2018-2019 field seasons of the Maize Genomes to Fields (G2F) G x E project.

OBJECTIVES: This report provides information about the public release of the 2018-2019 Maize G X E project of the Genomes to Fields (G2F) Initiative datasets. G2F is an umbrella initiative that evaluates maize hybrids and inbred lines across multiple environments and makes available phenotypic, genotypic, environmental, and metadata information. The initiative understands the necessity to characterize and deploy public sources of genetic diversity to face the challenges for more sustainable agriculture in the context of variable environmental conditions. DATA DESCRIPTION: Datasets include phenotypic, climatic, and soil measurements, metadata information, and inbred genotypic information for each combination of location and year. Collaborators in the G2F initiative collected data for each location and year; members of the group responsible for coordination and data processing combined all the collected information and removed obvious erroneous data. The collaborators received the data before the DOI release to verify and declare that the data generated in their own locations was accurate. ReadMe and description files are available for each dataset. Previous years of evaluation are already publicly available, with common hybrids present to connect across all locations and years evaluated since this project's inception.

Assessment of fitness and vector competence of a New Caledonia wMel Aedes aegypti strain before field-release.

BACKGROUND: Biological control programs involving Wolbachia-infected Aedes aegypti are currently deployed in different epidemiological settings. New Caledonia (NC) is an ideal location for the implementation and evaluation of such a strategy as the only proven vector for dengue virus (DENV) is Ae. aegypti and dengue outbreaks frequency and severity are increasing. We report the generation of a NC Wolbachia-infected Ae. aegypti strain and the results of experiments to assess the vector competence and fitness of this strain for future implementation as a disease control strategy in Noumea, NC. METHODS/PRINCIPAL FINDINGS: The NC Wolbachia strain (NC-wMel) was obtained by backcrossing Australian AUS-wMel females with New Caledonian Wild-Type (NC-WT) males. Blocking of DENV, chikungunya (CHIKV), and Zika (ZIKV) viruses were evaluated via mosquito oral feeding experiments and intrathoracic DENV challenge. Significant reduction in infection rates were observed for NC-wMel Ae. aegypti compared to WT Ae. aegypti. No transmission was observed for NC-wMel Ae. aegypti. Maternal transmission, cytoplasmic incompatibility, fertility, fecundity, wing length, and insecticide resistance were also assessed in laboratory experiments. Ae. aegypti NC-wMel showed complete cytoplasmic incompatibility and a strong maternal transmission. Ae. aegypti NC-wMel fitness seemed to be reduced compared to NC-WT Ae. aegypti and AUS-wMel Ae. aegypti regarding fertility and fecundity. However further experiments are required to assess it accurately. CONCLUSIONS/SIGNIFICANCE: Our results demonstrated that the NC-wMel Ae. aegypti strain is a strong inhibitor of DENV, CHIKV, and ZIKV infection and prevents transmission of infectious viral particles in mosquito saliva. Furthermore, our NC-wMel Ae. aegypti strain induces reproductive cytoplasmic incompatibility with minimal apparent fitness costs and high maternal transmission, supporting field-releases in Noumea, NC.

The importance of dominance and genotype-by-environment interactions on grain yield variation in a large-scale public cooperative maize experiment.

High-dimensional and high-throughput genomic, field performance, and environmental data are becoming increasingly available to crop breeding programs, and their integration can facilitate genomic prediction within and across environments and provide insights into the genetic architecture of complex traits and the nature of genotype-by-environment interactions. To partition trait variation into additive and dominance (main effect) genetic and corresponding genetic-by-environment variances, and to identify specific environmental factors that influence genotype-by-environment interactions, we curated and analyzed genotypic and phenotypic data on 1918 maize (Zea mays L.) hybrids and environmental data from 65 testing environments. For grain yield, dominance variance was similar in magnitude to additive variance, and genetic-by-environment variances were more important than genetic main effect variances. Models involving both additive and dominance relationships best fit the data and modeling unique genetic covariances among all environments provided the best characterization of the genotype-by-environment interaction patterns. Similarity of relative hybrid performance among environments was modeled as a function of underlying weather variables, permitting identification of weather covariates driving correlations of genetic effects across environments. The resulting models can be used for genomic prediction of mean hybrid performance across populations of environments tested or for environment-specific predictions. These results can also guide efforts to incorporate high-throughput environmental data into genomic prediction models and predict values in new environments characterized with the same environmental characteristics.

The Change in Winter Wheat Response to Deoxynivalenol and Fusarium Head Blight Through Technological and Agronomic Progress.

Fusarium head blight (FHB) in wheat causes yield loss, quality reduction, and mycotoxin contamination in temperate wheat production areas worldwide. The objective of this study was to quantify the progress of agronomic and FHB management strategies over the past two decades in FHB suppression and agronomic performance of winter wheat in environments favorable for FHB. Field experiments were conducted in environments typical of FHB epidemics to compare common agronomic and FHB management practices used in the 1996 era compared with those used in 2016. The experiments included a comparison of three different nitrogen (N) fertilizer application rates and six old (1996-era) and new (modern-era) winter wheat cultivars representing combinations of susceptibility and era to FHB, with and without a fungicide applied at flowering (pydiflumetofen + propiconazole). To mimic environments favorable for infection (similar to 1996 in Ontario, Canada), plots were challenged at 50% anthesis with F. graminearum macroconidia suspension followed by mist irrigation. The modern management strategy of using moderately resistant cultivars, a fungicide applied at flowering, and a high rate of N fertilizer reduced total deoxynivalenol by 67%, reduced Fusarium-damaged kernels by 49%, reduced FHB index by 86%, increased grain test weight by 11%, and increased grain yield by 31% compared with the standard management practice of seeding highly susceptible cultivars with no fungicide and a lower rate of N fertilizer recommended in the 1996 era. This study enabled a published economic assessment of the return on investment for the improvements in cultivars, fungicide, and N fertilizer applications since 1996.

Meta-Analysis of Soybean Yield Response to Foliar Fungicides Evaluated from 2005 to 2018 in the United States and Canada.

Random-effect meta-analyses were performed on data from 240 field trials conducted between 2005 and 2018 across nine U.S. states and Ontario, Canada, to quantify the yield response of soybean after application of foliar fungicides at beginning pod (R3) stage. Meta-analysis showed that the overall mean yield response when fungicide was used compared with not applying a fungicide was 2.7% (110 kg/ha). Moderator variables were also investigated and included fungicide group, growing season, planting date, and base yield, which all significantly influenced the yield response. There was also evidence that precipitation from the time of planting to the R3 growth stage influenced yield when fungicide was used (P = 0.059). Fungicides containing a premix of active ingredients from multiple groups (either two or three ingredients) increased the yield by 3.0% over not applying a fungicide. The highest and lowest yield responses were observed in 2005 and 2007, respectively. Better yield response to fungicides (a 3.0% increase) occurred when soybean crops were planted not later than 21 May and when total precipitation between planting and the R3 application date was above historic averages. Temperatures during the season did not influence the yield response. Yield response to fungicide was higher (a 4.7% increase) in average yield category (no spray control yield 2,878 to 3,758 kg/ha) and then gradually decreased with increasing base yield. Partial economic analyses indicated that use of foliar fungicides is less likely to be profitable when foliar diseases are absent or at low levels.

Maize genomes to fields (G2F): 2014-2017 field seasons: genotype, phenotype, climatic, soil, and inbred ear image datasets.

OBJECTIVES: Advanced tools and resources are needed to efficiently and sustainably produce food for an increasing world population in the context of variable environmental conditions. The maize genomes to fields (G2F) initiative is a multi-institutional initiative effort that seeks to approach this challenge by developing a flexible and distributed infrastructure addressing emerging problems. G2F has generated large-scale phenotypic, genotypic, and environmental datasets using publicly available inbred lines and hybrids evaluated through a network of collaborators that are part of the G2F's genotype-by-environment (G × E) project. This report covers the public release of datasets for 2014-2017. DATA DESCRIPTION: Datasets include inbred genotypic information; phenotypic, climatic, and soil measurements and metadata information for each testing location across years. For a subset of inbreds in 2014 and 2015, yield component phenotypes were quantified by image analysis. Data released are accompanied by README descriptions. For genotypic and phenotypic data, both raw data and a version without outliers are reported. For climatic data, a version calibrated to the nearest airport weather station and a version without outliers are reported. The 2014 and 2015 datasets are updated versions from the previously released files [1] while 2016 and 2017 datasets are newly available to the public.

Utility of Climatic Information via Combining Ability Models to Improve Genomic Prediction for Yield Within the Genomes to Fields Maize Project.

Genomic prediction provides an efficient alternative to conventional phenotypic selection for developing improved cultivars with desirable characteristics. New and improved methods to genomic prediction are continually being developed that attempt to deal with the integration of data types beyond genomic information. Modern automated weather systems offer the opportunity to capture continuous data on a range of environmental parameters at specific field locations. In principle, this information could characterize training and target environments and enhance predictive ability by incorporating weather characteristics as part of the genotype-by-environment (G×E) interaction component in prediction models. We assessed the usefulness of including weather data variables in genomic prediction models using a naïve environmental kinship model across 30 environments comprising the Genomes to Fields (G2F) initiative in 2014 and 2015. Specifically four different prediction scenarios were evaluated (i) tested genotypes in observed environments; (ii) untested genotypes in observed environments; (iii) tested genotypes in unobserved environments; and (iv) untested genotypes in unobserved environments. A set of 1,481 unique hybrids were evaluated for grain yield. Evaluations were conducted using five different models including main effect of environments; general combining ability (GCA) effects of the maternal and paternal parents modeled using the genomic relationship matrix; specific combining ability (SCA) effects between maternal and paternal parents; interactions between genetic (GCA and SCA) effects and environmental effects; and finally interactions between the genetics effects and environmental covariates. Incorporation of the genotype-by-environment interaction term improved predictive ability across all scenarios. However, predictive ability was not improved through inclusion of naive environmental covariates in G×E models. More research should be conducted to link the observed weather conditions with important physiological aspects in plant development to improve predictive ability through the inclusion of weather data.

Detecting wMel Wolbachia in field-collected Aedes aegypti mosquitoes using loop-mediated isothermal amplification (LAMP).

BACKGROUND: The World Mosquito Program uses Wolbachia pipientis for the biocontrol of arboviruses transmitted by Aedes aegypti mosquitoes. Diagnostic testing for Wolbachia in laboratory colonies and in field-caught mosquito populations has typically employed PCR. New, simpler methods to diagnose Wolbachia infection in mosquitoes are required for large-scale operational use. METHODS: Field-collected Ae. aegypti mosquitoes from North Queensland were tested using primers designed to detect the Wolbachia wsp gene, specific to the strain wMel. The results were analysed by colour change in the reaction mix. Furthermore, to confirm the efficiency of the LAMP assay, the results were compared to the gold-standard qPCR test. RESULTS: A novel loop-mediated isothermal amplification (LAMP) colorimetric test for the wMel strain of Wolbachia was designed, developed and validated for use in a high-throughput setting. Against the standard qPCR test, the analytical sensitivity, specificity and diagnostic metrics were: sensitivity (99.6%), specificity (92.2%), positive predictive value (97.08%) and negative predictive value (99.30%). CONCLUSIONS: We describe an alternative, novel and high-throughput method for diagnosing wMel Wolbachia infections in mosquitoes. This assay should support Wolbachia surveillance in both laboratory and field populations of Ae. aegypti.

Maize Genomes to Fields: 2014 and 2015 field season genotype, phenotype, environment, and inbred ear image datasets.

OBJECTIVES: Crop improvement relies on analysis of phenotypic, genotypic, and environmental data. Given large, well-integrated, multi-year datasets, diverse queries can be made: Which lines perform best in hot, dry environments? Which alleles of specific genes are required for optimal performance in each environment? Such datasets also can be leveraged to predict cultivar performance, even in uncharacterized environments. The maize Genomes to Fields (G2F) Initiative is a multi-institutional organization of scientists working to generate and analyze such datasets from existing, publicly available inbred lines and hybrids. G2F's genotype by environment project has released 2014 and 2015 datasets to the public, with 2016 and 2017 collected and soon to be made available. DATA DESCRIPTION: Datasets include DNA sequences; traditional phenotype descriptions, as well as detailed ear, cob, and kernel phenotypes quantified by image analysis; weather station measurements; and soil characterizations by site. Data are released as comma separated value spreadsheets accompanied by extensive README text descriptions. For genotypic and phenotypic data, both raw data and a version with outliers removed are reported. For weather data, two versions are reported: a full dataset calibrated against nearby National Weather Service sites and a second calibrated set with outliers and apparent artifacts removed.

Fusarium graminearum Mycotoxins in Maize Associated With Striacosta albicosta (Lepidoptera: Noctuidae) Injury.

Western bean cutworm, Striacosta albicosta (Smith; Lepidoptera: Noctuidae) has become a key pest of maize, Zea mays (L.), in Ontario, Canada which is challenging to control due to its lack of susceptibility to most Bt-maize events. Injury by S. albicosta may exacerbate Fusarium graminearum (Schwabe; Hypocreales: Nectriaceae) infection through provision of entry points on the ear. The objectives of this study were to: investigate the relationship between injury by S. albicosta and deoxynivalenol (DON) accumulation; evaluate non-Bt and Bt-maize hybrids, with and without insecticide and fungicide application; and determine optimal insecticide-fungicide application timing for reducing S. albicosta injury and DON accumulation. The incidence of injury by S. albicosta and ear rot severity were found to increase DON concentrations under favorable environmental conditions for F. graminearum infection. Incidence of S. albicosta injury was more important than severity of injury for DON accumulation which may be due to larval consumption of infected kernels. The Vip3A × Cry1Ab event provided superior protection from the incidence and severity of S. albicosta injury compared to non-Bt or Cry1F hybrids. Insecticide application to a Vip3A × Cry1Ab hybrid did not reduce injury further; however, lower severity of injury was observed for non-Bt and Cry1F hybrids when pyrethroids or diamides were applied at early VT or R1 stages. DON concentrations were reduced with application of prothioconazole fungicide tank-mixed with insecticide at late VT (before silk browning) or when insecticide was applied at early VT followed by prothioconazole at R1. The application of an insecticide/fungicide tank-mix is the most efficient approach for maize hybrids lacking high-dose insecticidal proteins against S. albicosta and F. graminearum tolerance. Results demonstrate that reducing the risk of DON accumulation requires a strategic approach to manage complex associations among S. albicosta, F. graminearum and the environment.

The effect of artificial selection on phenotypic plasticity in maize.

Remarkable productivity has been achieved in crop species through artificial selection and adaptation to modern agronomic practices. Whether intensive selection has changed the ability of improved cultivars to maintain high productivity across variable environments is unknown. Understanding the genetic control of phenotypic plasticity and genotype by environment (G × E) interaction will enhance crop performance predictions across diverse environments. Here we use data generated from the Genomes to Fields (G2F) Maize G × E project to assess the effect of selection on G × E variation and characterize polymorphisms associated with plasticity. Genomic regions putatively selected during modern temperate maize breeding explain less variability for yield G × E than unselected regions, indicating that improvement by breeding may have reduced G × E of modern temperate cultivars. Trends in genomic position of variants associated with stability reveal fewer genic associations and enrichment of variants 0-5000 base pairs upstream of genes, hypothetically due to control of plasticity by short-range regulatory elements.

Golgi-located NTPDase1 of Leishmania major is required for lipophosphoglycan elongation and normal lesion development whereas secreted NTPDase2 is dispensable for virulence.

Parasitic protozoa, such as Leishmania species, are thought to express a number of surface and secreted nucleoside triphosphate diphosphohydrolases (NTPDases) which hydrolyze a broad range of nucleoside tri- and diphosphates. However, the functional significance of NTPDases in parasite virulence is poorly defined. The Leishmania major genome was found to contain two putative NTPDases, termed LmNTPDase1 and 2, with predicted NTPDase catalytic domains and either an N-terminal signal sequence and/or transmembrane domain, respectively. Expression of both proteins as C-terminal GFP fusion proteins revealed that LmNTPDase1 was exclusively targeted to the Golgi apparatus, while LmNTPDase2 was predominantly secreted. An L. major LmNTPDase1 null mutant displayed increased sensitivity to serum complement lysis and exhibited a lag in lesion development when infections in susceptible BALB/c mice were initiated with promastigotes, but not with the obligate intracellular amastigote stage. This phenotype is characteristic of L. major strains lacking lipophosphoglycan (LPG), the major surface glycoconjugate of promastigote stages. Biochemical studies showed that the L. major NTPDase1 null mutant synthesized normal levels of LPG that was structurally identical to wild type LPG, with the exception of having shorter phosphoglycan chains. These data suggest that the Golgi-localized NTPase1 is involved in regulating the normal sugar-nucleotide dependent elongation of LPG and assembly of protective surface glycocalyx. In contrast, deletion of the gene encoding LmNTPDase2 had no measurable impact on parasite virulence in BALB/c mice. These data suggest that the Leishmania major NTPDase enzymes have potentially important roles in the insect stage, but only play a transient or non-major role in pathogenesis in the mammalian host.

A new way of measuring apoptosis by absolute quantitation of inter-nucleosomally fragmented genomic DNA.

Several critical events of apoptosis occur in the cell nucleus, including inter-nucleosomal DNA fragmentation (apoptotic DNA) and eventual chromatin condensation. The generation of apoptotic DNA has become a biochemical hallmark of apoptosis because it is a late 'point of no return' step in both the extrinsic (cell-death receptor) and intrinsic (mitochondrial) apoptotic pathways. Despite investigators observing apoptotic DNA and understanding its decisive role as a marker of apoptosis for over 20 years, measuring it has proved elusive. We have integrated ligation-mediated PCR and qPCR to design a new way of measuring apoptosis, termed ApoqPCR, which generates an absolute value for the amount (picogram) of apoptotic DNA per cell population. ApoqPCR's advances over current methods include a 1000-fold linear dynamic range yet sensitivity to distinguish subtle low-level changes, measurement with a 3- to 4-log improvement in sample economy, and capacity for archival or longitudinal studies combined with high-throughput capability. We demonstrate ApoqPCR's utility in both in vitro and in vivo contexts. Considering the fundamental role apoptosis has in vertebrate and invertebrate health, growth and disease, the reliable measurement of apoptotic nucleic acid by ApoqPCR will be of value in cell biology studies in basic and applied science.

Ubisol-Aqua: coenzyme Q10 prevents antiretroviral toxic neuropathy in an in vitro model.

BACKGROUND: Peripheral neuropathy is the dose-limiting toxicity of stavudine and didanosine (nucleoside analogs used in HIV treatment) and is attributed to mitochondrial toxicity from these drugs. Acetyl L-carnitine (ALC) and co-enzyme Q(10) are proposed as neuropathy treatments, but evidence to support these is limited. METHODS: We examined ALC and a water-soluble formulation of co-enzyme Q(10) (H(Q)O) for the prevention of d4T and ddI neurotoxicity using cultured fetal rat DRG as an in vitro model. RESULTS: DdI (33microM) and d4T (50microM) caused clear toxicity (impaired neurite growth) by day 8 of DRG culture. H(Q)O at concentrations 1-100microM completely prevented the toxicity of 33microM ddI in vitro and ALC at concentrations 1-100 microM substantially (but incompletely) prevented ddI toxicity in this model. In contrast, ALC was ineffective at all concentrations tested for preventing the toxicity of 50microM d4T. H(Q)O showed dose-dependent efficacy for preventing d4T toxicity. H(Q)O (1microM) partially prevented d4T toxicity while 10 and 100microM H(Q)O completely prevented d4T toxicity in this model. CONCLUSIONS: We find H(Q)O is superior to ALC for preventing the neurotoxicity of d4T (the HIV treatment most associated with neuropathy) and ddI in vitro. Further study is needed to clarify any clinical role for co-enzyme Q(10) co-administration with d4T and ddI and to assess whether this compound may have a role in treating established cases of neuropathy.

Apoptosis: a clinically useful measure of antiretroviral drug toxicity?

Antiretroviral therapy (ART) has improved life expectancy with HIV infection, but long-term toxicities associated with these medications are now a major global disease burden. There is a clear need to develop useful methods for monitoring patients on antiretroviral drugs for early signs of toxicity. Assays with predictive utility -- allowing therapy to be changed before serious end organ damage occurs -- would be ideal. Attempts to develop biochemical methods of monitoring ART toxicity have concentrated on the mitochondrial toxicity of nucleoside analogue reverse transcriptase inhibitors and have not generally lead to assays with widespread clinical applications. For example, plasma lactate and peripheral blood measurements of mitochondrial DNA associate with exposure to potentially toxic nucleoside analogue reverse transcriptase inhibitors but have not reliably predicted clinical toxicity. Better assays are needed, including markers of toxicity from additional drug classes. Apoptosis may be a potential marker of ART toxicity. Increased apoptosis has been demonstrated both in vitro and in vivo in association with various antiretroviral drug classes and a range of clinical toxicities. However, quantifying apoptosis on biopsy specimens of tissue (such as adipose tissue) is impractical for patient monitoring. Novel assays have been described that can quantify apoptosis using minute tissue samples and initial results from clinical samples suggest peripheral blood may have utility in predicting ART toxicities. The limitations and potential of such techniques for monitoring patients for drug side effects will be discussed.

Measuring and monitoring apoptosis and drug toxicity in HIV patients by ligation-mediated polymerase chain reaction.

Apoptosis has a critical role in normal physiology while its dysregulation has causal links with certain pathologies. A biochemical hallmark of apoptosis, internucleosomal genomic DNA fragmentation, is detectable by ligation-mediated polymerase chain reaction (LM-PCR). Here we converted LM-PCR into a new apoptosis quantifier by dividing trace quantities of 600 bp apoptotic amplicons into those of a single copy house-keeping gene, generating the LM-PCR 'value'. Dynamic range was approximately 17-fold correlating with a approximately 200-fold difference in degree of apoptotic fragmentation. Inter- and intra-gel reliability were both excellent, supporting LM-PCR's utility with large sample sets. Validation experiments comprising cell exposure to staurosporine over time revealed LM-PCR is as sensitive as caspase-3/ELISA and more sensitive than terminal deoxynucleotidyl transferase-mediated dUTP nick end labelling/flourescence-activated cell sorting (TUNEL/FACS) for distinguishing low degrees of apoptosis (the spectrum most relevant in vivo). The LM-PCR profile mirrored that of caspase-3/ELISA but not TUNEL/FACS. We then applied this molecular tool to clinical investigation. Increased apoptosis is implicated in lipoatrophy (subcutaneous fat wasting), a serious, persistent toxicity of some nucleoside analogue reverse transcriptase inhibitors (NRTIs) used in anti-HIV highly active antiretroviral therapy (HAART). We demonstrated in 105 peripheral blood mononuclear cell samples that elevated LM-PCR values are seen during therapy with stavudine (d4T), a particularly toxic NRTI (P< 0.0001 versus no HAART, unpaired t-test). Elevated values were also independently associated with clinical evidence of lipoatrophy (P= 0.007, multiple logistic regression modelling) but not with patient age, CD4 T-cell count nor HIV viral load (P> 0.8 for each). Together these data demonstrate that LM-PCR is a robust and reliable quantifier of apoptosis with potential for basic science and clinical investigation.

Modeling effects of environment, insect damage, and Bt genotypes on fumonisin accumulation in maize in Argentina and the Philippines.

Fumonisins are common contaminants of maize (Zea mays L.) grain products, especially in countries where maize is a major constituent of the diet and are harmful to human and animal health. There is a need to better define environmental conditions that favor fumonisin accumulation in the grain of maize. The impacts of biotic and abiotic factors, and hybrids containing the Cry1Ab protein from Bacillus thuringiensis (Bt), were associated with fumonisin accumulation in the grain of maize across contrasting environments in Argentina and the Philippines between 2000 and 2002. Average fumonisin concentrations in grain samples varied from 0.5 to 12 microg g(-1) across field locations in Argentina, and from 0.3 to 1.8 microg g(-1) across locations in the Philippines. The ratio of fumonisin B1 to fumonisin B2 was <3.0 in four of nine locations in Argentina, which proved to be due to a higher prevalence of Fusarium proliferatum in those locations. Most of the variability of total fumonisins among maize grain samples was explained by location or weather (47%), followed by insect damage severity in mature ears (17%), hybrid (14%), and with the use of Bt hybrids (11%). In Argentina, where conditions were more favorable for accumulation of fumonisin in the years considered, fumonisin concentrations were lower in Bt hybrids compared to their genetic isolines by an average of 40%. A model was developed to predict fumonisin concentration using insect damage to ears and weather variables as predictors in the model. Four periods of weather around silking were identified as critical for fumonisin concentrations at harvest. The model accounted for 82% of the variability of total fumonisin across all locations in 2 years of the study.