California Department of Food and Agriculture

The idea that noncrop habitat enhances pest control and represents a win-win opportunity to conserve biodiversity and bolster yields has emerged as an agroecological paradigm. However, while noncrop habitat in landscapes surrounding farms sometimes benefits pest predators, natural enemy responses remain heterogeneous across studies and effects on pests are inconclusive. The observed heterogeneity in species responses to noncrop habitat may be biological in origin or could result from variation in how habitat and biocontrol are measured. Here, we use a pest-control database encompassing 132 studies and 6,759 sites worldwide to model natural enemy and pest abundances, predation rates, and crop damage as a function of landscape composition. Our results showed that although landscape composition explained significant variation within studies, pest and enemy abundances, predation rates, crop damage, and yields each exhibited different responses across studies, sometimes increasing and sometimes decreasing in landscapes with more noncrop habitat but overall showing no consistent trend. Thus, models that used landscape-composition variables to predict pest-control dynamics demonstrated little potential to explain variation across studies, though prediction did improve when comparing studies with similar crop and landscape features. Overall, our work shows that surrounding noncrop habitat does not consistently improve pest management, meaning habitat conservation may bolster production in some systems and depress yields in others. Future efforts to develop tools that inform farmers when habitat conservation truly represents a win-win would benefit from increased understanding of how landscape effects are modulated by local farm management and the biology of pests and their enemies.

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Abstract Here we present a phylogeny of beetles ( I nsecta: C oleoptera) based on DNA sequence data from eight nuclear genes, including six single‐copy nuclear protein‐coding genes, for 367 species representing 172 of 183 extant families. Our results refine existing knowledge of relationships among major groups of beetles. Strepsiptera was confirmed as sister to C oleoptera and each of the suborders of C oleoptera was recovered as monophyletic. Interrelationships among the suborders, namely P olyphaga ( A dephaga ( A rchostemata, M yxophaga)), in our study differ from previous studies. Adephaga comprised two clades corresponding to H ydradephaga and G eadephaga. The series and superfamilies of P olyphaga were mostly monophyletic. The traditional C ucujoidea were recovered in three distantly related clades. Lymexyloidea was recovered within T enebrionoidea. Several of the series and superfamilies of P olyphaga received moderate to maximal clade support in most analyses, for example B uprestoidea, C hrysomeloidea, C occinelloidea, C ucujiformia, C urculionoidea, D ascilloidea, E lateroidea, H isteroidea and H ydrophiloidea. However, many of the relationships within P olyphaga lacked compatible resolution under maximum‐likelihood and B ayesian inference, and/or lacked consistently strong nodal support. Overall, we recovered slightly younger estimated divergence times than previous studies for most groups of beetles. The ordinal split between C oleoptera and S trepsiptera was estimated to have occurred in the E arly P ermian. Crown C oleoptera appeared in the Late P ermian, and only one or two lineages survived the end‐ P ermian mass extinction, with stem group representatives of all four suborders appearing by the end of the T riassic. The basal split in P olyphaga was estimated to have occurred in the T riassic, with the stem groups of most series and superfamilies originating during the T riassic or J urassic. Most extant families of beetles were estimated to have C retaceous origins. Overall, C oleoptera experienced an increase in diversification rate compared to the rest of N europteroidea. Furthermore, 10 family‐level clades, all in suborder P olyphaga, were identified as having experienced significant increases in diversification rate. These include most beetle species with phytophagous habits, but also several groups not typically or primarily associated with plants. Most of these groups originated in the C retaceous, which is also when a majority of the most species‐rich beetle families first appeared. An additional 12 clades showed evidence for significant decreases in diversification rate. These clades are species‐poor in the M odern fauna, but collectively exhibit diverse trophic habits. The apparent success of beetles, as measured by species numbers, may result from their associations with widespread and diverse substrates – especially plants, but also including fungi, wood and leaf litter – but what facilitated these associations in the first place or has allowed these associations to flourish likely varies within and between lineages. Our results provide a uniquely well‐resolved temporal and phylogenetic framework for studying patterns of innovation and diversification in C oleoptera, and a foundation for further sampling and resolution of the beetle tree of life.

BACKGROUND: Drosophila suzukii is an oriental species first reported outside Asia from Hawaii in 1980. The first confirmed records for the continental United States were made in 2008 in California. The identification of this pest is difficult because very few published resources exist. RESULTS: It has since been recorded in Oregon, Washington, British Columbia, Alberta, Manitoba, Ontario, Quebec, Utah, Michigan, Wisconsin, Louisiana, North Carolina, South Carolina and Florida. Males are relatively easy to identify by the black apical wing spots and the single row of combs on the first and second tarsal segment of the fore leg. The male genitalia are also very characteristic and will aid in identifying teneral specimens. Females can be identified by the large ovipositor, which is 6-7 times as long as the diameter of the spermatheca. Immature stages can only be identified by molecular techniques. CONCLUSION: Although this species has been recorded from many US states and Canadian provinces, it has not been established in all of these places, and the main economic damage is restricted to the western part of North America. With the characters laid out in this paper, it should be possible to identify the pest with high certainty.

The major pet food recall associated with acute renal failure in dogs and cats focused initially on melamine as the suspect toxicant. In the course of the investigation, cyanuric acid was identified in addition to melamine in the offending food. The purpose of this study was to characterize the toxicity potential of melamine, cyanuric acid, and a combination of melamine and cyanuric acid in cats. In this pilot study, melamine was added to the diet of 2 cats at 0.5% and 1%, respectively. Cyanuric acid was added to the diet of 1 cat at increasing doses of 0.2%, 0.5%, and 1% over the course of 10 days. Melamine and cyanuric acid were administered together at 0%, 0.2%, 0.5%, and 1% to 1 cat per dose group. No effect on renal function was observed in cats fed with melamine or cyanuric acid alone. Cats dosed with a combination were euthanized at 48 hours after dosing because of acute renal failure. Urine and touch impressions of kidneys from all cats dosed with the combination revealed the presence of fan-shaped, birefringent crystals. Histopathologic findings were limited to the kidneys and included crystals primarily within tubules of the distal nephron, severe renal interstitial edema, and hemorrhage at the corticomedullary junction. The kidneys contained estimated melamine concentrations of 496 to 734 mg/kg wet weight and estimated cyanuric acid concentrations of 487 to 690 mg/kg wet weight. The results demonstrate that the combination of melamine and cyanuric acid is responsible for acute renal failure in cats.

Inventory of the caterpillars, their food plants and parasitoids began in 1978 for today's Area de Conservacion Guanacaste (ACG), in northwestern Costa Rica. This complex mosaic of 120 000 ha of conserved and regenerating dry, cloud and rain forest over 0-2000 m elevation contains at least 10 000 species of non-leaf-mining caterpillars used by more than 5000 species of parasitoids. Several hundred thousand specimens of ACG-reared adult Lepidoptera and parasitoids have been intensively and extensively studied morphologically by many taxonomists, including most of the co-authors. DNA barcoding - the use of a standardized short mitochondrial DNA sequence to identify specimens and flush out undisclosed species - was added to the taxonomic identification process in 2003. Barcoding has been found to be extremely accurate during the identification of about 100 000 specimens of about 3500 morphologically defined species of adult moths, butterflies, tachinid flies, and parasitoid wasps. Less than 1% of the species have such similar barcodes that a molecularly based taxonomic identification is impossible. No specimen with a full barcode was misidentified when its barcode was compared with the barcode library. Also as expected from early trials, barcoding a series from all morphologically defined species, and correlating the morphological, ecological and barcode traits, has revealed many hundreds of overlooked presumptive species. Many but not all of these cryptic species can now be distinguished by subtle morphological and/or ecological traits previously ascribed to 'variation' or thought to be insignificant for species-level recognition. Adding DNA barcoding to the inventory has substantially improved the quality and depth of the inventory, and greatly multiplied the number of situations requiring further taxonomic work for resolution.

BACKGROUND: In the mega-diverse insect order Lepidoptera (butterflies and moths; 165,000 described species), deeper relationships are little understood within the clade Ditrysia, to which 98% of the species belong. To begin addressing this problem, we tested the ability of five protein-coding nuclear genes (6.7 kb total), and character subsets therein, to resolve relationships among 123 species representing 27 (of 33) superfamilies and 55 (of 100) families of Ditrysia under maximum likelihood analysis. RESULTS: Our trees show broad concordance with previous morphological hypotheses of ditrysian phylogeny, although most relationships among superfamilies are weakly supported. There are also notable surprises, such as a consistently closer relationship of Pyraloidea than of butterflies to most Macrolepidoptera. Monophyly is significantly rejected by one or more character sets for the putative clades Macrolepidoptera as currently defined (P < 0.05) and Macrolepidoptera excluding Noctuoidea and Bombycoidea sensu lato (P < or = 0.005), and nearly so for the superfamily Drepanoidea as currently defined (P < 0.08). Superfamilies are typically recovered or nearly so, but usually without strong support. Relationships within superfamilies and families, however, are often robustly resolved. We provide some of the first strong molecular evidence on deeper splits within Pyraloidea, Tortricoidea, Geometroidea, Noctuoidea and others.Separate analyses of mostly synonymous versus non-synonymous character sets revealed notable differences (though not strong conflict), including a marked influence of compositional heterogeneity on apparent signal in the third codon position (nt3). As available model partitioning methods cannot correct for this variation, we assessed overall phylogeny resolution through separate examination of trees from each character set. Exploration of "tree space" with GARLI, using grid computing, showed that hundreds of searches are typically needed to find the best-feasible phylogeny estimate for these data. CONCLUSION: Our results (a) corroborate the broad outlines of the current working phylogenetic hypothesis for Ditrysia, (b) demonstrate that some prominent features of that hypothesis, including the position of the butterflies, need revision, and (c) resolve the majority of family and subfamily relationships within superfamilies as thus far sampled. Much further gene and taxon sampling will be needed, however, to strongly resolve individual deeper nodes.

Americanae nace como un proyecto conjunto que surge dentro de la Red Europea de Información y Documentación sobre América Latina (REDIAL), y que ha afrontado la Biblioteca de la Agencia Española de Cooperación Internacional para el Desarrollo (AECID). Esta nueva biblioteca virtual hace más accesibles los libros digitales de tema americanista a los investigadores y usuarios interesados de cualquier parte del mundo.

Novel species of fungi described in the present study include the following from Malaysia: Castanediella eucalypti from Eucalyptus pellita, Codinaea acacia from Acacia mangium, Emarcea eucalyptigena from Eucalyptus brassiana, Myrtapenidiella eucalyptorum from Eucalyptus pellita, Pilidiella eucalyptigena from Eucalyptus brassiana and Strelitziana malaysiana from Acacia mangium. Furthermore, Stachybotrys sansevieriicola is described from Sansevieria ehrenbergii (Tanzania), Phacidium grevilleae from Grevillea robusta (Uganda), Graphium jumulu from Adansonia gregorii and Ophiostoma eucalyptigena from Eucalyptus marginata (Australia), Pleurophoma ossicola from bone and Plectosphaerella populi from Populus nigra (Germany), Colletotrichum neosansevieriae from Sansevieria trifasciata, Elsinoë othonnae from Othonna quinquedentata and Zeloasperisporium cliviae (Zeloasperisporiaceae fam. nov.) from Clivia sp. (South Africa), Neodevriesia pakbiae, Phaeophleospora hymenocallidis and Phaeophleospora hymenocallidicola on leaves of a fern (Thailand), Melanconium elaeidicola from Elaeis guineensis (Indonesia), Hormonema viticola from Vitis vinifera (Canary Islands), Chlorophyllum pseudoglobossum from a grassland (India), Triadelphia disseminata from an immunocompromised patient (Saudi Arabia), Colletotrichum abscissum from Citrus (Brazil), Polyschema sclerotigenum and Phialemonium limoniforme from human patients (USA), Cadophora vitícola from Vitis vinifera (Spain), Entoloma flavovelutinum and Bolbitius aurantiorugosus from soil (Vietnam), Rhizopogon granuloflavus from soil (Cape Verde Islands), Tulasnella eremophila from Euphorbia officinarum subsp. echinus (Morocco), Verrucostoma martinicensis from Danaea elliptica (French West Indies), Metschnikowia colchici from Colchicum autumnale (Bulgaria), Thelebolus microcarpus from soil (Argentina) and Ceratocystis adelpha from Theobroma cacao (Ecuador). Myrmecridium iridis (Myrmecridiales ord. nov., Myrmecridiaceae fam. nov.) is also described from Iris sp. (The Netherlands). Novel genera include (Ascomycetes): Budhanggurabania from Cynodon dactylon (Australia), Soloacrosporiella, Xenocamarosporium, Neostrelitziana and Castanediella from Acacia mangium and Sabahriopsis from Eucalyptus brassiana (Malaysia), Readerielliopsis from basidiomata of Fuscoporia wahlbergii (French Guyana), Neoplatysporoides from Aloe ferox (Tanzania), Wojnowiciella, Chrysofolia and Neoeriomycopsis from Eucalyptus (Colombia), Neophaeomoniella from Eucalyptus globulus (USA), Pseudophaeomoniella from Olea europaea (Italy), Paraphaeomoniella from Encephalartos altensteinii, Aequabiliella, Celerioriella and Minutiella from Prunus (South Africa). Tephrocybella (Basidiomycetes) represents a novel genus from wood (Italy). Morphological and culture characteristics along with ITS DNA barcodes are provided for all taxa.

1. The activities of lysosomal enzymes in the cortexes and medullas and the principal subcellular fractions of rat kidney were measured. 2. A method is described for the isolation of rat-kidney lysosomes and a detailed analysis of the enzymic composition of the lysosomes is reported. Enzyme analysis of the other principal subcellular fractions is included for comparison. 3. Studies of the distribution of alpha-glucosidase showed that the lysosomal fraction contained only 10% of the total enzyme activity. The microsomal fraction contained most of the particulate alpha-glucosidase. Lysozyme was concentrated mainly in the lysosomal fraction with only small amounts present in the microsomal fraction. Lysosomal alpha-glucosidase had optimum pH5 whereas the microsomal form had optimum pH6. Both lysosomal and microsomal lysozyme had optimum pH6.2. 4. The stability of lysosomal suspensions was studied. Incubation at 37 degrees and pH7 resulted in first an increased availability of enzymes without parallel release of enzyme. This was followed by a second stage during which the availability of enzymes was closely related to the release of enzymes. These changes were closely paralleled by changes in light-scattering properties of lysosomes. 5. The latent nature of the alpha-glucosidase and lysozyme of intact kidney lysosomes was demonstrated by their graded and parallel release with other typical lysosomal enzymes. 6. Isolated lysosomes were unstable at pH values lower than 5, most stable at pH6-7 and less stable at pH 8-9. Lysosomes were not disrupted when the osmolarity of the suspending medium was decreased from 0.6m to 0.25m. 7. The discussion compares the properties and composition of kidney lysosomes, liver lysosomes and the granules of macrophages. 8. The possible origin of the lysozyme in kidney lysosomes by reabsorption of the lysozyme in blood is discussed.

Parasite-specific antibody responses were detected using an indirect fluorescent antibody (IFA) test in cattle that were naturally or experimentally infected with Neospora parasites. The test was developed using Neospora tachyzoites isolated from an aborted bovine fetus and grown in bovine cell cultures (isolate BPA1). In all cases, infections were confirmed by the identification of Neospora tachyzoites and/or bradyzoite cysts in fetal or calf tissues using an immunoperoxidase test procedure. Fifty-five naturally infected cows that aborted Neospora-infected fetuses had titers of 320-5,120 at the time of abortion. The titer of 6 cows that were serologically monitored over a prolonged period decreased to 160–640 within 150 days after they aborted infected fetuses. Two of the cows showed an increase in their Neospora titers during their subsequent pregnancy, and they gave birth to congenitally infected calves that had precolostral titers of 10,240-20,480. Postcolostral titers of these calves and of 4 other calves with congenital Neospora infections were all 25,120, whereas calves with no detectable parasites had titers ≤ 160. Two pregnant heifers that were experimentally infected with the BPA1 isolate at approximately 120 days gestation seroconverted to Neospora antigens within 9 days and developed peak titers of 5,120 and 20,480 within 32 days of infection. The fetus taken by caesarian section 32 days postinfection from 1 heifer and the full-term calf born to the other had Neospora titers of 640 and 10,240, respectively. Nine cows that aborted uninfected fetuses and 61 adult cattle maintained under pasture or feedlot conditions, where risk of exposure to Neospora was considered to be low, had titers ≤ 320. Some of the feedlot cattle tested had serologic reactivity that was restricted to antigens at the apical end of both Neospora and Toxoplasma gondii tachyzoites. This type of reactivity, which may result from serologic cross-reactivity between conserved apical complex antigens of closely related sporozoan parasites, differed from the whole parasite fluorescence that was observed with sera from Neospora-infected animals. The significance of these results and the potential application of the IFA test for the diagnosis of Neospora infections in cattle are discussed.

In 2007, it was determined that melamine, ammeline, ammelide, and cyanuric acid (abbreviated as MARC for melamine and related contaminants) had been added to wheat gluten and rice protein that were subsequently incorporated into pet food. The consumption of food tainted by MARC compounds was implicated in numerous instances of renal failure in cats and dogs. A method for the analysis of MARC compounds in kidney tissue using high-performance liquid chromatography/tandem mass spectrometry (HPLC/MS/MS) has been developed. MARC analytes were extracted by homogenization of kidney tissue in 50/40/10 acetonitrile/water/diethylamine. The homogenate was centrifuged, and an aliquot of supernatant was diluted with acetonitrile, concentrated, and fortified with a stable isotope-labeled analogue of melamine. Analytes were detected using atmospheric pressure chemical ionization and multiple reaction monitoring. Quantitation of positive samples was performed using the internal standard method and five-point calibration curves ranging between 50 and 1000 ng/mL of each analyte. The method was validated by analysis of replicate kidney tissue samples fortified with the individual analytes and by analysis of kidney samples fortified with melamine cyanurate powder at two different concentrations. This method was successfully used for routine postmortem diagnosis of melamine toxicosis in animals. Melamine was also detected by this method in paraffin-embedded tissue from animals suspected to have died of melamine toxicosis.

Scientific communication is facilitated by a data-driven, scientifically sound taxonomy that considers the end-user’s needs and established successful practice. In 2013, the Fusarium community voiced near unanimous support for a concept of Fusarium that represented a clade comprising all agriculturally and clinically important Fusarium species, including the F. solani species complex (FSSC). Subsequently, this concept was challenged in 2015 by one research group who proposed dividing the genus Fusarium into seven genera, including the FSSC described as members of the genus Neocosmospora, with subsequent justification in 2018 based on claims that the 2013 concept of Fusarium is polyphyletic. Here, we test this claim and provide a phylogeny based on exonic nucleotide sequences of 19 orthologous protein-coding genes that strongly support the monophyly of Fusarium including the FSSC. We reassert the practical and scientific argument in support of a genus Fusarium that includes the FSSC and several other basal lineages, consistent with the longstanding use of this name among plant pathologists, medical mycologists, quarantine officials, regulatory agencies, students, and researchers with a stake in its taxonomy. In recognition of this monophyly, 40 species described as genus Neocosmospora were recombined in genus Fusarium, and nine others were renamed Fusarium. Here the global Fusarium community voices strong support for the inclusion of the FSSC in Fusarium, as it remains the best scientific, nomenclatural, and practical taxonomic option available.

Phylogenetics is the science of reconstructing the evolutionary history of life on Earth. Traditionally, phylogenies were constructed using morphological data only, but the introduction of Sanger sequencing and PCR in the late 1970s enabled genetic information to be incorporated into phylogenetic analyses. Early phylogenetic studies employing multilocus analyses contributed greatly to our knowledge of phylogenetic history and challenged some well-established views of the relationships among many groups of plants and animals. Since the publication of these pioneering studies, significant methodological advances in both sequencing and analytical techniques have been made, and molecular phylogenies are now broadly accepted to represent robust hypotheses of organismal relationships. Next-generation sequencing techniques, developed in the mid-2000s, revolutionized DNA sequencing and led to a dramatic reduction in sequencing cost per nucleotide and a sharp increase in data generation speed. As a result, the generation of unprecedented amounts of sequence data for both model and nonmodel organisms has become affordable. This development has transformed the field of molecular phylogenetics into phylogenomics—where genome-scale data are obtained from multiple samples at once at a much reduced cost (Mardis, 2011). The phylogenomic pipeline can be very complex, presenting an overwhelming array of methodologies available for the acquisition, manipulation, analysis and interpretation of massive datasets. Researchers also have to overcome the challenges of sequencing strategy design, identification of orthologous loci, model selection and phylogeny estimation. This can be particularly daunting for researchers new to the field—both students and established scientists—who wish to delve into novel methods and data to reconstruct the evolution of their study group. Here we present an entry-level overview of the theory and tools that are central to phylogenomics, with an emphasis on the appropriate application of techniques useful for phylogenetic analysis of genomic data. We focus on the sequencing technologies and statistical methods for phylogeny estimation, and the software implementing these methods and their application to large molecular datasets. We also discuss the tools and tradeoffs for improving the accuracy of phylogenomic analyses, including the biological and methodological sources of systematic error in phylogeny estimation. Finally, we provide a glossary of commonly encountered terms used in phylogenomics that may be useful for those entering the field and hoping to sort through the multitude of methods, analytical tools and terminology inherent to this relatively new, but rapidly advancing field. The word 'phylogenomics' was first introduced in the context of prediction of gene function for genome-scale data (Eisen, 1998), and soon after in the context of phylogenetic inference (O'Brien & Stanyon, 1999). The discipline of phylogenomics owes its existence to the advances made in DNA sequencing technology over the past two decades (Metzker, 2010). It comprises several areas of research at the interface between molecular and evolutionary biology and has two major goals: (i) to infer phylogenetic relationships between taxa and gain insights into the mechanisms of molecular evolution; and (ii) to use multispecies phylogenetic comparisons to infer putative functions for DNA or protein sequences. Traditional Sanger sequencing studies include relatively few loci and are therefore limited by stochastic or sampling error. As there is a relatively small number of phylogenetically informative characters available in one or a few genes, this random 'noise' influences the inference of backbone nodes, potentially leading to poorly resolved or poorly supported phylogenetic trees. This problem can be addressed successfully by using much larger amounts of sequence data. Modern phylogenomic analyses, which take advantage of hundreds to thousands of loci from across the genome, are, on average, orders of magnitude larger than traditional Sanger sequencing datasets. The size of these datasets therefore significantly reduces the impact of stochastic error and data availability as a limiting factor, offering great promise for resolving historically recalcitrant nodes in the tree of life. High-throughput sequencing technologies [also called next-generation sequencing (NGS)] (Fig. 1) have yielded genome-scale data in immense quantities. Next-generation sequencing technologies differ fundamentally from the Sanger method in that they allow for massively parallel DNA sequencing, providing extremely high throughput from multiple samples simultaneously and at a much reduced cost (Mardis, 2011). Millions to billions of DNA nucleotides can be sequenced in parallel, yielding orders of magnitude more data and minimizing the need for the fragment-cloning methods that are used with Sanger sequencing (Fig. 1). Recent progress in NGS technology and the rapid development of bioinformatics tools now allow research groups of any size to generate large amounts of genomic sequences for organisms of interest. High-throughput sequencing can be used for whole-genome sequencing (Lam, 2012), whole-transcriptome shotgun sequencing (also called RNA sequencing, RNA-seq, or transcriptomics; Wang, 2009), whole-exome sequencing (Rabbani, 2014), and reduced-representation genome sequencing (also called target enrichment) (e.g., Faircloth, 2012; Lemmon, 2012). Table 1 summarizes the most commonly used sequencing technologies in phylogenomics. For more details on these different technologies see the Beginner's Handbook of Next Generation Sequencing by Genohub (https://genohub.com/next-generation-sequencing-handbook/) (see also Ambardar, 2016; Besser et al., 2018, and references therein). Choosing the appropriate sequencing technology for a phylogenomic study has important effects on downstream workflows, especially in terms of read length, as library preparation in some phylogenomic techniques (e.g. ultraconserved elements and anchored hybrid enrichment, discussed later) requires a read size selection step. Strict experimental reproducibility is an integral—albeit uncommon—aspect of biological sciences, mainly due to varied technical challenges with implementation and curation of experimental methods and procedures. Despite the importance of phylogenetic analyses to most fields of biology, the reproducibility of phylogenetic experiments can be very low, with an estimated 60% of published phylogenetic analyses being 'lost to science' due to the unavailability of the underlying data and methods (Magee, 2014). Published phylogenetic studies can be difficult or impossible to replicate or expand upon, as the utilized analytical software, software versions, software parameters, dependencies and operating system versions can be very challenging to uncover or recreate. The promotion of open science and reproducible research can create a more productive and responsible scientific culture in phylogenomics, enabling researchers to build upon previous studies and continuously address larger and more complex questions. This philosophy encompasses the sharing of data and code used to produce the analysis, as well as open archiving of all raw data (Mork, 2015; Shade & Teal, 2015). Data provenance, the recording of the input and transformation of information used to generate a result, is a key issue in reproducibility. Several recommendations and guidelines to promote the best practices in reproducibility and data management in phylogenomics and bioinformatics have been proposed (Cranston et al., 2014; Magee, 2014; Debiasse & Ryan, 2019), and many tools for ensuring provenance and curation of both data and methods have been developed (e.g. Dunn, 2013; Oakley, 2014; Szitenberg, 2015). To ensure the best practices in phylogenomics and bioinformatics, it is vital that reproducibility checkpoints are enforced—places in a workflow devoted to scrutinizing its integrity, so results are validated across multiple iterations to ensure consistency of results. Additionally, adopting an iterative, branching workflow to systematically explore the methodological space is crucial. Linear methodology, with experimental and computational procedures lined up one after the other, as presented in most published studies, is rarely the reality of phylogenetic analysis. Instead, estimating phylogenetic trees is more often than not a messy enterprise, and a systematic exploration of the methods and data is recommended in order to select the best tools and pipelines to answer the question at hand. Finally, for good provenance of experimental procedures and computational tools utilized in a particular study, it is highly recommended that comprehensive notes are kept throughout the process. In particular, keeping a 'readme' file at every step can be extremely helpful in keeping track of the software versions used, parameter values utilized, goals of each step and how they relate to the software utilized, as well as indication of data format changes. All these can greatly contribute to standardization and ease of downstream efforts (Shade & Teal, 2015). Phylogenomic data are a precious scientific resource: molecular sequence alignments and phylogenies are expensive to generate, difficult to replicate and have seemingly infinite potential for synthesis and reuse. For most phylogenomic analyses, phylogeneticists are faced with a large combination of algorithms, models and data manipulation techniques. To address this issue, here we present a flowchart containing the major steps and tools utilized in phylogenomics (Fig. 2). The flowchart is not meant to be exhaustive, but merely a visualization of the commonly utilized methodologies and pipelines in recent phylogenomic studies. Taxon sampling is of extreme importance for phylogenetic inference, and increased sampling of taxa—coupled with increased sampling of loci—is commonly advocated as a solution to resolving recalcitrant nodes of the tree of life. Ideally, sampling of both taxa and sequences should be increased at the same pace, but the advances in high-throughput sequencing have caused increases in gene sampling to far outpace taxon sampling. As greater amounts of data are incorporated into phylogenetic studies, new evidence and hypotheses regarding relationships among taxa can emerge, and placement of lineages within clades can change dramatically. Taxon sampling can thus greatly influence hypotheses supported by phylogenetic inference (Rosenberg & Kumar, 2003; Nabhan & Sarkar, 2012). Taxon selection meant to address a specific research question should take place early in a phylogenomic study. 'Sufficient' taxon sampling is always dependent on the questions being addressed. Ideally, in order to unravel the phylogeny of an entire taxonomic unit, most, if not all, subordinate taxa in that unit should be sampled. Even though increasing the number of taxa results in a more complex computational problem for phylogenetic analysis, it has been demonstrated that denser taxon sampling improves phylogenetic accuracy (Heath et al., 2008). Taxon sampling, however, can be greatly limited by the phylogenomic method of choice. Transcriptomics, for instance, requires specimens collected and stored directly into liquid nitrogen or RNAlater, whereas other sequencing methods, including target enrichment, and shotgun and exome sequencing, will require molecular-grade specimens, preferably preserved in high-grade ethyl ethanol and stored in a laboratory ultrafreezer. A notable exception to this is target enrichment of ultraconserved elements (UCEs), a method that can successfully generate phylogenomic data from old, pinned insect museum specimens (Blaimer, 2016). Genome-scale projects may be particularly vulnerable to systematic error caused by nonproportional phylogenetic sampling. As dataset size increases, so does the accumulation of nonrandom systematic error and accompanying nonphylogenetic signal (Jeffroy, 2006). Bayesian analyses of macroevolutionary patterns—including divergence-time estimation, ancestral state reconstruction, and diversification rate estimation—assume proportional sampling of lineages within a clade, and deviations from it may potentially lead to biases (Stadler, 2009). However, some implementations enable 'corrections' for uneven taxon sampling (e.g. revbayes implements corrections for birth-death and various diversification rate models, except for fossilized birth-death). Before sequencing new specimens, it is also worth evaluating previously sequenced resources. The National Center for Biotechnology Information's Sequence Read Archive (NCBI SRA) contains user-uploaded raw sequence data and alignment information from high-throughput sequencing projects (Leinonen, 2011). Other resources include FlyBase (Thurmond, 2019), a large database of Drosophila genes and genomes, WormBase (https://www.wormbase.org), containing genomic data of Caenorhabditis elegans and related nematodes, and the UCSC Genome Browser (Kent et al., 2002), a large repository of mostly vertebrate genomes. Utilizing sequences from these databases can save money and/or increase taxon sampling in ongoing phylogenomic projects. For a comprehensive overview of insect DNA methods, see Moreau (2014), which offers a detailed description of DNA extraction methods using either commercial kits or phenol/chloroform protocols. After DNA extraction, specimens should be deposited in publicly accessible collections in association with their unique identifier, and publications utilizing these data should always include unique identifier, repository and specimen metadata (including specimen collector, date and method of collection, and geographic origin). Vouchering specimens with unique identifiers (alphanumeric database number) is crucial for all phylogenetic projects. Therefore, nondestructive or partially destructive DNA extraction methods should be used whenever possible, and in these cases, the extracted specimen itself becomes the voucher. By contrast, when nondestructive DNA extraction is not possible, such as in transcriptomic projects or small-bodied organisms, a photographic voucher can be associated with the sequence data. Moreover, when the destroyed specimen is part of a sample of conspecifics (e.g. in communal or social insects), another specimen from the same sample can serve as a voucher, provided it is made clear that it is not the extracted specimen. Properly vouchering specimens used for DNA extraction greatly increases reproducibility by alleviating issues related to sample identity and unstable taxonomy (Pleijel et al., 2008; Turney, 2015). Although large phylogenomic datasets have become increasingly more accessible and cost-efficient in recent years, it is now widely accepted that simply increasing the amount of sequence data will not unambiguously resolve some of the most difficult nodes in the tree of life, mainly due to systematic error from nonphylogenetic signal or model inadequacy. Appropriate locus selection is therefore crucial in phylogenomics, but knowledge of the best molecular markers for resolving difficult branches at various evolutionary depths is still incipient. Questions still remain about whether to use coding or noncoding sequence data, conserved or highly variable loci, and long or short alignments (Betancur-R. et al., 2014; Edwards et al., 2016; Chen et al., 2017). Therefore, one of the most critical decisions in a phylogenomic project is the sequencing method to be utilized, a decision that must be made a priori as each method will result in different types of genomic data sequenced. Different methods have their own characteristics, advantages, and limitations, including cost-effectiveness, ease of use, sample quality required, and downstream data filtering and analysis workflow. Phylogenomic sequencing methods (Table 2) can be broadly subdivided into shotgun sequencing and target enrichment sequencing. Shotgun sequencing is the process of sequencing from the entire fragmented genome at random, returning part or all of the genome depending on the sequencing depth achieved, whereas target enrichment uses bidirectional probes (analogous to primers in Sanger sequencing) to recover only genomic regions of interest. Popular methods of shotgun sequencing include genome skimming, whole-genome shotgun sequencing and transcriptome sequencing (i.e. RNA-seq). Popular methods of target enrichment for phylogenetics include anchored hybrid enrichment (AHE) (Lemmon, 2012) and UCEs (McCormack et al., 2012; Faircloth et al., 2012) [see also Mandel (2014) for an alternative method developed for plants in the family Compositae]. These techniques are reviewed briefly in Table 2 and have been covered in more detail elsewhere (e.g. Lemmon & Lemmon, 2013; McCormack, 2013; Wen et al., 2015; Zhang et al., 2019). Shotgun sequencing (Fig. 3) involves fragmenting template DNA into short pieces, which are then randomly sequenced to obtain reads. Next, various methods and software are used to overlap different reads and assemble them into a longer DNA sequence called a contig. RNA-seq can be considered a special form of shotgun sequencing, where whole mRNA is first extracted and reverse-transcribed into reverse-complement DNA, which is then sequenced. Sequencing depth, or the average number of times an individual base in the genome is sequenced, is a key concept in shotgun sequencing. Because the genome is sequenced at random, multiple-copy regions of the genome (i.e. mitochondrial, ribosomal, and plastid DNA) are sequenced more frequently than single-copy regions. Therefore, when a genome is sequenced at a relatively shallow depth, only fragments from multiple-copy regions of the genome are sequenced in sufficient quantities to be successfully recovered. Shallow-depth whole-genome shotgun sequencing is also called genome skimming (Straub et al., 2012), a time and cost-efficient method of sequencing mostly mitochondrial, ribosomal and plastid DNA. Conversely, when near-complete genomes are desired from whole-genome shotgun sequencing, a much greater sequencing depth is required in order to sequence sufficient numbers of fragments from single-copy regions of the genome. By contrast, in RNA-seq (or transcriptomics) the extracted mRNA is used as a template to generate reverse-complement DNA. This reverse-complement DNA is then sequenced, resulting in data generated from only the genomic regions undergoing active transcription at the time of tissue preservation. This method is therefore not only a genome-reduction strategy, but also facilitates the comparison of transcription activity between individual tissues, life stages, rearing conditions, etc. One of the major drawbacks of transcriptomics is the high tissue quality required—specimens must be flash-frozen in liquid nitrogen or collected directly into RNAlater, thus precluding the utilization of specimens already available in tissue collections or museums. Targeted sequence capture, or target enrichment (Fig. 4), is an umbrella term for multiple efficient, cost-effective methods for generating phylogenomic datasets for nonmodel organisms. These methods effectively reduce genomic DNA complexity through the use of short (60–120 bp), single-stranded nucleotide baits or probes that hybridize with template sequences, thus enabling the of particular sequences of with high As a result, mostly genomic regions of are DNA (including and can be present in the resulting of reads. samples can be and sequenced which the generation of DNA sequence data for hundreds of loci from over samples are two methods of target enrichment commonly used in et al., 2012) and UCEs (McCormack et al., 2012). methods are reduced-representation that on the utilization of a phylogenetically informative of the study genomes. Other target enrichment methods have been with varied locus selection including et al., 2014), loci et al., and 2019), but and UCEs have thus far been the most commonly used methods in phylogenomic studies of animals. In these methods, probes hybridize with sequences, which are the genomic regions by the probes and their regions are sequenced, such that both conserved and more variable thus more phylogenetically regions are sequenced at These reduced-representation methods enable researchers not only to the same of loci across all taxa of but also to or phylogenetically regions of the genome, including and A great of using the same of markers across studies is that it for as more phylogenomic data over By contrast, for RNA-seq datasets to be the must be from the same tissue and it can be challenging to and different analysis. hybrid enrichment sequencing mainly regions of the genome, that loci mostly and in some or other genomic elements (e.g. This that phylogenetic data can be more and both as nucleotides and as target loci are using genomes, or raw genomic reads from two or more sequences for the target loci for each to be in the are and an alignment for each locus is are developed on these for which a amount of quality is such that target loci are and have the appropriate amount of sequence to ensure both phylogenetic accuracy and enrichment (Lemmon, 2012). kits target loci on average, and genes (also called traditional or genes, are often in the target of loci, which facilitates with previous Sanger sequencing phylogenetic studies. in are highly conserved regions of the genome among As genetic markers for particular taxa of data are useful for reconstructing the evolutionary history and relationships of many organisms. In UCEs are by several genomes to each other, with and filtering of areas of very high sequence across all taxa of interest. are a number of different of UCEs for use as genetic markers and baits to target but the most commonly used in phylogenetics was in detail in Faircloth elements have been to well when data from museum samples et al., 2016; et al., which can greatly of taxon sampling as sequencing is longer to Despite specimens and loci in it is still to hundreds of markers from relatively specimens (McCormack et al., 2016). A major advantage of UCEs over data is an and software pipeline developed for the and analysis of data 2016). contains several software and that are extremely helpful and especially to These however, require some with in the of a or For a comprehensive and to on the see the For a comprehensive and informative overview of the theory and of UCEs for phylogenomics, see Zhang et on raw reads obtained from high-throughput sequencing is a step. should be for sequence of sequences and read (i.e. base and small Several have been proposed for quality of NGS data, including for reads and for reads from all other et al., 2010). These two resources a to and potential after which a software, such as et al., 2014), can be used to and offers an solution to sequence quality being the most commonly used in phylogenomics, other have been for instance, is used only for read but is especially useful for with sequences obtained from 2011). is a Bayesian for from the of sequences, where read quality is often is a used only for read quality & 2011). Finally, of a comprehensive quality pipeline that from application to application The software can simultaneously both and reads and process quality has been on raw the step is the of Sequence to and small DNA fragments obtained from a high-throughput sequencing in order to reconstruct longer DNA sequences. Sequence is whole genomes be sequenced in one but small of DNA of in are sequenced at a depending on the technology These short DNA are called and these reads are then into longer DNA sequences called are two techniques of genomic and methods of and resolving to and genome is [see for a introduction to In the of methods, a previously genome is used as a to which sequenced reads are every read is at its most and in to between reads is often the method in phylogenomics, as it does not require a genome. A multitude of methods for have been and the field is of different methods greatly on data and to be and each method has its own of including and computational complexity The most commonly used for in phylogenomics include & et al., 2012) and et al., 2017). For a comprehensive comparison of see & and & Finally, the software et al., manipulation of and transcriptomic data. The implements most of the in a pipeline that and of In the of of which can greatly reduce the computational of the problem with as well as introduced As is a major step in any phylogenomic analysis, at this and can lead to in downstream workflows, including and alignment and These increase the amount of data in the limiting the amount of useful data. For the best high long read and good read quality are all However, sequencing technologies not all for instance, sequencing results in but short length, sequencing very long but with quality (see Table 1). relationships should always be estimated on sequences that are related by as a result of than a from the inference of a tree on gene the gene tree the relationships among may differ from the has become a central problem for evolutionary and molecular In phylogenetic inference, it is a priori that the loci used to

OBJECTIVE: To estimate the minimum rate of abortion attributable to infection with Neospora sp in selected California dairy herds. DESIGN: Prospective study. ANIMALS: Twenty-six dairy herds containing 19,708 cows were studied. Fourteen herds had a history of abortions attributable to neosporosis, and 12 were herds in which neosporosis had not been identified as a cause of abortions. PROCEDURE: During a 1-year period, all available aborted fetuses were submitted to veterinary diagnostic laboratories to determine the cause of abortion. Reproductive records of cows that aborted were reviewed. RESULTS: Neospora sp infection was the major cause of abortion identified (113/266 abortions, 42.5%). The majority (232/266, 87.2%) of the aborted fetuses were submitted from herds with a history of abortions attributable to neosporosis, and Neospora sp infection was identified as the causative agent in 101 of 232 (43.5%) of the abortions from these herds. Fewer aborted fetuses were submitted from the 12 herds that did not have a history of abortion attributable to Neospora sp; however, neosporosis was confirmed as a cause of abortion in 6 of these 12 herds and was identified as the causative agent in 12 of 34 (35.3%) abortions from these herds. The disease was widespread throughout the state (19/26 herds in our study). Available reproductive histories of cows that had abortions attributed to neosporosis were evaluated, and 4 cows were identified that twice aborted Neospora-infected fetuses. CLINICAL IMPLICATIONS: Abortion attributable to Neospora sp infections can be expected to be a continuing major cause of abortion in dairy herds with a history of neosporosis as well as in dairy herds that have a history of sporadic abortions, but for which Neospora sp infections have not been previously identified as a cause of abortion. Subsequent pregnancies in cows that abort a Neospora sp-infected fetus also are at risk of infection, suggesting that the immunity provided by an initial infection is inadequate to prevent repeat infection or that cows can be persistently infected with Neospora sp.

Novel species of fungi described in the present study include the following from South Africa: Alanphillipsia aloeicola from Aloe sp., Arxiella dolichandrae from Dolichandra unguiscati, Ganoderma austroafricanum from Jacaranda mimosifolia, Phacidiella podocarpi and Phaeosphaeria podocarpi from Podocarpus latifolius, Phyllosticta mimusopisicola from Mimusops zeyheri and Sphaerulina pelargonii from Pelargonium sp. Furthermore, Barssia maroccana is described from Cedrus atlantica (Morocco), Codinaea pini from Pinus patula (Uganda), Crucellisporiopsis marquesiae from Marquesia acuminata (Zambia), Dinemasporium ipomoeae from Ipomoea pes-caprae (Vietnam), Diaporthe phragmitis from Phragmites australis (China), Marasmius vladimirii from leaf litter (India), Melanconium hedericola from Hedera helix (Spain), Pluteus albotomentosus and Pluteus extremiorientalis from a mixed forest (Russia), Rachicladosporium eucalypti from Eucalyptus globulus (Ethiopia), Sistotrema epiphyllum from dead leaves of Fagus sylvatica in a forest (The Netherlands), Stagonospora chrysopyla from Scirpus microcarpus (USA) and Trichomerium dioscoreae from Dioscorea sp. (Japan). Novel species from Australia include: Corynespora endiandrae from Endiandra introrsa, Gonatophragmium triuniae from Triunia youngiana, Penicillium coccotrypicola from Archontophoenix cunninghamiana and Phytophthora moyootj from soil. Novelties from Iran include Neocamarosporium chichastianum from soil and Seimatosporium pistaciae from Pistacia vera. Xenosonderhenia eucalypti and Zasmidium eucalyptigenum are newly described from Eucalyptus urophylla in Indonesia. Diaporthe acaciarum and Roussoella acacia are newly described from Acacia tortilis in Tanzania. New species from Italy include Comoclathris spartii from Spartium junceum and Phoma tamaricicola from Tamarix gallica. Novel genera include (Ascomycetes): Acremoniopsis from forest soil and Collarina from water sediments (Spain), Phellinocrescentia from a Phellinus sp. (French Guiana), Neobambusicola from Strelitzia nicolai (South Africa), Neocladophialophora from Quercus robur (Germany), Neophysalospora from Corymbia henryi (Mozambique) and Xenophaeosphaeria from Grewia sp. (Tanzania). Morphological and culture characteristics along with ITS DNA barcodes are provided for all taxa.

This study is the rst to statistically analyze the stylet probing/penetration behaviors of Lygus (Hemiptera: Miridae) bugs, and the external body movements associated with both probing and nonprobing, via electrical penetration graph (EPG) and videorecording, respectively. Behavioral quantication allows powerful statistical comparisons among host plants or other treatments. Thus, statistical analysis of data has played an important role in EPG research. However, few attempts have been made to standardize types and terminology used for statistical parameters. We provide here the rst complete system of organization and terminology for nonsequential EPG parameters. Widespread adoption of these terms will allow standardization in EPG research. Our EPG and video data reveal for the rst time the stylet penetration behaviors of nymphal L. hesperus that cause cotton square damage, and the mechanism involved. L. hesperus nymphs spent only 15% of their time on squares probing; the remainder was spent standing motionless in place, grooming, or in sensory exploration. While probing, two thirds of their time was spent in laceration/salivation and one third in ingestion. Thus, L. hesperus nymphs actively spread out numerous, minute injections of their macerating watery saliva, deeply drilled/lacerated into all parts of the developing square. After injection of saliva within the square, the insect then stands and waits for solubilization of the squares cell contents, and then quickly ingests the slurry. The extensive laceration by the stylets may, secondarily, potentiate salivary maceration by mechanically rupturing cell walls. The plant responses to such behavior are thus summarized as "mechanical cell rupture-enhanced maceration.

Volatile aroma compounds in plants are typically found both as “free” and “bound” to a sugar moiety. When bound, these compounds are not odor active; however, upon hydrolysis of the glycoside, these compounds may then be volatilized. In grapes and wine, a large proportion of volatile aroma compounds are found in the bound form. A review of glycosides in grapes and in wine is presented with a focus on identified glycoside structures, their biosynthesis, their potential roles in the plant, and methods for their analysis. Studies of these compounds and their concentration changes during the winemaking process are discussed.

Human milk lactoferrin (hmLF) is the most abundant glycoprotein present in human milk and displays a broad range of protective functions in the gut of newborn infants. hmLF is N-glycosylated, but little is known about the lactation stage-related development of the glycosylation phenotype. hmLF glycosylation from milk samples from five donors during the first 10 weeks of lactation was assessed and observed to be more diverse than previously reported. During this period dynamic changes in glycosylation were observed corresponding to a decrease in glycosylation in the second week followed by an increase in total glycosylation as well as higher order fucosylation thereafter. Gene expression analysis was performed in milk somatic cells from a sixth subject. It was found that fucosyltransferase expression increased during entire period, whereas expression of genes for the oligosaccharyl transferase complex decreased in the second week. The effect of hmLF glycosylation was examined for the protein's ability to affect bacterial binding to epithelial cells. hmLF significantly inhibited pathogen adhesion and purified hmLF glycans significantly reduced Salmonella invasion of colonic epithelial cells to levels associated with non-invasive deletion mutants. This study indicates that hmLF glycosylation is tightly regulated by gene expression and that glyco-variation is involved in modulating pathogen association. Human milk lactoferrin (hmLF) is the most abundant glycoprotein present in human milk and displays a broad range of protective functions in the gut of newborn infants. hmLF is N-glycosylated, but little is known about the lactation stage-related development of the glycosylation phenotype. hmLF glycosylation from milk samples from five donors during the first 10 weeks of lactation was assessed and observed to be more diverse than previously reported. During this period dynamic changes in glycosylation were observed corresponding to a decrease in glycosylation in the second week followed by an increase in total glycosylation as well as higher order fucosylation thereafter. Gene expression analysis was performed in milk somatic cells from a sixth subject. It was found that fucosyltransferase expression increased during entire period, whereas expression of genes for the oligosaccharyl transferase complex decreased in the second week. The effect of hmLF glycosylation was examined for the protein's ability to affect bacterial binding to epithelial cells. hmLF significantly inhibited pathogen adhesion and purified hmLF glycans significantly reduced Salmonella invasion of colonic epithelial cells to levels associated with non-invasive deletion mutants. This study indicates that hmLF glycosylation is tightly regulated by gene expression and that glyco-variation is involved in modulating pathogen association. Human milk constitutes the first source of nutrients for the newborn infant, but it has also evolved to endow several key physiological advantages to the neonate. Other than to provide the neonate with energy and amino acid building blocks, proteins possess a wide range of biological activities that promote the normal development and maturation of specific organs in the newborn, specifically, the functions of the gut mucosa and the growth of gut microbiota (1Lönnerdal B. Lien E.L. Nutritional and physiologic significance of alpha-lactalbumin in infants.Nutr. Rev. 2003; 61: 295-305Crossref PubMed Scopus (137) Google Scholar). Human milk proteins also display a protective effect against infectious diseases via antimicrobial and immuno-modulatory activities that confer passive immunity to the breast-fed infant (1Lönnerdal B. Lien E.L. Nutritional and physiologic significance of alpha-lactalbumin in infants.Nutr. Rev. 2003; 61: 295-305Crossref PubMed Scopus (137) Google Scholar, 2Kaufman D.A. Lactoferrin supplementation to prevent nosocomial infections in preterm infants.JAMA. 2009; 302: 1467-1468Crossref PubMed Scopus (10) Google Scholar, 3Venkatesh M.P. Rong L. Human recombinant lactoferrin acts synergistically with antimicrobials commonly used in neonatal practice against coagulase-negative staphylococci and Candida albicans causing neonatal sepsis.J. Med. Microbiol. 2008; 57: 1113-1121Crossref PubMed Scopus (53) Google Scholar). Many of these proteins are post-translationally modified and the possible roles of such modifications in mediating demonstrated bioactivities are largely unexplored. Lactoferrin (LF) 1The abbreviations used are:LFLactoferrinBSSLbile-salt stimulated lipaseDHB2,5-dihydroxybenzoic acidFUTfucosyltransferasehmLFhuman milk lactoferrinIRMPDinfrared multiphoton dissociationMWCOmolecular weight cut offOSToligosaccharide transferasePNGase Fpeptide N-glycosidase FRPKMreads per kilo base per million mapped readsSEMstandard error of the meanSWIFTstored-waveform inverse Fourier transformTFAtrifluoroacetic acidFTICRFourier Transform Ion Cyclotron Resonance. 1The abbreviations used are:LFLactoferrinBSSLbile-salt stimulated lipaseDHB2,5-dihydroxybenzoic acidFUTfucosyltransferasehmLFhuman milk lactoferrinIRMPDinfrared multiphoton dissociationMWCOmolecular weight cut offOSToligosaccharide transferasePNGase Fpeptide N-glycosidase FRPKMreads per kilo base per million mapped readsSEMstandard error of the meanSWIFTstored-waveform inverse Fourier transformTFAtrifluoroacetic acidFTICRFourier Transform Ion Cyclotron Resonance. is an iron-binding glycoprotein found in milk from most species, but human milk LF (hmLF) is the most abundant glycoprotein present in colostrum and mature milk (6–8 mg/ml and 2–4 mg/ml, respectively) (1Lönnerdal B. Lien E.L. Nutritional and physiologic significance of alpha-lactalbumin in infants.Nutr. Rev. 2003; 61: 295-305Crossref PubMed Scopus (137) Google Scholar, 4Garcia-Montoya I.A. Cendon T.S. Arevalo-Gallegos S. Rascon-Cruz Q. Lactoferrin a multiple bioactive protein: An overview.Biochim. Biophys. Acta. 2011; 1820: 226-236Crossref PubMed Scopus (312) Google Scholar). The presence of glycans on hmLF is long known (5Spik G. Strecker G. Fournet B. Bouquelet S. Montreuil J. Dorland L. van Halbeek H. Vliegenthart J.F. Primary structure of the glycans from human lactotransferrin.Eur. J. Biochem. 1982; 121: 413-419Crossref PubMed Scopus (159) Google Scholar), but so far, the only role identified is to protect the molecule from proteolysis (6van Veen H.A. Geerts M.E. van Berkel P.H. Nuijens J.H. The role of N-linked glycosylation in the protection of human and bovine lactoferrin against tryptic proteolysis.Eur. J. Biochem. 2004; 271: 678-684Crossref PubMed Scopus (95) Google Scholar). 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Levy Y. Folding of glycoproteins: toward understanding the biophysics of the glycosylation code.Curr. Opin. Struct. Biol. 2009; 19: 524-533Crossref PubMed Scopus (182) Google Scholar, 8Fukuda M.N. Sasaki H. Lopez L. Fukuda M. Survival of recombinant erythropoietin in the circulation: the role of carbohydrates.Blood. 1989; 73: 84-89Crossref PubMed Google Scholar, 9Marth J.D. Grewal P.K. Mammalian glycosylation in immunity.Nat. Rev. Immunol. 2008; 8: 874-887Crossref PubMed Scopus (527) Google Scholar, 10Ohtsubo K. Marth J.D. Glycosylation in cellular mechanisms of health and disease.Cell. 2006; 126: 855-867Abstract Full Text Full Text PDF PubMed Scopus (2086) Google Scholar). Carbohydrate structures attached to proteins play key roles in mediating cell signaling and cell-cell recognition events (11Takahashi M. Kuroki Y. Ohtsubo K. Taniguchi N. Core fucose and bisecting GlcNAc, the direct modifiers of the N-glycan core: their functions and target proteins.Carbohydr. 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Fernandes D.L. Leung A. Stanworth D. Rademacher T.W. Mizuochi T. Taniguchi T. Matsuta K. Association of rheumatoid arthritis and primary osteoarthritis with changes in the glycosylation pattern of total serum IgG.Nature. 1985; 316: 452-457Crossref PubMed Scopus (989) Google Scholar, 16Saldova R. Royle L. Radcliffe C.M. Abd Hamid U.M. Evans R. Arnold J.N. Banks R.E. Hutson R. Harvey D.J. Antrobus R. Petrescu S.M. Dwek R.A. Rudd P.M. Ovarian cancer is associated with changes in glycosylation in both acute-phase proteins and IgG.Glycobiology. 2007; 17: 1344-1356Crossref PubMed Scopus (331) Google Scholar, 17Arnold J.N. Saldova R. Galligan M.C. Murphy T.B. Mimura-Kimura Y. Telford J.E. Godwin A.K. Rudd P.M. Novel glycan biomarkers for the detection of lung cancer.J. Proteome Res. 2011; 10: 1755-1764Crossref PubMed Scopus (162) Google Scholar, 18Blomme B. Van Steenkiste C. Callewaert N. Van Vlierberghe H. Alteration of protein glycosylation in liver diseases.J. Hepatol. 2009; 50: 592-603Abstract Full Text Full Text PDF PubMed Scopus (176) Google Scholar, 19An H.J. Kronewitter S.R. de Leoz M.L. Lebrilla C.B. Glycomics and disease markers.Curr. Opin. Chem. Biol. 2009; 13: 601-607Crossref PubMed Scopus (222) Google Scholar). Additionally, glycosylation and glycan diversity are directly related to modulating microbial adhesion and invasion during infection (9Marth J.D. Grewal P.K. Mammalian glycosylation in immunity.Nat. Rev. Immunol. 2008; 8: 874-887Crossref PubMed Scopus (527) Google Scholar). Indeed, the first step in bacterial infection is the recognition of host glycans by bacterial lectins or vice versa. Thus, glycans attached to human milk to or pathogen to the epithelial in also be a key role in the protection of breast-fed against the in total protein as well as of human has been little the in glycosylation of milk the of bile-salt stimulated lipase was to have a dynamic glycosylation pattern the of lactation Y. M. Y. L. A. Changes in glycosylation of human lipase during Biochem. Biophys. PubMed Scopus Google Scholar). The biological significance of these so and unexplored. the role of lactoferrin in infant development and health and as the most abundant glycoprotein in examined the changes in glycosylation during the first of lactation with the that glycan is common the of lactation as a to pathogen during hmLF several J. Murphy Human milk lactoferrin by PubMed Scopus Google Scholar, S. G. P.H. of protein A as a protein of PubMed Google and M. Lactoferrin type in 2003; PubMed Scopus (95) Google Scholar, A. J. M. of and and their on and Microbiol. 2011; PubMed Scopus Google to antimicrobial of and/or bacterial the by a of amino found in the of the known as M. H. K. M. of a peptide from the of bovine 73: PubMed Scopus Google Scholar, M. K. H. K. M. of the of Biophys. Acta. PubMed Scopus Google Scholar). hmLF infections by and H. R.E. of 2009; PubMed Scopus Google Scholar). to the role of glycosylation of hmLF in these and activities has been the N-glycan of human milk lactoferrin by analysis in samples of hmLF purified from five donors during the first of The expression of genes associated with glycosylation in milk somatic cells was assessed to the regulation of the dynamic The biological and/or significance of glycans found in hmLF was in of with colonic epithelial cells and bacterial in the presence of hmLF and A purified human milk lactoferrin standard was from was from and 10 were from peptide N-glycosidase F was from was from and from was a from of and and were from and and were from and trifluoroacetic acid were or were by five from to human milk samples on and were in this milk samples were and were to a and LF from milk samples was performed in a by J. J. B. of lactoferrin from human milk by PubMed Scopus Google with as human milk samples were for The was in a and a was to a of The was and for were with of and with The were the and the was and the were and the samples were to with the for with of proteins were five of and were LF to and was with five of were against 10 and was the and were by LF and LF purified from samples from donors during the of lactation were reduced and in F or was to samples and glycans were by for glycans were purified by The were with of followed by of in trifluoroacetic acid and of The oligosaccharide samples were the for 10 and with of was performed to the to analysis in order to were with of in followed by of trifluoroacetic acid in and in were in 10 of were on an with an a a and a The was with of on the followed by the of of as a and of The were to was performed an with of to a of for a range of and were performed on in the The was with a and glycans were identified by with a glycan were the cell stored-waveform inverse Fourier transform to and infrared multiphoton dissociation the infrared was by a was by the between and was increased the of the was and hmLF were by of standard hmLF with the corresponding followed by of hmLF was in of with from was in with in and with was performed in The corresponding were to and with for in a the samples were to the and were from hmLF by and with a of 10 LF were in of and protein was hmLF were from of reduced and LF with F as and from the protein by a with of followed by of was and glycans were in of followed by in milk samples were from a on and to a infant the period, the a and on to the for were of for oligosaccharide analysis and from somatic cells. The of of the The was in with the in the with cells were by of to of milk and for 10 The of cells was with 10 of and 10 of and to The cells were for 10 The was and was from the milk somatic cell to the was by an and the and was assessed by the and by with an Gene expression analysis was on milk samples on and by was and purified and was to and first and second were followed by and The were purified and the Core the reads of were and in and expression analysis of Human was as the for the were by the per kilo base per million mapped for gene A. K. L. B. and by 2008; PubMed Scopus Google and with human epithelial cells were as per the in with for cells were to a of in a modified with bovine serum amino 10 acid 10 and were in for to S. for and their in A in 2003; 15: Scopus Google Scholar). The epithelial cells were with of to the with hmLF and bacterial to the adhesion bacterial was from growth for and for in the adhesion cells were from of the growth for with an of and to an of in modified amino 10 10 and bacterial was with hmLF or the purified by for and to the cells to a of infection of The cells with and hmLF were in an for to the the epithelial cells. were and the was with of L. M. L. of and cells to Res. 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Gene expression was for the genes and and is in The expression of genes the changes during the lactation a decrease from to and and increased between and during the the effect of genes on of hmLF is the changes observed by are with the changes in glycosylation observed for expression of the genes associated with glycosylation different during are in per kilo base per million mapped complex complex complex associated protein in a The of glycans on hmLF during The for the different are in are with the of fucosylation during The expression of fucosyltransferase genes were of the expression of genes increased during lactation and genes the lactation The observed increased expression of the genes in somatic cells from human milk with the increased levels of fucosylation that were observed on the of hmLF glycosylation on with human colonic epithelial cells and five were performed with of hmLF or hmLF in were and of Salmonella and were these are found in the gut during the first weeks of their the role of the hmLF protein as well as the hmLF were examined for total the of only was significantly reduced with of hmLF and S. of only the hmLF significantly reduced for Thus, was by the protein whereas binding of the was significantly reduced by the hmLF that the Salmonella used in this study are of a and were by hmLF and the of specific hmLF glycosylation on binding was examined by of of and from the Additionally, was adhesion and invasion to the role of of the was significantly by hmLF and of the of hmLF in a in of S. and S. to the of the of fucose from lactoferrin in a increase in adhesion of S. whereas it on S. and it reduced invasion of S. of increased but invasion of S. but on the that identified the in total protein and protein in human little has been in the glycosylation and in glycosylation of milk and the of The glycosylation of was in H. 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In addition to a nutritive role, human milk also guides the development of a protective intestinal microbiota in the infant. Human milk possesses an overabundance of complex oligosaccharides that are indigestible by the infant yet are consumed by microbial populations in the developing intestine. These oligosaccharides are believed to facilitate enrichment of a healthy infant gastrointestinal microbiota, often associated with bifidobacteria. Advances in glycomics have enabled precise determination of milk glycan structures as well as identification of the specific glycans consumed by various gut microbes. Furthermore, genomic analysis of bifidobacteria from infants has revealed specific genetic loci related to milk oligosaccharide import and processing, suggesting coevolution between the human host, milk glycans, and the microbes they enrich. This review discusses the current understanding of how human milk oligosaccharides interact with the infant microbiota and examines the opportunities for translating this knowledge to improve the functionality of infant formulas.