3BS QTL was detected across several environments.

Calves are born with a physically and metabolically underdeveloped rumen and initially rely on milk to meet nutrient demands for maintenance and growth. Initiation of solid feed consumption, acquisition of anaerobic microbes, establishment of rumen fermentation, expansion of rumen in volume, differentiation and growth of papillae, development of absorption and metabolic pathways, maturation of salivary apparatus and development of rumination behavior are all needed as the calf shifts from dependence on milk to solid feed. In nature and some production systems (e.g., most beef calves), young ruminants obtain nutrients from milk and fresh forages. In intensive dairying, calves are typically fed restricted amounts of milk and weaned onto starter feeds. Here we review the empirical work on the role of feeding and management during the transition from milk to solid feed in establishing the rumen ecosystem, rumen fermentation, rumen development, rumination behavior, and growth of dairy calves. In recent years, several studies have illustrated the benefits of feeding more milk and group rearing of dairy calves to take advantage of social facilitation (e.g., housing with peers or dam), and this review also examines the role of solid feed on rumen development and growth of calves fed large quantities of milk and reared under different housing situations. We conclude that the provision of high-starch and low-fiber starter feeds may negatively affect rumen development and that forage supplementation is beneficial for promoting development of the gut and rumination behavior in young calves. It is important to note that both the physical form of starter diets and their nutritional composition affect various aspects of development in calves. Further research is warranted to identify an optimal balance between physically effective fiber and readily degradable carbohydrates in starter diets to support development of a healthy gut and rumen, rumination behavior, and growth in young calves.

A rifampin-resistant Lactobacillus salivarius strain, CTC2197, was assessed as a probiotic in poultry, by studying its ability to prevent Salmonella enteritidis C-114 colonization in chickens. When the probiotic strain was dosed by oral gavage together with S. enteritidis C-114 directly into the proventriculus in 1-day-old Leghorn chickens, the pathogen was completely removed from the birds after 21 days. The same results were obtained when the probiotic strain was also administered through the feed and the drinking water apart from direct inoculation into the proventriculus. The inclusion of L. salivarius CTC2197 in the first day chicken feed revealed that a concentration of 10(5) CFU g(-1) was enough to ensure the colonization of the gastrointestinal tract of the birds after 1 week. However, between 21 and 28 days, L. salivarius CTC2197 was undetectable in the gastrointestinal tract of some birds, showing that more than one dose would be necessary to ensure its presence till the end of the rearing time. Freeze-drying and freezing with glycerol or skim milk as cryoprotective agents, appeared to be suitable methods to preserve the probiotic strain. The inclusion of the L. salivarius CTC2197 in a commercial feed mixture seemed to be a good way to supply it on the farm, although the strain showed sensitivity to the temperatures used during the feed mixture storage and in the chicken incubator rooms. Moreover, survival had been improved after several reinoculations in chicken feed mixture.

AbstractFood supply chains are increasingly associated with environmental and socio-economic impacts. An increasing global population, an evolution in consumers' needs, and changes in consumption models pose serious challenges to the overall sustainability of food production and consumption. Life cycle thinking (LCT) and assessment (LCA) are key elements in identifying more sustainable solutions for global food challenges. In defining solutions to major global challenges, it is fundamentally important to avoid burden shifting amongst supply chain stages and amongst typologies of impacts, and LCA should, therefore, be regarded as a reference method for the assessment of agri-food supply chains. Hence, this special volume has been prepared to present the role of life cycle thinking and life cycle assessment in: i) the identification of hotspots of impacts along food supply chains with a focus on major global challenges; ii) food supply chain optimisation (e.g. productivity increase, food loss reduction, etc.) that delivers sustainable solutions; and iii) assessment of future scenarios arising from both technological improvements and behavioural changes, and under different environmental conditions (e.g. climate change). This special volume consists of a collection of papers from a conference organized within the last Universal Exposition (EXPO2015) "LCA for Feeding the planet and energy for life" in Milan (Italy) in 2015 as well as other contributions that were submitted in the year after the conference that addressed the same key challenges presented at the conference. The papers in the special volume address some of the key challenges for optimizing food-related supply chains by using LCA as a reference method for environmental impact assessment. Beyond specific methodological improvements to better tailor LCA studies to food systems, there is a clear need for the LCA community to "think outside the box", exploring complementarity with other methods and domains. The concepts and the case studies presented in this special volume demonstrate how cross-fertilization among difference science domains (such as environmental, technological, social and economic ones) may be key elements of a sustainable "today and tomorrow" for feeding the planet.

ABSTRACT Beeswax, candelilla wax, carnauba wax, and a high‐melting fraction of anhydrous milkfat were homogenized with whey protein to produce edible emulsion films. Lipid type and amount were important in controlling the emulsion film water vapor permeability (WVP). The WVPs of the beeswax and milkfat emulsion films were significantly lower than that of films from lower moisture transmitters, carnauba and candelilla wax. Lipid WVP and degree of viscoelasticity determined the barrier properties of the films. A significant reduction in WVP of whey protein films could be achieved using large volume fractions of lipid depending on lipid type.