Genetic Dissection of Bloom Time in Low Chilling Sweet Cherry (Prunus avium L.) Using a Multi-Family QTL Approach

Bloom time in sweet cherry (Prunus avium L.) is a highly heritable trait that varies between genotypes and depends on the environmental conditions. Bud-break occurs after chill and heat requirements of each genotype are fulfilled, and dormancy is released. Bloom time is a critical trait for fruit production as matching cultivar adaptation to the growing area is essential for adequate fruit set. Additionally, low chilling cultivars are of interest to extend sweet cherry production to warmer regions, and for the crop adaptation to increasing winter and spring temperatures. The aim of this work is to investigate the genetic control of this trait by analyzing multiple families derived from the low chilling and extra-early flowering local Spanish cultivar ‘Cristobalina’ and other cultivars with higher chilling requirements and medium to late bloom times. Bloom time evaluation in six related sweet cherry populations confirmed a high heritability of this trait, and skewed distribution toward late flowering, revealing possible dominance of the late bloom alleles. SNP genotyping of the six populations (n = 406) resulted in a consensus map of 1269 SNPs. Quantitative trait loci (QTL) analysis using the Bayesian approach implemented by FlexQTL™ software revealed two major QTLs on linkage groups 1 and 2 (qP-BT1.1m and qP-BT2.1m) that explained 47.6% of the phenotypic variation. The QTL on linkage group 1 was mapped to a 0.26 Mbp region that overlaps with the DORMANCY ASSOCIATED MADS-BOX (DAM) genes. This finding is consistent with peach results that indicate that these genes are major determinants of chilling requirement in Prunus. Haplotype analysis of the linkage group 1 and 2 QTL regions showed that ‘Cristobalina’ was the only cultivar tested that contributed early bloom time alleles for these two QTLs. This work contributes to knowledge of the genetic control of chilling requirement and bloom date and will enable marker-assisted selection for low chilling in sweet cherry breeding programs

Genetic and molecular characterization of three novel S-haplotypes in sour cherry (Prunus cerasus L.)

Tetraploid sour cherry (Prunus cerasus L.) exhibits gametophytic self-incompatibility (GSI) whereby the specificity of self-pollen rejection is controlled by alleles of the stylar and pollen specificity genes, S-RNase and SFB (S haplotype-specific F-box protein gene), respectively. As sour cherry selections can be either self-compatible (SC) or self-incompatible (SI), polyploidy per se does not result in SC. Instead the genotype-dependent loss of SI in sour cherry is due to the accumulation of non-functional S-haplotypes. The presence of two or more non-functional S-haplotypes within sour cherry 2x pollen renders that pollen SC. Two new S-haplotypes from sour cherry, S33 and S34, that are presumed to be contributed by the P. fruticosa species parent, the complete S-RNase and SFB sequences of a third S-haplotype, S35, plus the presence of two previously identified sweet cherry S-haplotypes, S14 and S16 are described here. Genetic segregation data demonstrated that the S16-, S33-, S34-, and S35-haplotypes present in sour cherry are fully functional. This result is consistent with our previous finding that ‘hetero-allelic’ pollen is incompatible in sour cherry. Phylogenetic analyses of the SFB and S-RNase sequences from available Prunus species reveal that the relationships among S-haplotypes show no correspondence to known organismal relationships at any taxonomic level within Prunus, indicating that polymorphisms at the S-locus have been maintained throughout the evolution of the genus. Furthermore, the phylogenetic relationships among SFB sequences are generally incongruent with those among S-RNase sequences for the same S-haplotypes. Hypotheses compatible with these results are discussed

Development and bin mapping of a Rosaceae Conserved Ortholog Set (COS) of markers

12 pages, 5 tables, 6 figures, additional 3 files.Background: Detailed comparative genome analyses within the economically important Rosaceae family have not been conducted. This is largely due to the lack of conserved gene-based molecular markers that are transferable among the important crop genera within the family [e.g. Malus (apple), Fragaria (strawberry), and Prunus (peach, cherry, apricot and almond)]. The lack of molecular markers and comparative whole genome sequence analysis for this family severely hampers crop improvement efforts as well as QTL confirmation and validation studies. Results: We identified a set of 3,818 rosaceaous unigenes comprised of two or more ESTs that correspond to single copy Arabidopsis genes. From this Rosaceae Conserved Orthologous Set (RosCOS), 1039 were selected from which 857 were used for the development of intron-flanking primers and allele amplification. This led to successful amplification and subsequent mapping of 613 RosCOS onto the Prunus TxE reference map resulting in a genome-wide coverage of 0.67 to 1.06 gene-based markers per cM per linkage group. Furthermore, the RosCOS primers showed amplification success rates from 23 to 100% across the family indicating that a substantial part of the RosCOS primers can be directly employed in other less studied rosaceaous crops. Comparisons of the genetic map positions of the RosCOS with the physical locations of the orthologs in the Populus trichocarpa genome identified regions of colinearity between the genomes of Prunus-Rosaceae and Populus-Salicaceae. Conclusion: Conserved orthologous genes are extremely useful for the analysis of genome evolution among closely and distantly related species. The results presented in this study demonstrate the considerable potential of the mapped Prunus RosCOS for genome-wide marker employment and comparative whole genome studies within the Rosaceae family. Moreover, these markers will also function as useful anchor points for the genome sequencing efforts currently ongoing in this family as well as for comparative QTL analyses.This work is supported by USDA-NRI grants 2008-02259 and 2005-00743. AC was also supported by funds from the Department of Horticulture and Crop Science, The Ohio State University.Peer reviewe

Behavioral health emergencies encountered by community paramedics: lessons from the field and opportunities for skills advancement

INTRODUCTION: When an individual calls 911 because of a behavioral health emergency, paramedics may be the first health care professional on scene to address that crisis. Traditionally, the paramedic profession has been educated to respond to life-threatening medical emergencies.1 Paramedics are increasingly expected to respond to behavioral health crises but may not be adequately prepared to do so. 1 There has been limited research regarding paramedics’ experiences, attitudes, and perceptions about responding to behavioral health crises, with a particular dearth of literature on this subject from the USA. 1,2,3,4,5 As the first point of contact, paramedics may influence the quality of care individuals experiencing behavioral health emergencies receive, rendering this gap in knowledge concerning. 1,4 Paramedics encounter a range of behavioral health crises, from individuals who have self-harmed, substance use, to older adults experiencing social isolation. 6,7 This study aimed to answer the following questions: Do paramedics feel well prepared to respond to behavioral health crises? How do paramedics describe the challenges and facilitators involved in responding to these situations? What are the skills they use to respond to these situations? The present study examined paramedics’ experiences and perceptions regarding behavioral health emergencies, aiming to address a significant gap in the literature on paramedics’ ability to respond to the needs of individuals experiencing behavioral health crises. BACKGROUND: Limited research based in the UK, Europe, and Australia has examined the extent to which paramedics feel equipped by their training to respond to situations of a behavioral health nature, with a marked lack of USA-based research in this area. In a survey of members of the College of Paramedics (UK), 98% of the 623 respondents endorsed the need for increased mental health education and training. 2 Paramedic training curriculums in the UK have evolved to incorporate more of an emphasis on behavioral health, yet whether or not this behavioral health training is effective has not yet been investigated. 1 Several Australian studies investigated paramedic perceptions of their mental health training and decision-making during mental health emergencies.5,8. Paramedic participants reported inadequate mental health training and indicated a need for increased education on mental illness, particularly regarding possible treatment options that are feasible as part of prehospital care.8. Paramedics described relying more on intuition than education when faced with challenging cases.5 In a mixed methods study, Australian paramedics reported the need for more education on how to adequately respond to older adults experiencing complex mental health crises including depression, social isolation, and food insecurity.7 Limited research has explored paramedic perceptions, attitudes, and experiences responding to behavioral health emergencies, but the existing research indicates that many emergency healthcare providers consider physical health emergencies more valuable or important than behavioral health emergencies.8,9,10 Studies indicate that paramedic students do not consider training in mental health issues as relevant to the profession,1 with an Australian study of paramedic undergraduate students indicating a lower level of regard for those with intellectual disabilities, substance abuse disorders, or acute mental illness compared to students studying in other health professions.11 A multi-site, qualitative study based in Paris, France and New York, New York examined how social and professional values influenced prehospital emergency workers’ responses.9 The authors found that patients deemed to have lower social value were lower priority to emergency providers, with cases involving substance use or calls from lower socioeconomic areas, for instance, regarded as less legitimate uses of emergency services.9 Notably, situations that required more complex or heroic medical or surgical actions were also attributed higher value by emergency healthcare workers.9 Greater importance was attributed to medical (versus behavioral health) emergencies across additional studies, with paramedic participants implying that behavioral health calls were not valid emergencies,5 and with paramedic participants indicating that their role is mainly to transport—not to treat—individuals experiencing behavioral health crises.8 Additionally, paramedic participants perceived their services as often used inappropriately for behavioral health emergencies due to the limited availability of more appropriate behavioral health services.8 Related research regarding the perceptions of emergency department physicians and nurses in the USA also revealed a greater value placed on physical versus behavioral health emergencies, as well as perceptions that certain crises (such as self-harm) were less valid, or less worthy of emergency treatment.10 This is particularly concerning given that emergency department healthcare providers interact with a substantial number of persons seeking behavioral health emergency care: in 2015, approximately 4.1% of visits to an emergency department were related to diagnosed mental health disorders (approximately 5,666,000 visits).12 The number of emergency department visits related to opioid overdose increased by 29.7% in the USA between July 2016 and September 2017.13 Emergency healthcare workers are treating a significant number of behavioral health emergencies, rendering improved understanding of healthcare providers’ perceptions and attitudes towards this population crucial. Existing research into the perceptions of experienced paramedic practitioners regarding behavioral health emergencies is limited.1,3,4 With the exception of a multi-site study based in New York, New York and Paris, France,9 a thorough review of the extant literature failed to unearth research on the perceptions and experiences of paramedics responding to behavioral health crises in the USA. This is a particularly significant gap, as differences in paramedic education, relevant behavioral health legislation, and culture may exist across countries. The present study used qualitative analyses of in-depth interviews of practicing paramedics to address this gap by examining paramedics’ perceptions and experiences responding to behavioral health crises in the USA. METHODS: This study was part of a Patient-Centered Outcomes Research Institute (PCORI) contract funding an examination of the Acute Community Care Program (ACCP) at Commonwealth Care Alliance (CCA). The Acute Community Care Program is a state-funded community paramedicine intervention for the patients of CCA, in conjunction with EasCare Ambulance Company. INTERVIEW GUIDE AND RECRUITMENT: The research team, which included experienced paramedics at EasCare Ambulance Company, worked together to create an open-ended guide to interview paramedics. The interview protocol consisted of six sections: (1) history of career as paramedic, (2) current skills and job experiences, (3) views of paramedic experience, (4) career goals, (5) final questions and wrap up, and (6) demographics. The research team revised the original guide after several pilot interviews and based on feedback from key informants, namely seasoned paramedic supervisors from the research team. Interviews with paramedics lasted approximately 1 h and were conducted over the phone by either the PI or Co-PI, who are experienced in qualitative research. The research team intended to interview 25 paramedics who would be identified through two large ambulance companies in the Greater Boston area of Massachusetts. The director at these ambulance companies asked their paramedics on staff if they would be willing to be interviewed and, if they agreed, the director gave us their contact information. Six paramedics who had initially agreed could not be reached, and arrangements were made with one paramedic who did not follow through with the interview. In total, the researchers conducted telephone interviews with 23 paramedics, after receiving verbal informed consent to perform and record the interview. Participants were mailed a $50 gift card to thank them for their time. Professional transcription service transcribed digital recordings verbatim, and the project manager reviewed all transcripts against the digital recording, making small corrections as needed. DATA ANALYSIS: Audiotapes of the sessions were analyzed by three research team members. A thematic analysis approach was utilized, starting with familiarization with the data, generating initial codes, searching for themes, reviewing themes, defining and naming themes, and producing a final analysis.14 The researchers worked both separately and collaboratively, working individually at each stage of the process and then coming together to compare results in order to ensure the validity of identified themes. Early readings of the transcript data allowed the researchers to develop preliminary ideas regarding potential themes. Each researcher then thoroughly analyzed the individual transcripts, generating initial codes and beginning to search for themes both within and across documents. During this stage, the researchers met frequently to compare and contrast the results of each individual’s analysis, reviewing emerging themes and collaborating to refine themes into a coherent set. To asses inter-rater reliability, two un-read transcripts were reviewed by all team members and coded independently, followed by meetings as a research team to compare coding decisions on each transcript and reconcile any discrepancies. This rigorous process helped to ensure strong inter-rater reliability as we were able to gain consensus in our coding process for all other transcripts. Researchers individually arranged transcript data verbatim into groups of initial themes, and a master document was then created collaboratively, which outlined each finalized theme and included transcript excerpts embodying each theme. Memo-writing was employed throughout the analytic process to document coding decisions.15 As a final check for validity, the two experienced paramedic supervisors from the research team reviewed the themes and subthemes and provided feedback about the findings. The paramedic supervisors confirmed that these themes were what they might have expected the data to reveal, further validating the findings. RESULTS: Table 1 shows demographic characteristics of the 23 participants. Results for this paper will focus on three themes identified in the data analysis. Namely, paramedics (1) report frequently working with patients who are having a behavioral health crisis, (2) report having inadequate behavioral health training, and (3) have many difficulties managing these patients, relying primarily on their professional experiences and/or strong interpersonal skills rather than explicit training to address patients’ needs (see Table 2).Accepted manuscrip

Sweet cherry cultivar identification by using SSR markers

In this study, sweet cherry varieties and types grown in Turkey were described using SSR markers. This study was undertaken to develop DNA marker profiles that could be used to distinguish among the sweet cherry cultivars used in production in Turkey. For microsatellite analysis, 13SSR primers isolated from sweet cherry (P. avium L.) sour cherry (P. cerasus L.) and peach (P. persica L. Batsch) were used on sweet cherry cultivars and types. Two primer pairs did not give amplification with genotypes analyzed. Two-primer pairs amplified monomorphic fragment for the sweet cherry varieties therefore they were uninformative for the sweet cherry genetic analysis. Genetic similarities were calculated and a dendrogram has been established. All of the 8 SSR primers used and cherries have produced amplified bands. For each primer the alleles obtained has been between 1 and 6, in total 38 alleles have found. Through these analyses the similarities between these varieties have been converted into numerical values. This happens to be the first study kind towards the molecular identification of sweet cherry genetic resources in Turke

Prunus genetics and applications after de novo genome sequencing: achievements and prospects

Prior to the availability of whole-genome sequences, our understanding of the structural and functional aspects of Prunus tree genomes was limited mostly to molecular genetic mapping of important traits and development of EST resources. With public release of the peach genome and others that followed, significant advances in our knowledge of Prunus genomes and the genetic underpinnings of important traits ensued. In this review, we highlight key achievements in Prunus genetics and breeding driven by the availability of these whole-genome sequences. Within the structural and evolutionary contexts, we summarize: (1) the current status of Prunus whole-genome sequences; (2) preliminary and ongoing work on the sequence structure and diversity of the genomes; (3) the analyses of Prunus genome evolution driven by natural and man-made selection; and (4) provide insight into haploblocking genomes as a means to define genome-scale patterns of evolution that can be leveraged for trait selection in pedigree-based Prunus tree breeding programs worldwide. Functionally, we summarize recent and ongoing work that leverages whole-genome sequences to identify and characterize genes controlling 22 agronomically important Prunus traits. These include phenology, fruit quality, allergens, disease resistance, tree architecture, and self-incompatibility. Translationally, we explore the application of sequence-based marker-assisted breeding technologies and other sequence-guided biotechnological approaches for Prunus crop improvement. Finally, we present the current status of publically available Prunus genomics and genetics data housed mainly in the Genome Database for Rosaceae (GDR) and its updated functionalities for future bioinformatics-based Prunus genetics and genomics inquiry.info:eu-repo/semantics/publishedVersio

Development and evaluation of a 9K SNP array for peach by internationally coordinated SNP detection and validation in breeding germplasm

Although a large number of single nucleotide polymorphism (SNP) markers covering the entire genome are needed to enable molecular breeding efforts such as genome wide association studies, fine mapping, genomic selection and marker-assisted selection in peach [Prunus persica (L.) Batsch] and related Prunus species, only a limited number of genetic markers, including simple sequence repeats (SSRs), have been available to date. To address this need, an international consortium (The International Peach SNP Consortium; IPSC) has pursued a coordinated effort to perform genome-scale SNP discovery in peach using next generation sequencing platforms to develop and characterize a high-throughput Illumina Infinium® SNP genotyping array platform. We performed whole genome re-sequencing of 56 peach breeding accessions using the Illumina and Roche/454 sequencing technologies. Polymorphism detection algorithms identified a total of 1,022,354 SNPs. Validation with the Illumina GoldenGate® assay was performed on a subset of the predicted SNPs, verifying ∼75% of genic (exonic and intronic) SNPs, whereas only about a third of intergenic SNPs were verified. Conservative filtering was applied to arrive at a set of 8,144 SNPs that were included on the IPSC peach SNP array v1, distributed over all eight peach chromosomes with an average spacing of 26.7 kb between SNPs. Use of this platform to screen a total of 709 accessions of peach in two separate evaluation panels identified a total of 6,869 (84.3%) polymorphic SNPs.The almost 7,000 SNPs verified as polymorphic through extensive empirical evaluation represent an excellent source of markers for future studies in genetic relatedness, genetic mapping, and dissecting the genetic architecture of complex agricultural traits. The IPSC peach SNP array v1 is commercially available and we expect that it will be used worldwide for genetic studies in peach and related stone fruit and nut species

Comparative analysis of rosaceous genomes and the reconstruction of a putative ancestral genome for the family

Abstract Background Comparative genome mapping studies in Rosaceae have been conducted until now by aligning genetic maps within the same genus, or closely related genera and using a limited number of common markers. The growing body of genomics resources and sequence data for both Prunus and Fragaria permits detailed comparisons between these genera and the recently released Malus × domestica genome sequence. Results We generated a comparative analysis using 806 molecular markers that are anchored genetically to the Prunus and/or Fragaria reference maps, and physically to the Malus genome sequence. Markers in common for Malus and Prunus, and Malus and Fragaria, respectively were 784 and 148. The correspondence between marker positions was high and conserved syntenic blocks were identified among the three genera in the Rosaceae. We reconstructed a proposed ancestral genome for the Rosaceae. Conclusions A genome containing nine chromosomes is the most likely candidate for the ancestral Rosaceae progenitor. The number of chromosomal translocations observed between the three genera investigated was low. However, the number of inversions identified among Malus and Prunus was much higher than any reported genome comparisons in plants, suggesting that small inversions have played an important role in the evolution of these two genera or of the Rosaceae.Apple genome research at FEM is supported by the research office of the Provincia autonoma di Trento. DJS and ELG acknowledge a grant from the East Malling Trust. Fragaria genomics at EMR is funded by the BBSRC. JMB is supported by a grant by Plant & Food Research's Excellence Programme. Apple genomics at Plant & Food Research is partially supported by the New Zealand Foundation for Research Science and Technology project C06X0812 "Exploiting Opportunities from Horticultural Genomics". Research conducted at IRTA was partly funded by the CONSOLIDER-INGENIO 2010 Program (CSD2007-00036) and project INIA-RTA2007-00063-00-00, both from the Spanish Ministry of Science and Innovation. RosCOS development at OSU/MSU was funded by the National Research Initiative Competitive Grant 2005-35300-15454 of USDA's National Institute of Food and Agriculture.Peer Reviewe

Development and Evaluation of a 9K SNP Array for Peach by Internationally Coordinated SNP Detection and Validation in Breeding Germplasm

Correction 21 Jun 2012: Verde I, Bassil N, Scalabrin S, Gilmore B, Lawley CT, et al. (2012) Correction: Development and Evaluation of a 9K SNP Array for Peach by Internationally Coordinated SNP Detection and Validation in Breeding Germplasm. PLOS ONE 7(6): 10.1371/annotation/33f1ba92-c304-4757-91aa-555de64a0768.Although a large number of single nucleotide polymorphism (SNP) markers covering the entire genome are needed to enable molecular breeding efforts such as genome wide association studies, fine mapping, genomic selection and marker-assisted selection in peach [Prunus persica (L.) Batsch] and related Prunus species, only a limited number of genetic markers, including simple sequence repeats (SSRs), have been available to date. To address this need, an international consortium (The International Peach SNP Consortium; IPSC) has pursued a coordinated effort to perform genome-scale SNP discovery in peach using next generation sequencing platforms to develop and characterize a high-throughput Illumina Infinium® SNP genotyping array platform. We performed whole genome re-sequencing of 56 peach breeding accessions using the Illumina and Roche/454 sequencing technologies. Polymorphism detection algorithms identified a total of 1,022,354 SNPs. Validation with the Illumina GoldenGate® assay was performed on a subset of the predicted SNPs, verifying ~75% of genic (exonic and intronic) SNPs, whereas only about a third of intergenic SNPs were verified. Conservative filtering was applied to arrive at a set of 8,144 SNPs that were included on the IPSC peach SNP array v1, distributed over all eight peach chromosomes with an average spacing of 26.7 kb between SNPs. Use of this platform to screen a total of 709 accessions of peach in two separate evaluation panels identified a total of 6,869 (84.3%) polymorphic SNPs. The almost 7,000 SNPs verified as polymorphic through extensive empirical evaluation represent an excellent source of markers for future studies in genetic relatedness, genetic mapping, and dissecting the genetic architecture of complex agricultural traits. The IPSC peach SNP array v1 is commercially available and we expect that it will be used worldwide for genetic studies in peach and related stone fruit and nut species