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Sample records for bovine snp50 illumina

  1. Characteristics and use of the Illumina BovineLD BeadChip

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Genotypic information from the 6,909-SNP Illumina BovineLD (LD) Genotyping BeadChip, which replaced the Illumina GoldenGate Bovine3K (3K) Genotyping BeadChip, have been included in US genomic evaluations since November 2011. Of 73 LD single nucleotide polymorphisms (SNP) not used in genomic evaluati...

  2. Use of the Illumina Bovine3K BEAD chip in dairy genomic evaluation

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Genomic evaluations using genotypes from the Illumina Bovine3K BEAD chip became available in September 2010 and were made official in December 2010. Approximately 4,000 genotypes a month, (93% female) are from this low cost chip. To integrate the 3K genotypes into the evaluations, they are imputed f...

  3. First insights into the microbial diversity in the omasum and reticulum of bovine using Illumina sequencing.

    PubMed

    Peng, Shuai; Yin, Jigang; Liu, Xiaolei; Jia, Boyin; Chang, Zhiguang; Lu, Huijun; Jiang, Ning; Chen, Qijun

    2015-08-01

    The digestive systems of mammals harbor a complex gut microbiome, comprising bacteria and other microorganisms that confer metabolic and immunological benefits to the host. Ruminants that digest plant-based foods have a four-compartment stomach consisting of the rumen, reticulum, omasum, and abomasum. The microorganisms in the stomach are essential for providing the host with critical nutrients. However, the majority of these microorganisms are unknown species. The microbiome of the stomach is diverse, and the majority of these organisms cannot be cultured. Next-generation sequencing (NGS) combined with bioinformatic analysis tools have allowed the dissection of the composition of the microbiome in samples collected from a specific environment. In this study, for the first time, the bacterial composition in two compartments, the reticulum and the omasum, of bovine were analyzed using a metagenomic approach and compared to the bacterial composition of the rumen. These data will assist in understanding the biology of ruminants and benefit the agricultural industry. The diversity and composition of the bacterial community in samples collected from the rumen, reticulum, and omasum of bovines in the Changchun Region of Northeast China were analyzed by sequencing the V3 region of the 16S rRNA gene using a barcoded Illumina paired-end sequencing technique, and the primary composition of the microbiome in the rumen, reticulum, and omasum of the bovines was determined. These microbiomes contained 17 phyla and 107 genera in all three samples. Five phyla, Bacteroidetes, Firmicutes, Proteobacteria, Spirochaetes, and Lentisphaerae, were the most abundant taxonomic groups. Additionally, the different stomach compartments harbored different compositions of the microorganisms. PMID:25604266

  4. Technical note: Characteristics and use of the Illumina BovineLD and GeneSeek Genomic Profiler low-density bead chips for genomic evaluation

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The GeneSeek Genomic Profiler (GGP) BeadChip, which became available commercially in February 2012, is based on the Illumina BovineLD Genotyping BeadChip with 1,745 additional single nucleotide polymorphisms (SNPs) for genomic evaluation and SNPs for proprietary single-gene tests. The BovineLD chip ...

  5. Characterization of the Cattle HapMap Population using the Illumina Bovine-50K SNP Chip

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A Bovine 50K Illumina™ iSelect SNP chip (51,386 polymorphic SNP markers) was designed using a combination of publicly available SNPs along with highly informative novel SNPs discovered using a reduced representation and next-generation sequencing technology strategy. A total of 576 animals (426 mal...

  6. Short communication: Imputation of markers on the bovine X chromosome.

    PubMed

    Mao, Xiaowei; Johansson, Anna Maria; Sahana, Goutam; Guldbrandtsen, Bernt; De Koning, Dirk-Jan

    2016-09-01

    Imputation is a cost-effective approach to augment marker data for genomic selection and genome-wide association studies. However, most imputation studies have focused on autosomes. Here, we assessed the imputation of markers on the X chromosome in Holstein cattle for nongenotyped animals and animals genotyped with low-density (Illumina BovineLD, Illumina Inc., San Diego, CA) chips, using animals genotyped with medium-density (Illumina BovineSNP50) chips. A total of 26,884 genotyped Holstein individuals genotyped with medium-density chips were used in this study. Imputation was carried out using FImpute V2.2. The following parameters were examined: treating the pseudoautosomal region as autosomal or as X specific, different sizes of reference groups, different male/female proportions in the reference group, and cumulated degree of relationship between the reference group and target group. The imputation accuracy of markers on the X chromosome was improved if the pseudoautosomal region was treated as autosomal. Increasing the proportion of females in the reference group improved the imputation accuracy for the X chromosome. Imputation for nongenotyped animals in general had lower accuracy compared with animals genotyped with the low-density single nucleotide polymorphism array. In addition, higher cumulative pedigree relationships between the reference group and the target animal led to higher imputation accuracy. In the future, better marker coverage of the X chromosome should be developed to facilitate genomic studies involving the X chromosome. PMID:27423959

  7. Genomic Heritability of Bovine Growth Using a Mixed Model

    PubMed Central

    Ryu, Jihye; Lee, Chaeyoung

    2014-01-01

    This study investigated heritability for bovine growth estimated with genomewide single nucleotide polymorphism (SNP) information obtained from a DNA microarray chip. Three hundred sixty seven Korean cattle were genotyped with the Illumina BovineSNP50 BeadChip, and 39,112 SNPs of 364 animals filtered by quality assurance were analyzed to estimate heritability of body weights at 6, 9, 12, 15, 18, 21, and 24 months of age. Restricted maximum likelihood estimate of heritability was obtained using covariance structure of genomic relationships among animals in a mixed model framework. Heritability estimates ranged from 0.58 to 0.76 for body weights at different ages. The heritability estimates using genomic information in this study were larger than those which had been estimated previously using pedigree information. The results revealed a trend that the heritability for body weight increased at a younger age (6 months). This suggests an early genetic evaluation for bovine growth using genomic information to increase genetic merits of animals. PMID:25358309

  8. A genome-wide association study for somatic cell score using the Illumina high-density bovine beadchip identifies several novel QTL potentially related to mastitis susceptibility.

    PubMed

    Meredith, Brian K; Berry, Donagh P; Kearney, Francis; Finlay, Emma K; Fahey, Alan G; Bradley, Daniel G; Lynn, David J

    2013-01-01

    Mastitis is an inflammation-driven disease of the bovine mammary gland that occurs in response to physical damage or infection and is one of the most costly production-related diseases in the dairy industry worldwide. We performed a genome-wide association study (GWAS) to identify genetic loci associated with somatic cell score (SCS), an indicator trait of mammary gland inflammation. A total of 702 Holstein-Friesian bulls were genotyped for 777,962 single nucleotide polymorphisms (SNPs) and associated with SCS phenotypes. The SCS phenotypes were expressed as daughter yield deviations (DYD) based on a large number of progeny performance records. A total of 138 SNPs on 15 different chromosomes reached genome-wide significance (corrected p-value ≤ 0.05) for association with SCS (after correction for multiple testing). We defined 28 distinct QTL regions and a number of candidate genes located in these QTL regions were identified. The most significant association (p-value = 1.70 × 10(-7)) was observed on chromosome 6. This QTL had no known genes annotated within it, however, the Ensembl Genome Browser predicted the presence of a small non-coding RNA (a Y RNA gene) in this genomic region. This Y RNA gene was 99% identical to human RNY4. Y RNAs are a rare type of non-coding RNA that were originally discovered due to their association with the autoimmune disease, systemic lupus erythematosus. Examining small-RNA sequencing (RNAseq) data being generated by us in multiple different mastitis-pathogen challenged cell-types has revealed that this Y RNA is expressed (but not differentially expressed) in these cells. Other QTL regions identified in this study also encoded strong candidate genes for mastitis susceptibility. A QTL region on chromosome 13, for example, was found to contain a cluster of β-defensin genes, a gene family with known roles in innate immunity. Due to the increased SNP density, this study also refined the boundaries for several known QTL for SCS and

  9. Estimation of Genomic Inbreeding Coefficients Using BovineSNP50 genotypes from U.S. Jersey Cattle

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In dairy cattle, inbreeding coefficients have been estimated from pedigree information; however, recent advances in genotyping technology allow the calculation of inbreeding based on molecular pedigree information. Because strong selection and recurrent inbreeding have decreased genetic variation, ...

  10. Application of the BovineSNP50 assay for QTL mapping and prediction of genetic merit in Holstein cattle

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The previous fifteen years have produced numerous QTL mapping experiments aimed at the identification of causal or linked polymorphisms for use in marker assisted selection programs to increase the rate of genetic gain in livestock species. To date, very few causal mutations for QTL have been ident...

  11. Analysis of Illumina Microbial Assemblies

    SciTech Connect

    Clum, Alicia; Foster, Brian; Froula, Jeff; LaButti, Kurt; Sczyrba, Alex; Lapidus, Alla; Woyke, Tanja

    2010-05-28

    Since the emerging of second generation sequencing technologies, the evaluation of different sequencing approaches and their assembly strategies for different types of genomes has become an important undertaken. Next generation sequencing technologies dramatically increase sequence throughput while decreasing cost, making them an attractive tool for whole genome shotgun sequencing. To compare different approaches for de-novo whole genome assembly, appropriate tools and a solid understanding of both quantity and quality of the underlying sequence data are crucial. Here, we performed an in-depth analysis of short-read Illumina sequence assembly strategies for bacterial and archaeal genomes. Different types of Illumina libraries as well as different trim parameters and assemblers were evaluated. Results of the comparative analysis and sequencing platforms will be presented. The goal of this analysis is to develop a cost-effective approach for the increased throughput of the generation of high quality microbial genomes.

  12. Effective Population Size and Signatures of Selection Using Bovine 50K SNP Chips in Korean Native Cattle (Hanwoo)

    PubMed Central

    Li, Yi; Kim, Jong-Joo

    2015-01-01

    Inferring the effective population size and the pattern of selection signatures is of interest both from an evolutionary perspective and to improve models for mapping of quantitative trait genes. We used DNA samples of 61 sires and 486 progeny of the Hanwoo, genotyped by the Illumina Bovine SNP50 BeadChip, to analyze the genetic structure. Our study showed a persistent decline in effective population size throughout the period considered, but suggested a marked decline at one distinctive time point (100th generation) and two sharp decline intervals (50th–25th generation and 25th–10th generation). This pattern can be explained by Hanwoo formation and the modern breeding program. Our results revealed 95 regions exhibiting the footprint of recent positive selection at a threshold level of 0.01. We found an overlap of the 11 core regions presenting top P-values and those that had previously been identified as harboring quantitative trait loci from other breeds. The information generated from this study can be used to better understand the mechanism of selection in Hanwoo breeding, and provide important implications for the design and application of association studies in the Hanwoo population. PMID:26244003

  13. A Study of Applicability of SNP Chips Developed for Bovine and Ovine Species to Whole-Genome Analysis of Reindeer Rangifer tarandus.

    PubMed

    Kharzinova, Veronika R; Sermyagin, Alexander A; Gladyr, Elena A; Okhlopkov, Innokentiy M; Brem, Gottfried; Zinovieva, Natalia A

    2015-01-01

    Two sets of commercially available single nucleotide polymorphisms (SNPs) developed for cattle (BovineSNP50 BeadChip) and sheep (OvineSNP50 BeadChip) have been trialed for whole-genome analysis of 4 female samples of Rangifer tarandus inhabiting Russia. We found out that 43.0% of bovine and 47.0% of Ovine SNPs could be genotyped, while only 5.3% and 2.03% of them were respectively polymorphic. The scored and the polymorphic SNPs were identified on each bovine and each ovine chromosome, but their distribution was not unique. The maximal value of runs of homozygosity (ROH) was 30.93Mb (for SNPs corresponding to bovine chromosome 8) and 80.32Mb (for SNPs corresponding to ovine chromosome 7). Thus, the SNP chips developed for bovine and ovine species can be used as a powerful tool for genome analysis in reindeer R. tarandus. PMID:26447215

  14. Genome-wide association study for host response to bovine leukemia virus in Holstein cows.

    PubMed

    Brym, P; Bojarojć-Nosowicz, B; Oleński, K; Hering, D M; Ruść, A; Kaczmarczyk, E; Kamiński, S

    2016-07-01

    The mechanisms of leukemogenesis induced by bovine leukemia virus (BLV) and the processes underlying the phenomenon of differential host response to BLV infection still remain poorly understood. The aim of the study was to screen the entire cattle genome to identify markers and candidate genes that might be involved in host response to bovine leukemia virus infection. A genome-wide association study was performed using Holstein cows naturally infected by BLV. A data set included 43 cows (BLV positive) and 30 cows (BLV negative) genotyped for 54,609 SNP markers (Illumina Bovine SNP50 BeadChip). The BLV status of cows was determined by serum ELISA, nested-PCR and hematological counts. Linear Regression Analysis with a False Discovery Rate and kinship matrix (computed on the autosomal SNPs) was calculated to find out which SNP markers significantly differentiate BLV-positive and BLV-negative cows. Nine markers reached genome-wide significance. The most significant SNPs were located on chromosomes 23 (rs41583098), 3 (rs109405425, rs110785500) and 8 (rs43564499) in close vicinity of a patatin-like phospholipase domain containing 1 (PNPLA1); adaptor-related protein complex 4, beta 1 subunit (AP4B1); tripartite motif-containing 45 (TRIM45) and cell division cycle associated 2 (CDCA2) genes, respectively. Furthermore, a list of 41 candidate genes was composed based on their proximity to significant markers (within a distance of ca. 1 Mb) and functional involvement in processes potentially underlying BLV-induced pathogenesis. In conclusion, it was demonstrated that host response to BLV infection involves nine sub-regions of the cattle genome (represented by 9 SNP markers), containing many genes which, based on the literature, could be involved to enzootic bovine leukemia progression. New group of promising candidate genes associated with the host response to BLV infection were identified and could therefore be a target for future studies. The functions of candidate genes

  15. Illumina Unamplified Indexed Library Construction: An Automated Approach

    SciTech Connect

    Hack, Christopher A.; Sczyrba, Alexander; Cheng, Jan-Fang

    2011-03-21

    Manual library construction is a limiting factor in Illumina sequencing. Constructing libraries by hand is costly, time-consuming, low-throughput, and ergonomically hazardous, and constructing multiple libraries introduces risk of library failure due to pipetting errors. The ability to construct multiple libraries simultaneously in automated fashion represents significant cost and time savings. Here we present a strategy to construct up to 96 unamplified indexed libraries using Illumina TruSeq reagents and a Biomek FX robotic platform. We also present data to indicate that this library construction method has little or no risk of cross-contamination between samples.

  16. Illumina Spin-off to Develop Early-Detection Test.

    PubMed

    Colwell, Janet

    2016-04-01

    DNA-sequencing giant Illumina has formed a new company, called Grail, to develop liquid biopsies capable of spotting cancer before symptoms arise. The start-up is working on a low-cost "pan-cancer" test that can detect multiple cancer types early, which it hopes to introduce by 2019. PMID:26873346

  17. Current Status of Genotyping and Discovery Work at USMARC

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The Illumina BovineSNP50 DNA chip has substantially changed the genetic and genomic research program at USMARC. It has enhanced our commitment to produce genetic tools that can be exported to beef cattle producers to further their selection goals in hard-to-measure traits such as feed efficiency, co...

  18. Selection of single-nucleotide polymorphisms and quality of genotypes used in genomic evaluation of dairy cattle in the United States and Canada

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Nearly 57,000 single nucleotide polymorphisms (SNP) on the Illumina BovineSNP50™ chip were investigated to determine their usefulness for genomic prediction. Genotypes were obtained for 12,591 Holstein bulls and cows and SNP selection was done using 5,503 bulls that had genotypes on a larger set of ...

  19. Selection of single nucleotide polymorphisms and genotype quality for genomic prediction of genetic merit in dairy cattle

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A process to prepare high-density genotypic data for use in genomic prediction of genetic merit was developed. Marker genotypes from over 51,000 single nucleotide polymorphisms (SNP) were generated for 3,139 Holstein bulls on the Illumina Bovine SNP50™ chip. The SNP were categorized by minor allele ...

  20. A whole genome association analysis identified loci associated with Mycobacterium avium subsp. paratuberculosis infection status in U.S. Holstein cattle

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The purpose of this study was to identify loci associated with Mycobacterium avium subspecies paratuberculosis (Map) infection status in US Holsteins using the Illumina BovineSNP50 BeadChip whole genome SNP assay. Two hundred forty-five cows from dairies in New York, Pennsylvania and Vermont were fo...

  1. Selection and Management of DNA Markers for Use in Genomic Evaluation

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A database was constructed to store genotypes for 50,972 single-nucleotide polymorphisms (SNP) from the Illumina BovineSNP50 BeadChip for over 30,000 animals. The database allows storage of multiple samples per animal and stores all SNP genotypes for a sample in a single row. An indicator specifies ...

  2. HIGH-THROUGHPUT PHYLOGENOMICS: FROM ANCIENT DNA TO SIGNATURES OF HUMAN ANIMAL HUSBANDRY

    Technology Transfer Automated Retrieval System (TEKTRAN)

    We utilized the Illumina BovineSNP50 BeadChip with 54,693 single nucleotide polymorphism loci developed for Bos taurus taurus to rapidly genotype 677 individuals representing 61 Pecoran (horned ruminant) species diverged by up to 29 million years. We produced a completely bifurcating tree, the first...

  3. Accelerating genetic improvement with SNP chips and DNA sequencing

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The development of high-density single nucleotide polymorphism (SNP) assays is expected to have a profound impact on genetic progress in the U.S. dairy industry. In the 16 months since its initial availability, the Illumina BovineSNP50 BeadChip has been used to genotype nearly 20,000 Holsteins. Thes...

  4. QuorUM: An Error Corrector for Illumina Reads

    PubMed Central

    Marçais, Guillaume; Yorke, James A.; Zimin, Aleksey

    2015-01-01

    Motivation Illumina Sequencing data can provide high coverage of a genome by relatively short (most often 100 bp to 150 bp) reads at a low cost. Even with low (advertised 1%) error rate, 100 × coverage Illumina data on average has an error in some read at every base in the genome. These errors make handling the data more complicated because they result in a large number of low-count erroneous k-mers in the reads. However, there is enough information in the reads to correct most of the sequencing errors, thus making subsequent use of the data (e.g. for mapping or assembly) easier. Here we use the term “error correction” to denote the reduction in errors due to both changes in individual bases and trimming of unusable sequence. We developed an error correction software called QuorUM. QuorUM is mainly aimed at error correcting Illumina reads for subsequent assembly. It is designed around the novel idea of minimizing the number of distinct erroneous k-mers in the output reads and preserving the most true k-mers, and we introduce a composite statistic π that measures how successful we are at achieving this dual goal. We evaluate the performance of QuorUM by correcting actual Illumina reads from genomes for which a reference assembly is available. Results We produce trimmed and error-corrected reads that result in assemblies with longer contigs and fewer errors. We compared QuorUM against several published error correctors and found that it is the best performer in most metrics we use. QuorUM is efficiently implemented making use of current multi-core computing architectures and it is suitable for large data sets (1 billion bases checked and corrected per day per core). We also demonstrate that a third-party assembler (SOAPdenovo) benefits significantly from using QuorUM error-corrected reads. QuorUM error corrected reads result in a factor of 1.1 to 4 improvement in N50 contig size compared to using the original reads with SOAPdenovo for the data sets investigated

  5. Information Theoretical Analysis of a Bovine Gene Atlas Reveals Chromosomal Regions with Tissue Specific Gene Expression.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    An essential step to understanding the genomic biology of any organism is to comprehensively survey its transcriptome. We present the Bovine Gene Atlas (BGA) a compendium of over 7.2 million unique 20 base Illumina DGE tags representing 100 tissue transcriptomes collected primarily from L1 Dominette...

  6. Design of a bovine low-density SNP array optimized for imputation

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The Illumina BovineLD BeadChip was designed to support imputation to higher density genotypes in dairy and beef breeds by including single-nucleotide polymorphisms (SNPs) that had a high minor allele frequency as well as uniform spacing across the genome except at the ends of the chromosome where de...

  7. Illumina human exome genotyping array clustering and quality control

    PubMed Central

    Guo, Yan; He, Jing; Zhao, Shilin; Wu, Hui; Zhong, Xue; Sheng, Quanhu; Samuels, David C; Shyr, Yu; Long, Jirong

    2015-01-01

    With the rise of high-throughput sequencing technology, traditional genotyping arrays are gradually being replaced by sequencing technology. Against this trend, Illumina has introduced an exome genotyping array that provides an alternative approach to sequencing, especially suited to large-scale genome-wide association studies (GWASs). the exome genotyping array targets the exome plus rare single-nucleotide polymorphisms (SNPs), a feature that makes it substantially more challenging to process than previous genotyping arrays that targeted common SNPs. Researchers have struggled to generate a reliable protocol for processing exome genotyping array data. The Vanderbilt epidemiology center, in cooperation with Vanderbilt Technologies for Advanced Genomics Analysis and Research Design (VANGARD), has developed a thorough exome chip–processing protocol. The protocol was developed during the processing of several large exome genotyping array-based studies, which included over 60,000 participants combined. The protocol described herein contains detailed clustering techniques and robust quality control procedures, and it can benefit future exome genotyping array–based GWASs. PMID:25321409

  8. PANDAseq: paired-end assembler for illumina sequences

    PubMed Central

    2012-01-01

    Background Illumina paired-end reads are used to analyse microbial communities by targeting amplicons of the 16S rRNA gene. Publicly available tools are needed to assemble overlapping paired-end reads while correcting mismatches and uncalled bases; many errors could be corrected to obtain higher sequence yields using quality information. Results PANDAseq assembles paired-end reads rapidly and with the correction of most errors. Uncertain error corrections come from reads with many low-quality bases identified by upstream processing. Benchmarks were done using real error masks on simulated data, a pure source template, and a pooled template of genomic DNA from known organisms. PANDAseq assembled reads more rapidly and with reduced error incorporation compared to alternative methods. Conclusions PANDAseq rapidly assembles sequences and scales to billions of paired-end reads. Assembly of control libraries showed a 4-50% increase in the number of assembled sequences over naïve assembly with negligible loss of "good" sequence. PMID:22333067

  9. Acute hepatotoxicity: a predictive model based on focused illumina microarrays.

    PubMed

    Zidek, Nadine; Hellmann, Juergen; Kramer, Peter-Juergen; Hewitt, Philip G

    2007-09-01

    Drug-induced hepatotoxicity is a major issue for drug development, and toxicogenomics has the potential to predict toxicity during early toxicity screening. A bead-based Illumina oligonucleotide microarray containing 550 liver specific genes has been developed. We have established a predictive screening system for acute hepatotoxicity by analyzing differential gene expression profiles of well-known hepatotoxic and nonhepatotoxic compounds. Low and high doses of tetracycline, carbon tetrachloride (CCL4), 1-naphthylisothiocyanate (ANIT), erythromycin estolate, acetaminophen (AAP), or chloroform as hepatotoxicants, clofibrate, theophylline, naloxone, estradiol, quinidine, or dexamethasone as nonhepatotoxic compounds, were administered as a single dose to male Sprague-Dawley rats. After 6, 24, and 72 h, livers were taken for histopathological evaluation and for analysis of gene expression alterations. All hepatotoxic compounds tested generated individual gene expression profiles. Based on leave-one-out cross-validation analysis, gene expression profiling allowed the accurate discrimination of all model compounds, 24 h after high dose treatment. Even during the regeneration phase, 72 h after treatment, CCL4, ANIT, and AAP were predicted to be hepatotoxic, and only these three compounds showed histopathological changes at this time. Furthermore, we identified 64 potential marker genes responsible for class prediction, which reflected typical hepatotoxicity responses. These genes and pathways, commonly deregulated by hepatotoxicants, may be indicative of the early characterization of hepatotoxicity and possibly predictive of later hepatotoxicity onset. Two unknown test compounds were used for prevalidating the screening test system, with both being correctly predicted. We conclude that focused gene microarrays are sufficient to classify compounds with respect to toxicity prediction. PMID:17522070

  10. A large genome centre’s improvements to the Illumina sequencing system

    PubMed Central

    Quail, Michael A.; Kozarewa, Iwanka; Smith, Frances; Scally, Aylwyn; Stephens, Philip J.; Durbin, Richard; Swerdlow, Harold; Turner, Daniel J.

    2008-01-01

    Preface The Wellcome Trust Sanger Institute is one of the world’s largest genome centres, and a substantial amount of our sequencing is performed on ‘next generation’ massively parallel sequencing technologies: in June 2008 the quantity of purity filtered sequence data generated by our Genome Analyzer (Illumina) platforms reached 1 terabase, and our average weekly Illumina production output is currently 64gigabases (Gb). Here we describe a set of improvements we have made to the standard Illumina protocols to make the library preparation more reliable in a high throughput environment, to reduce bias, tighten insert size distribution, and reliably obtain high yields of data. PMID:19034268

  11. Complete genome sequence of Deltapapillomavirus 4 (bovine papillomavirus 2) from a bovine papillomavirus lesion in Amazon Region, Brazil

    PubMed Central

    Daudt, Cíntia; da Silva, Flavio RC; Cibulski, Samuel P; Weber, Matheus N; Mayer, Fabiana Q; Varela, Ana Paula M; Roehe, Paulo M; Canal, Cláudio W

    2016-01-01

    The complete genome sequence of bovine papillomavirus 2 (BPV2) from Brazilian Amazon Region was determined using multiple-primed rolling circle amplification followed by Illumina sequencing. The genome is 7,947 bp long, with 45.9% GC content. It encodes seven early (E1, E2,E4, E5, E6,E7, and E8) and two late (L1 and L2) genes. The complete genome of a BPV2 can help in future studies since this BPV type is highly reported worldwide although the lack of complete genome sequences available. PMID:27074259

  12. Complete genome sequence of Deltapapillomavirus 4 (bovine papillomavirus 2) from a bovine papillomavirus lesion in Amazon Region, Brazil.

    PubMed

    Daudt, Cíntia; Silva, Flavio Rc da; Cibulski, Samuel P; Weber, Matheus N; Mayer, Fabiana Q; Varela, Ana Paula M; Roehe, Paulo M; Canal, Cláudio W

    2016-04-01

    The complete genome sequence of bovine papillomavirus 2 (BPV2) from Brazilian Amazon Region was determined using multiple-primed rolling circle amplification followed by Illumina sequencing. The genome is 7,947 bp long, with 45.9% GC content. It encodes seven early (E1, E2,E4, E5, E6,E7, and E8) and two late (L1 and L2) genes. The complete genome of a BPV2 can help in future studies since this BPV type is highly reported worldwide although the lack of complete genome sequences available. PMID:27074259

  13. Analysis, Optimization and Verification of Illumina-Generated 16S rRNA Gene Amplicon Surveys

    PubMed Central

    Nelson, Michael C.; Morrison, Hilary G.; Benjamino, Jacquelynn; Grim, Sharon L.; Graf, Joerg

    2014-01-01

    The exploration of microbial communities by sequencing 16S rRNA genes has expanded with low-cost, high-throughput sequencing instruments. Illumina-based 16S rRNA gene sequencing has recently gained popularity over 454 pyrosequencing due to its lower costs, higher accuracy and greater throughput. Although recent reports suggest that Illumina and 454 pyrosequencing provide similar beta diversity measures, it remains to be demonstrated that pre-existing 454 pyrosequencing workflows can transfer directly from 454 to Illumina MiSeq sequencing by simply changing the sequencing adapters of the primers. In this study, we modified 454 pyrosequencing primers targeting the V4-V5 hyper-variable regions of the 16S rRNA gene to be compatible with Illumina sequencers. Microbial communities from cows, humans, leeches, mice, sewage, and termites and a mock community were analyzed by 454 and MiSeq sequencing of the V4-V5 region and MiSeq sequencing of the V4 region. Our analysis revealed that reference-based OTU clustering alone introduced biases compared to de novo clustering, preventing certain taxa from being observed in some samples. Based on this we devised and recommend an analysis pipeline that includes read merging, contaminant filtering, and reference-based clustering followed by de novo OTU clustering, which produces diversity measures consistent with de novo OTU clustering analysis. Low levels of dataset contamination with Illumina sequencing were discovered that could affect analyses that require highly sensitive approaches. While moving to Illumina-based sequencing platforms promises to provide deeper insights into the breadth and function of microbial diversity, our results show that care must be taken to ensure that sequencing and processing artifacts do not obscure true microbial diversity. PMID:24722003

  14. De novo genome sequence assembly of a filamentous fungus using Sanger, 454 and Illumina sequence data

    PubMed Central

    DiGuistini, Scott; Liao, Nancy Y; Platt, Darren; Robertson, Gordon; Seidel, Michael; Chan, Simon K; Docking, T Roderick; Birol, Inanc; Holt, Robert A; Hirst, Martin; Mardis, Elaine; Marra, Marco A; Hamelin, Richard C; Bohlmann, Jörg; Breuil, Colette; Jones, Steven JM

    2009-01-01

    Sequencing-by-synthesis technologies can reduce the cost of generating de novo genome assemblies. We report a method for assembling draft genome sequences of eukaryotic organisms that integrates sequence information from different sources, and demonstrate its effectiveness by assembling an approximately 32.5 Mb draft genome sequence for the forest pathogen Grosmannia clavigera, an ascomycete fungus. We also developed a method for assessing draft assemblies using Illumina paired end read data and demonstrate how we are using it to guide future sequence finishing. Our results demonstrate that eukaryotic genome sequences can be accurately assembled by combining Illumina, 454 and Sanger sequence data. PMID:19747388

  15. High-throughput illumina strand-specific RNA sequencing library preparation

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Conventional Illumina RNA-Seq does not have the resolution to decode the complex eukaryote transcriptome due to the lack of RNA polarity information. Strand-specific RNA sequencing (ssRNA-Seq) can overcome these limitations and as such is better suited for genome annotation, de novo transcriptome as...

  16. Using Illumina mRNA-Sequencing to understand Glycine max-Phakopsora pachyhrizi interaction

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Soybean is one of the top five agricultural products in the United States and is highly susceptible to soybean rust (SR), an exotic obligate fungus that arrived in the USA in 2004. We used mRNA-Seq by Illumina/Solexa to analyze gene expression pattern of the host and pathogen at different time point...

  17. Illumina sequencing of green stink bug nymph and adult cdna to identify potential rnai gene targets

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Whole-body transcriptomes for nymphs and adults of the green stink bug, Acrosternum hilare (Say), were sequenced on an Illumina® Genome Analyzer IIx sequencer. The insects were collected from sites in North Carolina and Virginia, USA. The cDNA library for each sample was sequenced on one lane of an...

  18. 77 FR 20319 - Bovine Spongiform Encephalopathy; Importation of Bovines and Bovine Products

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-04-04

    ...; ] DEPARTMENT OF AGRICULTURE Animal and Plant Health Inspection Service 9 CFR Part 93 RIN 0579-AC68 Bovine Spongiform Encephalopathy; Importation of Bovines and Bovine Products Correction In proposed rule...

  19. 78 FR 73993 - Bovine Spongiform Encephalopathy; Importation of Bovines and Bovine Products

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-12-10

    ... Health Inspection Service 9 CFR Parts 92, 93, 94, 95, 96, and 98 RIN 0579-AC68 Bovine Spongiform Encephalopathy; Importation of Bovines and Bovine Products Corrections In rule document 2013-28228 appearing...

  20. Preparing DNA Libraries for Multiplexed Paired-End Deep Sequencing for Illumina GA Sequencers

    PubMed Central

    Son, Mike S.; Taylor, Ronald K.

    2011-01-01

    Whole genome sequencing, also known as deep sequencing, is becoming a more affordable and efficient way to identify SNP mutations, deletions and insertions in DNA sequences across several different strains. Two major obstacles preventing the widespread use of deep sequencers are the costs involved in services used to prepare DNA libraries for sequencing and the overall accuracy of the sequencing data. This Unit describes the preparation of DNA libraries for multiplexed paired-end sequencing using the Illumina GA series sequencer. Self-preparation of DNA libraries can help reduce overall expenses, especially if optimization is required for the different samples, and use of the Illumina GA Sequencer can improve the quality of the data. PMID:21400673

  1. Ameliorated de novo transcriptome assembly using Illumina paired end sequence data with Trinity Assembler

    PubMed Central

    Bankar, Kiran Gopinath; Todur, Vivek Nagaraj; Shukla, Rohit Nandan; Vasudevan, Madavan

    2015-01-01

    Advent of Next Generation Sequencing has led to possibilities of de novo transcriptome assembly of organisms without availability of complete genome sequence. Among various sequencing platforms available, Illumina is the most widely used platform based on data quality, quantity and cost. Various de novo transcriptome assemblers are also available today for construction of de novo transcriptome. In this study, we aimed at obtaining an ameliorated de novo transcriptome assembly with sequence reads obtained from Illumina platform and assembled using Trinity Assembler. We found that, primary transcriptome assembly obtained as a result of Trinity can be ameliorated on the basis of transcript length, coverage, and depth and protein homology. Our approach to ameliorate is reproducible and could enhance the sensitivity and specificity of the assembled transcriptome which could be critical for validation of the assembled transcripts and for planning various downstream biological assays. PMID:26484285

  2. DADA2: High-resolution sample inference from Illumina amplicon data.

    PubMed

    Callahan, Benjamin J; McMurdie, Paul J; Rosen, Michael J; Han, Andrew W; Johnson, Amy Jo A; Holmes, Susan P

    2016-07-01

    We present the open-source software package DADA2 for modeling and correcting Illumina-sequenced amplicon errors (https://github.com/benjjneb/dada2). DADA2 infers sample sequences exactly and resolves differences of as little as 1 nucleotide. In several mock communities, DADA2 identified more real variants and output fewer spurious sequences than other methods. We applied DADA2 to vaginal samples from a cohort of pregnant women, revealing a diversity of previously undetected Lactobacillus crispatus variants. PMID:27214047

  3. The effect of strand bias in Illumina short-read sequencing data

    PubMed Central

    2012-01-01

    Background When using Illumina high throughput short read data, sometimes the genotype inferred from the positive strand and negative strand are significantly different, with one homozygous and the other heterozygous. This phenomenon is known as strand bias. In this study, we used Illumina short-read sequencing data to evaluate the effect of strand bias on genotyping quality, and to explore the possible causes of strand bias. Result We collected 22 breast cancer samples from 22 patients and sequenced their exome using the Illumina GAIIx machine. By comparing the consistency between the genotypes inferred from this sequencing data with the genotypes inferred from SNP chip data, we found that, when using sequencing data, SNPs with extreme strand bias did not have significantly lower consistency rates compared to SNPs with low or no strand bias. However, this result may be limited by the small subset of SNPs present in both the exome sequencing and the SNP chip data. We further compared the transition and transversion ratio and the number of novel non-synonymous SNPs between the SNPs with low or no strand bias and those with extreme strand bias, and found that SNPs with low or no strand bias have better overall quality. We also discovered that the strand bias occurs randomly at genomic positions across these samples, and observed no consistent pattern of strand bias location across samples. By comparing results from two different aligners, BWA and Bowtie, we found very consistent strand bias patterns. Thus strand bias is unlikely to be caused by alignment artifacts. We successfully replicated our results using two additional independent datasets with different capturing methods and Illumina sequencers. Conclusion Extreme strand bias indicates a potential high false-positive rate for SNPs. PMID:23176052

  4. Unlocking the bovine genome

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The draft genome sequence of cattle (Bos taurus) has now been analyzed by the Bovine Genome Sequencing and Analysis Consortium and the Bovine HapMap Consortium, which together represent an extensive collaboration involving more than 300 scientists from 25 different countries. ...

  5. Bovine viral diarrhea viruses

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Infections with bovine viral diarrhea viruses (BVDV) result in significant economic losses for beef and dairy producers worldwide. BVDV is actually an umbrella term for two species of viruses, BVDV1 and BVDV2, within the Pestivirus genus of the Flavivirus family. While denoted as a bovine pathogen...

  6. Evaluation of Different Normalization and Analysis Procedures for Illumina Gene Expression Microarray Data Involving Small Changes

    PubMed Central

    Johnstone, Daniel M.; Riveros, Carlos; Heidari, Moones; Graham, Ross M.; Trinder, Debbie; Berretta, Regina; Olynyk, John K.; Scott, Rodney J.; Moscato, Pablo; Milward, Elizabeth A.

    2013-01-01

    While Illumina microarrays can be used successfully for detecting small gene expression changes due to their high degree of technical replicability, there is little information on how different normalization and differential expression analysis strategies affect outcomes. To evaluate this, we assessed concordance across gene lists generated by applying different combinations of normalization strategy and analytical approach to two Illumina datasets with modest expression changes. In addition to using traditional statistical approaches, we also tested an approach based on combinatorial optimization. We found that the choice of both normalization strategy and analytical approach considerably affected outcomes, in some cases leading to substantial differences in gene lists and subsequent pathway analysis results. Our findings suggest that important biological phenomena may be overlooked when there is a routine practice of using only one approach to investigate all microarray datasets. Analytical artefacts of this kind are likely to be especially relevant for datasets involving small fold changes, where inherent technical variation—if not adequately minimized by effective normalization—may overshadow true biological variation. This report provides some basic guidelines for optimizing outcomes when working with Illumina datasets involving small expression changes.

  7. Viral metagenomics: analysis of begomoviruses by illumina high-throughput sequencing.

    PubMed

    Idris, Ali; Al-Saleh, Mohammed; Piatek, Marek J; Al-Shahwan, Ibrahim; Ali, Shahjahan; Brown, Judith K

    2014-01-01

    Traditional DNA sequencing methods are inefficient, lack the ability to discern the least abundant viral sequences, and ineffective for determining the extent of variability in viral populations. Here, populations of single-stranded DNA plant begomoviral genomes and their associated beta- and alpha-satellite molecules (virus-satellite complexes) (genus, Begomovirus; family, Geminiviridae) were enriched from total nucleic acids isolated from symptomatic, field-infected plants, using rolling circle amplification (RCA). Enriched virus-satellite complexes were subjected to Illumina-Next Generation Sequencing (NGS). CASAVA and SeqMan NGen programs were implemented, respectively, for quality control and for de novo and reference-guided contig assembly of viral-satellite sequences. The authenticity of the begomoviral sequences, and the reproducibility of the Illumina-NGS approach for begomoviral deep sequencing projects, were validated by comparing NGS results with those obtained using traditional molecular cloning and Sanger sequencing of viral components and satellite DNAs, also enriched by RCA or amplified by polymerase chain reaction. As the use of NGS approaches, together with advances in software development, make possible deep sequence coverage at a lower cost; the approach described herein will streamline the exploration of begomovirus diversity and population structure from naturally infected plants, irrespective of viral abundance. This is the first report of the implementation of Illumina-NGS to explore the diversity and identify begomoviral-satellite SNPs directly from plants naturally-infected with begomoviruses under field conditions. PMID:24625811

  8. Illumina-based analysis of bacterial diversity related to halophytes Salicornia europaea and Sueada aralocaspica.

    PubMed

    Shi, Ying-wu; Lou, Kai; Li, Chun; Wang, Lei; Zhao, Zhen-yong; Zhao, Shuai; Tian, Chang-yan

    2015-10-01

    We used Illumina-based 16S rRNA V3 amplicon pyrosequencing to investigate the community structure of soil bacteria from the rhizosphere surrounding Salicornia europaea, and endophytic bacteria living in Salicornia europaea plants and Sueada aralocaspica seeds growing at the Fukang Desert Ecosystem Observation and Experimental Station (FDEOES) in Xinjiang Province, China, using an Illumina genome analyzer. A total of 89.23 M effective sequences of the 16S rRNA gene V3 region were obtained from the two halophyte species. These sequences revealed a number of operational taxonomic units (OTUs) in the halophytes. There were between 22-2,206 OTUs in the halophyte plant sample, at the 3% cutoff level, and a sequencing depth of 30,000 sequences. We identified 25 different phyla, 39 classes and 141 genera from the resulting 134,435 sequences. The most dominant phylum in all the samples was Proteobacteria (41.61%-99.26%; average, 43.30%). The other large phyla were Firmicutes (0%- 7.19%; average, 1.15%), Bacteroidetes (0%-1.64%; average, 0.44%) and Actinobacteria (0%-0.46%; average, 0.24%). This result suggested that the diversity of bacteria is abundant in the rhizosphere soil, while the diversity of bacteria was poor within Salicornia europaea plant samples. To the extent of our knowledge, this study is the first to characterize and compare the endophytic bacteria found within different halophytic plant species roots using PCR-based Illumina pyrosequencing method. PMID:26428918

  9. High Throughput Plasmid Sequencing with Illumina and CLC Bio (Seventh Annual Sequencing, Finishing, Analysis in the Future (SFAF) Meeting 2012)

    SciTech Connect

    Athavale, Ajay

    2012-06-01

    Ajay Athavale (Monsanto) presents "High Throughput Plasmid Sequencing with Illumina and CLC Bio" at the 7th Annual Sequencing, Finishing, Analysis in the Future (SFAF) Meeting held in June, 2012 in Santa Fe, NM.

  10. High Throughput Plasmid Sequencing with Illumina and CLC Bio (Seventh Annual Sequencing, Finishing, Analysis in the Future (SFAF) Meeting 2012)

    ScienceCinema

    Athavale, Ajay [Monsanto

    2013-01-25

    Ajay Athavale (Monsanto) presents "High Throughput Plasmid Sequencing with Illumina and CLC Bio" at the 7th Annual Sequencing, Finishing, Analysis in the Future (SFAF) Meeting held in June, 2012 in Santa Fe, NM.

  11. Use of the Illumina GoldenGate assay for single nucleotide polymorphism (SNP) genotyping in cereal crops.

    PubMed

    Chao, Shiaoman; Lawley, Cindy

    2015-01-01

    Highly parallel genotyping assays, such as the GoldenGate assay developed by Illumina, capable of interrogating up to 3,072 single nucleotide polymorphisms (SNPs) simultaneously, have greatly facilitated genome-wide studies, particularly for crops with large and complex genome structures. In this report, we provide detailed information and guidelines regarding genomic DNA preparation, SNP assay design, SNP assay protocols, and genotype calling using Illumina's GenomeStudio software. PMID:25373766

  12. Development of High Throughput Process for Constructing 454 Titanium and Illumina Libraries

    SciTech Connect

    Deshpande, Shweta; Hack, Christopher; Tang, Eric; Malfatti, Stephanie; Ewing, Aren; Lucas, Susan; Cheng, Jan-Fang

    2010-05-28

    We have developed two processes with the Biomek FX robot to construct 454 titanium and Illumina libraries in order to meet the increasing library demands. All modifications in the library construction steps were made to enable the adaptation of the entire processes to work with the 96-well plate format. The key modifications include the shearing of DNA with Covaris E210 and the enzymatic reaction cleaning and fragment size selection with SPRI beads and magnetic plate holders. The construction of 96 Titanium libraries takes about 8 hours from sheared DNA to ssDNA recovery. The processing of 96 Illumina libraries takes less time than that of the Titanium library process. Although both processes still require manual transfer of plates from robot to other work stations such as thermocyclers, these robotic processes represent about 12- to 24-folds increase of library capacity comparing to the manual processes. To enable the sequencing of many libraries in parallel, we have also developed sets of molecular barcodes for both library types. The requirements for the 454 library barcodes include 10 bases, 40-60percent GC, no consecutive same base, and no less than 3 bases difference between barcodes. We have used 96 of the resulted 270 barcodes to construct libraries and pool to test the ability of accurately assigning reads to the right samples. When allowing 1 base error occurred in the 10 base barcodes, we could assign 99.6percent of the total reads and 100percent of them were uniquely assigned. As for the Illumina barcodes, the requirements include 4 bases, balanced GC, and at least 2 bases difference between barcodes. We have begun to assess the ability to assign reads after pooling different number of libraries. We will discuss the progress and the challenges of these scale-up processes.

  13. Whole genome sequencing of enriched chloroplast DNA using the Illumina GAII platform

    PubMed Central

    2010-01-01

    Background Complete chloroplast genome sequences provide a valuable source of molecular markers for studies in molecular ecology and evolution of plants. To obtain complete genome sequences, recent studies have made use of the polymerase chain reaction to amplify overlapping fragments from conserved gene loci. However, this approach is time consuming and can be more difficult to implement where gene organisation differs among plants. An alternative approach is to first isolate chloroplasts and then use the capacity of high-throughput sequencing to obtain complete genome sequences. We report our findings from studies of the latter approach, which used a simple chloroplast isolation procedure, multiply-primed rolling circle amplification of chloroplast DNA, Illumina Genome Analyzer II sequencing, and de novo assembly of paired-end sequence reads. Results A modified rapid chloroplast isolation protocol was used to obtain plant DNA that was enriched for chloroplast DNA, but nevertheless contained nuclear and mitochondrial DNA. Multiply-primed rolling circle amplification of this mixed template produced sufficient quantities of chloroplast DNA, even when the amount of starting material was small, and improved the template quality for Illumina Genome Analyzer II (hereafter Illumina GAII) sequencing. We demonstrate, using independent samples of karaka (Corynocarpus laevigatus), that there is high fidelity in the sequence obtained from this template. Although less than 20% of our sequenced reads could be mapped to chloroplast genome, it was relatively easy to assemble complete chloroplast genome sequences from the mixture of nuclear, mitochondrial and chloroplast reads. Conclusions We report successful whole genome sequencing of chloroplast DNA from karaka, obtained efficiently and with high fidelity. PMID:20920211

  14. Mouse strain specific gene expression differences for illumina microarray expression profiling in embryos

    PubMed Central

    2012-01-01

    Background In the field of mouse genetics the advent of technologies like microarray based expression profiling dramatically increased data availability and sensitivity, yet these advanced methods are often vulnerable to the unavoidable heterogeneity of in vivo material and might therefore reflect differentially expressed genes between mouse strains of no relevance to a targeted experiment. The aim of this study was not to elaborate on the usefulness of microarray analysis in general, but to expand our knowledge regarding this potential “background noise” for the widely used Illumina microarray platform surpassing existing data which focused primarily on the adult sensory and nervous system, by analyzing patterns of gene expression at different embryonic stages using wild type strains and modern transgenic models of often non-isogenic backgrounds. Results Wild type embryos of 11 mouse strains commonly used in transgenic and molecular genetic studies at three developmental time points were subjected to Illumina microarray expression profiling in a strain-by-strain comparison. Our data robustly reflects known gene expression patterns during mid-gestation development. Decreasing diversity of the input tissue and/or increasing strain diversity raised the sensitivity of the array towards the genetic background. Consistent strain sensitivity of some probes was attributed to genetic polymorphisms or probe design related artifacts. Conclusion Our study provides an extensive reference list of gene expression profiling background noise of value to anyone in the field of developmental biology and transgenic research performing microarray expression profiling with the widely used Illumina microarray platform. Probes identified as strain specific background noise further allow for microarray expression profiling on its own to be a valuable tool for establishing genealogies of mouse inbred strains. PMID:22583621

  15. Analysis of Illumina MiSeq Metabarcoding Data: Application to Benthic Indices for Environmental Monitoring.

    PubMed

    Aylagas, Eva; Rodríguez-Ezpeleta, Naiara

    2016-01-01

    This protocol details the analysis of Illumina MiSeq amplicon libraries derived from marine benthic macroinvertebrate samples and based on two barcodes of the mitochondrial cytochrome oxidase 1 (CO1) gene: a "short region," covered by overlapping forward and reverse reads and a "long region" for which forward and reverse reads do not overlap. Aside from providing guidelines for analyzing both types of amplicons, we show how amplicon reads can be used for the calculation of benthic indices for environmental monitoring. PMID:27460382

  16. Insights into the bovine rumen plasmidome

    PubMed Central

    Kav, Aya Brown; Sasson, Goor; Jami, Elie; Doron-Faigenboim, Adi; Benhar, Itai; Mizrahi, Itzhak

    2012-01-01

    Plasmids are self-replicating genetic elements capable of mobilization between different hosts. Plasmids often serve as mediators of lateral gene transfer, a process considered to be a strong and sculpting evolutionary force in microbial environments. Our aim was to characterize the overall plasmid population in the environment of the bovine rumen, which houses a complex and dense microbiota that holds enormous significance for humans. We developed a procedure for the isolation of total rumen plasmid DNA, termed rumen plasmidome, and subjected it to deep sequencing using the Illumina paired-end protocol and analysis using public and custom-made bioinformatics tools. A large number of plasmidome contigs aligned with plasmids of rumen bacteria isolated from different locations and at various time points, suggesting that not only the bacterial taxa, but also their plasmids, are defined by the ecological niche. The bacterial phylum distribution of the plasmidome was different from that of the rumen bacterial taxa. Nevertheless, both shared a dominance of the phyla Firmicutes, Bacteroidetes, and Proteobacteria. Evidently, the rumen plasmidome is of a highly mosaic nature that can cross phyla. Interestingly, when we compared the functional profile of the rumen plasmidome to two plasmid databases and two recently published rumen metagenomes, it became apparent that the rumen plasmidome codes for functions, which are enriched in the rumen ecological niche and could confer advantages to their hosts, suggesting that the functional profiles of mobile genetic elements are associated with their environment, as has been previously implied for viruses. PMID:22431592

  17. Illumina Production Sequencing at the DOE Joint Genome Institute - Workflow and Optimizations

    SciTech Connect

    Tarver, Angela; Fern, Alison; Diego, Matthew San; Kennedy, Megan; Zane, Matthew; Daum, Christopher; Hack, Christopher; Tang, Eric; Deshpande, Shweta; Cheng, Jan-Fang; Roberts, Simon; Alexandre, Melanie; Harmon-Smith, Miranda; Lucas, Susan

    2010-06-18

    The U.S. Department of Energy (DOE) Joint Genome Institute?s (JGI) Production Sequencing group is committed to the generation of high-quality genomic DNA sequence to support the DOE mission areas of renewable energy generation, global carbon management, and environmental characterization and clean-up. Within the JGI?s Production Sequencing group, the Illumina Genome Analyzer pipeline has been established as one of three sequencing platforms, along with Roche/454 and ABI/Sanger. Optimization of the Illumina pipeline has been ongoing with the aim of continual process improvement of the laboratory workflow. These process improvement projects are being led by the JGI?s Process Optimization, Sequencing Technologies, Instrumentation& Engineering, and the New Technology Production groups. Primary focus has been on improving the procedural ergonomics and the technicians? operating environment, reducing manually intensive technician operations with different tools, reducing associated production costs, and improving the overall process and generated sequence quality. The U.S. DOE JGI was established in 1997 in Walnut Creek, CA, to unite the expertise and resources of five national laboratories? Lawrence Berkeley, Lawrence Livermore, Los Alamos, Oak Ridge, and Pacific Northwest ? along with HudsonAlpha Institute for Biotechnology. JGI is operated by the University of California for the U.S. DOE.

  18. Analysis of Polygala tenuifolia Transcriptome and Description of Secondary Metabolite Biosynthetic Pathways by Illumina Sequencing.

    PubMed

    Tian, Hongling; Xu, Xiaoshuang; Zhang, Fusheng; Wang, Yaoqin; Guo, Shuhong; Qin, Xuemei; Du, Guanhua

    2015-01-01

    Radix polygalae, the dried roots of Polygala tenuifolia and P. sibirica, is one of the most well-known traditional Chinese medicinal plants. Radix polygalae contains various saponins, xanthones, and oligosaccharide esters and these compounds are responsible for several pharmacological properties. To provide basic breeding information, enhance molecular biological analysis, and determine secondary metabolite biosynthetic pathways of P. tenuifolia, we applied Illumina sequencing technology and de novo assembly. We also applied this technique to gain an overview of P. tenuifolia transcriptome from samples with different years. Using Illumina sequencing, approximately 67.2% of unique sequences were annotated by basic local alignment search tool similarity searches against public sequence databases. We classified the annotated unigenes by using Nr, Nt, GO, COG, and KEGG databases compared with NCBI. We also obtained many candidates CYP450s and UGTs by the analysis of genes in the secondary metabolite biosynthetic pathways, including putative terpenoid backbone and phenylpropanoid biosynthesis pathway. With this transcriptome sequencing, future genetic and genomics studies related to the molecular mechanisms associated with the chemical composition of P. tenuifolia may be improved. Genes involved in the enrichment of secondary metabolite biosynthesis-related pathways could enhance the potential applications of P. tenuifolia in pharmaceutical industries. PMID:26543847

  19. Analysis of Polygala tenuifolia Transcriptome and Description of Secondary Metabolite Biosynthetic Pathways by Illumina Sequencing

    PubMed Central

    Tian, Hongling; Xu, Xiaoshuang; Zhang, Fusheng; Wang, Yaoqin; Guo, Shuhong; Qin, Xuemei; Du, Guanhua

    2015-01-01

    Radix polygalae, the dried roots of Polygala tenuifolia and P. sibirica, is one of the most well-known traditional Chinese medicinal plants. Radix polygalae contains various saponins, xanthones, and oligosaccharide esters and these compounds are responsible for several pharmacological properties. To provide basic breeding information, enhance molecular biological analysis, and determine secondary metabolite biosynthetic pathways of P. tenuifolia, we applied Illumina sequencing technology and de novo assembly. We also applied this technique to gain an overview of P. tenuifolia transcriptome from samples with different years. Using Illumina sequencing, approximately 67.2% of unique sequences were annotated by basic local alignment search tool similarity searches against public sequence databases. We classified the annotated unigenes by using Nr, Nt, GO, COG, and KEGG databases compared with NCBI. We also obtained many candidates CYP450s and UGTs by the analysis of genes in the secondary metabolite biosynthetic pathways, including putative terpenoid backbone and phenylpropanoid biosynthesis pathway. With this transcriptome sequencing, future genetic and genomics studies related to the molecular mechanisms associated with the chemical composition of P. tenuifolia may be improved. Genes involved in the enrichment of secondary metabolite biosynthesis-related pathways could enhance the potential applications of P. tenuifolia in pharmaceutical industries. PMID:26543847

  20. Massively parallel multiplex DNA sequencing for specimen identification using an Illumina MiSeq platform.

    PubMed

    Shokralla, Shadi; Porter, Teresita M; Gibson, Joel F; Dobosz, Rafal; Janzen, Daniel H; Hallwachs, Winnie; Golding, G Brian; Hajibabaei, Mehrdad

    2015-01-01

    Genetic information is a valuable component of biosystematics, especially specimen identification through the use of species-specific DNA barcodes. Although many genomics applications have shifted to High-Throughput Sequencing (HTS) or Next-Generation Sequencing (NGS) technologies, sample identification (e.g., via DNA barcoding) is still most often done with Sanger sequencing. Here, we present a scalable double dual-indexing approach using an Illumina Miseq platform to sequence DNA barcode markers. We achieved 97.3% success by using half of an Illumina Miseq flowcell to obtain 658 base pairs of the cytochrome c oxidase I DNA barcode in 1,010 specimens from eleven orders of arthropods. Our approach recovers a greater proportion of DNA barcode sequences from individuals than does conventional Sanger sequencing, while at the same time reducing both per specimen costs and labor time by nearly 80%. In addition, the use of HTS allows the recovery of multiple sequences per specimen, for deeper analysis of genetic variation in target gene regions. PMID:25884109

  1. Diversity of Frankia in soil assessed by Illumina sequencing of nifH gene fragments.

    PubMed

    Rodriguez, David; Guerra, Trina M; Forstner, Michael R J; Hahn, Dittmar

    2016-09-01

    Targeted Illumina sequencing of nitrogenase reductase (nifH) gene fragments and analyses of pair-end reads through a modified QIIME pipeline were used to assess the diversity of the actinomyceteous genus Frankia in three soils. Soils were vegetated with host or non-host plants, and included locations in Illinois (ABA, host), Colorado (CoMt, non-host), and Wisconsin (FMWI, non-host). After filtering, seven unique sequences were recovered for soil ABA, six for CoMt, and four sequences for FMWI. These sequences were included in a Bayesian topology anchored by published sequence data from pure cultures of Frankia. Sequences from all three soils showed affinities to Frankia strains from both the Alnus and Elaeagnus host infection groups. Reads representing Casuarina-infective strains were not detected. Four sequences from soil CoMt and five sequences from soil ABA did not cluster, at 97% similarity, into a shared OTU that contained a cultured relative. These results demonstrate that targeted Illumina sequencing provides an efficient and economical method for assessing haplotype diversity of ecofunctional genes (e.g. nifH) at the genus level in microorganisms that perform important ecosystem functions. PMID:27485903

  2. Genome-Wide Microsatellite Identification in the Fungus Anisogramma anomala Using Illumina Sequencing and Genome Assembly

    PubMed Central

    Cai, Guohong; Leadbetter, Clayton W.; Muehlbauer, Megan F.; Molnar, Thomas J.; Hillman, Bradley I.

    2013-01-01

    High-throughput sequencing has been dramatically accelerating the discovery of microsatellite markers (also known as Simple Sequence Repeats). Both 454 and Illumina reads have been used directly in microsatellite discovery and primer design (the “Seq-to-SSR” approach). However, constraints of this approach include: 1) many microsatellite-containing reads do not have sufficient flanking sequences to allow primer design, and 2) difficulties in removing microsatellite loci residing in longer, repetitive regions. In the current study, we applied the novel “Seq-Assembly-SSR” approach to overcome these constraints in Anisogramma anomala. In our approach, Illumina reads were first assembled into a draft genome, and the latter was then used in microsatellite discovery. A. anomala is an obligate biotrophic ascomycete that causes eastern filbert blight disease of commercial European hazelnut. Little is known about its population structure or diversity. Approximately 26 M 146 bp Illumina reads were generated from a paired-end library of a fungal strain from Oregon. The reads were assembled into a draft genome of 333 Mb (excluding gaps), with contig N50 of 10,384 bp and scaffold N50 of 32,987 bp. A bioinformatics pipeline identified 46,677 microsatellite motifs at 44,247 loci, including 2,430 compound loci. Primers were successfully designed for 42,923 loci (97%). After removing 2,886 loci close to assembly gaps and 676 loci in repetitive regions, a genome-wide microsatellite database of 39,361 loci was generated for the fungus. In experimental screening of 236 loci using four geographically representative strains, 228 (96.6%) were successfully amplified and 214 (90.7%) produced single PCR products. Twenty-three (9.7%) were found to be perfect polymorphic loci. A small-scale population study using 11 polymorphic loci revealed considerable gene diversity. Clustering analysis grouped isolates of this fungus into two clades in accordance with their geographic origins

  3. The complete mitogenome of Cylindrus obtusus (Helicidae, Ariantinae) using Illumina next generation sequencing

    PubMed Central

    2012-01-01

    Background This study describes how the complete mitogenome of a terrestrial snail, Cylindrus obtusus (Draparnaud, 1805) was sequenced without PCRs from a collection specimen that had been in 70% ethanol for 8 years. The mitogenome was obtained with Illumina GAIIx shot gun sequencing. Although the used specimen was collected relatively recently and kept in a DNA-friendly preservative (not formalin as frequently used with old museum specimens), we believe that the exclusion of PCRs as facilitated by NGS (Next Generation Sequencing) removes a great obstacle in DNA sequencing of collection specimens. A brief comparison is made between our Illumina GAIIx approach and a similar study that made use of the Roche 454-FLX platform. Results The mtDNA sequence of C. obtusus is 14,610 bases in length (about 0.5 kb larger than other stylommatophoran mitogenomes reported hitherto) and contains the 37 genes (13 protein coding genes, two rRNAs and 22 tRNAs) typical for metazoans. Except for a swap between the position of tRNA-Pro and tRNA-Ala, the gene arrangement of C. obtusus is identical to that reported for Cepaea nemoralis. The 'aberrant' rearrangement of tRNA-Thr and COIII compared to that of other Sigmurethra (and the majority of gastropods), is not unique for C. nemoralis (subfamily Helicinae), but is also shown to occur in C. obtusus (subfamily Ariantinae) and might be a synapomorphy for the family Helicidae. Conclusions Natural history collections potentially harbor a wealth of information for the field of evolutionary genetics, but it can be difficult to amplify DNA from such specimens (due to DNA degradation for instance). Because NGS techniques do not rely on primer-directed amplification (PCR) and allow DNA to be fragmented (DNA gets sheared during library preparation), NGS could be a valuable tool for retrieving DNA sequence data from such specimens. A comparison between Illumina GAIIx and the Roche 454 platform suggests that the former might be more suited for de

  4. The solvent action of sodium hypochlorite on bovine tendon collagen, bovine pulp, and bovine gingiva.

    PubMed

    Nakamura, H; Asai, K; Fujita, H; Nakazato, H; Nishimura, Y; Furuse, Y; Sahashi, E

    1985-09-01

    The purpose of this study was to determine the optimum temperature and concentration of sodium hypochlorite solution required to dissolve bovine tendon collagen, pulp, and gingiva. The 10% concentration of sodium hypochlorite solution at 37 degrees C was found to be most effective in dissolving bovine tendon collagen, pulp, and gingiva. Sodium hypochlorite solution was more effective in dissolving bovine pulp or tendon collagen than in dissolving bovine gingiva. PMID:3862046

  5. Analyzing Illumina Gene Expression Microarray Data Obtained From Human Whole Blood Cell and Blood Monocyte Samples.

    PubMed

    Teumer, Alexander; Schurmann, Claudia; Schillert, Arne; Schramm, Katharina; Ziegler, Andreas; Prokisch, Holger

    2016-01-01

    Microarray profiling of gene expression is widely applied to studies in molecular biology and functional genomics. Experimental and technical variations make not only the statistical analysis of single studies but also meta-analyses of different studies very challenging. Here, we describe the analytical steps required to substantially reduce the variations of gene expression data without affecting true effect sizes. A software pipeline has been established using gene expression data from a total of 3358 whole blood cell and blood monocyte samples, all from three German population-based cohorts, measured on the Illumina HumanHT-12 v3 BeadChip array. In summary, adjustment for a few selected technical factors greatly improved reliability of gene expression analyses. Such adjustments are particularly required for meta-analyses of different studies. PMID:26614070

  6. Removing unwanted variation in a differential methylation analysis of Illumina HumanMethylation450 array data

    PubMed Central

    Maksimovic, Jovana; Gagnon-Bartsch, Johann A.; Speed, Terence P.; Oshlack, Alicia

    2015-01-01

    Due to their relatively low-cost per sample and broad, gene-centric coverage of CpGs across the human genome, Illumina's 450k arrays are widely used in large scale differential methylation studies. However, by their very nature, large studies are particularly susceptible to the effects of unwanted variation. The effects of unwanted variation have been extensively documented in gene expression array studies and numerous methods have been developed to mitigate these effects. However, there has been much less research focused on the appropriate methodology to use for accounting for unwanted variation in methylation array studies. Here we present a novel 2-stage approach using RUV-inverse in a differential methylation analysis of 450k data and show that it outperforms existing methods. PMID:25990733

  7. Development of genomic microsatellites in Gleditsia triacanthos (Fabaceae) using Illumina sequencing1

    PubMed Central

    Owusu, Sandra A.; Staton, Margaret; Jennings, Tara N.; Schlarbaum, Scott; Coggeshall, Mark V.; Romero-Severson, Jeanne; Carlson, John E.; Gailing, Oliver

    2013-01-01

    • Premise of the study: Fourteen genomic microsatellite markers were developed and characterized in honey locust, Gleditsia triacanthos, using Illumina sequencing. Due to their high variability, these markers can be applied in analyses of genetic diversity and structure, and in mating system and gene flow studies. • Methods and Results: Thirty-six individuals from across the species range were included in a genetic diversity analysis and yielded three to 20 alleles per locus. Observed heterozygosity and expected heterozygosity ranged from 0.214 to 0.944 and from 0.400 to 0.934, respectively, with minimal occurrence of null alleles. Regular segregation of maternal alleles was observed at seven loci and moderate segregation distortion at four of 11 loci that were heterozygous in the seed parent. • Conclusions: Honey locust is an important agroforestry tree capable of very fast growth and tolerance of poor site conditions. This is the first report of genomic microsatellites for this species. PMID:25202504

  8. Illumina MiSeq sequencing disfavours a sequence motif in the GFP reporter gene

    PubMed Central

    Van den Hoecke, Silvie; Verhelst, Judith; Saelens, Xavier

    2016-01-01

    Green fluorescent protein (GFP) is one of the most used reporter genes. We have used next-generation sequencing (NGS) to analyse the genetic diversity of a recombinant influenza A virus that expresses GFP and found a remarkable coverage dip in the GFP coding sequence. This coverage dip was present when virus-derived RT-PCR product or the parental plasmid DNA was used as starting material for NGS and regardless of whether Nextera XT transposase or Covaris shearing was used for DNA fragmentation. Therefore, the sequence coverage dip in the GFP coding sequence was not the result of emerging GFP mutant viruses or a bias introduced by Nextera XT fragmentation. Instead, we found that the Illumina MiSeq sequencing method disfavours the ‘CCCGCC’ motif in the GFP coding sequence. PMID:27193250

  9. Illumina MiSeq sequencing disfavours a sequence motif in the GFP reporter gene.

    PubMed

    Van den Hoecke, Silvie; Verhelst, Judith; Saelens, Xavier

    2016-01-01

    Green fluorescent protein (GFP) is one of the most used reporter genes. We have used next-generation sequencing (NGS) to analyse the genetic diversity of a recombinant influenza A virus that expresses GFP and found a remarkable coverage dip in the GFP coding sequence. This coverage dip was present when virus-derived RT-PCR product or the parental plasmid DNA was used as starting material for NGS and regardless of whether Nextera XT transposase or Covaris shearing was used for DNA fragmentation. Therefore, the sequence coverage dip in the GFP coding sequence was not the result of emerging GFP mutant viruses or a bias introduced by Nextera XT fragmentation. Instead, we found that the Illumina MiSeq sequencing method disfavours the 'CCCGCC' motif in the GFP coding sequence. PMID:27193250

  10. The complete mitochondrial genome of the ruby-topaz hummingbird Chrysolampis mosquitus through Illumina sequencing.

    PubMed

    Souto, Helena Magarinos; Ruschi, Piero Angeli; Furtado, Carolina; Jennings, W Bryan; Prosdocimi, Francisco

    2016-01-01

    The complete mitochondrial genome of the Ruby-Topaz Hummingbird, Chrysolampis mosquitus, was determined using 1/11 of an Illumina Hi-seq lane ran with a Nextera kit. We assembled the mitogenome in a two-step approach using both (i) de novo (SOAPdenovo-Trans) and (ii) reference-based (MITObim) genome assembly software. A circular molecule containing 17,767 bp was assembled. As expected for most vertebrates, the C. mosquitus mitogenome contained 13 protein-coding genes, 22 transfer RNA, 2 ribosomal RNA genes, and 1 non-coding control region. We assembled the whole mitogenome using 0.45% of the total amount of reads and produced a high-coverage mitochondrial genome (>1000×). We deposited the assembled mitogenome into GenBank (accession number KJ619585). PMID:24841437

  11. Quality Control for the Illumina HumanExome BeadChip.

    PubMed

    Igo, Robert P; Cooke Bailey, Jessica N; Romm, Jane; Haines, Jonathan L; Wiggs, Janey L

    2016-01-01

    The Illumina HumanExome BeadChip and other exome-based genotyping arrays offer inexpensive genotyping of some 240,000 mostly nonsynonymous coding variants across the human genome. The HumanExome chip, with its highly non-uniform distribution of markers and emphasis on rare coding variants, presents some unique challenges for quality control (QC) and data cleaning. Here, we describe QC procedures for HumanExome data, with examples of challenges specific to exome arrays from our experience cleaning a data set of ∼7,500 samples from the NEIGHBORHOOD Consortium. We focus on standard procedures for QC of genome-wide array data including genotype calling, sex verification, sample identity verification, relationship checking, and population structure that are complicated by the HumanExome panel's enrichment in rare, exonic variation. © 2016 by John Wiley & Sons, Inc. PMID:27367164

  12. Assessing Illumina technology for the high-throughput sequencing of bacteriophage genomes

    PubMed Central

    Rihtman, Branko; Meaden, Sean; Clokie, Martha R.J.; Koskella, Britt

    2016-01-01

    Bacteriophages are the most abundant biological entities on the planet, playing crucial roles in the shaping of bacterial populations. Phages have smaller genomes than their bacterial hosts, yet there are currently fewer fully sequenced phage than bacterial genomes. We assessed the suitability of Illumina technology for high-throughput sequencing and subsequent assembly of phage genomes. In silico datasets reveal that 30× coverage is sufficient to correctly assemble the complete genome of ~98.5% of known phages, with experimental data confirming that the majority of phage genomes can be assembled at 30× coverage. Furthermore, in silico data demonstrate it is possible to co-sequence multiple phages from different hosts, without introducing assembly errors. PMID:27280068

  13. Metagenomic study of the oral microbiota by Illumina high-throughput sequencing

    PubMed Central

    Lazarevic, Vladimir; Whiteson, Katrine; Huse, Susan; Hernandez, David; Farinelli, Laurent; Østerås, Magne; Schrenzel, Jacques; François, Patrice

    2013-01-01

    To date, metagenomic studies have relied on the utilization and analysis of reads obtained using 454 pyrosequencing to replace conventional Sanger sequencing. After extensively scanning the 16S ribosomal RNA (rRNA) gene, we identified the V5 hypervariable region as a short region providing reliable identification of bacterial sequences available in public databases such as the Human Oral Microbiome Database. We amplified samples from the oral cavity of three healthy individuals using primers covering an ~82-base segment of the V5 loop, and sequenced using the Illumina technology in a single orientation. We identified 135 genera or higher taxonomic ranks from the resulting 1,373,824 sequences. While the abundances of the most common phyla (Firmicutes, Proteobacteria, Actinobacteria, Fusobacteria and TM7) are largely comparable to previous studies, Bacteroidetes were less present. Potential sources for this difference include classification bias in this region of the 16S rRNA gene, human sample variation, sample preparation and primer bias. Using an Illumina sequencing approach, we achieved a much greater depth of coverage than previous oral microbiota studies, allowing us to identify several taxa not yet discovered in these types of samples, and to assess that at least 30,000 additional reads would be required to identify only one additional phylotype. The evolution of high-throughput sequencing technologies, and their subsequent improvements in read length enable the utilization of different platforms for studying communities of complex flora. Access to large amounts of data is already leading to a better representation of sample diversity at a reasonable cost. PMID:19796657

  14. Protein Crystal Bovine Insulin

    NASA Technical Reports Server (NTRS)

    1991-01-01

    The comparison of protein crystal, Bovine Insulin space-grown (left) and earth-grown (right). Facilitates the incorporation of glucose into cells. In diabetics, there is either a decrease in or complete lack of insulin, thereby leading to several harmful complications. Principal Investigator is Larry DeLucas.

  15. Bovine Spongiform Encephalopathy

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bovine spongiform encephalopathy (BSE), also referred to as “mad cow disease” is a chronic, non-febrile, neuro-degenerative disease affecting the central nervous system. The transmissible spongiform encephalopathies (TSEs) of domestic animals, of which BSE is a member includes scrapie of sheep...

  16. [Bovine spongiform encephalopathy].

    PubMed

    Suárez Fernández, G

    2001-01-01

    An histórical and conceptual review is made about Bovine Spongiform Encephalopathy or mad cows disease and an epidemiological analysis as a present and future health problem. This analysis of BSE should not be negative, considering the truths that we know today. PMID:11783042

  17. BOVINE VIRAL DIARRHEA VIRUSES

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bovine viral diarrhea virus (BVDV) is an umbrella term for two species of viruses, BVDV1 and BVDV2, within the Pestivirus genus of the Flavivirus family. BVDV viruses are further subclassified as cytopathic and noncytopathic based on their activity in cultured epithelial cells. Noncytopathic BVDV p...

  18. Bovine milk exosome proteome

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Exosomes are 40-100 nm membrane vesicles of endocytic origin and are found in blood, urine, amniotic fluid, bronchoalveolar lavage (BAL) fluid, as well as human and bovine milk. Exosomes are extracellular organelles important in intracellular communication/signaling, immune function, and biomarkers ...

  19. Genotyping bovine coronaviruses.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bovine coronaviruses (BoCV) are enveloped, single-stranded, positive-sense RNA viruses of the Coronaviridae family. Infection is associated with enteritis and pneumonia in calves and Winter Dysentery in adult cattle. Strains, isolated more than 50 years ago, are used in vaccines and as laboratory ...

  20. Use of the Illumina GoldenGate assay for single nucleotide polymorphism (SNP) genotyping in cereal crops

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The highly parallel genotyping assay, such as the GoldenGate assay developed by Illumina, capable of interrogating up to 3,072 single nucleotide polymorphisms (SNPs) simultaneously, has greatly facilitated the genome-wide studies particularly for crops with large and complex genome structures. In th...

  1. The First Illumina-Based De Novo Transcriptome Sequencing and Analysis of Safflower Flowers

    PubMed Central

    Lulin, Huang; Xiao, Yang; Pei, Sun; Wen, Tong; Shangqin, Hu

    2012-01-01

    Background The safflower, Carthamus tinctorius L., is a worldwide oil crop, and its flowers, which have a high flavonoid content, are an important medicinal resource against cardiovascular disease in traditional medicine. Because the safflower has a large and complex genome, the development of its genomic resources has been delayed. Second-generation Illumina sequencing is now an efficient route for generating an enormous volume of sequences that can represent a large number of genes and their expression levels. Methodology/Principal Findings To investigate the genes and pathways that might control flavonoids and other secondary metabolites in the safflower, we used Illumina sequencing to perform a de novo assembly of the safflower tubular flower tissue transcriptome. We obtained a total of 4.69 Gb in clean nucleotides comprising 52,119,104 clean sequencing reads, 195,320 contigs, and 120,778 unigenes. Based on similarity searches with known proteins, we annotated 70,342 of the unigenes (about 58% of the identified unigenes) with cut-off E-values of 10−5. In total, 21,943 of the safflower unigenes were found to have COG classifications, and BLAST2GO assigned 26,332 of the unigenes to 1,754 GO term annotations. In addition, we assigned 30,203 of the unigenes to 121 KEGG pathways. When we focused on genes identified as contributing to flavonoid biosynthesis and the biosynthesis of unsaturated fatty acids, which are important pathways that control flower and seed quality, respectively, we found that these genes were fairly well conserved in the safflower genome compared to those of other plants. Conclusions/Significance Our study provides abundant genomic data for Carthamus tinctorius L. and offers comprehensive sequence resources for studying the safflower. We believe that these transcriptome datasets will serve as an important public information platform to accelerate studies of the safflower genome, and may help us define the mechanisms of flower tissue

  2. Comparison of the Equine Reference Sequence with Its Sanger Source Data and New Illumina Reads

    PubMed Central

    Rebolledo-Mendez, Jovan; Hestand, Matthew S.; Coleman, Stephen J.; Zeng, Zheng; Orlando, Ludovic; MacLeod, James N.; Kalbfleisch, Ted

    2015-01-01

    The reference assembly for the domestic horse, EquCab2, published in 2009, was built using approximately 30 million Sanger reads from a Thoroughbred mare named Twilight. Contiguity in the assembly was facilitated using nearly 315 thousand BAC end sequences from Twilight’s half brother Bravo. Since then, it has served as the foundation for many genome-wide analyses that include not only the modern horse, but ancient horses and other equid species as well. As data mapped to this reference has accumulated, consistent variation between mapped datasets and the reference, in terms of regions with no read coverage, single nucleotide variants, and small insertions/deletions have become apparent. In many cases, it is not clear whether these differences are the result of true sequence variation between the research subjects’ and Twilight’s genome or due to errors in the reference. EquCab2 is regarded as “The Twilight Assembly.” The objective of this study was to identify inconsistencies between the EquCab2 assembly and the source Twilight Sanger data used to build it. To that end, the original Sanger and BAC end reads have been mapped back to this equine reference and assessed with the addition of approximately 40X coverage of new Illumina Paired-End sequence data. The resulting mapped datasets identify those regions with low Sanger read coverage, as well as variation in genomic content that is not consistent with either the original Twilight Sanger data or the new genomic sequence data generated from Twilight on the Illumina platform. As the haploid EquCab2 reference assembly was created using Sanger reads derived largely from a single individual, the vast majority of variation detected in a mapped dataset comprised of those same Sanger reads should be heterozygous. In contrast, homozygous variations would represent either errors in the reference or contributions from Bravo's BAC end sequences. Our analysis identifies 720,843 homozygous discrepancies between new

  3. Successful Recovery of Nuclear Protein-Coding Genes from Small Insects in Museums Using Illumina Sequencing.

    PubMed

    Kanda, Kojun; Pflug, James M; Sproul, John S; Dasenko, Mark A; Maddison, David R

    2015-01-01

    In this paper we explore high-throughput Illumina sequencing of nuclear protein-coding, ribosomal, and mitochondrial genes in small, dried insects stored in natural history collections. We sequenced one tenebrionid beetle and 12 carabid beetles ranging in size from 3.7 to 9.7 mm in length that have been stored in various museums for 4 to 84 years. Although we chose a number of old, small specimens for which we expected low sequence recovery, we successfully recovered at least some low-copy nuclear protein-coding genes from all specimens. For example, in one 56-year-old beetle, 4.4 mm in length, our de novo assembly recovered about 63% of approximately 41,900 nucleotides in a target suite of 67 nuclear protein-coding gene fragments, and 70% using a reference-based assembly. Even in the least successfully sequenced carabid specimen, reference-based assembly yielded fragments that were at least 50% of the target length for 34 of 67 nuclear protein-coding gene fragments. Exploration of alternative references for reference-based assembly revealed few signs of bias created by the reference. For all specimens we recovered almost complete copies of ribosomal and mitochondrial genes. We verified the general accuracy of the sequences through comparisons with sequences obtained from PCR and Sanger sequencing, including of conspecific, fresh specimens, and through phylogenetic analysis that tested the placement of sequences in predicted regions. A few possible inaccuracies in the sequences were detected, but these rarely affected the phylogenetic placement of the samples. Although our sample sizes are low, an exploratory regression study suggests that the dominant factor in predicting success at recovering nuclear protein-coding genes is a high number of Illumina reads, with success at PCR of COI and killing by immersion in ethanol being secondary factors; in analyses of only high-read samples, the primary significant explanatory variable was body length, with small beetles

  4. Transcriptomic Analysis of Grapevine (cv. Summer Black) Leaf, Using the Illumina Platform

    PubMed Central

    Pervaiz, Tariq; Haifeng, Jia; Salman Haider, Muhammad; Cheng, Zhang; Cui, Mengjie; Wang, Mengqi; Cui, Liwen; Wang, Xicheng; Fang, Jinggui

    2016-01-01

    Proceeding to illumina sequencing, determining RNA integrity numbers for poly RNA were separated from each of the four developmental stages of cv. Summer Black leaves by using Illumina HiSeq™ 2000. The sums of 272,941,656 reads were generated from vitis vinifera leaf at four different developmental stages, with more than 27 billion nucleotides of the sequence data. At each growth stage, RNA samples were indexed through unique nucleic acid identifiers and sequenced. KEGG annotation results depicted that the highest number of transcripts in 2,963 (2Avs4A) followed by 1Avs4A (2,920), and 3Avs4A (2,294) out of 15,614 (71%) transcripts were recorded. In comparison, a total of 1,532 transcripts were annotated in GOs, including Cellular component, with the highest number in “Cell part” 251 out of 353 transcripts (71.1%), followed by intracellular organelle 163 out of 353 transcripts (46.2%), while in molecular function and metabolic process 375 out of 525 (71.4%) transcripts, multicellular organism process 40 out of 525 (7.6%) transcripts in biological process were most common in 1Avs2A. While in case of 1Avs3A, cell part 476 out of 662 transcripts (71.9%), and membrane-bounded organelle 263 out of 662 transcripts (39.7%) were recorded in Cellular component. In the grapevine transcriptome, during the initial stages of leaf development 1Avs2A showed single transcript was down-regulated and none of them were up-regulated. While in comparison of 1A to 3A showed one up-regulated (photosystem II reaction center protein C) and one down regulated (conserved gene of unknown function) transcripts, during the hormone regulating pathway namely SAUR-like auxin-responsive protein family having 2 up-regulated and 7 down-regulated transcripts, phytochrome-associated protein showed 1 up-regulated and 9 down-regulated transcripts, whereas genes associated with the Leucine-rich repeat protein kinase family protein showed 7 up-regulated and 1 down-regulated transcript, meanwhile Auxin

  5. Successful Recovery of Nuclear Protein-Coding Genes from Small Insects in Museums Using Illumina Sequencing

    PubMed Central

    Dasenko, Mark A.

    2015-01-01

    In this paper we explore high-throughput Illumina sequencing of nuclear protein-coding, ribosomal, and mitochondrial genes in small, dried insects stored in natural history collections. We sequenced one tenebrionid beetle and 12 carabid beetles ranging in size from 3.7 to 9.7 mm in length that have been stored in various museums for 4 to 84 years. Although we chose a number of old, small specimens for which we expected low sequence recovery, we successfully recovered at least some low-copy nuclear protein-coding genes from all specimens. For example, in one 56-year-old beetle, 4.4 mm in length, our de novo assembly recovered about 63% of approximately 41,900 nucleotides in a target suite of 67 nuclear protein-coding gene fragments, and 70% using a reference-based assembly. Even in the least successfully sequenced carabid specimen, reference-based assembly yielded fragments that were at least 50% of the target length for 34 of 67 nuclear protein-coding gene fragments. Exploration of alternative references for reference-based assembly revealed few signs of bias created by the reference. For all specimens we recovered almost complete copies of ribosomal and mitochondrial genes. We verified the general accuracy of the sequences through comparisons with sequences obtained from PCR and Sanger sequencing, including of conspecific, fresh specimens, and through phylogenetic analysis that tested the placement of sequences in predicted regions. A few possible inaccuracies in the sequences were detected, but these rarely affected the phylogenetic placement of the samples. Although our sample sizes are low, an exploratory regression study suggests that the dominant factor in predicting success at recovering nuclear protein-coding genes is a high number of Illumina reads, with success at PCR of COI and killing by immersion in ethanol being secondary factors; in analyses of only high-read samples, the primary significant explanatory variable was body length, with small beetles

  6. SEME: A Fast Mapper of Illumina Sequencing Reads with Statistical Evaluation

    PubMed Central

    Chen, Shijian; Wang, Anqi

    2013-01-01

    Abstract Mapping reads to a reference genome is a routine yet computationally intensive task in research based on high-throughput sequencing. In recent years, the sequencing reads of the Illumina platform have become longer and their quality scores higher. According to our calculation, this allows perfect k-mer seed match for almost all reads when a close reference genome is available subject to reasonable specificity. Our other observation is that the majority reads contain at most one short INDEL polymorphism. Based on these observations, we propose a fast-mapping approach, referred to as “SEME,” which has two core steps: First it scans a read sequentially in a specific order for a k-mer exact match seed; next it extends the alignment on both sides allowing, at most, one short INDEL each using a novel method called “auto-match function.” We decompose the evaluation of the sensitivity and specificity into two parts corresponding to the seed and extension step, and the composite result provides an approximate overall reliability estimate of each mapping. We compare SEME with some existing mapping methods on several datasets, and SEME shows better performance in terms of both running time and mapping rates. PMID:24195707

  7. ENmix: a novel background correction method for Illumina HumanMethylation450 BeadChip

    PubMed Central

    Xu, Zongli; Niu, Liang; Li, Leping; Taylor, Jack A.

    2016-01-01

    The Illumina HumanMethylation450 BeadChip is increasingly utilized in epigenome-wide association studies, however, this array-based measurement of DNA methylation is subject to measurement variation. Appropriate data preprocessing to remove background noise is important for detecting the small changes that may be associated with disease. We developed a novel background correction method, ENmix, that uses a mixture of exponential and truncated normal distributions to flexibly model signal intensity and uses a truncated normal distribution to model background noise. Depending on data availability, we employ three approaches to estimate background normal distribution parameters using (i) internal chip negative controls, (ii) out-of-band Infinium I probe intensities or (iii) combined methylated and unmethylated intensities. We evaluate ENmix against other available methods for both reproducibility among duplicate samples and accuracy of methylation measurement among laboratory control samples. ENmix out-performed other background correction methods for both these measures and substantially reduced the probe-design type bias between Infinium I and II probes. In reanalysis of existing EWAS data we show that ENmix can identify additional CpGs, and results in smaller P-value estimates for previously-validated CpGs. We incorporated the method into R package ENmix, which is freely available from Bioconductor website. PMID:26384415

  8. Illumina-based analysis of core actinobacteriome in roots, stems, and grains of rice.

    PubMed

    Wang, Wenfeng; Zhai, Yanyan; Cao, Lixiang; Tan, Hongming; Zhang, Renduo

    2016-09-01

    Seed-borne microbiota can transmit vertically from generation to generation and be a favour mutualism between the endosymbionts and hosts. The aim of this study was to investigate the rice-associated actinobacterial taxa in roots, stems, and grains and explore vertically transmitted core actinobacteriome of rice plants. Illumina sequencing analyses of samples of rice grains, stems, and roots showed that the roots contained the most diverse actinobacteria among the tissues. The grains contained 78 actinobacterial operational taxonomic units (OTUs), among which 44 were shared with those in the stems, 30 shared with those in the roots. The coexisted OTUs in the three types of samples mainly belong to genera of Pseudonocardia, Dietzia, Nocardioides, Streptomyces, Mycobacterium, Corynebacterium, Citricoccus, Salinibacterium, and Agrococcus, and other unclassified taxa. The dominant actinobacterial genera Pseudonocardia and Dietzia in the stems and roots were still detected in relatively high abundance in the grains. The Streptomyces isolated from surface sterilized grains could improve nitrogen use efficiency of rice seedlings and the resistance to rice blast fungus. The results suggested that the rice grains contained diverse actinobacterial taxa deriving from stems and roots and showed intimate correlation with the host plants. PMID:27393994

  9. Diversity of prokaryotic community at a shallow marine hydrothermal site elucidated by Illumina sequencing technology.

    PubMed

    Lentini, Valeria; Gugliandolo, Concetta; Bunk, Boyke; Overmann, Jörg; Maugeri, Teresa L

    2014-10-01

    To investigate the prokaryotic community structure and composition in an active hydrothermal site, named Black Point, off Panarea Island (Eolian Islands, Italy), we examined sediment and fluid samples, differing in temperature, by a massive parallel sequencing (Illumina) technique targeting the V3 region of the 16S rRNA gene. The used technique enabled us to detect a greater prokaryotic diversity than that until now observed and to reveal also microorganisms occurring at very low abundance (≤0.01 %). Most of sequences were assigned to Bacteria while Archaea were a minor component of the microbial community in both low- and high-temperature samples. Proteobacteria (mainly consisting of Alpha-, Gamma-, and Epsilonproteobacteria) dominated among all samples followed by Actinobacteria and Bacteroidetes. Analyzed DNA obtained from samples taken at different temperatures indicated the presence of members of different dominant genera. The main differences were observed between sediment samples where Rhodovulum and Thiohalospira prevailed at high temperature, while Thalassomonas and Sulfurimonas at low temperature. Chlorobium, Acinetobacter, Sulfurimonas, and Brevundimonas were abundant in both low- and high-temperature fluid samples. Euryarchaeota dominated the archaeal community in all samples. Classes of Euryarchaeota embracing hyperthermophilic members (Thermococci and Thermoplasmata) and of Crenarchaeota (Thermoprotei) were more abundant in high-temperature samples. A great number of sequences referred to Bacteria and Archaea still remained unaffiliated, indicating that Black Point site represents a rich source of so-far uncharted prokaryotic diversity. PMID:24849732

  10. Whole-transcriptome sequencing of Pinellia ternata using the Illumina platform.

    PubMed

    Huang, X; Jing, Y; Liu, D J; Yang, B Y; Chen, H; Li, M

    2016-01-01

    Pinelliae rhizoma is the dried tuber of Pinellia ternata (Thunb.) Breit., and has been used for thousands of years as a traditional Chinese medicine. However, its genomic background is little known. With the development of high-throughput genomic sequencing, it is now easy and cheap to obtain genomic information. In this study, 193,032,910 high-quality clean reads were generated using the Illumina Hiseq 2000 platform. A total of 53,544 unigenes were identified from the contigs assembled. Functional annotation analysis annotated 37,318, 27,697, 23,043, 22,869, 23,328, and 27,415 unigenes. KEGG analysis revealed that five pathways (169 genes) were associated with alkaloid synthesis, 201 unigenes were related to fatty acid biosynthesis (ko00061), and 133 unigenes were involved in the biosynthesis of unsaturated fatty acids (ko01040). In addition, 6703 simple sequence repeats were designed based on the unigene sequences for screening germplasm resources in the future. These data are a valuable resource for genomic studies on Pinellia plants. PMID:27420994

  11. 32 species validation of a new Illumina paired-end approach for the development of microsatellites.

    PubMed

    Lance, Stacey L; Love, Cara N; Nunziata, Schyler O; O'Bryhim, Jason R; Scott, David E; Flynn, R Wesley; Jones, Kenneth L

    2013-01-01

    Development and optimization of novel species-specific microsatellites, or simple sequence repeats (SSRs) remains an important step for studies in ecology, evolution, and behavior. Numerous approaches exist for identifying new SSRs that vary widely in terms of both time and cost investments. A recent approach of using paired-end Illumina sequence data in conjunction with the bioinformatics pipeline, PAL_FINDER, has the potential to substantially reduce the cost and labor investment while also improving efficiency. However, it does not appear that the approach has been widely adopted, perhaps due to concerns over its broad applicability across taxa. Therefore, to validate the utility of the approach we developed SSRs for 32 species representing 30 families, 25 orders, 11 classes, and six phyla and optimized SSRs for 13 of the species. Overall the IPE method worked extremely well and we identified 1000s of SSRs for all species (mean = 128,485), with 17% of loci being potentially amplifiable loci, and 25% of these met our most stringent criteria designed to that avoid SSRs associated with repetitive elements. Approximately 61% of screened primers yielded strong amplification of a single locus. PMID:24312368

  12. A glance at quality score: implication for de novo transcriptome reconstruction of Illumina reads.

    PubMed

    Mbandi, Stanley Kimbung; Hesse, Uljana; Rees, D Jasper G; Christoffels, Alan

    2014-01-01

    Downstream analyses of short-reads from next-generation sequencing platforms are often preceded by a pre-processing step that removes uncalled and wrongly called bases. Standard approaches rely on their associated base quality scores to retain the read or a portion of it when the score is above a predefined threshold. It is difficult to differentiate sequencing error from biological variation without a reference using quality scores. The effects of quality score based trimming have not been systematically studied in de novo transcriptome assembly. Using RNA-Seq data produced from Illumina, we teased out the effects of quality score based filtering or trimming on de novo transcriptome reconstruction. We showed that assemblies produced from reads subjected to different quality score thresholds contain truncated and missing transfrags when compared to those from untrimmed reads. Our data supports the fact that de novo assembling of untrimmed data is challenging for de Bruijn graph assemblers. However, our results indicates that comparing the assemblies from untrimmed and trimmed read subsets can suggest appropriate filtering parameters and enable selection of the optimum de novo transcriptome assembly in non-model organisms. PMID:24575122

  13. Method to detect differentially methylated loci with case-control designs using Illumina arrays.

    PubMed

    Wang, Shuang

    2011-11-01

    It is now understood that many human cancer types are the result of the accumulation of both genetic and epigenetic changes. DNA methylation is a molecular modification of DNA that is crucial for normal development. Genes that are rich in CpG dinucleotides are usually not methylated in normal tissues, but are frequently hypermethylated in cancer. With the advent of high-throughput platforms, large-scale structure of genomic methylation patterns is available through genome-wide scans and tremendous amount of DNA methylation data have been recently generated. However, sophisticated statistical methods to handle complex DNA methylation data are very limited. Here, we developed a likelihood based Uniform-Normal-mixture model to select differentially methylated loci between case and control groups using Illumina arrays. The idea is to model the data as three types of methylation loci, one unmethylated, one completely methylated, and one partially methylated. A three-component mixture model with two Uniform distributions and one truncated normal distribution was used to model the three types. The mixture probabilities and the mean of the normal distribution were used to make inference about differentially methylated loci. Through extensive simulation studies, we demonstrated the feasibility and power of the proposed method. An application to a recently published study on ovarian cancer identified several methylation loci that are missed by the existing method. PMID:21818777

  14. Characterization of 13 microsatellite markers for Calochortus gunnisonii (Liliaceae) from Illumina MiSeq sequencing1

    PubMed Central

    Fuller, Ryan S.; Frietze, Seth; McGlaughlin, Mitchell E.

    2015-01-01

    Premise of the study: Microsatellite primers were designed for Calochortus gunnisonii (Liliaceae), a montane lily species of the central and southern Rocky Mountains, using next-generation DNA sequencing. The markers will be used to investigate population structure, genetic diversity, and demographic history. Methods and Results: Thirteen polymorphic microsatellite loci were isolated from C. gunnisonii using Illumina MiSeq next-generation DNA sequencing and bioinformatic screening. The mean number of alleles per locus ranged from 4.15 to 5.92 (avg. = 4.97). Observed and expected heterozygosity ranged from 0.077 to 0.871 and 0.213 to 0.782, respectively. The primers were also tested for cross-species amplification value with C. flexuosus, C. nuttallii, C. kennedyi var. kennedyi, and C. subalpinus. Conclusions: These primers will be useful for genetic and evolutionary studies across C. gunnisonii’s range within the southern and central Rocky Mountains. Furthermore, these markers have proven valuable for cross-species amplifications within Calochortus. PMID:26312200

  15. Sonication-based isolation and enrichment of Chlorella protothecoides chloroplasts for illumina genome sequencing

    SciTech Connect

    Angelova, Angelina; Park, Sang-Hycuk; Kyndt, John; Fitzsimmons, Kevin; Brown, Judith K

    2013-09-01

    With the increasing world demand for biofuel, a number of oleaginous algal species are being considered as renewable sources of oil. Chlorella protothecoides Krüger synthesizes triacylglycerols (TAGs) as storage compounds that can be converted into renewable fuel utilizing an anabolic pathway that is poorly understood. The paucity of algal chloroplast genome sequences has been an important constraint to chloroplast transformation and for studying gene expression in TAGs pathways. In this study, the intact chloroplasts were released from algal cells using sonication followed by sucrose gradient centrifugation, resulting in a 2.36-fold enrichment of chloroplasts from C. protothecoides, based on qPCR analysis. The C. protothecoides chloroplast genome (cpDNA) was determined using the Illumina HiSeq 2000 sequencing platform and found to be 84,576 Kb in size (8.57 Kb) in size, with a GC content of 30.8 %. This is the first report of an optimized protocol that uses a sonication step, followed by sucrose gradient centrifugation, to release and enrich intact chloroplasts from a microalga (C. prototheocoides) of sufficient quality to permit chloroplast genome sequencing with high coverage, while minimizing nuclear genome contamination. The approach is expected to guide chloroplast isolation from other oleaginous algal species for a variety of uses that benefit from enrichment of chloroplasts, ranging from biochemical analysis to genomics studies.

  16. Bacterial community compositions in sediment polluted by perfluoroalkyl acids (PFAAs) using Illumina high-throughput sequencing.

    PubMed

    Sun, Yajun; Wang, Tieyu; Peng, Xiawei; Wang, Pei; Lu, Yonglong

    2016-06-01

    The characterization of bacterial community compositions and the change in perfluoroalkyl acids (PFAAs) along a natural river distribution system were explored in the present study. Illumina high-throughput sequencing was used to explore bacterial community diversity and structure in sediment polluted by PFAAs from the Xiaoqing River, the area with concentrated fluorochemical facilities in China. The concentration of PFAAs was in the range of 8.44-465.60 ng/g dry weight (dw) in sediment. Perfluorooctanoic acid (PFOA) was the dominant PFAA in all samples, which accounted for 94.2 % of total PFAAs. High-level PFOA could lead to an obvious increase in relative abundance of Proteobacteria, ε-Proteobacteria, Thiobacillus, and Sulfurimonas and the decrease in relative abundance of other bacteria. Redundancy analysis revealed that PFOA played an important role in the formation of bacterial community, and PFOA at higher concentration could reduce the diversity of bacterial community. When the concentration of PFOA was below 100 ng/g dw in sediment, no significant effect on microbial community structure was observed. Thiobacillus and Sulfurimonas were positively correlated with the concentration of PFOA, suggesting that both genera were resistant to PFOA contamination. PMID:26780047

  17. RNA-Seq Analysis and Gene Discovery of Andrias davidianus Using Illumina Short Read Sequencing

    PubMed Central

    Li, Fenggang; Wang, Lixin; Lan, Qingjing; Yang, Hui; Li, Yang; Liu, Xiaolin; Yang, Zhaoxia

    2015-01-01

    The Chinese giant salamander, Andrias davidianus, is an important species in the course of evolution; however, there is insufficient genomic data in public databases for understanding its immunologic mechanisms. High-throughput transcriptome sequencing is necessary to generate an enormous number of transcript sequences from A. davidianus for gene discovery. In this study, we generated more than 40 million reads from samples of spleen and skin tissue using the Illumina paired-end sequencing technology. De novo assembly yielded 87,297 transcripts with a mean length of 734 base pairs (bp). Based on the sequence similarities, searching with known proteins, 38,916 genes were identified. Gene enrichment analysis determined that 981 transcripts were assigned to the immune system. Tissue-specific expression analysis indicated that 443 of transcripts were specifically expressed in the spleen and skin. Among these transcripts, 147 transcripts were found to be involved in immune responses and inflammatory reactions, such as fucolectin, β-defensins and lymphotoxin beta. Eight tissue-specific genes were selected for validation using real time reverse transcription quantitative PCR (qRT-PCR). The results showed that these genes were significantly more expressed in spleen and skin than in other tissues, suggesting that these genes have vital roles in the immune response. This work provides a comprehensive genomic sequence resource for A. davidianus and lays the foundation for future research on the immunologic and disease resistance mechanisms of A. davidianus and other amphibians. PMID:25874626

  18. Comparison of normalization methods for Illumina BeadChip HumanHT-12 v3

    PubMed Central

    2010-01-01

    Background Normalization of microarrays is a standard practice to account for and minimize effects which are not due to the controlled factors in an experiment. There is an overwhelming number of different methods that can be applied, none of which is ideally suited for all experimental designs. Thus, it is important to identify a normalization method appropriate for the experimental setup under consideration that is neither too negligent nor too stringent. Major aim is to derive optimal results from the underlying experiment. Comparisons of different normalization methods have already been conducted, none of which, to our knowledge, comparing more than a handful of methods. Results In the present study, 25 different ways of pre-processing Illumina Sentrix BeadChip array data are compared. Among others, methods provided by the BeadStudio software are taken into account. Looking at different statistical measures, we point out the ideal versus the actual observations. Additionally, we compare qRT-PCR measurements of transcripts from different ranges of expression intensities to the respective normalized values of the microarray data. Taking together all different kinds of measures, the ideal method for our dataset is identified. Conclusions Pre-processing of microarray gene expression experiments has been shown to influence further downstream analysis to a great extent and thus has to be carefully chosen based on the design of the experiment. This study provides a recommendation for deciding which normalization method is best suited for a particular experimental setup. PMID:20525181

  19. Transcriptome Analysis of Dastarcus helophoroides (Coleoptera: Bothrideridae) Using Illumina HiSeq Sequencing

    PubMed Central

    Zhang, Wei; Song, Wang; Zhang, Zhengqing; Wang, Haidong; Yang, Miaomiao; Guo, Ruijian; Li, Menglou

    2014-01-01

    Background Dastarcus helophoroides is known as the most valuable natural enemy insect against many large-body longhorned beetles. The molecular mechanism of its long lifespan and reproduction makes it a unique resource for genomic research. However, molecular biological studies on this parasitic beetle are scarce, and genomic information for D. helophoroides is not currently available. Thus, transcriptome information for this species is an important resource that is required for a better understanding of the molecular mechanisms of D. helophoroides. In this study, we obtained transcriptome information of D. helophoroides using high-throughput RNA sequencing. Results Using Illumina HiSeq 2000 sequencing, 27,543,746 clean reads corresponding to a total of 2.48 Gb nucleotides were obtained from a single run. These reads were assembled into 42,810 unigenes with a mean length of 683 bp. Using a sequence similarity search against the five public databases (NR, Swiss-Prot, GO, COG, KEGG) with a cut-off E-value of 10−5 using Blastx, a total of 31,293 unigenes were annotated with gene description, gene ontology terms, or metabolic pathways. Conclusions To the best of our knowledge, this is the first study on the transcriptome information of D. helophoroides. The transcriptome data presented in this study provide comprehensive information for future studies in D. helophoroides, particularly for functional genomic studies in this parasitic beetle. PMID:24979346

  20. Evaluating methods for purifying cyanobacterial cultures by qPCR and high-throughput Illumina sequencing.

    PubMed

    Heck, Karina; Machineski, Gabriela Silva; Alvarenga, Danillo Oliveira; Vaz, Marcelo Gomes Marçal Vieira; Varani, Alessandro de Mello; Fiore, Marli Fátima

    2016-10-01

    Cyanobacteria are commonly found in association with other microorganisms, which constitutes a great challenge during the isolation of cyanobacterial strains. Although several methods have been published for obtaining axenic cyanobacterial cultures, their efficiency is usually evaluated by observing the growth of non-cyanobacteria in culture media. In order to verify whether uncultured bacteria should be a concern during cyanobacterial isolation, this work aimed to detect by molecular methods sequences from cyanobacteria and other bacteria present before and after a technique for obtaining axenic cultures from plating and exposure of Fischerella sp. CENA161 akinetes to the Extran detergent and sodium hypochlorite. Solutions containing 0.5, 1, and 2% sodium hypochlorite were able to remove contaminant bacterial CFUs from the culture. However, qPCR pointed that the quantity of sequences amplified with universal bacteria primers was higher than the number of cyanobacteria-specific sequences before and after treatments. The presence of uncultured bacteria in post-hypochlorite cultures was confirmed by high-throughput Illumina sequencing. These results suggest that culturing may overlook the presence of uncultured bacteria associated to cyanobacterial strains and is not sufficient for monitoring the success of cyanobacterial isolation by itself. Molecular methods such as qPCR could be employed as an additional measure for evaluating axenity in cyanobacterial strains. PMID:27476485

  1. Illumina next generation sequencing data and expression microarrays data from retinoblastoma and medulloblastoma tissues.

    PubMed

    García-Chequer, A J; Méndez-Tenorio, A; Olguín-López, G; Sánchez-Vallejo, C; Isa, P; Arias, C F; Torres, J; Hernández-Angeles, A; Ramírez-Ortiz, M A; Lara, C; Cabrera-Muñoz, Ma de L; Sadowinski-Pine, S; Bravo-Ortiz, J C; Ramón-García, G; Diegopérez-Ramírez, J; Ramírez-Reyes, G; Casarrubias-Islas, R; Ramírez, J; Orjuela, M; Ponce-Castañeda, M V

    2016-03-01

    Retinoblastoma (Rb) is a pediatric intraocular malignancy and probably the most robust clinical model on which genetic predisposition to develop cancer has been demonstrated. Since deletions in chromosome 13 have been described in this tumor, we performed next generation sequencing to test whether recurrent losses could be detected in low coverage data. We used Illumina platform for 13 tumor tissue samples: two pools of 4 retinoblastoma cases each and one pool of 5 medulloblastoma cases (raw data can be found at http://www.ebi.ac.uk/ena/data/view/PRJEB6630). We first created an in silico reference profile generated from a human sequenced genome (GRCh37p5). From this data we calculated an integrity score to get an overview of gains and losses in all chromosomes; we next analyzed each chromosome in windows of 40 kb length, calculating for each window the log2 ratio between reads from tumor pool and in silico reference. Finally we generated panoramic maps with all the windows whether lost or gained along each chromosome associated to its cytogenetic bands to facilitate interpretation. Expression microarrays was done for the same samples and a list of over and under expressed genes is presented here. For this detection a significance analysis was done and a log2 fold change was chosen as significant (raw data can be found at http://www.ncbi.nlm.nih.gov/geo/accession number GSE11488). The complete research article can be found at Cancer Genetics journal (Garcia-Chequer et al., in press) [1]. In summary here we provide an overview with visual graphics of gains and losses chromosome by chromosome in retinoblastoma and medulloblastoma, also the integrity score analysis and a list of genes with relevant expression associated. This material can be useful to researchers that may want to explore gains and losses in other malignant tumors with this approach or compare their data with retinoblastoma. PMID:26937470

  2. Quasispecies Analyses of the HIV-1 Near-full-length Genome With Illumina MiSeq.

    PubMed

    Ode, Hirotaka; Matsuda, Masakazu; Matsuoka, Kazuhiro; Hachiya, Atsuko; Hattori, Junko; Kito, Yumiko; Yokomaku, Yoshiyuki; Iwatani, Yasumasa; Sugiura, Wataru

    2015-01-01

    Human immunodeficiency virus type-1 (HIV-1) exhibits high between-host genetic diversity and within-host heterogeneity, recognized as quasispecies. Because HIV-1 quasispecies fluctuate in terms of multiple factors, such as antiretroviral exposure and host immunity, analyzing the HIV-1 genome is critical for selecting effective antiretroviral therapy and understanding within-host viral coevolution mechanisms. Here, to obtain HIV-1 genome sequence information that includes minority variants, we sought to develop a method for evaluating quasispecies throughout the HIV-1 near-full-length genome using the Illumina MiSeq benchtop deep sequencer. To ensure the reliability of minority mutation detection, we applied an analysis method of sequence read mapping onto a consensus sequence derived from de novo assembly followed by iterative mapping and subsequent unique error correction. Deep sequencing analyses of aHIV-1 clone showed that the analysis method reduced erroneous base prevalence below 1% in each sequence position and discarded only < 1% of all collected nucleotides, maximizing the usage of the collected genome sequences. Further, we designed primer sets to amplify the HIV-1 near-full-length genome from clinical plasma samples. Deep sequencing of 92 samples in combination with the primer sets and our analysis method provided sufficient coverage to identify >1%-frequency sequences throughout the genome. When we evaluated sequences of pol genes from 18 treatment-naïve patients' samples, the deep sequencing results were in agreement with Sanger sequencing and identified numerous additional minority mutations. The results suggest that our deep sequencing method would be suitable for identifying within-host viral population dynamics throughout the genome. PMID:26617593

  3. Illumina next generation sequencing data and expression microarrays data from retinoblastoma and medulloblastoma tissues

    PubMed Central

    García-Chequer, A.J.; Méndez-Tenorio, A.; Olguín-López, G.; Sánchez-Vallejo, C.; Isa, P.; Arias, C.F.; Torres, J.; Hernández-Angeles, A.; Ramírez-Ortiz, M.A.; Lara, C.; Cabrera-Muñoz, Ma.de.L.; Sadowinski-Pine, S.; Bravo-Ortiz, J.C.; Ramón-García, G.; Diegopérez-Ramírez, J.; Ramírez-Reyes, G.; Casarrubias-Islas, R.; Ramírez, J.; Orjuela, M.; Ponce-Castañeda, M.V.

    2016-01-01

    Retinoblastoma (Rb) is a pediatric intraocular malignancy and probably the most robust clinical model on which genetic predisposition to develop cancer has been demonstrated. Since deletions in chromosome 13 have been described in this tumor, we performed next generation sequencing to test whether recurrent losses could be detected in low coverage data. We used Illumina platform for 13 tumor tissue samples: two pools of 4 retinoblastoma cases each and one pool of 5 medulloblastoma cases (raw data can be found at http://www.ebi.ac.uk/ena/data/view/PRJEB6630). We first created an in silico reference profile generated from a human sequenced genome (GRCh37p5). From this data we calculated an integrity score to get an overview of gains and losses in all chromosomes; we next analyzed each chromosome in windows of 40 kb length, calculating for each window the log2 ratio between reads from tumor pool and in silico reference. Finally we generated panoramic maps with all the windows whether lost or gained along each chromosome associated to its cytogenetic bands to facilitate interpretation. Expression microarrays was done for the same samples and a list of over and under expressed genes is presented here. For this detection a significance analysis was done and a log2 fold change was chosen as significant (raw data can be found at http://www.ncbi.nlm.nih.gov/geo/accession number GSE11488). The complete research article can be found at Cancer Genetics journal (Garcia-Chequer et al., in press) [1]. In summary here we provide an overview with visual graphics of gains and losses chromosome by chromosome in retinoblastoma and medulloblastoma, also the integrity score analysis and a list of genes with relevant expression associated. This material can be useful to researchers that may want to explore gains and losses in other malignant tumors with this approach or compare their data with retinoblastoma. PMID:26937470

  4. Quasispecies Analyses of the HIV-1 Near-full-length Genome With Illumina MiSeq

    PubMed Central

    Ode, Hirotaka; Matsuda, Masakazu; Matsuoka, Kazuhiro; Hachiya, Atsuko; Hattori, Junko; Kito, Yumiko; Yokomaku, Yoshiyuki; Iwatani, Yasumasa; Sugiura, Wataru

    2015-01-01

    Human immunodeficiency virus type-1 (HIV-1) exhibits high between-host genetic diversity and within-host heterogeneity, recognized as quasispecies. Because HIV-1 quasispecies fluctuate in terms of multiple factors, such as antiretroviral exposure and host immunity, analyzing the HIV-1 genome is critical for selecting effective antiretroviral therapy and understanding within-host viral coevolution mechanisms. Here, to obtain HIV-1 genome sequence information that includes minority variants, we sought to develop a method for evaluating quasispecies throughout the HIV-1 near-full-length genome using the Illumina MiSeq benchtop deep sequencer. To ensure the reliability of minority mutation detection, we applied an analysis method of sequence read mapping onto a consensus sequence derived from de novo assembly followed by iterative mapping and subsequent unique error correction. Deep sequencing analyses of aHIV-1 clone showed that the analysis method reduced erroneous base prevalence below 1% in each sequence position and discarded only < 1% of all collected nucleotides, maximizing the usage of the collected genome sequences. Further, we designed primer sets to amplify the HIV-1 near-full-length genome from clinical plasma samples. Deep sequencing of 92 samples in combination with the primer sets and our analysis method provided sufficient coverage to identify >1%-frequency sequences throughout the genome. When we evaluated sequences of pol genes from 18 treatment-naïve patients' samples, the deep sequencing results were in agreement with Sanger sequencing and identified numerous additional minority mutations. The results suggest that our deep sequencing method would be suitable for identifying within-host viral population dynamics throughout the genome. PMID:26617593

  5. Identification of neglected cestode Taenia multiceps microRNAs by illumina sequencing and bioinformatic analysis

    PubMed Central

    2013-01-01

    Background Worldwide, but especially in developing countries, coenurosis of sheep and other livestock is caused by Taenia multiceps larvae, and zoonotic infections occur in humans. Infections frequently lead to host death, resulting in huge socioeconomic losses. MicroRNAs (miRNAs) have important roles in the post-transcriptional regulation of a large number of animal genes by imperfectly binding target mRNAs. To date, there have been no reports of miRNAs in T. multiceps. Results In this study, we obtained 12.8 million high quality raw reads from adult T. multiceps small RNA library using Illumina sequencing technology. A total of 796 conserved miRNA families (containing 1,006 miRNAs) from 170,888 unique miRNAs were characterized using miRBase (Release 17.0). Here, we selected three conserved miRNA/miRNA* (antisense strand) duplexes at random and amplified their corresponding precursors using a PCR-based method. Furthermore, 20 candidate novel miRNA precursors were verified by genomic PCR. Among these, six corresponding T. multiceps miRNAs are considered specific for Taeniidae because no homologs were found in other species annotated in miRBase. In addition, 181,077 target sites within T. multiceps transcriptome were predicted for 20 candidate newly miRNAs. Conclusions Our large-scale investigation of miRNAs in adult T. multiceps provides a substantial platform for improving our understanding of the molecular regulation of T. multiceps and other cestodes development. PMID:23941076

  6. Using Illumina next generation sequencing technologies to sequence multigene families in de novo species.

    PubMed

    Hughes, Graham M; Gang, Li; Murphy, William J; Higgins, Desmond G; Teeling, Emma C

    2013-05-01

    The advent of Next Generation Sequencing Technology (NGST) has revolutionized molecular biology research, allowing for rapid gene/genome sequencing from a multitude of diverse species. As high throughput sequencing becomes more accessible, more efficient workflows must be developed to deal with the amounts of data produced and better assemble the genomes of de novo lineages. We combine traditional laboratory methods with Illumina NGST to amplify and sequence the largest mammalian multigene family, the Olfactory Receptor gene family, for species with and without a reference genome. We develop novel assembly methods to annotate and filter these data, which can be utilized for any gene family or any species. We find no significant difference between the ratio of genes within their respective gene families of our data compared with available genomic data. Using simulated data we explore the limitations of short-read sequence data and our assembly in recovering this gene family. We highlight the benefits and shortcomings of these methods. Compared with data generated from traditional polymerase chain reaction, cloning and Sanger sequencing methodologies, sequence data generated using our pipeline increases yield and sequencing efficiency without reducing the number of unique genes amplified. A cloning step is not required, therefore shortening data generation time. The novel downstream methodologies and workflows described provide a tool to be utilized by many fields of biology, to access and analyze the vast quantities of data generated. By combining laboratory and in silico methods, we provide a means of extracting genomic information for multigene families without complete genome sequencing. PMID:23480365

  7. De Novo Assembly and Annotation of Salvia splendens Transcriptome Using the Illumina Platform

    PubMed Central

    Shi, Aiping; Liu, Kefeng

    2014-01-01

    Background As an important perennial herbaceous flower, Salvia splendens possesses high ornamental value. Understanding its branching processes may help scientists select the best plant type. Although Salvia splendens is a frequently-used horticultural flower, only limited transcriptomic or genomic research is available in public databases. In the present study, we, for the first time, constructed a comprehensive dataset for Salvia splendens through de novo high-throughput transcriptome sequencing. Methodology/Principal Findings We performed de novo transcriptome sequencing on two different branching type plants (Strain 35 and Cailinghong) using the Illumina paired-end sequencing technology. For Strain 35, a total of 16,488,829 reads were generated and assembled into 38,498 unigenes, with a mean length of approximately 779 bp. For Cailinghong, 16,464,713 reads were generated and assembled into 34,302 unigenes, with a mean length of approximately 812 bp. Moreover, a total of 49,310 unigenes for Salvia splendens were identified, among them 33,925 (68.80%) were annotated in the non-redundant NCBI database, 25,371 (51.45%) were annotated in the Swiss-Prot database, while 24,888 (50.47%) and 9,896 (20.07%) unigenes were assigned to gene ontology categories and clusters of orthologous groups, respectively. Using the Kyoto Encyclopedia of Genes and Genomes pathway database, we identified 134 differently expressed unigenes between Strain 35 and Cailinghong, and then these unigenes were mapped to 79 pathways. In addition, we detected 2,453 simple sequence repeats (SSRs). Conclusions We obtained a comprehensive transcriptomic information from this work and provided a valuable resource of transcript sequences of Salvia splendens in public databases. Moreover, some candidate genes potentially involved in branching were identified. Furthermore, numerous obtained SSRs might contribute to marker-assisted selection. These data could be further utilized in functional genomics

  8. Illumina sequencing of fungi associated with manganese oxide deposits in cave systems

    NASA Astrophysics Data System (ADS)

    Zorn, B. T.; Santelli, C. M.; Carmichael, S. K.; Pepe-Ranney, C. P.; Roble, L.; Carmichael, M.; Bräuer, S.

    2013-12-01

    The environmental cycling of manganese (Mn) remains relatively poorly characterized when compared with other metals such as iron. However, fungi have been observed to produce Mn(III/IV) oxides resembling buserite, birnessite, and todorokite on the periphery of vegetative hyphae, hyphal branching points and at the base of fruiting bodies. Recent studies indicate that some of these oxides may be generated by a two-stage reaction with soluble Mn(II) and biogenic reactive oxygen species for some groups of fungi, in particular the Ascomycota. These oxides can provide a versatile protective barrier or aid in the capture of trace metals in the environment, although the exact evolutionary function and trigger is unclear. In this study, two caves in the southern Appalachians, a pristine cave and an anthropogenically impacted cave, were compared by analyzing fungal community assemblages in manganese oxide rich deposits. Quantitative PCR data indicated that fungi are present in a low abundance (<1%) in all locations sampled within the caves. Among amplified DNA sequences retrieved in an 18S rDNA clone library, over 88% were representative of the phylum Basidiomycota (predominantly Agaricomycetes), 2.74% of Ascomycota, 2.28% of Blastocladiomycota and Chytridiomycota, 0.46% of Zygomycota, and 3.65% of Eukarya or Fungi incertae sedis. Using Illumina's MiSeq to sequence amplicons of the fungal ITS1 gene has yielded roughly 100,000-200,000 paired-end reads per sample. These data are currently being analyzed to compare fungal communities before and after induced Mn oxidation in the field. In addition, sites within the pristine cave are being compared with analogous sites in the impacted cave. Culturing efforts have thus far yielded Mn oxide producing members of the orders Glomerales and Pleosporales as well as two Genus incertae sedis (Fungal sp. YECT1, and Fungal sp. YECT3, growing on discarded electrical tape) that do not appear to be closely related to any other known Mn

  9. Bovine myocardial epithelial inclusions.

    PubMed

    Baker, D C; Schmidt, S P; Langheinrich, K A; Cannon, L; Smart, R A

    1993-01-01

    Light microscopic, histochemical, immunohistochemical, and ultrastructural methods were used to examine myocardial epithelial masses in the hearts of ten cattle. The tissues consisted of paraffin-embedded or formalin-fixed samples from eight hearts that were being inspected in slaughter houses and from two hearts from calves that died of septicemia. The ages of the cattle ranged from 4 days to 12 years; the breeds were unspecified for all but one Hereford female and the two Holstein calves; and there were three males, four females, and three steers. The masses in these cases were compared with similar appearing lesions found in other animal species. The lesions in the bovine hearts were single to multiple, well circumscribed, found in the left ventricle wall, and composed of squamous to cuboidal epithelial cells that formed tubular, ductular, and acinar structures with lumens that were void or filled with amorphous protein globules. Electron microscopic examination revealed epithelial cells that had sparse apical microvilli, tight apical intercellular junctions, perinuclear bundles of filaments, and rare cilia. Almost half of the bovine epithelial masses (4/9) had occasional diastase-resistant periodic acid-Schiff-positive granules in their cytoplasm, and few had hyaluronidase-resistant alcian blue-positive granules (2/9) or colloidal iron-positive granules (1/9). All myocardial masses had abundant collagen surrounding the tubular and acinar structures, and 2/9 had elastin fibers as well. None of the myocardial masses had Churukian-Schenk or Fontana Masson's silver staining granules in epithelial cells. Immunohistochemically, all bovine myocardial tumors stained positively for cytokeratin (8/8), and occasional masses stained positively for vimentin (3/8) or carcinoembryonic antigen (3/8). None of the masses stained positively for desmin. The myocardial epithelial tumors most likely represent endodermal rests of tissue misplaced during organogenesis. PMID:7680178

  10. Enzootic Bovine Leukosis

    PubMed Central

    Reed, V. Ivan

    1981-01-01

    The author emphasizes the significance of enzootic bovine leukosis in Canada. He describes in detail diagnostic methods, various types of the disease and methods of transmission. Various aspects of the disease in Canada are compared with those in other countries. Prevention and control are discussed in a Canadian context and include the current policies of the Government of Canada in relationship to this disease. The possibility of developing a certification program for herds free of the disease is also discussed. The paper includes incidence in various parts of Canada. PMID:6265053

  11. Determining Microeukaryotic Plankton Community around Xiamen Island, Southeast China, Using Illumina MiSeq and PCR-DGGE Techniques.

    PubMed

    Yu, Lingyu; Zhang, Wenjing; Liu, Lemian; Yang, Jun

    2015-01-01

    Microeukaryotic plankton are important components of aquatic environments and play key roles in marine microbial food webs; however, little is known about their genetic diversity in subtropical offshore areas. Here we examined the community composition and genetic diversity of the microeukaryotic plankton in Xiamen offshore water by PCR-DGGE (polymerase chain reaction-denaturing gradient gel electrophoresis), clone-based sequencing and Illumina based sequencing. The Illumina MiSeq sequencing revealed a much (approximately two orders of magnitude) higher species richness of the microeukaryotic community than DGGE, but there were no significant difference in species richness and diversity among the northern, eastern, southern or western stations based on both methods. In this study, Copepoda, Ciliophora, Chlorophyta, Dinophyceae, Cryptophyta and Bacillariophyta (diatoms) were the dominant groups even though diatoms were not detected by DGGE. Our Illumina based results indicated that two northern communities (sites N2 and N3) were significantly different from others in having more protozoa and fewer diatoms. Redundancy analysis (RDA) showed that both temperature and salinity were the significant environmental factors influencing dominant species communities, whereas the full microeukaryotic community appeared to be affected by a complex of environmental factors. Our results suggested that extensive sampling combined with more deep sequencing are needed to obtain the complete diversity of the microeukaryotic community, and different diversity patterns for both abundant and rare taxa may be important in evaluating the marine ecosystem health. PMID:26020532

  12. Determining Microeukaryotic Plankton Community around Xiamen Island, Southeast China, Using Illumina MiSeq and PCR-DGGE Techniques

    PubMed Central

    Yu, Lingyu; Zhang, Wenjing; Liu, Lemian; Yang, Jun

    2015-01-01

    Microeukaryotic plankton are important components of aquatic environments and play key roles in marine microbial food webs; however, little is known about their genetic diversity in subtropical offshore areas. Here we examined the community composition and genetic diversity of the microeukaryotic plankton in Xiamen offshore water by PCR-DGGE (polymerase chain reaction-denaturing gradient gel electrophoresis), clone-based sequencing and Illumina based sequencing. The Illumina MiSeq sequencing revealed a much (approximately two orders of magnitude) higher species richness of the microeukaryotic community than DGGE, but there were no significant difference in species richness and diversity among the northern, eastern, southern or western stations based on both methods. In this study, Copepoda, Ciliophora, Chlorophyta, Dinophyceae, Cryptophyta and Bacillariophyta (diatoms) were the dominant groups even though diatoms were not detected by DGGE. Our Illumina based results indicated that two northern communities (sites N2 and N3) were significantly different from others in having more protozoa and fewer diatoms. Redundancy analysis (RDA) showed that both temperature and salinity were the significant environmental factors influencing dominant species communities, whereas the full microeukaryotic community appeared to be affected by a complex of environmental factors. Our results suggested that extensive sampling combined with more deep sequencing are needed to obtain the complete diversity of the microeukaryotic community, and different diversity patterns for both abundant and rare taxa may be important in evaluating the marine ecosystem health. PMID:26020532

  13. SSR_pipeline: a bioinformatic infrastructure for identifying microsatellites from paired-end Illumina high-throughput DNA sequencing data

    USGS Publications Warehouse

    Miller, Mark P.; Knaus, Brian J.; Mullins, Thomas D.; Haig, Susan M.

    2013-01-01

    SSR_pipeline is a flexible set of programs designed to efficiently identify simple sequence repeats (e.g., microsatellites) from paired-end high-throughput Illumina DNA sequencing data. The program suite contains 3 analysis modules along with a fourth control module that can automate analyses of large volumes of data. The modules are used to 1) identify the subset of paired-end sequences that pass Illumina quality standards, 2) align paired-end reads into a single composite DNA sequence, and 3) identify sequences that possess microsatellites (both simple and compound) conforming to user-specified parameters. The microsatellite search algorithm is extremely efficient, and we have used it to identify repeats with motifs from 2 to 25bp in length. Each of the 3 analysis modules can also be used independently to provide greater flexibility or to work with FASTQ or FASTA files generated from other sequencing platforms (Roche 454, Ion Torrent, etc.). We demonstrate use of the program with data from the brine fly Ephydra packardi (Diptera: Ephydridae) and provide empirical timing benchmarks to illustrate program performance on a common desktop computer environment. We further show that the Illumina platform is capable of identifying large numbers of microsatellites, even when using unenriched sample libraries and a very small percentage of the sequencing capacity from a single DNA sequencing run. All modules from SSR_pipeline are implemented in the Python programming language and can therefore be used from nearly any computer operating system (Linux, Macintosh, and Windows).

  14. Analysis of the cystic fibrosis lung microbiota via serial Illumina sequencing of bacterial 16S rRNA hypervariable regions.

    PubMed

    Maughan, Heather; Wang, Pauline W; Diaz Caballero, Julio; Fung, Pauline; Gong, Yunchen; Donaldson, Sylva L; Yuan, Lijie; Keshavjee, Shaf; Zhang, Yu; Yau, Yvonne C W; Waters, Valerie J; Tullis, D Elizabeth; Hwang, David M; Guttman, David S

    2012-01-01

    The characterization of bacterial communities using DNA sequencing has revolutionized our ability to study microbes in nature and discover the ways in which microbial communities affect ecosystem functioning and human health. Here we describe Serial Illumina Sequencing (SI-Seq): a method for deep sequencing of the bacterial 16S rRNA gene using next-generation sequencing technology. SI-Seq serially sequences portions of the V5, V6 and V7 hypervariable regions from barcoded 16S rRNA amplicons using an Illumina short-read genome analyzer. SI-Seq obtains taxonomic resolution similar to 454 pyrosequencing for a fraction of the cost, and can produce hundreds of thousands of reads per sample even with very high multiplexing. We validated SI-Seq using single species and mock community controls, and via a comparison to cystic fibrosis lung microbiota sequenced using 454 FLX Titanium. Our control runs show that SI-Seq has a dynamic range of at least five orders of magnitude, can classify >96% of sequences to the genus level, and performs just as well as 454 and paired-end Illumina methods in estimation of standard microbial ecology diversity measurements. We illustrate the utility of SI-Seq in a pilot sample of central airway secretion samples from cystic fibrosis patients. PMID:23056217

  15. [Bovine immunodeficiency virus: short review].

    PubMed

    Bouillant, A M; Archambault, D

    1990-01-01

    A bovine visna-like virus was isolated by Van Der Maaten et al (1972) but it did not draw attention since, at that time, most efforts were directed towards research on bovine leukemia virus. However, new interest was shown on the bovine visna-like virus after the isolation of the human immunodeficiency virus (HIV), because of the urgent need for developing animal models for the acquired immunodeficiency syndrome (AIDS). The purpose of this paper is to describe the different stages of the identification of the bovine virus and to up-date knowledge about it. The bovine visna-like virus has recently been named the bovine immuno-deficiency-like virus (BIV) and is the sole bovine lentivirus known to-date. BIV shares morphologic, antigenic and genomic characteristics with other lentiviruses. It grows and induces large syncytia in vitro and generates virus-productive and latent infections in cell culture. It causes persistent infection and slow progressive disease in cattle and probably in sheep. As target cells of the virus are leukocytes, the type of which is unknown, perturbations of the immune system are expected. Consequently, BIV may potentiate the occurrence of secondary infections and play a role in retroviral, multiple infections. It is not oncogenic. Transmission appears to occur in cattle by contact, but evidence of transmission in human beings has not been shown. Finally, BIV may be a potential model in vitro and in vivo for HIV and AIDS. PMID:1963056

  16. Parallel tagged amplicon sequencing of relatively long PCR products using the Illumina HiSeq platform and transcriptome assembly.

    PubMed

    Feng, Yan-Jie; Liu, Qing-Feng; Chen, Meng-Yun; Liang, Dan; Zhang, Peng

    2016-01-01

    In phylogenetics and population genetics, a large number of loci are often needed to accurately resolve species relationships. Normally, loci are enriched by PCR and sequenced by Sanger sequencing, which is expensive when the number of amplicons is large. Next-generation sequencing (NGS) techniques are increasingly used for parallel amplicon sequencing, which reduces sequencing costs tremendously, but has not reduced preparation costs very much. Moreover, for most current NGS methods, amplicons need to be purified and quantified before sequencing and their lengths are also restricted (normally <700 bp). Here, we describe an approach to sequence pooled amplicons of any length using the Illumina platform. Using this method, amplicons are pooled at equal volume rather than at equal concentration, thus eliminating the laborious purification and quantification steps. We then shear the pooled amplicons, repair the ends, add sample identifying linkers and pool multiple samples prior to Illumina library preparation. Data are then assembled using the transcriptome assembly program trinity, which is optimized to deal with templates of highly varying quantities. We demonstrated the utility of our approach by recovering 93.5% of the target amplicons (size up to 1650 bp) in full length for a 16 taxa × 101 loci project, using ~2.0 GB of Illumina HiSeq paired-end 90-bp data. Overall, we validate a rapid, cost-effective and scalable approach to sequence a large number of targeted loci from a large number of samples that is particularly suitable for both phylogenetics and population genetics studies that require a modest scale of data. PMID:25959587

  17. RNAseq versus genome-predicted transcriptomes: a large population of novel transcripts identified in an Illumina-454 Hydra transcriptome

    PubMed Central

    2013-01-01

    Background Evolutionary studies benefit from deep sequencing technologies that generate genomic and transcriptomic sequences from a variety of organisms. Genome sequencing and RNAseq have complementary strengths. In this study, we present the assembly of the most complete Hydra transcriptome to date along with a comparative analysis of the specific features of RNAseq and genome-predicted transcriptomes currently available in the freshwater hydrozoan Hydra vulgaris. Results To produce an accurate and extensive Hydra transcriptome, we combined Illumina and 454 Titanium reads, giving the primacy to Illumina over 454 reads to correct homopolymer errors. This strategy yielded an RNAseq transcriptome that contains 48’909 unique sequences including splice variants, representing approximately 24’450 distinct genes. Comparative analysis to the available genome-predicted transcriptomes identified 10’597 novel Hydra transcripts that encode 529 evolutionarily-conserved proteins. The annotation of 170 human orthologs points to critical functions in protein biosynthesis, FGF and TOR signaling, vesicle transport, immunity, cell cycle regulation, cell death, mitochondrial metabolism, transcription and chromatin regulation. However, a majority of these novel transcripts encodes short ORFs, at least 767 of them corresponding to pseudogenes. This RNAseq transcriptome also lacks 11’270 predicted transcripts that correspond either to silent genes or to genes expressed below the detection level of this study. Conclusions We established a simple and powerful strategy to combine Illumina and 454 reads and we produced, with genome assistance, an extensive and accurate Hydra transcriptome. The comparative analysis of the RNAseq transcriptome with genome-predicted transcriptomes lead to the identification of large populations of novel as well as missing transcripts that might reflect Hydra-specific evolutionary events. PMID:23530871

  18. Microbial Community Composition and Diversity via 16S rRNA Gene Amplicons: Evaluating the Illumina Platform

    PubMed Central

    Sinclair, Lucas; Osman, Omneya Ahmed; Bertilsson, Stefan; Eiler, Alexander

    2015-01-01

    As new sequencing technologies become cheaper and older ones disappear, laboratories switch vendors and platforms. Validating the new setups is a crucial part of conducting rigorous scientific research. Here we report on the reliability and biases of performing bacterial 16S rRNA gene amplicon paired-end sequencing on the MiSeq Illumina platform. We designed a protocol using 50 barcode pairs to run samples in parallel and coded a pipeline to process the data. Sequencing the same sediment sample in 248 replicates as well as 70 samples from alkaline soda lakes, we evaluated the performance of the method with regards to estimates of alpha and beta diversity. Using different purification and DNA quantification procedures we always found up to 5-fold differences in the yield of sequences between individually barcodes samples. Using either a one-step or a two-step PCR preparation resulted in significantly different estimates in both alpha and beta diversity. Comparing with a previous method based on 454 pyrosequencing, we found that our Illumina protocol performed in a similar manner – with the exception for evenness estimates where correspondence between the methods was low. We further quantified the data loss at every processing step eventually accumulating to 50% of the raw reads. When evaluating different OTU clustering methods, we observed a stark contrast between the results of QIIME with default settings and the more recent UPARSE algorithm when it comes to the number of OTUs generated. Still, overall trends in alpha and beta diversity corresponded highly using both clustering methods. Our procedure performed well considering the precisions of alpha and beta diversity estimates, with insignificant effects of individual barcodes. Comparative analyses suggest that 454 and Illumina sequence data can be combined if the same PCR protocol and bioinformatic workflows are used for describing patterns in richness, beta-diversity and taxonomic composition. PMID:25647581

  19. Global Transcriptome Sequencing Using the Illumina Platform and the Development of EST-SSR Markers in Autotetraploid Alfalfa

    PubMed Central

    Liu, Zhipeng; Chen, Tianlong; Ma, Lichao; Zhao, Zhiguang; Zhao, Patrick X.; Nan, Zhibiao; Wang, Yanrong

    2013-01-01

    Background Alfalfa is the most widely cultivated forage legume and one of the most economically valuable crops in the world. The large size and complexity of the alfalfa genome has delayed the development of genomic resources for alfalfa research. Second-generation Illumina transcriptome sequencing is an efficient method for generating a global transcriptome sequence dataset for gene discovery and molecular marker development in alfalfa. Methodology/Principal Findings More than 28 million sequencing reads (5.64 Gb of clean nucleotides) were generated by Illumina paired-end sequencing from 15 different alfalfa tissue samples. In total, 40,433 unigenes with an average length of 803 bp were obtained by de novo assembly. Based on a sequence similarity search of known proteins, a total of 36,684 (90.73%) unigenes were annotated. In addition, 1,649 potential EST-SSRs were identified as potential molecular markers from unigenes with lengths exceeding 1 kb. A total of 100 pairs of PCR primers were randomly selected to validate the assembly quality and develop EST-SSR markers from genomic DNA. Of these primer pairs, 82 were able to amplify sequences in initial screening tests, and 27 primer pairs successfully amplified DNA fragments and detected significant amounts of polymorphism among 10 alfalfa accessions. Conclusions/Significance The present study provided global sequence data for autotetraploid alfalfa and demonstrates the Illumina platform is a fast and effective approach to EST-SSR markers development in alfalfa. The use of these transcriptome datasets will serve as a valuable public information platform to accelerate studies of the alfalfa genome. PMID:24349529

  20. Species Identification and Profiling of Complex Microbial Communities Using Shotgun Illumina Sequencing of 16S rRNA Amplicon Sequences

    PubMed Central

    Lay, Christophe; Ho, Eliza Xin Pei; Low, Louie; Hibberd, Martin Lloyd; Nagarajan, Niranjan

    2013-01-01

    The high throughput and cost-effectiveness afforded by short-read sequencing technologies, in principle, enable researchers to perform 16S rRNA profiling of complex microbial communities at unprecedented depth and resolution. Existing Illumina sequencing protocols are, however, limited by the fraction of the 16S rRNA gene that is interrogated and therefore limit the resolution and quality of the profiling. To address this, we present the design of a novel protocol for shotgun Illumina sequencing of the bacterial 16S rRNA gene, optimized to amplify more than 90% of sequences in the Greengenes database and with the ability to distinguish nearly twice as many species-level OTUs compared to existing protocols. Using several in silico and experimental datasets, we demonstrate that despite the presence of multiple variable and conserved regions, the resulting shotgun sequences can be used to accurately quantify the constituents of complex microbial communities. The reconstruction of a significant fraction of the 16S rRNA gene also enabled high precision (>90%) in species-level identification thereby opening up potential application of this approach for clinical microbial characterization. PMID:23579286

  1. Bovine respiratory disease model based on dual infections with infection with bovine viral diarrhea virus and bovine corona virus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bovine respiratory disease complex (BRDC) is the leading cause of economic loss in the U.S. cattle industry. BRDC likely results from simultaneous or sequential infections with multiple pathogens including both viruses and bacteria. Bovine viral diarrhea virus (BVDV) and bovine corona virus (BoCV...

  2. Material Properties of Inorganic Bovine Cancellous Bovine: Nukbone

    NASA Astrophysics Data System (ADS)

    Piña, Cristina; Palma, Benito; Munguía, Nadia

    2006-09-01

    In this work, inorganic cancellous bovine bone implants prepared in the Instituto de Investigaciones en Materiales — UNAM were characterized. Elementary chemical analysis was made, toxic elements concentration were measured and the content of organic matter also. These implants fulfill all the requirements of the ASTM standards, and therefore it is possible their use in medical applications.

  3. Illumina GA IIx& HiSeq 2000 Production Sequenccing and QC Analysis Pipelines at the DOE Joint Genome Institute

    SciTech Connect

    Daum, Christopher; Zane, Matthew; Han, James; Kennedy, Megan; San Diego, Matthew; Copeland, Alex; Li, Mingkun; Lucas, Susan

    2011-01-31

    The U.S. Department of Energy (DOE) Joint Genome Institute's (JGI) Production Sequencing group is committed to the generation of high-quality genomic DNA sequence to support the mission areas of renewable energy generation, global carbon management, and environmental characterization and clean-up. Within the JGI's Production Sequencing group, a robust Illumina Genome Analyzer and HiSeq pipeline has been established. Optimization of the sesequencer pipelines has been ongoing with the aim of continual process improvement of the laboratory workflow, reducing operational costs and project cycle times to increases ample throughput, and improving the overall quality of the sequence generated. A sequence QC analysis pipeline has been implemented to automatically generate read and assembly level quality metrics. The foremost of these optimization projects, along with sequencing and operational strategies, throughput numbers, and sequencing quality results will be presented.

  4. Illumina next-generation sequencing reveals the complete mitochondrial genome of Psenopsis anomala (Perciformes: Centrolophidae) with phylogenetic consideration.

    PubMed

    Chen, Huapu; Che, Zhiwei; Li, Jiantao; Dai, Mingli; Xiang, Ling; Deng, Siping; Zhu, Chunhua; Huang, Hai; Li, Guangli

    2016-09-01

    Using Illumina next-generation sequencing (NGS), the complete mitochondrial genome of the Psenopsis anomala was sequenced in the present study. The mitochondrial genome of P. anomala is 16,528 bp long and consists of 13 protein-coding genes, 2 rRNA genes, 22 tRNA genes, and a control region. The structure about gene order and composition of P. anomala mitochondrial genome is similar to those of most other vertebrates. The nucleotide compositions of the light strand in descending order is 29.18% of T, 27.97% of G, 27.06% of A, and 15.79% of C. With the exception of the NADH dehydrogenase subunit 6 (ND6) and eight tRNA genes, other mitochondrial genes are encoded on the heavy strand. The phylogenetic analysis by maximum-likelihood (ML) method shown that the Psenopsis anomala was closer to Peprilus triacanthus in the phylogenetic relationship. PMID:25974666

  5. Gene expression profiling of Sinapis alba leaves under drought stress and rewatering growth conditions with Illumina deep sequencing.

    PubMed

    Dong, Cai-Hua; Li, Chen; Yan, Xiao-Hong; Huang, Shun-Mou; Huang, Jin-Yong; Wang, Li-Jun; Guo, Rui-Xing; Lu, Guang-Yuan; Zhang, Xue-Kun; Fang, Xiao-Ping; Wei, Wen-Hui

    2012-05-01

    Sinapis alba has many desirable agronomic traits including tolerance to drought. In this investigation, we performed the genome-wide transcriptional profiling of S. alba leaves under drought stress and rewatering growth conditions in an attempt to identify candidate genes involved in drought tolerance, using the Illumina deep sequencing technology. The comparative analysis revealed numerous changes in gene expression level attributable to the drought stress, which resulted in the down-regulation of 309 genes and the up-regulation of 248 genes. Gene ontology analysis revealed that the differentially expressed genes were mainly involved in cell division and catalytic and metabolic processes. Our results provide useful information for further analyses of the drought stress tolerance in Sinapis, and will facilitate molecular breeding for Brassica crop plants. PMID:22207172

  6. Analyzing illumina gene expression microarray data from different tissues: methodological aspects of data analysis in the metaxpress consortium.

    PubMed

    Schurmann, Claudia; Heim, Katharina; Schillert, Arne; Blankenberg, Stefan; Carstensen, Maren; Dörr, Marcus; Endlich, Karlhans; Felix, Stephan B; Gieger, Christian; Grallert, Harald; Herder, Christian; Hoffmann, Wolfgang; Homuth, Georg; Illig, Thomas; Kruppa, Jochen; Meitinger, Thomas; Müller, Christian; Nauck, Matthias; Peters, Annette; Rettig, Rainer; Roden, Michael; Strauch, Konstantin; Völker, Uwe; Völzke, Henry; Wahl, Simone; Wallaschofski, Henri; Wild, Philipp S; Zeller, Tanja; Teumer, Alexander; Prokisch, Holger; Ziegler, Andreas

    2012-01-01

    Microarray profiling of gene expression is widely applied in molecular biology and functional genomics. Experimental and technical variations make meta-analysis of different studies challenging. In a total of 3358 samples, all from German population-based cohorts, we investigated the effect of data preprocessing and the variability due to sample processing in whole blood cell and blood monocyte gene expression data, measured on the Illumina HumanHT-12 v3 BeadChip array.Gene expression signal intensities were similar after applying the log(2) or the variance-stabilizing transformation. In all cohorts, the first principal component (PC) explained more than 95% of the total variation. Technical factors substantially influenced signal intensity values, especially the Illumina chip assignment (33-48% of the variance), the RNA amplification batch (12-24%), the RNA isolation batch (16%), and the sample storage time, in particular the time between blood donation and RNA isolation for the whole blood cell samples (2-3%), and the time between RNA isolation and amplification for the monocyte samples (2%). White blood cell composition parameters were the strongest biological factors influencing the expression signal intensities in the whole blood cell samples (3%), followed by sex (1-2%) in both sample types. Known single nucleotide polymorphisms (SNPs) were located in 38% of the analyzed probe sequences and 4% of them included common SNPs (minor allele frequency >5%). Out of the tested SNPs, 1.4% significantly modified the probe-specific expression signals (Bonferroni corrected p-value<0.05), but in almost half of these events the signal intensities were even increased despite the occurrence of the mismatch. Thus, the vast majority of SNPs within probes had no significant effect on hybridization efficiency.In summary, adjustment for a few selected technical factors greatly improved reliability of gene expression analyses. Such adjustments are particularly required for meta

  7. Development and validation of microsatellite markers for Brachiaria ruziziensis obtained by partial genome assembly of Illumina single-end reads

    PubMed Central

    2013-01-01

    Background Brachiaria ruziziensis is one of the most important forage species planted in the tropics. The application of genomic tools to aid the selection of superior genotypes can provide support to B. ruziziensis breeding programs. However, there is a complete lack of information about the B. ruziziensis genome. Also, the availability of genomic tools, such as molecular markers, to support B. ruziziensis breeding programs is rather limited. Recently, next-generation sequencing technologies have been applied to generate sequence data for the identification of microsatellite regions and primer design. In this study, we present a first validated set of SSR markers for Brachiaria ruziziensis, selected from a de novo partial genome assembly of single-end Illumina reads. Results A total of 85,567 perfect microsatellite loci were detected in contigs with a minimum 10X coverage. We selected a set of 500 microsatellite loci identified in contigs with minimum 100X coverage for primer design and synthesis, and tested a subset of 269 primer pairs, 198 of which were polymorphic on 11 representative B. ruziziensis accessions. Descriptive statistics for these primer pairs are presented, as well as estimates of marker transferability to other relevant brachiaria species. Finally, a set of 11 multiplex panels containing the 30 most informative markers was validated and proposed for B. ruziziensis genetic analysis. Conclusions We show that the detection and development of microsatellite markers from genome assembled Illumina single-end DNA sequences is highly efficient. The developed markers are readily suitable for genetic analysis and marker assisted selection of Brachiaria ruziziensis. The use of this approach for microsatellite marker development is promising for species with limited genomic information, whose breeding programs would benefit from the use of genomic tools. To our knowledge, this is the first set of microsatellite markers developed for this important species

  8. A tale of three next generation sequencing platforms: comparison of Ion Torrent, Pacific Biosciences and Illumina MiSeq sequencers

    PubMed Central

    2012-01-01

    Background Next generation sequencing (NGS) technology has revolutionized genomic and genetic research. The pace of change in this area is rapid with three major new sequencing platforms having been released in 2011: Ion Torrent’s PGM, Pacific Biosciences’ RS and the Illumina MiSeq. Here we compare the results obtained with those platforms to the performance of the Illumina HiSeq, the current market leader. In order to compare these platforms, and get sufficient coverage depth to allow meaningful analysis, we have sequenced a set of 4 microbial genomes with mean GC content ranging from 19.3 to 67.7%. Together, these represent a comprehensive range of genome content. Here we report our analysis of that sequence data in terms of coverage distribution, bias, GC distribution, variant detection and accuracy. Results Sequence generated by Ion Torrent, MiSeq and Pacific Biosciences technologies displays near perfect coverage behaviour on GC-rich, neutral and moderately AT-rich genomes, but a profound bias was observed upon sequencing the extremely AT-rich genome of Plasmodium falciparum on the PGM, resulting in no coverage for approximately 30% of the genome. We analysed the ability to call variants from each platform and found that we could call slightly more variants from Ion Torrent data compared to MiSeq data, but at the expense of a higher false positive rate. Variant calling from Pacific Biosciences data was possible but higher coverage depth was required. Context specific errors were observed in both PGM and MiSeq data, but not in that from the Pacific Biosciences platform. Conclusions All three fast turnaround sequencers evaluated here were able to generate usable sequence. However there are key differences between the quality of that data and the applications it will support. PMID:22827831

  9. Strategies for Achieving High Sequencing Accuracy for Low Diversity Samples and Avoiding Sample Bleeding Using Illumina Platform

    PubMed Central

    Mitra, Abhishek; Skrzypczak, Magdalena; Ginalski, Krzysztof; Rowicka, Maga

    2015-01-01

    Sequencing microRNA, reduced representation sequencing, Hi-C technology and any method requiring the use of in-house barcodes result in sequencing libraries with low initial sequence diversity. Sequencing such data on the Illumina platform typically produces low quality data due to the limitations of the Illumina cluster calling algorithm. Moreover, even in the case of diverse samples, these limitations are causing substantial inaccuracies in multiplexed sample assignment (sample bleeding). Such inaccuracies are unacceptable in clinical applications, and in some other fields (e.g. detection of rare variants). Here, we discuss how both problems with quality of low-diversity samples and sample bleeding are caused by incorrect detection of clusters on the flowcell during initial sequencing cycles. We propose simple software modifications (Long Template Protocol) that overcome this problem. We present experimental results showing that our Long Template Protocol remarkably increases data quality for low diversity samples, as compared with the standard analysis protocol; it also substantially reduces sample bleeding for all samples. For comprehensiveness, we also discuss and compare experimental results from alternative approaches to sequencing low diversity samples. First, we discuss how the low diversity problem, if caused by barcodes, can be avoided altogether at the barcode design stage. Second and third, we present modified guidelines, which are more stringent than the manufacturer’s, for mixing low diversity samples with diverse samples and lowering cluster density, which in our experience consistently produces high quality data from low diversity samples. Fourth and fifth, we present rescue strategies that can be applied when sequencing results in low quality data and when there is no more biological material available. In such cases, we propose that the flowcell be re-hybridized and sequenced again using our Long Template Protocol. Alternatively, we discuss how

  10. Design of a 9K illumina BeadChip for polar bears (Ursus maritimus) from RAD and transcriptome sequencing.

    PubMed

    Malenfant, René M; Coltman, David W; Davis, Corey S

    2015-05-01

    Single-nucleotide polymorphisms (SNPs) offer numerous advantages over anonymous markers such as microsatellites, including improved estimation of population parameters, finer-scale resolution of population structure and more precise genomic dissection of quantitative traits. However, many SNPs are needed to equal the resolution of a single microsatellite, and reliable large-scale genotyping of SNPs remains a challenge in nonmodel species. Here, we document the creation of a 9K Illumina Infinium BeadChip for polar bears (Ursus maritimus), which will be used to investigate: (i) the fine-scale population structure among Canadian polar bears and (ii) the genomic architecture of phenotypic traits in the Western Hudson Bay subpopulation. To this end, we used restriction-site associated DNA (RAD) sequencing from 38 bears across their circumpolar range, as well as blood/fat transcriptome sequencing of 10 individuals from Western Hudson Bay. Six-thousand RAD SNPs and 3000 transcriptomic SNPs were selected for the chip, based primarily on genomic spacing and gene function respectively. Of the 9000 SNPs ordered from Illumina, 8042 were successfully printed, and - after genotyping 1450 polar bears - 5441 of these SNPs were found to be well clustered and polymorphic. Using this array, we show rapid linkage disequilibrium decay among polar bears, we demonstrate that in a subsample of 78 individuals, our SNPs detect known genetic structure more clearly than 24 microsatellites genotyped for the same individuals and that these results are not driven by the SNP ascertainment scheme. Here, we present one of the first large-scale genotyping resources designed for a threatened species. PMID:25187336

  11. Analysis of the global transcriptome of longan (Dimocarpus longan Lour.) embryogenic callus using Illumina paired-end sequencing

    PubMed Central

    2013-01-01

    Background Longan is a tropical/subtropical fruit tree of great economic importance in Southeast Asia. Progress in understanding molecular mechanisms of longan embryogenesis, which is the primary influence on fruit quality and yield, is slowed by lack of transcriptomic and genomic information. Illumina second generation sequencing, which is suitable for generating enormous numbers of transcript sequences that can be used for functional genomic analysis of longan. Results In this study, a longan embryogenic callus (EC) cDNA library was sequenced using an Illumina HiSeq 2000 system. A total of 64,876,258 clean reads comprising 5.84 Gb of nucleotides were assembled into 68,925 unigenes of 448-bp mean length, with unigenes ≥1000 bp accounting for 8.26% of the total. Using BLASTx, 40,634 unigenes were found to have significant similarity with accessions in Nr and Swiss- Prot databases. Of these, 38,845 unigenes were assigned to 43 GO sub-categories and 17,118 unigenes were classified into 25 COG sub-groups. In addition, 17,306 unigenes mapped to 199 KEGG pathways, with the categories of Metabolic pathways, Plant-pathogen interaction, Biosynthesis of secondary metabolites, and Genetic information processing being well represented. Analyses of unigenes ≥1000 bp revealed 328 embryogenesis-related unigenes as well as numerous unigenes expressed in EC associated with functions of reproductive growth, such as flowering, gametophytogenesis, and fertility, and vegetative growth, such as root and shoot growth. Furthermore, 23 unigenes related to embryogenesis and reproductive and vegetative growth were validated by quantitative real time PCR (qPCR) in samples from different stages of longan somatic embryogenesis (SE); their differentially expressions in the various embryogenic cultures indicated their possible roles in longan SE. Conclusions The quantity and variety of expressed EC genes identified in this study is sufficient to serve as a global transcriptome dataset for

  12. The Use of Kosher Phenotyping for Mapping QTL Affecting Susceptibility to Bovine Respiratory Disease

    PubMed Central

    Eitam, Harel; Yishay, Moran; Schiavini, Fausta; Soller, Morris; Bagnato, Alessandro; Shabtay, Ariel

    2016-01-01

    Bovine respiratory disease (BRD) is the leading cause of morbidity and mortality in feedlot cattle, caused by multiple pathogens that become more virulent in response to stress. As clinical signs often go undetected and various preventive strategies failed, identification of genes affecting BRD is essential for selection for resistance. Selective DNA pooling (SDP) was applied in a genome wide association study (GWAS) to map BRD QTLs in Israeli Holstein male calves. Kosher scoring of lung adhesions was used to allocate 122 and 62 animals to High (Glatt Kosher) and Low (Non-Kosher) resistant groups, respectively. Genotyping was performed using the Illumina BovineHD BeadChip according to the Infinium protocol. Moving average of -logP was used to map QTLs and Log drop was used to define their boundaries (QTLRs). The combined procedure was efficient for high resolution mapping. Nineteen QTLRs distributed over 13 autosomes were found, some overlapping previous studies. The QTLRs contain polymorphic functional and expression candidate genes to affect kosher status, with putative immunological and wound healing activities. Kosher phenotyping was shown to be a reliable means to map QTLs affecting BRD morbidity. PMID:27077383

  13. The Use of Kosher Phenotyping for Mapping QTL Affecting Susceptibility to Bovine Respiratory Disease.

    PubMed

    Lipkin, Ehud; Strillacci, Maria Giuseppina; Eitam, Harel; Yishay, Moran; Schiavini, Fausta; Soller, Morris; Bagnato, Alessandro; Shabtay, Ariel

    2016-01-01

    Bovine respiratory disease (BRD) is the leading cause of morbidity and mortality in feedlot cattle, caused by multiple pathogens that become more virulent in response to stress. As clinical signs often go undetected and various preventive strategies failed, identification of genes affecting BRD is essential for selection for resistance. Selective DNA pooling (SDP) was applied in a genome wide association study (GWAS) to map BRD QTLs in Israeli Holstein male calves. Kosher scoring of lung adhesions was used to allocate 122 and 62 animals to High (Glatt Kosher) and Low (Non-Kosher) resistant groups, respectively. Genotyping was performed using the Illumina BovineHD BeadChip according to the Infinium protocol. Moving average of -logP was used to map QTLs and Log drop was used to define their boundaries (QTLRs). The combined procedure was efficient for high resolution mapping. Nineteen QTLRs distributed over 13 autosomes were found, some overlapping previous studies. The QTLRs contain polymorphic functional and expression candidate genes to affect kosher status, with putative immunological and wound healing activities. Kosher phenotyping was shown to be a reliable means to map QTLs affecting BRD morbidity. PMID:27077383

  14. Transcriptome Analysis of Bovine Ovarian Follicles at Predeviation and Onset of Deviation Stages of a Follicular Wave

    PubMed Central

    Li, Pengfei; Meng, Jinzhu; Liu, Wenzhong; Smith, George W.; Yao, Jianbo; Lyu, Lihua

    2016-01-01

    For two libraries (PDF1 and ODF1) using Illumina sequencing 44,082,301 and 43,708,132 clean reads were obtained, respectively. After being mapped to the bovine RefSeq database, 15,533 genes were identified to be expressed in both types of follicles (cut-off RPKM > 0.5), of which 719 were highly expressed in bovine follicles (cut-off RPKM > 100). Furthermore, 83 genes were identified as being differentially expressed in ODF1 versus PDF1, where 42 genes were upregulated and 41 genes were downregulated. KEGG pathway analysis revealed two upregulated genes in ODF1 versus PDF1, CYP11A1, and CYP19A1, which are important genes in the steroid hormone biosynthesis pathway. This study represents the first investigation of transcriptome of bovine follicles at predeviation and onset of deviation stages and provides a foundation for future investigation of the regulatory mechanisms involved in follicular development in cattle. PMID:27088081

  15. 78 FR 72979 - Bovine Spongiform Encephalopathy; Importation of Bovines and Bovine Products

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-12-04

    ..., 2012, we published in the Federal Register (77 FR 15848-15913, Docket No. APHIS-2008-0010) a proposal... January 4, 2005 (70 FR 460-553, Docket No. 03-080-3). The delay of applicability was removed in a final... Products Derived from Bovines,'' published in the Federal Register on September 18, 2007 (72 FR...

  16. Design of the Illumina Porcine 50K+ SNP Iselect(TM) Beadchip and Characterization of the Porcine HapMap Population

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Using next generation sequencing technology the International Swine SNP Consortium has identified 500,000 SNPs and used these to design an Illumina Infinium iSelect™ SNP BeadChip with a selection of 60,218 SNPs. The selected SNPs include previously validated SNPs and SNPs identified de novo using se...

  17. Comparison of Illumina de novo assembled and Sanger sequenced viral genomes: A case study for RNA viruses recovered from the plant pathogenic fungus Sclerotinia sclerotiorum.

    PubMed

    Khalifa, Mahmoud E; Varsani, Arvind; Ganley, Austen R D; Pearson, Michael N

    2016-07-01

    The advent of 'next generation sequencing' (NGS) technologies has led to the discovery of many novel mycoviruses, the majority of which are sufficiently different from previously sequenced viruses that there is no appropriate reference sequence on which to base the sequence assembly. Although many new genome sequences are generated by NGS, confirmation of the sequence by Sanger sequencing is still essential for formal classification by the International Committee for the Taxonomy of Viruses (ICTV), although this is currently under review. To empirically test the validity of de novo assembled mycovirus genomes from dsRNA extracts, we compared the results from Illumina sequencing with those from random cloning plus targeted PCR coupled with Sanger sequencing for viruses from five Sclerotinia sclerotiorum isolates. Through Sanger sequencing we detected nine viral genomes while through Illumina sequencing we detected the same nine viruses plus one additional virus from the same samples. Critically, the Illumina derived sequences share >99.3 % identity to those obtained by cloning and Sanger sequencing. Although, there is scope for errors in de novo assembled viral genomes, our results demonstrate that by maximising the proportion of viral sequence in the data and using sufficiently rigorous quality controls, it is possible to generate de novo genome sequences of comparable accuracy from Illumina sequencing to those obtained by Sanger sequencing. PMID:26581665

  18. Evaluation of Multiplexed 16S rRNA Microbial Population Surveys Using Illumina MiSeq Platform (Seventh Annual Sequencing, Finishing, Analysis in the Future (SFAF) Meeting 2012)

    SciTech Connect

    Tremblay, Julien

    2012-06-01

    Julien Tremblay from DOE JGI presents "Evaluation of Multiplexed 16S rRNA Microbial Population Surveys Using Illumina MiSeq Platorm" at the 7th Annual Sequencing, Finishing, Analysis in the Future (SFAF) Meeting held in June, 2012 in Santa Fe, NM.

  19. Investigation of bacterial and archaeal communities: novel protocols using modern sequencing by Illumina MiSeq and traditional DGGE-cloning.

    PubMed

    Kraková, Lucia; Šoltys, Katarína; Budiš, Jaroslav; Grivalský, Tomáš; Ďuriš, František; Pangallo, Domenico; Szemes, Tomáš

    2016-09-01

    Different protocols based on Illumina high-throughput DNA sequencing and denaturing gradient gel electrophoresis (DGGE)-cloning were developed and applied for investigating hot spring related samples. The study was focused on three target genes: archaeal and bacterial 16S rRNA and mcrA of methanogenic microflora. Shorter read lengths of the currently most popular technology of sequencing by Illumina do not allow analysis of the complete 16S rRNA region, or of longer gene fragments, as was the case of Sanger sequencing. Here, we demonstrate that there is no need for special indexed or tailed primer sets dedicated to short variable regions of 16S rRNA since the presented approach allows the analysis of complete bacterial 16S rRNA amplicons (V1-V9) and longer archaeal 16S rRNA and mcrA sequences. Sample augmented with transposon is represented by a set of approximately 300 bp long fragments that can be easily sequenced by Illumina MiSeq. Furthermore, a low proportion of chimeric sequences was observed. DGGE-cloning based strategies were performed combining semi-nested PCR, DGGE and clone library construction. Comparing both investigation methods, a certain degree of complementarity was observed confirming that the DGGE-cloning approach is not obsolete. Novel protocols were created for several types of laboratories, utilizing the traditional DGGE technique or using the most modern Illumina sequencing. PMID:27338271

  20. Evaluation of Multiplexed 16S rRNA Microbial Population Surveys Using Illumina MiSeq Platform (Seventh Annual Sequencing, Finishing, Analysis in the Future (SFAF) Meeting 2012)

    ScienceCinema

    Tremblay, Julien [DOE JGI

    2013-01-25

    Julien Tremblay from DOE JGI presents "Evaluation of Multiplexed 16S rRNA Microbial Population Surveys Using Illumina MiSeq Platorm" at the 7th Annual Sequencing, Finishing, Analysis in the Future (SFAF) Meeting held in June, 2012 in Santa Fe, NM.

  1. Evaluation of the Illumina(®) Beta Version ForenSeq™ DNA Signature Prep Kit for use in genetic profiling.

    PubMed

    Churchill, Jennifer D; Schmedes, Sarah E; King, Jonathan L; Budowle, Bruce

    2016-01-01

    While capillary electrophoresis-based technologies have been the mainstay for human identity typing applications, there are limitations with this methodology's resolution, scalability, and throughput. Massively parallel sequencing (MPS) offers the capability to multiplex multiple types of forensically-relevant markers and multiple samples together in one run all at an overall lower cost per nucleotide than traditional capillary electrophoresis-based methods; thus, addressing some of these limitations. MPS also is poised to expand forensic typing capabilities by providing new strategies for mixture deconvolution with the identification of intra-STR allele sequence variants and the potential to generate new types of investigative leads with an increase in the overall number and types of genetic markers being analyzed. The beta version of the Illumina ForenSeq DNA Signature Prep Kit is a MPS library preparation method with a streamlined workflow that allows for targeted amplification and sequencing of 63 STRs and 95 identity SNPs, with the option to include an additional 56 ancestry SNPs and 22 phenotypic SNPs depending on the primer mix chosen for amplification, on the MiSeq desktop sequencer (Illumina). This study was divided into a series of experiments that evaluated reliability, sensitivity of detection, mixture analysis, concordance, and the ability to analyze challenged samples. Genotype accuracy, depth of coverage, and allele balance were used as informative metrics for the quality of the data produced. The ForenSeq DNA Signature Prep Kit produced reliable, reproducible results and obtained full profiles with DNA input amounts of 1ng. Data were found to be concordant with current capillary electrophoresis methods, and mixtures at a 1:19 ratio were resolved accurately. Data from the challenged samples showed concordant results with current DNA typing methods with markers in common and minimal allele drop out from the large number of markers typed on these

  2. Circulating microRNA Profiles during the Bovine Oestrous Cycle

    PubMed Central

    Ioannidis, Jason; Donadeu, F. Xavier

    2016-01-01

    Up to 50% of ovulations go undetected in modern dairy herds due to attenuated oestrus behavior and a lack of high-accuracy methods for detection of fertile oestrus. This significantly reduces overall herd productivity and constitutes a high economic burden to the dairy industry. MicroRNAs (miRNAs) are ubiquitous regulators of gene expression during both health and disease and they have been shown to regulate different reproductive processes. Extracellular miRNAs are stable and can provide useful biomarkers of tissue function; changes in circulating miRNA profiles have been reported during menstrual cycles. This study sought to establish the potential of circulating miRNAs as biomarkers of oestrus in cattle. We collected plasma samples from 8 Holstein-Friesian heifers on days Days 0, 8 and 16 of an oestrous cycle and analysed small RNA populations on each Day using two independent high-throughput approaches, namely, Illumina sequencing (n = 24 samples) and Qiagen PCR arrays (n = 9 sample pools, 3–4 samples / pool). Subsequently, we used RT-qPCR (n = 24 samples) to validate the results of high-throughput analyses, as well as to establish the expression profiles of additional miRNAs previously reported to be differentially expressed during reproductive cycles. Overall, we identified four miRNAs (let-7f, miR-125b, miR-145 and miR-99a-5p), the plasma levels of which distinctly increased (up to 2.2-fold, P < 0.05) during oestrus (Day 0) relative to other stages of the cycle (Days 8 and 16). Moreover, we identified several hundred different isomiRs and established their relative abundance in bovine plasma. In summary, our results reveal the dynamic nature of plasma miRNAs during the oestrous cycle and provide evidence of the feasibility of using circulating miRNAs as biomarkers of reproductive function in livestock in the future. PMID:27340826

  3. The Role of Viral Population Diversity in Adaptation of Bovine Coronavirus to New Host Environments

    PubMed Central

    Borucki, Monica K.; Allen, Jonathan E.; Chen-Harris, Haiyin; Zemla, Adam; Vanier, Gilda; Mabery, Shalini; Torres, Clinton; Hullinger, Pamela; Slezak, Tom

    2013-01-01

    The high mutation rate of RNA viruses enables a diverse genetic population of viral genotypes to exist within a single infected host. In-host genetic diversity could better position the virus population to respond and adapt to a diverse array of selective pressures such as host-switching events. Multiple new coronaviruses, including SARS, have been identified in human samples just within the last ten years, demonstrating the potential of coronaviruses as emergent human pathogens. Deep sequencing was used to characterize genomic changes in coronavirus quasispecies during simulated host-switching. Three bovine nasal samples infected with bovine coronavirus were used to infect human and bovine macrophage and lung cell lines. The virus reproduced relatively well in macrophages, but the lung cell lines were not infected efficiently enough to allow passage of non lab-adapted samples. Approximately 12 kb of the genome was amplified before and after passage and sequenced at average coverages of nearly 950×(454 sequencing) and 38,000×(Illumina). The consensus sequence of many of the passaged samples had a 12 nucleotide insert in the consensus sequence of the spike gene, and multiple point mutations were associated with the presence of the insert. Deep sequencing revealed that the insert was present but very rare in the unpassaged samples and could quickly shift to dominate the population when placed in a different environment. The insert coded for three arginine residues, occurred in a region associated with fusion entry into host cells, and may allow infection of new cell types via heparin sulfate binding. Analysis of the deep sequencing data indicated that two distinct genotypes circulated at different frequency levels in each sample, and support the hypothesis that the mutations present in passaged strains were “selected” from a pre-existing pool rather than through de novo mutation and subsequent population fixation. PMID:23308119

  4. Bovine ephemeral fever: a review.

    PubMed

    Nandi, S; Negi, B S

    1999-04-01

    Bovine ephemeral fever is a viral disease of cattle and buffaloes besides subclinical involvement of a variety of ruminant species. The subtropical and temperate regions of Africa, Asia and Australia have experienced the major epidemic of the bovine ephemeral fever but the occurrence in the tropics can not be overlooked. Although the substantial role played by the vectors viz., mosquitoes and culicoides in bovine ephemeral fever perpetuation and dissemination, other vector involvement if any should be extensively studied. The clinical severity of the disease is not apparent and the mortality is low. However, high morbidity, enormous economic losses in terms of significant reduction in production, disruption of national and international trade and finally a variety of complications resulting from the disease have drawn appreciable attention from the researchers around the world to resolve the unsolved questions in this area. In this review, detailed informations of all the aspects of the disease has been provided in a simple, lucid and easily understandable manner. PMID:10051179

  5. Development and cross-species transferability of EST-SSR markers in Siberian wildrye (Elymus sibiricus L.) using Illumina sequencing

    PubMed Central

    Zhou, Qiang; Luo, Dong; Ma, Lichao; Xie, Wengang; Wang, Yu; Wang, Yanrong; Liu, Zhipeng

    2016-01-01

    Siberian wildrye (Elymus sibiricus L.) is a perennial, self-fertilizing grass that plays an important role in animal husbandry and environmental sustenance. However, the transcriptomic and genomic information on this species is very limited, which hinders genetic and breeding studies. In the present study, 76,686,804 clean reads were generated from 11 different tissue samples of E. sibiricus by Illumina paired-end sequencing, and the reads were deposited into the NCBI SRA database (SRX574376). A total of 8,769 EST-SSRs were identified from 94,458 unigene sequences, which were obtained by de novo assembly. Moreover, 1,078 primer pairs were successfully designed, and 500 pairs were randomly selected to assess polymorphisms in 15 E. sibiricus accessions. A total of 112 primer pairs were polymorphic, and the polymorphism information content (PIC) values ranged from 0.39 to 0.81, indicating a high level of informativeness. Furthermore, these 112 polymorphic primer pairs were used to evaluate the transferability to 13 other related species, and 55 EST-SSR markers were found to be polymorphic among these 13 Elymus species. This study collected the global sequence data for E. sibiricus, and the newly developed markers will prove valuable in facilitating genetic diversity in E. sibiricus and related Elymus species. PMID:26853106

  6. Illumina Synthetic Long Read Sequencing Allows Recovery of Missing Sequences even in the "Finished" C. elegans Genome.

    PubMed

    Li, Runsheng; Hsieh, Chia-Ling; Young, Amanda; Zhang, Zhihong; Ren, Xiaoliang; Zhao, Zhongying

    2015-01-01

    Most next-generation sequencing platforms permit acquisition of high-throughput DNA sequences, but the relatively short read length limits their use in genome assembly or finishing. Illumina has recently released a technology called Synthetic Long-Read Sequencing that can produce reads of unusual length, i.e., predominately around 10 Kb. However, a systematic assessment of their use in genome finishing and assembly is still lacking. We evaluate the promise and deficiency of the long reads in these aspects using isogenic C. elegans genome with no gap. First, the reads are highly accurate and capable of recovering most types of repetitive sequences. However, the presence of tandem repetitive sequences prevents pre-assembly of long reads in the relevant genomic region. Second, the reads are able to reliably detect missing but not extra sequences in the C. elegans genome. Third, the reads of smaller size are more capable of recovering repetitive sequences than those of bigger size. Fourth, at least 40 Kbp missing genomic sequences are recovered in the C. elegans genome using the long reads. Finally, an N50 contig size of at least 86 Kbp can be achieved with 24 × reads but with substantial mis-assembly errors, highlighting a need for novel assembly algorithm for the long reads. PMID:26039588

  7. De Novo Assembly and Annotation of the Chinese Chive (Allium tuberosum Rottler ex Spr.) Transcriptome Using the Illumina Platform

    PubMed Central

    Zhou, Shu-Mei; Chen, Li-Mei; Liu, Shi-Qi; Wang, Xiu-Feng; Sun, Xiu-Dong

    2015-01-01

    Chinese chive (A. tuberosum Rottler ex Spr.) is one of the most widely cultivated Allium species in China. However, minimal transcriptomic and genomic data are available to reveal its evolution and genetic diversity. In this study, de novo transcriptome sequencing was performed to produce large transcript sequences using an Illumina HiSeq 2000 instrument. We produced 51,968,882 high-quality clean reads and assembled them into 150,154 contigs. A total of 60,031 unigenes with an average length of 631 bp were identified. Of these, 36,523 unigenes were homologous to existing database sequences, 35,648 unigenes were annotated in the NCBI non-redundant (Nr) sequence database, and 23,509 unigenes were annotated in the Swiss-Prot database. A total of 26,798 unigenes were assigned to 57 Gene Ontology (GO) terms, and 13,378 unigenes were assigned to Cluster of Orthologous Group categories. Using the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway database, we mapped 21,361 unigenes onto 128 pathways. Furthermore, 2,125 sequences containing simple sequence repeats (SSRs) were identified. This new dataset provides the most comprehensive resource currently available for gene expression, gene discovery, and future genomic research on Chinese chive. The sequence resources developed in this study can be used to develop molecular markers that will facilitate further genetic research on Chinese chive and related species. PMID:26204518

  8. Phyllosphere Bacterial Community of Floating Macrophytes in Paddy Soil Environments as Revealed by Illumina High-Throughput Sequencing

    PubMed Central

    Xie, Wan-Ying

    2014-01-01

    The phyllosphere of floating macrophytes in paddy soil ecosystems, a unique habitat, may support large microbial communities but remains largely unknown. We took Wolffia australiana as a representative floating plant and investigated its phyllosphere bacterial community and the underlying driving forces of community modulation in paddy soil ecosystems using Illumina HiSeq 2000 platform-based 16S rRNA gene sequence analysis. The results showed that the phyllosphere of W. australiana harbored considerably rich communities of bacteria, with Proteobacteria and Bacteroidetes as the predominant phyla. The core microbiome in the phyllosphere contained genera such as Acidovorax, Asticcacaulis, Methylibium, and Methylophilus. Complexity of the phyllosphere bacterial communities in terms of class number and α-diversity was reduced compared to those in corresponding water and soil. Furthermore, the bacterial communities exhibited structures significantly different from those in water and soil. These findings and the following redundancy analysis (RDA) suggest that species sorting played an important role in the recruitment of bacterial species in the phyllosphere. The compositional structures of the phyllosphere bacterial communities were modulated predominantly by water physicochemical properties, while the initial soil bacterial communities had limited impact. Taken together, the findings from this study reveal the diversity and uniqueness of the phyllosphere bacterial communities associated with the floating macrophytes in paddy soil environments. PMID:25362067

  9. Building two indica rice reference genomes with PacBio long-read and Illumina paired-end sequencing data.

    PubMed

    Zhang, Jianwei; Chen, Ling-Ling; Sun, Shuai; Kudrna, Dave; Copetti, Dario; Li, Weiming; Mu, Ting; Jiao, Wen-Biao; Xing, Feng; Lee, Seunghee; Talag, Jayson; Song, Jia-Ming; Du, Bogu; Xie, Weibo; Luo, Meizhong; Maldonado, Carlos Ernesto; Goicoechea, Jose Luis; Xiong, Lizhong; Wu, Changyin; Xing, Yongzhong; Zhou, Dao-Xiu; Yu, Sibin; Zhao, Yu; Wang, Gongwei; Yu, Yeisoo; Luo, Yijie; Hurtado, Beatriz Elena Padilla; Danowitz, Ann; Wing, Rod A; Zhang, Qifa

    2016-01-01

    Over the past 30 years, we have performed many fundamental studies on two Oryza sativa subsp. indica varieties, Zhenshan 97 (ZS97) and Minghui 63 (MH63). To improve the resolution of many of these investigations, we generated two reference-quality reference genome assemblies using the most advanced sequencing technologies. Using PacBio SMRT technology, we produced over 108 (ZS97) and 174 (MH63) Gb of raw sequence data from 166 (ZS97) and 209 (MH63) pools of BAC clones, and generated ~97 (ZS97) and ~74 (MH63) Gb of paired-end whole-genome shotgun (WGS) sequence data with Illumina sequencing technology. With these data, we successfully assembled two platinum standard reference genomes that have been publicly released. Here we provide the full sets of raw data used to generate these two reference genome assemblies. These data sets can be used to test new programs for better genome assembly and annotation, aid in the discovery of new insights into genome structure, function, and evolution, and help to provide essential support to biological research in general. PMID:27622467

  10. Identification of polymorphic and off-target probe binding sites on the Illumina Infinium MethylationEPIC BeadChip.

    PubMed

    McCartney, Daniel L; Walker, Rosie M; Morris, Stewart W; McIntosh, Andrew M; Porteous, David J; Evans, Kathryn L

    2016-09-01

    Genome-wide analysis of DNA methylation has now become a relatively inexpensive technique thanks to array-based methylation profiling technologies. The recently developed Illumina Infinium MethylationEPIC BeadChip interrogates methylation at over 850,000 sites across the human genome, covering 99% of RefSeq genes. This array supersedes the widely used Infinium HumanMethylation450 BeadChip, which has permitted insights into the relationship between DNA methylation and a wide range of conditions and traits. Previous research has identified issues with certain probes on both the HumanMethylation450 BeadChip and its predecessor, the Infinium HumanMethylation27 BeadChip, which were predicted to affect array performance. These issues concerned probe-binding specificity and the presence of polymorphisms at target sites. Using in silico methods, we have identified probes on the Infinium MethylationEPIC BeadChip that are predicted to (i) measure methylation at polymorphic sites and (ii) hybridise to multiple genomic regions. We intend these resources to be used for quality control procedures when analysing data derived from this platform. PMID:27330998

  11. 3' tag digital gene expression profiling of human brain and universal reference RNA using Illumina Genome Analyzer

    PubMed Central

    2009-01-01

    Background Massive parallel sequencing has the potential to replace microarrays as the method for transcriptome profiling. Currently there are two protocols: full-length RNA sequencing (RNA-SEQ) and 3'-tag digital gene expression (DGE). In this preliminary effort, we evaluated the 3' DGE approach using two reference RNA samples from the MicroArray Quality Control Consortium (MAQC). Results Using Brain RNA sample from multiple runs, we demonstrated that the transcript profiles from 3' DGE were highly reproducible between technical and biological replicates from libraries constructed by the same lab and even by different labs, and between two generations of Illumina's Genome Analyzers. Approximately 65% of all sequence reads mapped to mitochondrial genes, ribosomal RNAs, and canonical transcripts. The expression profiles of brain RNA and universal human reference RNA were compared which demonstrated that DGE was also highly quantitative with excellent correlation of differential expression with quantitative real-time PCR. Furthermore, one lane of 3' DGE sequencing, using the current sequencing chemistry and image processing software, had wider dynamic range for transcriptome profiling and was able to detect lower expressed genes which are normally below the detection threshold of microarrays. Conclusion 3' tag DGE profiling with massive parallel sequencing achieved high sensitivity and reproducibility for transcriptome profiling. Although it lacks the ability of detecting alternative splicing events compared to RNA-SEQ, it is much more affordable and clearly out-performed microarrays (Affymetrix) in detecting lower abundant transcripts. PMID:19917133

  12. De novo transcriptome of the desert beetle Microdera punctipennis (Coleoptera: Tenebrionidae) using Illumina RNA-seq technology.

    PubMed

    Lu, Xueying; Li, Jieqiong; Yang, Jianhuan; Liu, Xiaoning; Ma, Ji

    2014-11-01

    Insects in Tenebrionidae have unique stress adaptations that allow them to survive temperature extremes. We report here a gene expression profiling of Microdera punctipennis, a beetle in desert region, to gain a global view of its environmental adaptations. A total of 48,158,004 reads were obtained by transcriptome sequencing, and the de novo assembly yielded 56,348 unigenes with an average length of 666 bp. Based on similarity searches with a cut-off E-value of 10(-5) against two protein sequence databases, 41,109 of the unigenes (about 72.96%) were matched to known proteins. An in-depth analysis of the data revealed a large number of genes were associated with environmental stress, including genes that encode heat shock proteins, antifreeze proteins, and enzymes such as chitinase, trehalose, and trehalose-6-phosphate synthase. This study generated a substantial number of M. punctipennis transcript sequences that can be used to discover novel genes associated with stress adaptation. These sequences are a valuable resource for future studies of the desert beetle and other insects in Tenebrionidae. Transcriptome analysis based on Illumina paired-end sequencing is a powerful approach for gene discovery and molecular marker development for non-model species. PMID:25142502

  13. Bacterial community compositions of coking wastewater treatment plants in steel industry revealed by Illumina high-throughput sequencing.

    PubMed

    Ma, Qiao; Qu, Yuanyuan; Shen, Wenli; Zhang, Zhaojing; Wang, Jingwei; Liu, Ziyan; Li, Duanxing; Li, Huijie; Zhou, Jiti

    2015-03-01

    In this study, Illumina high-throughput sequencing was used to reveal the community structures of nine coking wastewater treatment plants (CWWTPs) in China for the first time. The sludge systems exhibited a similar community composition at each taxonomic level. Compared to previous studies, some of the core genera in municipal wastewater treatment plants such as Zoogloea, Prosthecobacter and Gp6 were detected as minor species. Thiobacillus (20.83%), Comamonas (6.58%), Thauera (4.02%), Azoarcus (7.78%) and Rhodoplanes (1.42%) were the dominant genera shared by at least six CWWTPs. The percentages of autotrophic ammonia-oxidizing bacteria and nitrite-oxidizing bacteria were unexpectedly low, which were verified by both real-time PCR and fluorescence in situ hybridization analyses. Hierarchical clustering and canonical correspondence analysis indicated that operation mode, flow rate and temperature might be the key factors in community formation. This study provides new insights into our understanding of microbial community compositions and structures of CWWTPs. PMID:25569032

  14. High coverage of the complete mitochondrial genome of the rare Gray's beaked whale (Mesoplodon grayi) using Illumina next generation sequencing.

    PubMed

    Thompson, Kirsten F; Patel, Selina; Williams, Liam; Tsai, Peter; Constantine, Rochelle; Baker, C Scott; Millar, Craig D

    2016-01-01

    Using an Illumina platform, we shot-gun sequenced the complete mitochondrial genome of Gray's beaked whale (Mesoplodon grayi) to an average coverage of 152X. We performed a de novo assembly using SOAPdenovo2 and determined the total mitogenome length to be 16,347 bp. The nucleotide composition was asymmetric (33.3% A, 24.6% C, 12.6% G, 29.5% T) with an overall GC content of 37.2%. The gene organization was similar to that of other cetaceans with 13 protein-coding genes, 2 rRNAs (12S and 16S), 22 predicted tRNAs and 1 control region or D-loop. We found no evidence of heteroplasmy or nuclear copies of mitochondrial DNA in this individual. Beaked whales within the genus Mesoplodon are rarely seen at sea and their basic biology is poorly understood. These data will contribute to resolving the phylogeography and population ecology of this speciose group. PMID:24450706

  15. Somatic Mutation Screening Using Archival Formalin-Fixed, Paraffin-Embedded Tissues by Fluidigm Multiplex PCR and Illumina Sequencing.

    PubMed

    Wang, Ming; Escudero-Ibarz, Leire; Moody, Sarah; Zeng, Naiyan; Clipson, Alexandra; Huang, Yuanxue; Xue, Xuemin; Grigoropoulos, Nicholas F; Barrans, Sharon; Worrillow, Lisa; Forshew, Tim; Su, Jing; Firth, Andrew; Martin, Howard; Jack, Andrew; Brugger, Kim; Du, Ming-Qing

    2015-09-01

    High-throughput somatic mutation screening using FFPE tissues is a major challenge because of a lack of established methods and validated variant calling algorithms. We aimed to develop a targeted sequencing protocol by Fluidigm multiplex PCR and Illumina sequencing and to establish a companion variant calling algorithm. The experimental protocol and variant calling algorithm were first developed and optimized against a series of somatic mutations (147 substitutions, 12 indels ranging from 1 to 33 bp) in seven genes, previously detected by Sanger sequencing of DNA from 163 FFPE lymphoma biopsy specimens. The optimized experimental protocol and variant calling algorithm were further ascertained in two separate experiments by including the seven genes as a part of larger gene panels (22 or 13 genes) using FFPE and high-molecular-weight lymphoma DNAs, respectively. We found that most false-positive variants were due to DNA degradation, deamination, and Taq polymerase errors, but they were nonreproducible and could be efficiently eliminated by duplicate experiments. A small fraction of false-positive variants appeared in duplicate, but they were at low alternative allele frequencies and could be separated from mutations when appropriate threshold value was used. In conclusion, we established a robust practical approach for high-throughput mutation screening using archival FFPE tissues. PMID:26165823

  16. Characterization of genome-wide microsatellites of Saccharina japonica based on a preliminary assembly of Illumina sequencing reads

    NASA Astrophysics Data System (ADS)

    Zhang, Linan; Peng, Jie; Li, Xiaojie; Cui, Cuiju; Sun, Juan; Yang, Guanpin

    2016-06-01

    Microsatellites or simple sequence repeats (SSR) function widely and locate dependently in genome. However, their characteristics are often ignored due to the lack of genomic sequences of most species. Kelp ( Saccharina japonica), a brown macroalga, is extensively cultured in China. In this study, the genome of S. japonica was surveyed using an Illumina sequencing platform, and its microsatellites were characterized. The preliminarily assembled genome was 469.4 Mb in size, with a scaffold N50 of 20529 bp. Among the 128370 identified microsatellites, 90671, 25726 and 11973 were found in intergenic regions, introns and exons, averaging 339.3, 178.8 and 205.4 microsatellites per Mb, respectively. These microsatellites distributed unevenly in S. japonica genome. Mononucleotide motifs were the most abundant in the genome, while trinucleotide ones were the most prevalent in exons. The microsatellite abundance decreased significantly with the increase of motif repeat numbers, and the microsatellites with a small number of repeats accounted for a higher proportion of the exons than those of the intergenic regions and introns. C/G-rich motifs were more common in exons than in intergenic regions and introns. These characteristics of microsatellites in S. japonica genome may associate with their functions, and ultimately their adaptation and evolution. Among the 120140 pairs of designed microsatellite primers, approximately 75% were predicted to be able to amplify S. japonica DNA. These microsatellite markers will be extremely useful for the genetic breeding and population evolution studies of kelp.

  17. Illumina Synthetic Long Read Sequencing Allows Recovery of Missing Sequences even in the “Finished” C. elegans Genome

    PubMed Central

    Li, Runsheng; Hsieh, Chia-Ling; Young, Amanda; Zhang, Zhihong; Ren, Xiaoliang; Zhao, Zhongying

    2015-01-01

    Most next-generation sequencing platforms permit acquisition of high-throughput DNA sequences, but the relatively short read length limits their use in genome assembly or finishing. Illumina has recently released a technology called Synthetic Long-Read Sequencing that can produce reads of unusual length, i.e., predominately around 10 Kb. However, a systematic assessment of their use in genome finishing and assembly is still lacking. We evaluate the promise and deficiency of the long reads in these aspects using isogenic C. elegans genome with no gap. First, the reads are highly accurate and capable of recovering most types of repetitive sequences. However, the presence of tandem repetitive sequences prevents pre-assembly of long reads in the relevant genomic region. Second, the reads are able to reliably detect missing but not extra sequences in the C. elegans genome. Third, the reads of smaller size are more capable of recovering repetitive sequences than those of bigger size. Fourth, at least 40 Kbp missing genomic sequences are recovered in the C. elegans genome using the long reads. Finally, an N50 contig size of at least 86 Kbp can be achieved with 24×reads but with substantial mis-assembly errors, highlighting a need for novel assembly algorithm for the long reads. PMID:26039588

  18. Use of Illumina sequencing to identify transposon insertions underlying mutant phenotypes in high-copy Mutator lines of maize.

    PubMed

    Williams-Carrier, Rosalind; Stiffler, Nicholas; Belcher, Susan; Kroeger, Tiffany; Stern, David B; Monde, Rita-Ann; Coalter, Robert; Barkan, Alice

    2010-07-01

    High-copy transposons have been effectively exploited as mutagens in a variety of organisms. However, their utility for phenotype-driven forward genetics has been hampered by the difficulty of identifying the specific insertions responsible for phenotypes of interest. We describe a new method that can substantially increase the throughput of linking a disrupted gene to a known phenotype in high-copy Mutator (Mu) transposon lines in maize. The approach uses the Illumina platform to obtain sequences flanking Mu elements in pooled, bar-coded DNA samples. Insertion sites are compared among individuals of suitable genotype to identify those that are linked to the mutation of interest. DNA is prepared for sequencing by mechanical shearing, adapter ligation, and selection of DNA fragments harboring Mu flanking sequences by hybridization to a biotinylated oligonucleotide corresponding to the Mu terminal inverted repeat. This method yields dense clusters of sequence reads that tile approximately 400 bp flanking each side of each heritable insertion. The utility of the approach is demonstrated by identifying the causal insertions in four genes whose disruption blocks chloroplast biogenesis at various steps: thylakoid protein targeting (cpSecE), chloroplast gene expression (polynucleotide phosphorylase and PTAC12), and prosthetic group attachment (HCF208/CCB2). This method adds to the tools available for phenotype-driven Mu tagging in maize, and could be adapted for use with other high-copy transposons. A by-product of the approach is the identification of numerous heritable insertions that are unrelated to the targeted phenotype, which can contribute to community insertion resources. PMID:20409008

  19. Use of the melting curve assay as a means for high-throughput quantification of Illumina sequencing libraries

    PubMed Central

    Forster, John W.

    2016-01-01

    Background. Multiplexed sequencing is commonly performed on massively parallel short-read sequencing platforms such as Illumina, and the efficiency of library normalisation can affect the quality of the output dataset. Although several library normalisation approaches have been established, none are ideal for highly multiplexed sequencing due to issues of cost and/or processing time. Methods. An inexpensive and high-throughput library quantification method has been developed, based on an adaptation of the melting curve assay. Sequencing libraries were subjected to the assay using the Bio-Rad Laboratories CFX ConnectTM Real-Time PCR Detection System. The library quantity was calculated through summation of reduction of relative fluorescence units between 86 and 95 °C. Results.PCR-enriched sequencing libraries are suitable for this quantification without pre-purification of DNA. Short DNA molecules, which ideally should be eliminated from the library for subsequent processing, were differentiated from the target DNA in a mixture on the basis of differences in melting temperature. Quantification results for long sequences targeted using the melting curve assay were correlated with those from existing methods (R2 > 0.77), and that observed from MiSeq sequencing (R2 = 0.82). Discussion.The results of multiplexed sequencing suggested that the normalisation performance of the described method is equivalent to that of another recently reported high-throughput bead-based method, BeNUS. However, costs for the melting curve assay are considerably lower and processing times shorter than those of other existing methods, suggesting greater suitability for highly multiplexed sequencing applications. PMID:27602269

  20. Sequencing, De novo Assembly, Functional Annotation and Analysis of Phyllanthus amarus Leaf Transcriptome Using the Illumina Platform

    PubMed Central

    Bose Mazumdar, Aparupa; Chattopadhyay, Sharmila

    2016-01-01

    Phyllanthus amarus Schum. and Thonn., a widely distributed annual medicinal herb has a long history of use in the traditional system of medicine for over 2000 years. However, the lack of genomic data for P. amarus, a non-model organism hinders research at the molecular level. In the present study, high-throughput sequencing technology has been employed to enhance better understanding of this herb and provide comprehensive genomic information for future work. Here P. amarus leaf transcriptome was sequenced using the Illumina Miseq platform. We assembled 85,927 non-redundant (nr) “unitranscript” sequences with an average length of 1548 bp, from 18,060,997 raw reads. Sequence similarity analyses and annotation of these unitranscripts were performed against databases like green plants nr protein database, Gene Ontology (GO), Clusters of Orthologous Groups (COG), PlnTFDB, KEGG databases. As a result, 69,394 GO terms, 583 enzyme codes (EC), 134 KEGG maps, and 59 Transcription Factor (TF) families were generated. Functional and comparative analyses of assembled unitranscripts were also performed with the most closely related species like Populus trichocarpa and Ricinus communis using TRAPID. KEGG analysis showed that a number of assembled unitranscripts were involved in secondary metabolites, mainly phenylpropanoid, flavonoid, terpenoids, alkaloids, and lignan biosynthetic pathways that have significant medicinal attributes. Further, Fragments Per Kilobase of transcript per Million mapped reads (FPKM) values of the identified secondary metabolite pathway genes were determined and Reverse Transcription PCR (RT-PCR) of a few of these genes were performed to validate the de novo assembled leaf transcriptome dataset. In addition 65,273 simple sequence repeats (SSRs) were also identified. To the best of our knowledge, this is the first transcriptomic dataset of P. amarus till date. Our study provides the largest genetic resource that will lead to drug development and pave

  1. Slim-Filter: an interactive windows-based application for illumina genome analyzer data assessment and manipulation

    PubMed Central

    2012-01-01

    Background The emergence of Next Generation Sequencing technologies has made it possible for individual investigators to generate gigabases of sequencing data per week. Effective analysis and manipulation of these data is limited due to large file sizes, so even simple tasks such as data filtration and quality assessment have to be performed in several steps. This requires (potentially problematic) interaction between the investigator and a bioinformatics/computational service provider. Furthermore, such services are often performed using specialized computational facilities. Results We present a Windows-based application, Slim-Filter designed to interactively examine the statistical properties of sequencing reads produced by Illumina Genome Analyzer and to perform a broad spectrum of data manipulation tasks including: filtration of low quality and low complexity reads; filtration of reads containing undesired subsequences (such as parts of adapters and PCR primers used during the sample and sequencing libraries preparation steps); excluding duplicated reads (while keeping each read’s copy number information in a specialized data format); and sorting reads by copy numbers allowing for easy access and manual editing of the resulting files. Slim-Filter is organized as a sequence of windows summarizing the statistical properties of the reads. Each data manipulation step has roll-back abilities, allowing for return to previous steps of the data analysis process. Slim-Filter is written in C++ and is compatible with fasta, fastq, and specialized AS file formats presented in this manuscript. Setup files and a user’s manual are available for download at the supplementary web site ( https://www.bioinfo.uh.edu/Slim_Filter/). Conclusion The presented Windows-based application has been developed with the goal of providing individual investigators with integrated sequencing reads analysis, curation, and manipulation capabilities. PMID:22800377

  2. De novo Transcriptome Analysis of Chinese Citrus Fly, Bactrocera minax (Diptera: Tephritidae), by High-Throughput Illumina Sequencing.

    PubMed

    Wang, Jia; Xiong, Ke-Cai; Liu, Ying-Hong

    2016-01-01

    The Chinese citrus fly, Bactrocera minax (Enderlein), is one of the most devastating pests of citrus in the temperate areas of Asia. So far, studies involving molecular biology and physiology of B. minax are still scarce, partly because of the lack of genomic information and inability to rear this insect in laboratory. In this study, de novo assembly of a transcriptome was performed using Illumina sequencing technology. A total of 20,928,907 clean reads were obtained and assembled into 33,324 unigenes, with an average length of 908.44 bp. Unigenes were annotated by alignment against NCBI non-redundant protein (Nr), Swiss-Prot, Clusters of Orthologous Groups (COG), Gene Ontology (GO), and Kyoto Encyclopedia of Genes and Genomes Pathway (KEGG) database. Genes potentially involved in stress tolerance, including 20 heat shock protein (Hsps) genes, 26 glutathione S-transferases (GSTs) genes, and 2 ferritin subunit genes, were identified. These genes may play roles in stress tolerance in B. minax diapause stage. It has previously been found that 20E application on B. minax pupae could avert diapause, but the underlying mechanisms remain unknown. Thus, genes encoding enzymes in 20E biosynthesis pathway, including Neverland, Spook, Phantom, Disembodied, Shadow, Shade, and Cyp18a1, and genes encoding 20E receptor proteins, ecdysone receptor (EcR) and ultraspiracle (USP), were identified. The expression patterns of 20E-related genes among developmental stages and between 20E-treated and untreated pupae demonstrated their roles in diapause program. In addition, 1,909 simple sequence repeats (SSRs) were detected, which will contribute to molecular marker development. The findings in this study greatly improve our genetic understanding of B. minax, and lay the foundation for future studies on this species. PMID:27331903

  3. Sequencing and De Novo Assembly of the Asian Clam (Corbicula fluminea) Transcriptome Using the Illumina GAIIx Method

    PubMed Central

    Chen, Huihui; Zha, Jinmiao; Liang, Xuefang; Bu, Jihong; Wang, Miao; Wang, Zijian

    2013-01-01

    Background The Asian clam (Corbicula fluminea) is currently one of the most economically important aquatic species in China and has been used as a test organism in many environmental studies. However, the lack of genomic resources, such as sequenced genome, expressed sequence tags (ESTs) and transcriptome sequences has hindered the research on C. fluminea. Recent advances in large-scale RNA-Seq enable generation of genomic resources in a short time, and provide large expression datasets for functional genomic analysis. Methodology/Principal Findings We used a next-generation high-throughput DNA sequencing technique with an Illumina GAIIx method to analyze the transcriptome from the whole bodies of C. fluminea. More than 62,250,336 high-quality reads were generated based on the raw data, and 134,684 unigenes with a mean length of 791 bp were assembled using the Velvet and Oases software. All of the assembly unigenes were annotated by running BLASTx and BLASTn similarity searches on the Nt, Nr, Swiss-Prot, COG and KEGG databases. In addition, the Clusters of Orthologous Groups (COGs), Gene Ontology (GO) terms and Kyoto Encyclopedia of Gene and Genome (KEGG) annotations were also assigned to each unigene transcript. To provide a preliminary verification of the assembly and annotation results, and search for potential environmental pollution biomarkers, 15 functional genes (five antioxidase genes, two cytochrome P450 genes, three GABA receptor-related genes and five heat shock protein genes) were cloned and identified. Expressions of the 15 selected genes following fluoxetine exposure confirmed that the genes are indeed linked to environmental stress. Conclusions/Significance The C. fluminea transcriptome advances the underlying molecular understanding of this freshwater clam, provides a basis for further exploration of C. fluminea as an environmental test organism and promotes further studies on other bivalve organisms. PMID:24244519

  4. Use of the melting curve assay as a means for high-throughput quantification of Illumina sequencing libraries.

    PubMed

    Shinozuka, Hiroshi; Forster, John W

    2016-01-01

    Background. Multiplexed sequencing is commonly performed on massively parallel short-read sequencing platforms such as Illumina, and the efficiency of library normalisation can affect the quality of the output dataset. Although several library normalisation approaches have been established, none are ideal for highly multiplexed sequencing due to issues of cost and/or processing time. Methods. An inexpensive and high-throughput library quantification method has been developed, based on an adaptation of the melting curve assay. Sequencing libraries were subjected to the assay using the Bio-Rad Laboratories CFX Connect(TM) Real-Time PCR Detection System. The library quantity was calculated through summation of reduction of relative fluorescence units between 86 and 95 °C. Results.PCR-enriched sequencing libraries are suitable for this quantification without pre-purification of DNA. Short DNA molecules, which ideally should be eliminated from the library for subsequent processing, were differentiated from the target DNA in a mixture on the basis of differences in melting temperature. Quantification results for long sequences targeted using the melting curve assay were correlated with those from existing methods (R (2) > 0.77), and that observed from MiSeq sequencing (R (2) = 0.82). Discussion.The results of multiplexed sequencing suggested that the normalisation performance of the described method is equivalent to that of another recently reported high-throughput bead-based method, BeNUS. However, costs for the melting curve assay are considerably lower and processing times shorter than those of other existing methods, suggesting greater suitability for highly multiplexed sequencing applications. PMID:27602269

  5. Transcriptome Characterization for Non-Model Endangered Lycaenids, Protantigius superans and Spindasis takanosis, Using Illumina HiSeq 2500 Sequencing

    PubMed Central

    Patnaik, Bharat Bhusan; Hwang, Hee-Ju; Kang, Se Won; Park, So Young; Wang, Tae Hun; Park, Eun Bi; Chung, Jong Min; Song, Dae Kwon; Kim, Changmu; Kim, Soonok; Lee, Jae Bong; Jeong, Heon Cheon; Park, Hong Seog; Han, Yeon Soo; Lee, Yong Seok

    2015-01-01

    The Lycaenidae butterflies, Protantigius superans and Spindasis takanosis, are endangered insects in Korea known for their symbiotic association with ants. However, necessary genomic and transcriptomics data are lacking in these species, limiting conservation efforts. In this study, the P. superans and S. takanosis transcriptomes were deciphered using Illumina HiSeq 2500 sequencing. The P. superans and S. takanosis transcriptome data included a total of 254,340,693 and 245,110,582 clean reads assembled into 159,074 and 170,449 contigs and 107,950 and 121,140 unigenes, respectively. BLASTX hits (E-value of 1.0 × 10−5) against the known protein databases annotated a total of 46,754 and 51,908 transcripts for P. superans and S. takanosis. Approximately 41.25% and 38.68% of the unigenes for P. superans and S. takanosis found homologous sequences in Protostome DB (PANM-DB). BLAST2GO analysis confirmed 18,611 unigenes representing Gene Ontology (GO) terms and a total of 5259 unigenes assigned to 116 pathways for P. superans. For S. takanosis, a total of 6697 unigenes were assigned to 119 pathways using the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway database. Additionally, 382,164 and 390,516 Simple Sequence Repeats (SSRs) were compiled from the unigenes of P. superans and S. takanosis, respectively. This is the first report to record new genes and their utilization for conservation of lycaenid species population and as a reference information for closely related species. PMID:26694362

  6. De Novo Assembly of Auricularia polytricha Transcriptome Using Illumina Sequencing for Gene Discovery and SSR Marker Identification

    PubMed Central

    Zhou, Yan; Chen, Lianfu; Fan, Xiuzhi; Bian, Yinbing

    2014-01-01

    Auricularia polytricha (Mont.) Sacc., a type of edible black-brown mushroom with a gelatinous and modality-specific fruiting body, is in high demand in Asia due to its nutritional and medicinal properties. Illumina Solexa sequenceing technology was used to generate very large transcript sequences from the mycelium and the mature fruiting body of A. polytricha for gene discovery and molecular marker development. De novo assembly generated 36,483 ESTs with an N50 length of 636 bp. A total of 28,108 ESTs demonstrated significant hits with known proteins in the nr database, and 94.03% of the annotated ESTs showed the greatest similarity to A. delicata, a related species of A. polytricha. Functional categorization of the Gene Ontology (GO), Clusters of Orthologous Groups (COG) and Kyoto Encyclopedia of Genes and Genomes (KEGG) metabolic pathways revealed the conservation of genes involved in various biological processes in A. polytricha. Gene expression profile analysis indicated that a total of 2,057 ESTs were differentially expressed, including 1,020 ESTs that were up-regulated in the mycelium and 1,037 up-regulated in the fruiting body. Functional enrichment showed that the ESTs associated with biosynthesis, metabolism and assembly of proteins were more active in fruiting body development. The expression patterns of homologous transcription factors indicated that the molecular mechanisms of fruiting body formation and development were not exactly the same as for other agarics. Interestingly, an EST encoding tyrosinase was significantly up-regulated in the fruiting body, indicating that melanins accumulated during the processes of the formation of the black-brown color of the fruiting body in A. polytricha development. In addition, a total of 1,715 potential SSRs were detected in this transcriptome. The transcriptome analysis of A. polytricha provides valuable sequence resources and numerous molecular markers to facilitate further functional genomics studies and

  7. De Novo Assembly and Characterization of the Transcriptome of Seagrass Zostera marina Using Illumina Paired-End Sequencing

    PubMed Central

    Kong, Fanna; Li, Hong; Sun, Peipei; Zhou, Yang; Mao, Yunxiang

    2014-01-01

    Background The seagrass Zostera marina is a monocotyledonous angiosperm belonging to a polyphyletic group of plants that can live submerged in marine habitats. Zostera marina L. is one of the most common seagrasses and is considered a cornerstone of marine plant molecular ecology research and comparative studies. However, the mechanisms underlying its adaptation to the marine environment still remain poorly understood due to limited transcriptomic and genomic data. Principal Findings Here we explored the transcriptome of Z. marina leaves under different environmental conditions using Illumina paired-end sequencing. Approximately 55 million sequencing reads were obtained, representing 58,457 transcripts that correspond to 24,216 unigenes. A total of 14,389 (59.41%) unigenes were annotated by blast searches against the NCBI non-redundant protein database. 45.18% and 46.91% of the unigenes had significant similarity with proteins in the Swiss-Prot database and Pfam database, respectively. Among these, 13,897 unigenes were assigned to 57 Gene Ontology (GO) terms and 4,745 unigenes were identified and mapped to 233 pathways via functional annotation against the Kyoto Encyclopedia of Genes and Genomes pathway database (KEGG). We compared the orthologous gene family of the Z. marina transcriptome to Oryza sativa and Pyropia yezoensis and 11,667 orthologous gene families are specific to Z. marina. Furthermore, we identified the photoreceptors sensing red/far-red light and blue light. Also, we identified a large number of genes that are involved in ion transporters and channels including Na+ efflux, K+ uptake, Cl− channels, and H+ pumping. Conclusions Our study contains an extensive sequencing and gene-annotation analysis of Z. marina. This information represents a genetic resource for the discovery of genes related to light sensing and salt tolerance in this species. Our transcriptome can be further utilized in future studies on molecular adaptation to abiotic stress in

  8. Diversity of endophytic and rhizoplane bacterial communities associated with exotic Spartina alterniflora and native mangrove using Illumina amplicon sequencing.

    PubMed

    Hong, Youwei; Liao, Dan; Hu, Anyi; Wang, Han; Chen, Jinsheng; Khan, Sardar; Su, Jianqiang; Li, Hu

    2015-10-01

    Root-associated microbial communities are very important for biogeochemical cycles in wetland ecosystems and help to elaborate the mechanisms of plant invasions. In the estuary of Jiulong River (China), Spartina alterniflora has widely invaded Kandelia obovata-dominated habitats, offering an opportunity to study the influence of root-associated bacteria. The community structures of endophytic and rhizosphere bacteria associated with selected plant species were investigated using the barcoded Illumina paired-end sequencing technique. The diversity indices of bacteria associated with the roots of S. alterniflora were higher than those of the transition stands and K. obovata monoculture. Using principal coordinate analysis with UniFrac metrics, the comparison of β-diversity showed that all samples could be significantly clustered into 3 major groups, according to the bacteria communities of origin. Four phyla, namely Proteobacteria, Bacteroidetes, Chloroflexi, and Firmicutes, were enriched in the rhizoplane of both salt marsh plants, while they shared higher abundances of Cyanobacteria and Proteobacteria among endophytic bacteria. Members of the phyla Spirochaetes and Chloroflexi were found among the endophytic bacteria of S. alterniflora and K. obovata, respectively. One of the interesting findings was that endophytes were more sensitive in response to plant invasion than were rhizosphere bacteria. With linear discriminate analysis, we found some predominant rhizoplane and endophytic bacteria, including Methylococcales, Pseudoalteromonadacea, Clostridium, Vibrio, and Desulfovibrio, which have the potential to affect the carbon, nitrogen, and sulfur cycles. Thus, the results provide clues to the isolation of functional bacteria and the effects of root-associated microbial groups on S. alterniflora invasions. PMID:26223001

  9. De novo Transcriptome Analysis of Chinese Citrus Fly, Bactrocera minax (Diptera: Tephritidae), by High-Throughput Illumina Sequencing

    PubMed Central

    Wang, Jia; Xiong, Ke-Cai; Liu, Ying-Hong

    2016-01-01

    The Chinese citrus fly, Bactrocera minax (Enderlein), is one of the most devastating pests of citrus in the temperate areas of Asia. So far, studies involving molecular biology and physiology of B. minax are still scarce, partly because of the lack of genomic information and inability to rear this insect in laboratory. In this study, de novo assembly of a transcriptome was performed using Illumina sequencing technology. A total of 20,928,907 clean reads were obtained and assembled into 33,324 unigenes, with an average length of 908.44 bp. Unigenes were annotated by alignment against NCBI non-redundant protein (Nr), Swiss-Prot, Clusters of Orthologous Groups (COG), Gene Ontology (GO), and Kyoto Encyclopedia of Genes and Genomes Pathway (KEGG) database. Genes potentially involved in stress tolerance, including 20 heat shock protein (Hsps) genes, 26 glutathione S-transferases (GSTs) genes, and 2 ferritin subunit genes, were identified. These genes may play roles in stress tolerance in B. minax diapause stage. It has previously been found that 20E application on B. minax pupae could avert diapause, but the underlying mechanisms remain unknown. Thus, genes encoding enzymes in 20E biosynthesis pathway, including Neverland, Spook, Phantom, Disembodied, Shadow, Shade, and Cyp18a1, and genes encoding 20E receptor proteins, ecdysone receptor (EcR) and ultraspiracle (USP), were identified. The expression patterns of 20E-related genes among developmental stages and between 20E-treated and untreated pupae demonstrated their roles in diapause program. In addition, 1,909 simple sequence repeats (SSRs) were detected, which will contribute to molecular marker development. The findings in this study greatly improve our genetic understanding of B. minax, and lay the foundation for future studies on this species. PMID:27331903

  10. Serum susceptibility of bovine pasteurellas.

    PubMed Central

    Blau, K A; Ward, A C; Prieur, D J; Corbeil, L B

    1987-01-01

    In this study, the serum sensitivity of 23 P. haemolytica isolates and 18 P. multocida isolates was determined by incubating dilutions of bacteria with equal volumes of fresh or heat-inactivated bovine serum for one, two, or three hours. Clinical isolates of both Pasteurella species were resistant to serum, whereas isolates from asymptomatic cattle varied in serum susceptibility. The classical pathway of complement appeared to be the principal means of complement mediated killing as detected by incubation in the presence or absence of EGTA-MgCl2. Lyzozyme and iron saturation of serum did not greatly affect serum susceptibility with either of the Pasteurella species. PMID:3300919

  11. [Toxinology of bovine paraplegic syndrome].

    PubMed

    Sevcik, C; Brito, J C; D'Suze, G; Mijares, A J; Domínguez, M G

    1993-01-01

    A clinical entity named "Bovine Paraplegic Syndrome" ("Síndrome Parapléjico de los Bovinos") has spread alarmingly, in the cattle growing areas of the central and eastern plains of Venezuela. Approximately four million cattle are bread in the area were the disease occurs. The mortality index due to the disease ranges 5 to 25% of the animals at risk, mostly cows, pregnant or lactating. The principal characteristic of the bovine paraplegic syndrome is decubitus, ventral or sternal, in animals that make vane efforts to stand when stimulated. The diagnosis is established ruling out, clinically and with laboratory findings, that the animals are suffering known diseases with similar symptoms such as paralytic rabies, botulism and blood parasites such Trypanosoma sp., Babesia sp., and Anaplasma sp.. Death occurs always, usually after few days, and to this date there is no known treatment able to save the sick cows. In this work, we describe results that suggest the presence of a toxin in the cattle suffering and prone to suffer the syndrome; it is a natural toxin produced by ruminal bacteria. In squid giant axons under voltage clamp conditions, this toxin is very specific to block sodium current during nerve electrical activity. PMID:8085407

  12. Adipogenesis of bovine perimuscular preadipocytes

    SciTech Connect

    Taniguchi, Masaaki; Le Luo Guan; Zhang Bing; Dodson, Michael V.; Okine, Erasmus; Moore, Stephen S.

    2008-02-01

    In this study, non-transformed progeny adipofibroblasts, derived from mature adipocyte dedifferentiation, was used as a novel in vitro model to study adipogenic gene expression in cattle. Adipofibroblasts from dedifferentiated mature perimuscular fat (PMF) tissue were cultured with differentiation stimulants until the cells exhibited morphological differentiation. Treated cells were harvested from day 2 to 16 for RNA extraction, whereas control cells were cultured without addition of stimulants. Results from time course gene expression assays by quantitative real-time PCR revealed that peroxisome proliferator-activated receptor gamma (PPAR-{gamma}), sterol regulatory element binding protein 1 (SREBP-1) and their six down-stream genes were co-expressed at day 2 post-differentiation induction. When compared to other adipogenesis culture systems, the adipogenic gene expression of bovine PMF adipofibroblasts culture was different, especially to the rodent model. Collectively, these results demonstrated PPAR-{gamma} and SREBP-1 cooperatively play a key role to regulate the re-differentiation of bovine adipofibroblasts, during early conversion stages in vitro.

  13. Bovine somatic cell nuclear transfer.

    PubMed

    Ross, Pablo J; Cibelli, Jose B

    2010-01-01

    Somatic cell nuclear transfer (SCNT) is a technique by which the nucleus of a differentiated cell is introduced into an oocyte from which its genetic material has been removed by a process called enucleation. In mammals, the reconstructed embryo is artificially induced to initiate embryonic development (activation). The oocyte turns the somatic cell nucleus into an embryonic nucleus. This process is called nuclear reprogramming and involves an important change of cell fate, by which the somatic cell nucleus becomes capable of generating all the cell types required for the formation of a new individual, including extraembryonic tissues. Therefore, after transfer of a cloned embryo to a surrogate mother, an offspring genetically identical to the animal from which the somatic cells where isolated, is born. Cloning by nuclear transfer has potential applications in agriculture and biomedicine, but is limited by low efficiency. Cattle were the second mammalian species to be cloned after Dolly the sheep, and it is probably the most widely used species for SCNT experiments. This is, in part due to the high availability of bovine oocytes and the relatively higher efficiency levels usually obtained in cattle. Given the wide utilization of this species for cloning, several alternatives to this basic protocol can be found in the literature. Here we describe a basic protocol for bovine SCNT currently being used in our laboratory, which is amenable for the use of the nuclear transplantation technique for research or commercial purposes. PMID:20336522

  14. Snat: a SNP annotation tool for bovine by integrating various sources of genomic information

    PubMed Central

    2011-01-01

    Background Most recently, with maturing of bovine genome sequencing and high throughput SNP genotyping technologies, a large number of significant SNPs associated with economic important traits can be identified by genome-wide association studies (GWAS). To further determine true association findings in GWAS, the common strategy is to sift out most promising SNPs for follow-up replication studies. Hence it is crucial to explore the functional significance of the candidate SNPs in order to screen and select the potential functional ones. To systematically prioritize these statistically significant SNPs and facilitate follow-up replication studies, we developed a bovine SNP annotation tool (Snat) based on a web interface. Results With Snat, various sources of genomic information are integrated and retrieved from several leading online databases, including SNP information from dbSNP, gene information from Entrez Gene, protein features from UniProt, linkage information from AnimalQTLdb, conserved elements from UCSC Genome Browser Database and gene functions from Gene Ontology (GO), KEGG PATHWAY and Online Mendelian Inheritance in Animals (OMIA). Snat provides two different applications, including a CGI-based web utility and a command-line version, to access the integrated database, target any single nucleotide loci of interest and perform multi-level functional annotations. For further validation of the practical significance of our study, SNPs involved in two commercial bovine SNP chips, i.e., the Affymetrix Bovine 10K chip array and the Illumina 50K chip array, have been annotated by Snat, and the corresponding outputs can be directly downloaded from Snat website. Furthermore, a real dataset involving 20 identified SNPs associated with milk yield in our recent GWAS was employed to demonstrate the practical significance of Snat. Conclusions To our best knowledge, Snat is one of first tools focusing on SNP annotation for livestock. Snat confers researchers with a

  15. Identification of miRNAs associated with sexual maturity in chicken ovary by Illumina small RNA deep sequencing

    PubMed Central

    2013-01-01

    Background MicroRNAs have been suggested to play important roles in the regulation of gene expression in various biological processes. To investigate the function of miRNAs in chicken ovarian development and folliculogenesis, two small RNA libraries constructed from sexually mature (162-day old) and immature (42-day old) ovary tissues of Single Comb White Leghorn chicken were sequenced using Illumina small RNA deep sequencing. Results In the present study, 14,545,100 and 14,774,864 clean reads were obtained from sexually mature (162-d) and sexually immature (42-d) ovaries, respectively. In total, 202 known miRNAs were identified, and 93 of them were found to be significantly differentially expressed: 42 miRNAs were up-regulated and 51 miRNAs were down-regulated in the mature ovary compared to the immature ovary. Among the up-regulated miRNAs, gga-miR-1a has the largest fold-change (6.405-fold), while gga-miR-375 has the largest fold-change (11.345-fold) among the down-regulated miRNAs. The three most abundant miRNAs in the chicken ovary are gga-miR-10a, gga-let-7 and gga-miR-21. Five differentially expressed miRNAs (gga-miR-1a, 21, 26a, 137 and 375) were validated by real-time quantitative RT-PCR (qRT-PCR). Furthermore, the expression patterns of the five miRNAs were analyzed in different developmental stages of chicken ovary and follicles of various sizes. Conclusion The present study provides the first miRNA profile in sexually immature and mature chicken ovaries. Some miRNAs such as gga-miR-1a and gga-miR-21are expressed differentially in immature and mature chicken ovaries as well as among different sized follicles, suggesting an important role in the follicular growth or ovulation mechanism in the chicken. PMID:23705682

  16. Muscarine binding sites in bovine adrenal medulla.

    PubMed

    Barron, B A; Murrin, L C; Hexum, T D

    1986-03-18

    The presence of muscarinic binding sites in the bovine adrenal medulla was investigated using [3H]QNB and the bovine adrenal medulla. Scatchard analysis combined with computer analysis yielded data consistent with a two binding site configuration. KDs of 0.15 and 14 nM and Bmax s of 29 and 210 fmol/mg protein, respectively, were observed. Displacement of [3H]QNB by various cholinergic agents is, in order of decreasing potency: QNB, dexetimide, atropine, scopolamine, imipramine, desipramine, oxotremorine, pilocarpine, acetylcholine, methacholine and carbachol. These results demonstrate the presence of more than one muscarine binding site in the bovine adrenal gland. PMID:3709656

  17. Growth and metabolism of murine and bovine embryos in bovine uterine flushing-supplemented culture media.

    PubMed Central

    Rondeau, M; Guay, P; Goff, A K; Cooke, G M

    1996-01-01

    The aim of this study was to compare the development and metabolic activity of cultured murine and bovine embryos in 2 standard media (HAM F-10 and RPMI) in the presence or absence of bovine uterine flushings. Murine morulae (n = 653) and day 7 bovine embryos (n = 273) were cultured for 18 h or 36 h in either HAM F-10 or RPMI in the presence or absence of bovine uterine flushings. After culture, the development, quality, and metabolic activity (glucose utilization or methionine uptake and incorporation) of embryos was assessed. It was found that HAM F-10 (without uterine flushings) was a more suitable medium than RPMI for optimal development and metabolism of murine and bovine embryos. Poor quality and development, as well as decreased metabolism, were evident after culture of murine embryos in RPMI; in contrast, this medium had no adverse effects on bovine embryos in culture. Supplementation of HAM F-10 with bovine uterine flushings improved the growth of murine embryos and the protein synthesis (as measured by an increased methionine incorporation) for both murine and bovine embryos. However, supplementation with bovine uterine flushings could not overcome deficiencies of an inappropriate medium (RPMI) for murine embryos. Supplementation of a well-defined culture medium with uterine flushings increased metabolism of embryos in culture, and thus might help to increase pregnancy rates after transfer of such embryos to recipient cows. PMID:8825988

  18. Elastomechanical properties of bovine veins.

    PubMed

    Rossmann, Jenn Stroud

    2010-02-01

    Veins have historically been discussed in qualitative, relative terms: "more compliant" than arteries, subject to "lower pressures". The structural and compositional differences between arteries and veins are directly related to the different functions of these vessels. Veins are often used as grafts to reroute flow from atherosclerotic arteries, and venous elasticity plays a role in the development of conditions such as varicose veins and valvular insufficiency. It is therefore of clinical interest to determine the elastomechanical properties of veins. In the current study, both tensile and vibration testing are used to obtain elastic moduli of bovine veins. Representative stress-strain data are shown, and the mechanical and failure properties reported. Nonlinear and viscoelastic behavior is observed, though most properties show little strain rate dependence. These data suggest parameters for constitutive modeling of veins and may inform the design and testing of prosthetic venous valves as well as vein grafts. PMID:20129420

  19. Bovine Mastitis: Frontiers in Immunogenetics

    PubMed Central

    Thompson-Crispi, Kathleen; Atalla, Heba; Miglior, Filippo; Mallard, Bonnie A.

    2014-01-01

    Mastitis is one of the most prevalent and costly diseases in the dairy industry with losses attributable to reduced milk production, discarded milk, early culling, veterinary services, and labor costs. Typically, mastitis is an inflammation of the mammary gland most often, but not limited to, bacterial infection, and is characterized by the movement of leukocytes and serum proteins from the blood to the site of infection. It contributes to compromised milk quality and the potential spread of antimicrobial resistance if antibiotic treatment is not astutely applied. Despite the implementation of management practises and genetic selection approaches, bovine mastitis control continues to be inadequate. However, some novel genetic strategies have recently been demonstrated to reduce mastitis incidence by taking advantage of a cow’s natural ability to make appropriate immune responses against invading pathogens. Specifically, dairy cattle with enhanced and balanced immune responses have a lower occurrence of disease, including mastitis, and they can be identified and selected for using the high immune response (HIR) technology. Enhanced immune responsiveness is also associated with improved response to vaccination, increased milk, and colostrum quality. Since immunity is an important fitness trait, beneficial associations with longevity and reproduction are also often noted. This review highlights the genetic regulation of the bovine immune system and its vital contributions to disease resistance. Genetic selection approaches currently used in the dairy industry to reduce the incidence of disease are reviewed, including the HIR technology, genomics to improve disease resistance or immune response, as well as the Immunity+™ sire line. Improving the overall immune responsiveness of cattle is expected to provide superior disease resistance, increasing animal welfare and food quality while maintaining favorable production levels to feed a growing population. PMID

  20. Bovine mastitis: frontiers in immunogenetics.

    PubMed

    Thompson-Crispi, Kathleen; Atalla, Heba; Miglior, Filippo; Mallard, Bonnie A

    2014-01-01

    Mastitis is one of the most prevalent and costly diseases in the dairy industry with losses attributable to reduced milk production, discarded milk, early culling, veterinary services, and labor costs. Typically, mastitis is an inflammation of the mammary gland most often, but not limited to, bacterial infection, and is characterized by the movement of leukocytes and serum proteins from the blood to the site of infection. It contributes to compromised milk quality and the potential spread of antimicrobial resistance if antibiotic treatment is not astutely applied. Despite the implementation of management practises and genetic selection approaches, bovine mastitis control continues to be inadequate. However, some novel genetic strategies have recently been demonstrated to reduce mastitis incidence by taking advantage of a cow's natural ability to make appropriate immune responses against invading pathogens. Specifically, dairy cattle with enhanced and balanced immune responses have a lower occurrence of disease, including mastitis, and they can be identified and selected for using the high immune response (HIR) technology. Enhanced immune responsiveness is also associated with improved response to vaccination, increased milk, and colostrum quality. Since immunity is an important fitness trait, beneficial associations with longevity and reproduction are also often noted. This review highlights the genetic regulation of the bovine immune system and its vital contributions to disease resistance. Genetic selection approaches currently used in the dairy industry to reduce the incidence of disease are reviewed, including the HIR technology, genomics to improve disease resistance or immune response, as well as the Immunity(+)™ sire line. Improving the overall immune responsiveness of cattle is expected to provide superior disease resistance, increasing animal welfare and food quality while maintaining favorable production levels to feed a growing population. PMID

  1. Aerosol stability of bovine adenovirus type 3.

    PubMed Central

    Elazhary, M A; Derbyshire, J B

    1979-01-01

    The WBR-1 strain of bovine adenovirus type 3 was suspended in Eagle's medium or bovine nasal secretion and atomized into a rotating drum at temperatures of 6 degrees C or 32 degrees C and relative humidities of 30% or 90%. Impinger samples of the aerosols were collected seven minutes, one, two and three hours postgeneration, and titrated for infectivity in embryonic bovine kidney cell cultures. Under certain conditions of temperature and relative humidity, the virus was more stable in aerosols of Eagle's medium than in nasal secretion. The bovine adenovirus was usually inactivated more rapidly at 30% relative humidity than at 90% relative humidity and during aging of the aerosols the virus was inactivated more rapidly at 32 degrees C than at 6 degrees C. PMID:226247

  2. Genome-wide association study identifies novel loci associated with resistance to bovine tuberculosis

    PubMed Central

    Bermingham, M L; Bishop, S C; Woolliams, J A; Pong-Wong, R; Allen, A R; McBride, S H; Ryder, J J; Wright, D M; Skuce, R A; McDowell, S WJ; Glass, E J

    2014-01-01

    Tuberculosis (TB) caused by Mycobacterium bovis is a re-emerging disease of livestock that is of major economic importance worldwide, as well as being a zoonotic risk. There is significant heritability for host resistance to bovine TB (bTB) in dairy cattle. To identify resistance loci for bTB, we undertook a genome-wide association study in female Holstein–Friesian cattle with 592 cases and 559 age-matched controls from case herds. Cases and controls were categorised into distinct phenotypes: skin test and lesion positive vs skin test negative on multiple occasions, respectively. These animals were genotyped with the Illumina BovineHD 700K BeadChip. Genome-wide rapid association using linear and logistic mixed models and regression (GRAMMAR), regional heritability mapping (RHM) and haplotype-sharing analysis identified two novel resistance loci that attained chromosome-wise significance, protein tyrosine phosphatase receptor T (PTPRT; P=4.8 × 10−7) and myosin IIIB (MYO3B; P=5.4 × 10−6). We estimated that 21% of the phenotypic variance in TB resistance could be explained by all of the informative single-nucleotide polymorphisms, of which the region encompassing the PTPRT gene accounted for 6.2% of the variance and a further 3.6% was associated with a putative copy number variant in MYO3B. The results from this study add to our understanding of variation in host control of infection and suggest that genetic marker-based selection for resistance to bTB has the potential to make a significant contribution to bTB control. PMID:24496092

  3. Bovine TLR2 and TLR4 mediate Cryptosporidium parvum recognition in bovine intestinal epithelial cells.

    PubMed

    Yang, Zhengtao; Fu, Yunhe; Gong, Pengtao; Zheng, Jingtong; Liu, Li; Yu, Yuqiang; Li, Jianhua; Li, He; Yang, Ju; Zhang, Xichen

    2015-08-01

    Cryptosporidium parvum (C. parvum) is an intestinal parasite that causes diarrhea in neonatal calves. It results in significant morbidity of neonatal calves and economic losses for producers worldwide. Innate resistance against C. parvum is thought to depend on engagement of pattern recognition receptors. However, the role of innate responses to C. parvum has not been elucidated in bovine. The aim of this study was to evaluate the role of TLRs in host-cell responses during C. parvum infection of cultured bovine intestinal epithelial cells. The expressions of TLRs in bovine intestinal epithelial cells were detected by qRT-PCR. To determine which, if any, TLRs may play a role in the response of bovine intestinal epithelial cells to C. parvum, the cells were stimulated with C. parvum and the expression of TLRs were tested by qRT-PCR. The expression of NF-κB was detected by western blotting. Further analyses were carried out in bovine TLRs transfected HEK293 cells and by TLRs-DN transfected bovine intestinal epithelial cells. The results showed that bovine intestinal epithelial cells expressed all known TLRs. The expression of TLR2 and TLR4 were up-regulated when bovine intestinal epithelial cells were treated with C. parvum. Meanwhile, C. parvum induced IL-8 production in TLR2 or TLR4/MD-2 transfected HEK293 cells. Moreover, C. parvum induced NF-κB activation and cytokine expression in bovine intestinal epithelial cells. The induction of NF-κB activation and cytokine expression by C. parvum were reduced in TLR2-DN and TLR4-DN transfected cells. The results showed that bovine intestinal epithelial cells expressed all known TLRs, and bovine intestinal epithelial cells recognized and responded to C. parvum via TLR2 and TLR4. PMID:26048276

  4. Activation of bovine neutrophils by Brucella spp.

    PubMed

    Keleher, Lauren L; Skyberg, Jerod A

    2016-09-01

    Brucellosis is a globally important zoonotic infectious disease caused by gram negative bacteria of the genus Brucella. While many species of Brucella exist, Brucella melitensis, Brucella abortus, and Brucella suis are the most common pathogens of humans and livestock. The virulence of Brucella is largely influenced by its ability to evade host factors, including phagocytic killing mechanisms, which are critical for the host response to infection. The aim of this study was to characterize the bovine neutrophil response to virulent Brucella spp. Here, we found that virulent strains of smooth B. abortus, B. melitensis, B. suis, and virulent, rough, strains of Brucella canis possess similar abilities to resist killing by resting, or IFN-γ-activated, bovine neutrophils. Bovine neutrophils responded to infection with a time-dependent oxidative burst that varied little between Brucella spp. Inhibition of TAK1, or SYK kinase blunted the oxidative burst of neutrophils in response to Brucella infection. Interestingly, Brucella spp. did not induce robust death of bovine neutrophils. These results indicate that bovine neutrophils respond similarly to virulent Brucella spp. In addition, virulent Brucella spp., including naturally rough strains of B. canis, have a conserved ability to resist killing by bovine neutrophils. PMID:27436438

  5. Characterization of a novel citrus tristeza virus genotype within three cross-protecting source GFMS12 sub-isolates in South Africa by means of Illumina sequencing.

    PubMed

    Zablocki, Olivier; Pietersen, Gerhard

    2014-08-01

    Tristeza disease (caused by citrus tristeza virus, CTV) is currently controlled in South Africa by means of cross-protection. In this study, we characterized the CTV populations of three grapefruit mild strain 12 (GFMS12) single-aphid-transmission-derived sub-isolates at the whole-genome level using Illumina sequencing technology. A novel South African isolate (CT-ZA3, of the T68 genotype) was shown to be the dominant genotype in all GFMS12 sub-isolates tested, along with reads unique to various other genotypes occurring as minor components. Uncertainty remains as to the significance of these minor components. PMID:24623089

  6. RNA-Seq profiling of single bovine oocyte transcript abundance and its modulation by cytoplasmic polyadenylation

    PubMed Central

    Reyes, Juan M; Chitwood, James L; Ross, Pablo J

    2014-01-01

    Molecular changes occurring during mammalian oocyte maturation are partly regulated by cytoplasmic polyadenylation (CP) and affect oocyte quality, yet the extent of CP activity during oocyte maturation remains unknown. Single bovine oocyte RNA sequencing (RNA-Seq) was performed to examine changes in transcript abundance during in vitro oocyte maturation in cattle. Polyadenylated RNA from individual germinal-vesicle and metaphase-II oocytes was amplified and processed for Illumina sequencing, producing approximately 30 million reads per replicate for each sample type. A total of 10,494 genes were found to be expressed, of which 2,455 were differentially expressed (adjusted P<0.05 and fold change >2) between stages, with 503 and 1,952 genes respectively increasing and decreasing in abundance. Differentially expressed genes with complete 3’-untranslated-region sequence (279 increasing and 918 decreasing in polyadenylated transcript abundance) were examined for the presence, position, and distribution of motifs mediating CP, revealing enrichment (85%) and lack there of (18%) in up- and down-regulated genes, respectively. Examination of total and polyadenylated RNA abundance by quantitative PCR validated these RNA-Seq findings. The observed increases in polyadenylated transcript abundance within the RNA-Seq data are likely due to CP, providing novel insight into targeted transcripts and resultant differential gene expression profiles that contribute to oocyte maturation. PMID:25560149

  7. Prevalence of bovine virus diarrhoea and infectious bovine rhinotracheitis antibodies in Nigerian sheep and goats.

    PubMed

    Taylor, W P; Okeke, A N; Shidali, N N

    1977-08-01

    Neutralising antibodies to bovine virus diarrhoea virus were commoner in Nigerian sheep than goats while precipitating antibodies offered an alternative but less reliable indicator of previous infection. In contrast, neutralising antibodies to infectious bovine rhinotracheitis virus were more common in goats than sheep. These findings are discussed in relation to infectivity rates in cattle and general husbandry practices. PMID:410130

  8. Complete genome sequence of a bovine viral diarrhea virus 2 from commercial fetal bovine serum.

    PubMed

    Liu, Hua; Li, Yan; Gao, Mingchun; Wen, Kai; Jia, Ying; Liu, Xiaomei; Zhang, Wenlong; Ma, Bo; Wang, Junwei

    2012-09-01

    We isolated a bovine viral diarrhea virus (BVDV) from commercial fetal bovine serum and designated it HLJ-10. The complete genome is 12,284 nucleotides (nt); the open reading frame is 11,694 nt, coding 3,898 amino acids. Phylogenetic analysis indicated that this strain belongs to BVDV group 2. PMID:22923795

  9. Bovine viral diarrhea virus: involvement in bovine respiratory disease and diagnostic challenges

    Technology Transfer Automated Retrieval System (TEKTRAN)

    This paper reviews the contribution of bovine viral diarrhea viruses (BVDV) to the development of Bovine Respiratory Disease (BRD). Veterinarians and producers generally consider BRD as one of the most significant diseases affecting production in the cattle industry. BRD can affect the performance (...

  10. The Contribution of Infections with Bovine Viral Diarrhea Viruses to Bovine Respiratory Disease

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The contribution of bovine viral diarrhea viruses (BVDV) to the development of bovine respiratory disease is the sum of a number of different factors. These factors include the contribution of acute uncomplicated BVDV infections, the high incidence of respiratory disease in animals persistently inf...

  11. Ultrastructure of bovine sperm chromatin.

    PubMed

    Filho, Romualdo Morandi; Beletti, Marcelo Emilio; de Oliveira, Fabio

    2015-12-01

    Mammalian semen chromatin comprises DNA, protamine, and, at lower levels, other proteins. This constitution confers intense compaction to the chromatin, helping to protect the DNA and causing the head of the sperm to be very small, facilitating the safe transport of its genetic contents. It is known that changes in the sperm chromatin compaction lead to fertility problems in bulls, justifying studies of this structure. Although there are theoretical models of sperm chromatin because of its high compaction, there is no morphological evidence of such models. The aim of this study was to demonstrate the ultrastructure of bovine sperm chromatin in an attempt to corroborate the theoretical chromatin models existing today. The isolated bull sperm heads had their chromatin partially unpacked by chemical treatment using sodium dodecyl sulfate (SDS) and dithiothreitol (DTT) and were then embedded in Epon resin. Using an ultramicrotome, ultrathin sections were obtained, which were contrasted with uranyl acetate and lead citrate, and then viewed under transmission electron microscopy. The methodology used allowed the visualization of toroidal structures interconnected by a filamentous nuclear matrix, which is entirely consistent with the most current theoretical models. PMID:26515508

  12. Bovine colostrum: an emerging nutraceutical.

    PubMed

    Bagwe, Siddhi; Tharappel, Leo J P; Kaur, Ginpreet; Buttar, Harpal S

    2015-09-01

    Nutraceutical, a term combining the words "nutrition" and "pharmaceuticals", is a food or food product that provides health benefits as an adjuvant or alternative therapy, including the treatment and prevention of infectious diseases in children and adults. There is emerging evidence that bovine colostrum (BC) may be one of the promising nutraceuticals which can prevent or mitigate various diseases in newborns and adults. Immunity-related disorders are one of the leading causes of mortality in the world. BC is rich in immunity, growth and antimicrobial factors, which promote tissue growth and the maturation of digestive tract and immune function in neonatal animals and humans. The immunoglobulins and lactoferrin present in colostrum are known to build natural immunity in newborns which helps to reduce the mortality rate in this population. Also, the side-effect profile of colostrum proteins and possible lactose intolerance is relatively less in comparison with milk. In general, BC is considered safe and well tolerated. Since colostrum has several important nutritional constituents, well-designed, double-blind, placebo-controlled studies with colostrum products should be conducted to widen its therapeutic use. The objectives of this review are to create awareness about the nutraceutical properties of colostrum and to discuss the various ongoing alternative treatments of colostrum and its active ingredients as well as to address colostrum's future nutraceutical and therapeutic implications in humans. PMID:25781716

  13. Genetics of bovine abomasal displacement.

    PubMed

    Zerbin, Ina; Lehner, Stefanie; Distl, Ottmar

    2015-04-01

    Displacement of the abomasum (DA) is a common inherited condition in Holstein cows. This article reviews the genetics of DA including risk factors, genetic parameters and molecular genetic results. Breeds other than Holsteins affected by DA include Guernseys, Jerseys, Brown Swiss, Ayrshires and Simmental-Red Holsteins. In most DA cases, left displacements of the abomasum (LDA) are seen. Lactation incidence rates are higher for DA in first lactation Holsteins compared to later lactations. For Holstein cows, heritability estimates for DA are between 0.03 and 0.53. Genetic correlation estimates among DA and milk production traits range from positive to negative. Genome-wide significant genomic regions associated with LDA are located on bovine chromosomes (BTA) 1, 3, 11, 20 and 23. Motilin-associated single nucleotide polymorphisms on BTA23 exhibit a functional relationship with LDA. Pathways for deposition of calcium, insulin-dependent diabetes mellitus and synaptic transmission are significantly related to LDA in Holsteins. Deciphering the DA-associated genomic regions and genes may be an important step in the quest to understand the underlying disease-causing mechanisms and in unravelling mutations with a causal relationship to DA. PMID:25840863

  14. Transcriptome Profile of the Asian Giant Hornet (Vespa mandarinia) Using Illumina HiSeq 4000 Sequencing: De Novo Assembly, Functional Annotation, and Discovery of SSR Markers.

    PubMed

    Patnaik, Bharat Bhusan; Park, So Young; Kang, Se Won; Hwang, Hee-Ju; Wang, Tae Hun; Park, Eun Bi; Chung, Jong Min; Song, Dae Kwon; Kim, Changmu; Kim, Soonok; Lee, Jae Bong; Jeong, Heon Cheon; Park, Hong Seog; Han, Yeon Soo; Lee, Yong Seok

    2016-01-01

    Vespa mandarinia found in the forests of East Asia, including Korea, occupies the highest rank in the arthropod food web within its geographical range. It serves as a source of nutrition in the form of Vespa amino acid mixture and is listed as a threatened species, although no conservation measures have been implemented. Here, we performed de novo assembly of the V. mandarinia transcriptome by Illumina HiSeq 4000 sequencing. Over 60 million raw reads and 59,184,811 clean reads were obtained. After assembly, a total of 66,837 unigenes were clustered, 40,887, 44,455, and 22,390 of which showed homologous matches against the PANM, Unigene, and KOG databases, respectively. A total of 15,675 unigenes were assigned to Gene Ontology terms, and 5,132 unigenes were mapped to 115 KEGG pathways. The zinc finger domain (C2H2-like), serine/threonine/dual specificity protein kinase domain, and RNA recognition motif domain were among the top InterProScan domains predicted for V. mandarinia sequences. Among the unigenes, we identified 534,922 cDNA simple sequence repeats as potential markers. This is the first transcriptomic analysis of the wasp V. mandarinia using Illumina HiSeq 4000. The obtained datasets should promote the search for new genes to understand the physiological attributes of this wasp. PMID:26881195

  15. Comparative clinical sample preparation of DNA and RNA viral nucleic acids for a commercial deep sequencing system (Illumina MiSeq(®)).

    PubMed

    Ullmann, Leila Sabrina; de Camargo Tozato, Claudia; Malossi, Camila Dantas; da Cruz, Tais Fukuta; Cavalcante, Raíssa Vasconcelos; Kurissio, Jacqueline Kazue; Cagnini, Didier Quevedo; Rodrigues, Marianna Vaz; Biondo, Alexander Welker; Araujo, João Pessoa

    2015-08-01

    Sequence-independent methods for viral discovery have been widely used for whole genome sequencing of viruses. Different protocols for viral enrichment, library preparation and sequencing have increasingly been more available and at lower costs. However, no study to date has focused on optimization of viral sample preparation for commercial deep sequencing. Accordingly, the aim of the present study was to evaluate an In-House enzymatic protocol for double-stranded DNA (dsDNA) synthesis and also compare the use of a commercially available kit protocol (Nextera XT, Illumina Inc, San Diego, CA, USA) and its combination with a library quantitation kit (Kapa, Kapa Biosystems, Wilmington, MA, USA) for deep sequencing (Illumina Miseq). Two RNA viruses (canine distemper virus and dengue virus) and one ssDNA virus (porcine circovirus type 2) were tested with the optimized protocols. The tested method for dsDNA synthesis has shown satisfactory results and may be used in laboratory setting, particularly when enzymes are already available. Library preparation combining commercial kits (Nextera XT and Kapa) has yielded more reads and genome coverage, probably due to a lack of small fragment recovering at the normalization step of Nextera XT. In addition, libraries may be diluted or concentrated to provide increase on genome coverage with Kapa quantitation. PMID:25901649

  16. Transcriptome Profile of the Asian Giant Hornet (Vespa mandarinia) Using Illumina HiSeq 4000 Sequencing: De Novo Assembly, Functional Annotation, and Discovery of SSR Markers

    PubMed Central

    Park, So Young; Kang, Se Won; Hwang, Hee-Ju; Wang, Tae Hun; Park, Eun Bi; Chung, Jong Min; Song, Dae Kwon; Kim, Changmu; Kim, Soonok; Lee, Jae Bong; Jeong, Heon Cheon; Park, Hong Seog; Han, Yeon Soo; Lee, Yong Seok

    2016-01-01

    Vespa mandarinia found in the forests of East Asia, including Korea, occupies the highest rank in the arthropod food web within its geographical range. It serves as a source of nutrition in the form of Vespa amino acid mixture and is listed as a threatened species, although no conservation measures have been implemented. Here, we performed de novo assembly of the V. mandarinia transcriptome by Illumina HiSeq 4000 sequencing. Over 60 million raw reads and 59,184,811 clean reads were obtained. After assembly, a total of 66,837 unigenes were clustered, 40,887, 44,455, and 22,390 of which showed homologous matches against the PANM, Unigene, and KOG databases, respectively. A total of 15,675 unigenes were assigned to Gene Ontology terms, and 5,132 unigenes were mapped to 115 KEGG pathways. The zinc finger domain (C2H2-like), serine/threonine/dual specificity protein kinase domain, and RNA recognition motif domain were among the top InterProScan domains predicted for V. mandarinia sequences. Among the unigenes, we identified 534,922 cDNA simple sequence repeats as potential markers. This is the first transcriptomic analysis of the wasp V. mandarinia using Illumina HiSeq 4000. The obtained datasets should promote the search for new genes to understand the physiological attributes of this wasp. PMID:26881195

  17. Ruminant alphaherpesviruses related to bovine herpesvirus 1.

    PubMed

    Thiry, Julien; Keuser, Véronique; Muylkens, Benoît; Meurens, François; Gogev, Sacha; Vanderplasschen, Alain; Thiry, Etienne

    2006-01-01

    Herpesviruses have mainly co-evolved with their hosts for millions of years. Consequently, different related host species may have been infected by various genetically related herpesviruses. Illustrating this concept, several ruminant alphaherpesviruses have been shown to form a cluster of viruses closely related to bovine herpesvirus 1 (BoHV-1): namely bovine herpesvirus 5, bubaline herpesvirus 1, caprine herpesvirus 1, cervid herpesviruses 1 and 2 and elk herpesvirus 1. These viruses share common antigenic properties and the serological relationships between them can be considered as a threat to BoHV-1 eradication programmes. BoHV-1 is a herpesvirus responsible for infectious bovine rhinotracheitis, which is a disease of major economic concern. In this article, the genetic properties of these ruminant alphaherpesviruses are reviewed on a comparative basis and the issue of interspecific recombination is assessed. The pathogenesis of these infections is described with emphasis on the host range and crossing of the host species barrier. Indeed, the non bovine ruminant species susceptible to these ruminant alphaherpesviruses may be potential BoHV-1 reservoirs. The differential diagnosis of these related infections is also discussed. In addition, available epidemiological data are used to assess the potential of cross-infection in ruminant populations. A better knowledge of these ruminant alphaherpesvirus infections is essential to successfully control infectious bovine rhinotracheitis. PMID:16472518

  18. Bovine Fetal Inoculations with Calf Rotavirus

    PubMed Central

    Schlafer, D. H.; Schultz, R. D.; Scott, F. W.; Duncan, J. R.

    1979-01-01

    The serological and histopathological responses of bovine fetuses to in utero inoculation with virulent and attenuated strains of the calf rotavirus (reovirus-like agent of neonatal calf diarrhea) are described. Thirteen bovine fetuses, 63 to 190 days of gestation, were inoculated in utero with attenuated (three fetuses) or field strain virus (nine fetuses) or both (one fetus). Serum-neutralizing antibody titers ranging from 1:16 to > 1:256 were detected in six of eight fetuses tested, demonstrating the ability of the bovine fetus to respond immunologically to this agent. The youngest fetus in the series was inoculated at 63 days of gestation and developed a titer of 128 in 64 days. This represents the earliest stage of gestation at which a bovine fetus has been inoculated with a bovine virus and found to produce antibody to it. Serum neutralizing titers in six of the eight dams tested increased significantly following the inoculations of their fetuses in utero. Histological changes associated with viral replication and antigenic stimulation of the lymphoreticular system were observed. Pneumonic lesions consisting of both local and diffuse lymphoreticular proliferation were present in five of the nine fetuses that were alive at slaughter. Gliosis and perivascular cuffing were noted in the brains of two of these fetuses and meningitis was seen in one. No evidence of teratogenic change was found. ImagesFig. 2.Fig. 3.Fig. 4.Fig. 5.Fig. 6.Fig. 7. PMID:232853

  19. Recent Progress in Cryopreservation of Bovine Oocytes

    PubMed Central

    Hochi, Shinichi

    2014-01-01

    Principle of oocyte cryoinjury is first overviewed and then research history of cryopreservation using bovine oocytes is summarized for the last two decades with a few special references to recent progresses. Various types of cryodevices have been developed to accelerate the cooling rate and applied to the oocytes from large domestic species enriched with cytoplasmic lipid droplets. Two recent approaches include the qualitative improvement of IVM oocytes prior to the vitrification and the short-term recovery culture of vitrified-warmed oocytes prior to the subsequent IVF. Supplementation of L-carnitine to IVM medium of bovine oocytes has been reported to reduce the amount of cytoplasmic lipid droplets and improve the cryotolerance of the oocytes, but it is still controversial whether the positive effect of L-carnitine is reproducible. Incidence of multiple aster formation, a possible cause for low developmental potential of vitrified-warmed bovine oocytes, was inhibited by a short-term culture of the postwarm oocytes in the presence of Rho-associated coiled-coil kinase (ROCK) inhibitor. Use of an antioxidant α-tocopherol, instead of the ROCK inhibitor, also supported the revivability of the postwarm bovine oocytes. Further improvements of the vitrification procedure, combined with pre- and postvitrification chemical treatment, would overcome the high sensitivity of bovine oocytes to cryopreservation. PMID:24738063

  20. Evaluation of experimental vaccines for bovine viral diarrhea in bovines, ovines and guinea pigs.

    PubMed

    Fernández, F; Costantini, V; Barrandeguy, M; Parreño, V; Schiappacassi, G; Maliandi, F; Leunda, M; Odeón, A

    2009-01-01

    The bovine viral diarrhea virus (BVDV) infection control should be based on elimination of persistently infected animals and on immunization through vaccination, to prevent fetal infection. However, the efficacy of inactivated BVDV vaccines is variable due to its low immunogenicity. This study evaluated the humoral immune response against homologous and heterologous strains of 7 inactivated BVDV vaccines, in bovines and two experimental models (ovine and guinea pig) which might be used to test candidate vaccines. Vaccines formulated with BVDV Singer, Oregon, NADL, and multivalent, induced seroconversion in the three animal models studied, reaching antibody titres higher than 2. Vaccine containing 125C -genotype 2- only induced a low antibody response in ovine, while VS-115 NCP vaccine was not immunogenic. Furthermore, bovine sera at 60 dpv presented homologous as well as heterologous antibody response, indicating a high degree of cross-reactivity among the strains studied. However, when bovine sera were tested against the Argentine field strain 00-693, they showed the lowest levels of cross-reactivity, suggesting the need of continued surveillance to identify and characterize emerging field BVDV strains. Finally, optimal correlations between bovine-ovine and bovine-guinea pig models were observed, indicating that two alternative species could replace bovines when testing the immunogenicity of BVDV candidate vaccines. PMID:19623897

  1. Parthenogenetic activation of bovine oocytes using bovine and murine phospholipase C zeta

    PubMed Central

    Ross, Pablo J; Beyhan, Zeki; Iager, Amy E; Yoon, Sook-Young; Malcuit, Christopher; Schellander, Karl; Fissore, Rafael A; Cibelli, Jose B

    2008-01-01

    Background During natural fertilization, sperm fusion with the oocyte induces long lasting intracellular calcium oscillations which in turn are responsible for oocyte activation. PLCZ1 has been identified as the factor that the sperm delivers into the egg to induce such a response. We tested the hypothesis that PLCZ1 cRNA injection can be used to activate bovine oocytes. Results Mouse and bovine PLCZ1 cRNAs were injected into matured bovine oocytes at different concentrations. Within the concentrations tested, mouse PLCZ1 injection activated bovine oocytes at a maximum rate when the pipette concentration of cRNA ranged from 0.25 to 1 μg/μL, while bovine PLCZ1 was optimal at 0.1 μg/μL. At their most effective concentrations, PLCZ1 induced parthenogenetic development at rates similar to those observed using other activation stimuli such as Ionomycin/CHX and Ionomycin/DMAP. Injection of mouse and bovine PLCZ1 cRNA induced dose-dependent sperm-like calcium oscillations whose frequency increased over time. Injection of bovine and mouse PLCZ1 cRNA also induced IP3R-1 degradation, although bovine PLCZ1 cRNA evoked greater receptor degradation than its mouse counterpart. Conclusion Injection of PLCZ1 cRNA efficiently activated bovine oocytes by inducing a sperm-like calcium oscillatory pattern. Importantly, the high rate of aneuploidy encountered in parthenogenetic embryos activated by certain chemical means was not observed in PLCZ1 activated embryos. PMID:18284699

  2. Understanding and evaluating bovine testes.

    PubMed

    Kastelic, John P

    2014-01-01

    The objective is to briefly review bovine testes and how they are assessed, with an emphasis on articles from Theriogenology. Scrotal circumference (SC) is the most common method to assess testicular size; it varies among individual bulls and breeds and is highly heritable. In general, a large SC is associated with early puberty, more sperm, a higher percentage of morphologically normal sperm, and better reproductive performance in closely related females. Consequently, there are minimum requirements for SC for breeding soundness. In prepubertal bull calves, there is an early rise (10-20 weeks of age) in LH, which is critically related to onset of puberty and testicular development. Feeding bulls approximately 130% of maintenance requirements of energy and protein from approximately 8 to 30 weeks of age increased LH release during the early rise, hastened puberty (approximately 1 month), and increased mature testis size and sperm production (approximately 20%-30%). However, high-energy diets after weaning (>200 days) often reduced sperm production and semen quality. A bull's testes and scrotum have opposing (complementary) temperature gradients, which keep the testicular temperature 2 °C to 6 °C cooler than core body temperature for production of fertile sperm (increased testicular temperature reduces semen quality). Infrared thermography, a quick and noninvasive method of assessing scrotal surface temperature, may be beneficial for evaluations of breeding soundness. The primary clinical use of ultrasonography in assessment of reproductive function in the bull is characterization of grossly detectable lesions in the testes and scrotum. In conclusion, testis size and function are critical for bull fertility, affected by nutrition, and readily assessed clinically. PMID:24274406

  3. New antibacterial peptide derived from bovine hemoglobin.

    PubMed

    Daoud, Rachid; Dubois, Veronique; Bors-Dodita, Loredana; Nedjar-Arroume, Naima; Krier, Francois; Chihib, Nour-Eddine; Mary, Patrice; Kouach, Mostafa; Briand, Gilbert; Guillochon, Didier

    2005-05-01

    Peptic digestion of bovine hemoglobin at low degree of hydrolysis yields an intermediate peptide fraction exhibiting antibacterial activity against Micrococcus luteus A270, Listeria innocua, Escherichia coli and Salmonella enteritidis after separation by reversed-phase HPLC. From this fraction a pure peptide was isolated and analyzed by matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) and electrospray ionization tandem mass spectrometry (ESI-MS/MS). This peptide correspond to the 107-136 fragment of the alpha chain of bovine hemoglobin. The minimum inhibitory concentrations (MIC) towards the four strains and its hemolytic activity towards bovine erythrocytes were determined. A MIC of 38 microM was reported against L. innocua and 76 microM for other various bacterial species. This peptide had no hemolytic activity up to 380 microM concentration. PMID:15808900

  4. Arachidonate metabolism in bovine gallbladder muscle

    SciTech Connect

    Nakano, M.; Hidaka, T.; Ueta, T.; Ogura, R.

    1983-04-01

    Incubation of (1-/sup 14/C)arachidonic acid (AA) with homogenates of bovine gallbladder muscle generated a large amount of radioactive material having the chromatographic mobility of 6-keto-PGF1 alpha (stable product of PGI2) and smaller amounts of products that comigrated with PGF2 alpha PGE2. Formation of these products was inhibited by the cyclooxygenase inhibitor indomethacin. The major radioactive product identified by thin-layer chromatographic mobility and by gas chromatography - mass spectrometric analysis was found to be 6-keto-PGF1 alpha. The quantitative metabolic pattern of (1-/sup 14/C)PGH2 was virtually identical to that of (1-/sup 14/C)AA. Incubation of arachidonic acid with slices of bovine gallbladder muscle released labile anti-aggregatory material in the medium, which was inhibited by aspirin or 15-hydroperoxy-AA. These results indicate that bovine gallbladder muscle has a considerable enzymatic capacity to produce PGI2 from arachidonic acid.

  5. Comparison of human and bovine protoporphyria.

    PubMed

    Brenner, D A; Bloomer, J R

    1979-01-01

    Protoporphyria (PP) is an inherited disorder of porphyrin metabolism in man in which there is excessive accumulation and excretion of protoporphyrin. Recently, a similar disorder has been described in cattle. In this report, the clinical, biochemical, and genetic features of bovine and human PP are compared. Human and bovine PP are characterized by photosensitivity and elevation of erythrocyte and fecal protoporphyrin levels. In both disorders, a deficiency of heme synthase activity is present in all tissues which have been examined. The diseases differ clinically in that hepatobiliary disease has been found thus far only in human PP. They also have different inheritance patterns. Human PP is an autosomal dominant disease, while initial studies strongly suggest that there is an autosomal recessive pattern of inheritance in bovine PP. PMID:392959

  6. 21 CFR 522.1125 - Hemoglobin glutamer-200 (bovine).

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 6 2014-04-01 2014-04-01 false Hemoglobin glutamer-200 (bovine). 522.1125 Section... § 522.1125 Hemoglobin glutamer-200 (bovine). (a) Specifications. Each 125 milliliter bag contains 13 grams per deciliter of polymerized hemoglobin of bovine origin in modified Lactated Ringer's...

  7. 21 CFR 522.1125 - Hemoglobin glutamer-200 (bovine).

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 6 2013-04-01 2013-04-01 false Hemoglobin glutamer-200 (bovine). 522.1125 Section... § 522.1125 Hemoglobin glutamer-200 (bovine). (a) Specifications. Each 125 milliliter bag contains 13 grams per deciliter of polymerized hemoglobin of bovine origin in modified Lactated Ringer's...

  8. 21 CFR 522.1125 - Hemoglobin glutamer-200 (bovine).

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 6 2011-04-01 2011-04-01 false Hemoglobin glutamer-200 (bovine). 522.1125 Section... § 522.1125 Hemoglobin glutamer-200 (bovine). (a) Specifications. Each 125 milliliter bag contains 13 grams per deciliter of polymerized hemoglobin of bovine origin in modified Lactated Ringer's...

  9. 21 CFR 522.1125 - Hemoglobin glutamer-200 (bovine).

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 6 2012-04-01 2012-04-01 false Hemoglobin glutamer-200 (bovine). 522.1125 Section... § 522.1125 Hemoglobin glutamer-200 (bovine). (a) Specifications. Each 125 milliliter bag contains 13 grams per deciliter of polymerized hemoglobin of bovine origin in modified Lactated Ringer's...

  10. 76 FR 26239 - Bovine Tuberculosis and Brucellosis; Public Meetings

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-05-06

    ...; ] DEPARTMENT OF AGRICULTURE Animal and Plant Health Inspection Service Bovine Tuberculosis and Brucellosis... framework being developed for the bovine tuberculosis and brucellosis programs in the United States. The... tuberculosis (TB) and bovine brucellosis in the United States. In keeping with its commitment to...

  11. Characterization of the bovine C alpha gene.

    PubMed Central

    Brown, W R; Rabbani, H; Butler, J E; Hammarström, L

    1997-01-01

    The complete genomic sequence of a bovine C alpha gene is reported here. The genomic sequence was obtained from a C alpha phage clone that had been cloned from a genomic EMBL4 phage vector library. The C alpha sequence had previously been expressed as a chimeric antibody and identified as IgA using IgA-specific antibodies. Intron/exon boundaries were determined by comparison of the genomic sequence with an expressed bovine C alpha sequence obtained from spleen by reverse transcription-polymerase chain reaction (RT-PCR). Analysis of 50 Swedish bovine genomic DNA samples using genomic blots and five different restriction enzymes failed to detect evidence of polymorphism. However, PstI digests of Brown Swiss DNA showed a restriction fragment length polymorphism (RFLP), suggesting that at least two allelic variants of bovine IgA exist. Comparison of the deduced amino acid sequence of bovine IgA with sequences available for other species indicated that the highest homology was with that of swine, another artiodactyl. This was the highest homology observed for all mammalian IgA compared except for that between IgA1 and IgA2 in humans. Bovine IgA shares with rabbit IgA3 and IgA4, an additional N-linked glycosylation site at position 282. However, the collective data indicate that cattle are like swine and rodents and unlike rabbits in having a single locus of the gene encoding IgA of this species. Images Figure 4 PMID:9203958

  12. Development and evaluation of SoySNP50K, a high-density genotyping array for soybean

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The objective of this research was to identify single nucleotide polymorphisms (SNPs) and to efficiently develop an Infinium iSelect beadchip that contained over 50,000 SNPs from soybean. A total of 498,921,777 reads 35-45bp in length were obtained from DNA sequence analysis of reduced representati...

  13. [Investigation on bovine leukocyte adhesion deficiency].

    PubMed

    Ma, Jin-Zhu; Cui, Yu-Dong; Zhu, Zhan-Bo; Cao, Hong-Wei; Piao, Fan-Ze

    2006-10-01

    Bovine leukocyte adhesion deficiency (BLAD) is autosomal recessive disease. The pathogeny of BLAD is genic mutation of CD18-integrins on the leukocyte. In order to know the carrier and occurrence of bovine leukocyte adhesion deficiency (BLAD) among cows age from one to six years old in China, 1,000 cows were investigated by means of amplifying a CD18 gene fragment via reverse transcriptase-PCR followed by restriction digestion with Taq I. Results showed that 19 cows were BLAD carriers, indicating that the BLAD carrier rate was 1.9 percent. In addition, one cow was found to have BLAD. PMID:17035180

  14. Economics of bovine leukemia virus infection.

    PubMed

    Pelzer, K D

    1997-03-01

    A herd infected with bovine leukemia virus suffers a direct economic loss due to clinical lymphosarcoma. A major indirect cost associated with infection is restriction of the sale of animals and germplasma to foreign markets. Reports on the economic effects of infection on production have been variable and are reviewed in this article. In order to develop cost-effective bovine leukemia virus control programs, costs associated with the disease, the cost of prevention, and expected economic returns from a program need to be considered. PMID:9071750

  15. The interaction between bovine herpesvirus type 1 and activated bovine T lymphocytes.

    PubMed

    Griebel, P J; Ohmann, H B; Lawman, M J; Babiuk, L A

    1990-02-01

    The interaction between activated bovine T lymphocytes (BTLs) and bovine herpesvirus type 1 (BHV-1) was investigated. BHV-1 infection of BTLs reduced the amplitude of recombinant bovine interleukin 2-induced proliferative responses. This decreased proliferation was caused by a virus-induced lymphocytolysis which was dependent on viable virus and was not inhibited by recombinant bovine interferon-alpha I1. Furthermore, lymphocytolysis was not associated with virus replication or with the synthesis of detectable levels of viral proteins. Electron microscopic examination of virus-infected cells revealed that lymphocytolysis was characterized by early nuclear disintegration resembling apoptosis. These observations suggest that activated T cells, localized at the site of BHV-1 infection, may be susceptible to virus-induced cytolysis. PMID:2155290

  16. Molecular differentiation of bovine sarcocysts.

    PubMed

    Akhlaghi, Majedeh; Razavi, Mostafa; Hosseini, Arsalan

    2016-07-01

    Cattle are common intermediate hosts of Sarcocystis, and the prevalence in adult bovine muscle is close to 100 % in most regions of the world. Three Sarcocystis spp. are known to infect cattle as intermediate hosts, namely, S. cruzi, S. hirsuta, and S. hominis. The aim of the present study was the molecular identification and differentiation of these three species, Neospora caninum and Besnoitia by PCR and RFLP methods. Tissue samples were obtained from diaphragmatic muscle of 101 cattle slaughtered in Shiraz, Fars Province, Iran, for both smear preparation and DNA extraction. The samples were digested by Pepsin, washed three times with PBS solution before taking smears, fixed in absolute methanol and stained with 10 % Giemsa. The slides were examined microscopically for Sarcocystis bradyzoites and DNA was extracted from 100 mg of Sarcocystis-infected meat samples. Since the primers also bind to 18S rRNA gene of some tissue cyst-forming coccidian protozoa, DNA was also extracted from 100 μl of tachyzoite-containing suspension of N. caninum and Besnoitia isolated from goat to compare RFLP pattern. Polymerase chain reaction (PCR) was performed on DNA of samples which were microscopically positive for Sarcocystis. Five restriction enzymes Dra1, EcoRV, RsaI, AvaI, and SspI were used for RFLP and DNA of one sample from protozoa was sequenced. Based on the RFLP results, 87 (98.9 %) DNA samples were cut with DraI, indicating infection by S. cruzi. One sample (1.1 %) of PCR products of infected samples was cut only with EcoRV which showed S. hominis infection. Forty-eight samples (53.3 %) of PCR products were cut with both DraI, EcoRV, or with DraI, EcoRV, and RsaI while none of them was cut with SspI, which shows the mixed infection of both S. cruzi and S. hominis and no infection with S. hirsuta. It seems by utilizing these restriction enzymes, RLFP could be a suitable method not only for identification of Sarcocystis species but also for differentiating them

  17. Development and characterization of 26 novel microsatellite loci for the trochid gastropod Gibbula divaricata (Linnaeus, 1758), using Illumina MiSeq next generation sequencing technology

    PubMed Central

    García-Jiménez, Ricardo; Templado, José; Machordom, Annie

    2016-01-01

    In the present study we used the high-throughput sequencing technology Illumina MiSeq to develop 26 polymorphic microsatellite loci for the marine snail Gibbula divaricata. Four to 32 alleles were detected per locus across 30 samples analyzed. Observed and expected heterozygosities ranged from 0.130 to 0.933 and from 0.294 to 0.956, respectively. No significant linkage disequilibrium existed. Seven loci deviated from Hardy-Weinberg equilibrium that could not totally be explained by the presence of null alleles. Sympatric distribution with other species of the genus Gibbula, as G. rarilineata and G. varia, lead us to test the cross utility of the developed markers in these two species, which could be useful to test common biogeographic patterns or potential hybridization phenomena, since morphological intermediate specimens were found. PMID:27042392

  18. Development of microsatellite loci of pod mahogany, Afzelia quanzensis (Fabaceae), by Illumina shotgun sequencing, and cross-amplification in A. africana1

    PubMed Central

    Jinga, Percy; Palagi, Jason; Ashley, Mary V.

    2016-01-01

    Premise of the study: Microsatellite loci were developed for Afzelia quanzensis (Fabaceae) as a first step toward investigating genetic diversity and population structure of the species in its native range. Methods and Results: Illumina shotgun sequencing was used to generate raw sequence reads, which were searched for potential microsatellite loci. A total of 70 potential microsatellite loci were tested for amplification and polymorphism, and 39 successfully amplified. Of the 39 loci that amplified, 12 were polymorphic while 27 were monomorphic. The 12 polymorphic loci were cross-amplified in A. africana, and eight successfully amplified. Conclusions: The 12 polymorphic microsatellite loci can be used for genetic studies of A. quanzensis, which can help determine its conservation status. Eight loci can also be used for genotyping in A. africana. PMID:27347453

  19. The mitochondrial genome of a Texas outbreak strain of the cattle tick, Rhipicephalus (Boophilus) microplus, derived from whole genome sequencing Pacific Biosciences and Illumina reads.

    PubMed

    McCooke, John K; Guerrero, Felix D; Barrero, Roberto A; Black, Michael; Hunter, Adam; Bell, Callum; Schilkey, Faye; Miller, Robert J; Bellgard, Matthew I

    2015-10-15

    The cattle fever tick, Rhipicephalus (Boophilus) microplus is one of the most significant medical veterinary pests in the world, vectoring several serious livestock diseases negatively impacting agricultural economies of tropical and subtropical countries around the world. In our study, we assembled the complete R. microplus mitochondrial genome from Illumina and Pac Bio sequencing reads obtained from the ongoing R. microplus (Deutsch strain from Texas, USA) genome sequencing project. We compared the Deutsch strain mitogenome to the mitogenome from a Brazilian R. microplus and from an Australian cattle tick that has recently been taxonomically designated as Rhipicephalus australis after previously being considered R. microplus. The sequence divergence of the Texas and Australia ticks is much higher than the divergence between the Texas and Brazil ticks. This is consistent with the idea that the Australian ticks are distinct from the R. microplus of the Americas. PMID:26117172

  20. Concurrent Bovine Virus Diarrhea and Bovine Papular Stomatitis Infection in a Calf

    PubMed Central

    Bohac, J. G.; Yates, W. D. G.

    1980-01-01

    A case of concurrent infection with the viruses of bovine virus diarrhea and papular stomatitis in a calf is reported. The difficulties posed by such situations are described and the criteria used for diagnosis outlined. The two diseases are reviewed briefly and the possible mechanisms whereby bovine virus diarrhea virus is suspected of facilitating infection by other agents are discussed. ImagesFigure 1.Figure 2.Figure 3.Figure 4. PMID:7459795

  1. Transcriptome of the Female Synganglion of the Black-Legged Tick Ixodes scapularis (Acari: Ixodidae) with Comparison between Illumina and 454 Systems

    PubMed Central

    Egekwu, Noble; Sonenshine, Daniel E.; Bissinger, Brooke W.; Roe, R. Michael

    2014-01-01

    Illumina and 454 pyrosequencing were used to characterize genes from the synganglion of female Ixodes scapularis. GO term searching success for biological processes was similar for samples sequenced by both methods. However, for molecular processes, it was more successful for the Illumina samples than for 454 samples. Functional assignments of transcripts predicting neuropeptides, neuropeptide receptors, neurotransmitter receptors and other genes of interest was done, supported by strong e-values (<−6), and high consensus sequence alignments. Transcripts predicting 15 putative neuropeptide prepropeptides ((allatostatin, allatotropin, bursicon α, corticotropin releasing factor (CRF), CRF-binding protein, eclosion hormone, FMRFamide, glycoprotein A, insulin-like peptide, ion transport peptide, myoinhibitory peptide, inotocin ( =  neurophysin-oxytocin), Neuropeptide F, sulfakinin and SIFamide)) and transcripts predicting receptors for 14 neuropeptides (allatostatin, calcitonin, cardioacceleratory peptide, corazonin, CRF, eclosion hormone, gonadotropin-releasing hormone/AKH-like, insulin-like peptide, neuropeptide F, proctolin, pyrokinin, SIFamide, sulfakinin and tachykinin) are reported. Similar to Dermacentor variabilis, we found transcripts matching pro-protein convertase, essential for converting neuropeptide hormones to their mature form. Additionally, transcripts predicting 6 neurotransmitter/neuromodulator receptors (acetylcholine, GABA, dopamine, glutamate, octopamine and serotonin) and 3 neurotransmitter transporters (GABA transporter, noradrenalin-norepinephrine transporter and Na+-neurotransmitter/symporter) are described. Further, we found transcripts predicting genes for pheromone odorant receptor, gustatory receptor, novel GPCR messages, ecdysone nuclear receptor, JH esterase binding protein, steroidogenic activating protein, chitin synthase, chitinase, and other genes of interest. Also found were transcripts predicting genes for spermatogenesis

  2. NGS-QCbox and Raspberry for Parallel, Automated and Rapid Quality Control Analysis of Large-Scale Next Generation Sequencing (Illumina) Data

    PubMed Central

    Katta, Mohan A. V. S. K.; Khan, Aamir W.; Doddamani, Dadakhalandar; Thudi, Mahendar; Varshney, Rajeev K.

    2015-01-01

    Rapid popularity and adaptation of next generation sequencing (NGS) approaches have generated huge volumes of data. High throughput platforms like Illumina HiSeq produce terabytes of raw data that requires quick processing. Quality control of the data is an important component prior to the downstream analyses. To address these issues, we have developed a quality control pipeline, NGS-QCbox that scales up to process hundreds or thousands of samples. Raspberry is an in-house tool, developed in C language utilizing HTSlib (v1.2.1) (http://htslib.org), for computing read/base level statistics. It can be used as stand-alone application and can process both compressed and uncompressed FASTQ format files. NGS-QCbox integrates Raspberry with other open-source tools for alignment (Bowtie2), SNP calling (SAMtools) and other utilities (bedtools) towards analyzing raw NGS data at higher efficiency and in high-throughput manner. The pipeline implements batch processing of jobs using Bpipe (https://github.com/ssadedin/bpipe) in parallel and internally, a fine grained task parallelization utilizing OpenMP. It reports read and base statistics along with genome coverage and variants in a user friendly format. The pipeline developed presents a simple menu driven interface and can be used in either quick or complete mode. In addition, the pipeline in quick mode outperforms in speed against other similar existing QC pipeline/tools. The NGS-QCbox pipeline, Raspberry tool and associated scripts are made available at the URL https://github.com/CEG-ICRISAT/NGS-QCbox and https://github.com/CEG-ICRISAT/Raspberry for rapid quality control analysis of large-scale next generation sequencing (Illumina) data. PMID:26460497

  3. NGS-QCbox and Raspberry for Parallel, Automated and Rapid Quality Control Analysis of Large-Scale Next Generation Sequencing (Illumina) Data.

    PubMed

    Katta, Mohan A V S K; Khan, Aamir W; Doddamani, Dadakhalandar; Thudi, Mahendar; Varshney, Rajeev K

    2015-01-01

    Rapid popularity and adaptation of next generation sequencing (NGS) approaches have generated huge volumes of data. High throughput platforms like Illumina HiSeq produce terabytes of raw data that requires quick processing. Quality control of the data is an important component prior to the downstream analyses. To address these issues, we have developed a quality control pipeline, NGS-QCbox that scales up to process hundreds or thousands of samples. Raspberry is an in-house tool, developed in C language utilizing HTSlib (v1.2.1) (http://htslib.org), for computing read/base level statistics. It can be used as stand-alone application and can process both compressed and uncompressed FASTQ format files. NGS-QCbox integrates Raspberry with other open-source tools for alignment (Bowtie2), SNP calling (SAMtools) and other utilities (bedtools) towards analyzing raw NGS data at higher efficiency and in high-throughput manner. The pipeline implements batch processing of jobs using Bpipe (https://github.com/ssadedin/bpipe) in parallel and internally, a fine grained task parallelization utilizing OpenMP. It reports read and base statistics along with genome coverage and variants in a user friendly format. The pipeline developed presents a simple menu driven interface and can be used in either quick or complete mode. In addition, the pipeline in quick mode outperforms in speed against other similar existing QC pipeline/tools. The NGS-QCbox pipeline, Raspberry tool and associated scripts are made available at the URL https://github.com/CEG-ICRISAT/NGS-QCbox and https://github.com/CEG-ICRISAT/Raspberry for rapid quality control analysis of large-scale next generation sequencing (Illumina) data. PMID:26460497

  4. Microbial Populations in Naked Neck Chicken Ceca Raised on Pasture Flock Fed with Commercial Yeast Cell Wall Prebiotics via an Illumina MiSeq Platform.

    PubMed

    Park, Si Hong; Lee, Sang In; Ricke, Steven C

    2016-01-01

    Prebiotics are non-digestible carbohydrate dietary supplements that selectively stimulate the growth of one or more beneficial bacteria in the gastrointestinal tract of the host. These bacteria can inhibit colonization of pathogenic bacteria by producing antimicrobial substances such as short chain fatty acids (SCFAs) and competing for niches with pathogens within the gut. Pasture flock chickens are generally raised outdoors with fresh grass, sunlight and air, which represents different environmental growth conditions compared to conventionally raised chickens. The purpose of this study was to evaluate the difference in microbial populations from naked neck chicken ceca fed with commercial prebiotics derived from brewer's yeast cell wall via an Illumina MiSeq platform. A total of 147 day-of-hatch naked neck chickens were distributed into 3 groups consisted of 1) C: control (no prebiotic), 2) T1: Biolex® MB40 with 0.2%, and 3) T2: Leiber® ExCel with 0.2%, consistently supplemented prebiotics during the experimental period. At 8 weeks, a total of 15 birds from each group were randomly selected and ceca removed for DNA extraction. The Illumina Miseq platform based on V4 region of 16S rRNA gene was applied for microbiome analysis. Both treatments exhibited limited impact on the microbial populations at the phylum level, with no significant differences in the OTU number of Bacteroidetes among groups and an increase of Proteobacteria OTUs for the T1 (Biolex® MB40) group. In addition there was a significant increase of genus Faecalibacterium OTU, phylum Firmicutes. According to the development of next generation sequencing (NGS), microbiome analysis based on 16S rRNA gene proved to be informative on the prebiotic impact on poultry gut microbiota in pasture-raised naked neck birds. PMID:26992104

  5. Microbial Populations in Naked Neck Chicken Ceca Raised on Pasture Flock Fed with Commercial Yeast Cell Wall Prebiotics via an Illumina MiSeq Platform

    PubMed Central

    Park, Si Hong; Lee, Sang In; Ricke, Steven C.

    2016-01-01

    Prebiotics are non-digestible carbohydrate dietary supplements that selectively stimulate the growth of one or more beneficial bacteria in the gastrointestinal tract of the host. These bacteria can inhibit colonization of pathogenic bacteria by producing antimicrobial substances such as short chain fatty acids (SCFAs) and competing for niches with pathogens within the gut. Pasture flock chickens are generally raised outdoors with fresh grass, sunlight and air, which represents different environmental growth conditions compared to conventionally raised chickens. The purpose of this study was to evaluate the difference in microbial populations from naked neck chicken ceca fed with commercial prebiotics derived from brewer’s yeast cell wall via an Illumina MiSeq platform. A total of 147 day-of-hatch naked neck chickens were distributed into 3 groups consisted of 1) C: control (no prebiotic), 2) T1: Biolex® MB40 with 0.2%, and 3) T2: Leiber® ExCel with 0.2%, consistently supplemented prebiotics during the experimental period. At 8 weeks, a total of 15 birds from each group were randomly selected and ceca removed for DNA extraction. The Illumina Miseq platform based on V4 region of 16S rRNA gene was applied for microbiome analysis. Both treatments exhibited limited impact on the microbial populations at the phylum level, with no significant differences in the OTU number of Bacteroidetes among groups and an increase of Proteobacteria OTUs for the T1 (Biolex® MB40) group. In addition there was a significant increase of genus Faecalibacterium OTU, phylum Firmicutes. According to the development of next generation sequencing (NGS), microbiome analysis based on 16S rRNA gene proved to be informative on the prebiotic impact on poultry gut microbiota in pasture-raised naked neck birds. PMID:26992104

  6. Recombinant Bovine Growth Hormone Criticism Grows.

    ERIC Educational Resources Information Center

    Gaard, Greta

    1995-01-01

    Discusses concerns related to the use of recombinant bovine growth hormone in the United States and other countries. Analyses the issue from the perspectives of animal rights, human health, world hunger, concerns of small and organic farmers, costs to the taxpayer, and environmental questions. A sidebar discusses Canadian review of the hormone.…

  7. Unraveling bovin phylogeny: accomplishments and challenges.

    PubMed

    Bibi, Faysal; Vrba, Elisabeth S

    2010-01-01

    The phylogenetic systematics of bovin species forms a common basis for studies at multiple scales, from the level of domestication in populations to major cladogenesis. The main big-picture accomplishments of this productive field, including two recent works, one in BMC Genomics, are reviewed with an eye for some of the limitations and challenges impeding progress. PMID:20525112

  8. Bovine Viral Diarrhea Virus: Global Status

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Despite the success of regional bovine viral diarrhea viruses (BVDV) eradication programs, infections with this diverse group of viruses remain a source of economic loss for producers worldwide. There is a wide range of variation among BVDV results in differences in genotype (BVDV1 and BVDV2), biot...

  9. Epidemiology and control of bovine ephemeral fever.

    PubMed

    Walker, Peter J; Klement, Eyal

    2015-01-01

    Bovine ephemeral fever (or 3-day sickness) is an acute febrile illness of cattle and water buffaloes. Caused by an arthropod-borne rhabdovirus, bovine ephemeral fever virus (BEFV), the disease occurs seasonally over a vast expanse of the globe encompassing much of Africa, the Middle East, Asia and Australia. Although mortality rates are typically low, infection prevalence and morbidity rates during outbreaks are often very high, causing serious economic impacts through loss of milk production, poor cattle condition at sale and loss of traction power at harvest. There are also significant impacts on trade to regions in which the disease does not occur, including the Americas and most of Europe. In recent years, unusually severe outbreaks of bovine ephemeral fever have been reported from several regions in Asia and the Middle East, with mortality rates through disease or culling in excess of 10-20%. There are also concerns that, like other vector-borne diseases of livestock, the geographic distribution of bovine ephemeral fever could expand into regions that have historically been free of the disease. Here, we review current knowledge of the virus, including its molecular and antigenic structure, and the epidemiology of the disease across its entire geographic range. We also discuss the effectiveness of vaccination and other strategies to prevent or control infection. PMID:26511615

  10. Bovine respiratory disease research (1983-2009).

    PubMed

    Fulton, Robert W

    2009-12-01

    Bovine respiratory disease (BRD) research has provided significant understanding of the disease over the past 26 years. Modern research tools that have been used include monoclonal antibodies, genomics, polymerase chain reaction, immunohistochemistry (IHC), DNA vaccines and viral vectors coding for immunogens. Emerging/reemerging viruses and new antigenic strains of viruses and bacteria have been identified. Methods of detection and the role for cattle persistently infected bovine viral diarrhea virus (BVDV) were identified; viral subunits, cellular components and bacterial products have been characterized. Product advances have included vaccines for bovine respiratory syncytial virus, Mannheimia haemolytica and Pasteurella multocida; the addition of BVDV2 to the existing vaccines and new antibiotics. The role of Mycoplasma spp., particularly Mycoplasma bovis in BRD, has been more extensively studied. Bovine immunology research has provided more specific information on immune responses, T cell subsets and cytokines. The molecular and genetic basis for viral-bacterial synergy in BRD has been described. Attempts have been made to document how prevention of BRD by proper vaccination and management prior to exposure to infectious agents can minimize disease and serve as economic incentives for certified health programs. PMID:20003649

  11. Bovine viral diarrhea virus: biosecurity and control

    Technology Transfer Automated Retrieval System (TEKTRAN)

    This paper discusses the recommended procedures involved in setting up biosecurity and control programs designed to limit bovine viral diarrhea virus infections in beef cattle operations. For the purpose of these discussions, a working definition of a biosecurity plan was considered to be an organiz...

  12. Molecular biology of bovine viral diarrhea virus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bovine viral diarrhea viruses (BVDV) are arguably the most important viral pathogen of ruminants worldwide and can cause severe economic loss. Clinical symptoms of the disease caused by BVDV range from subclinical to severe acute hemorrhagic syndrome, with the severity of disease being strain depend...

  13. A physical map of the bovine genome

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Background Cattle are important agriculturally and relevant as a model organism. Previously described genetic and radiation hybrid (RH) maps of the bovine genome have been used to identify genomic regions and genes affecting specific traits. Application of these maps to identify influential geneti...

  14. NUTRIENTS AND EPIGENETICS IN BOVINE CELLS

    Technology Transfer Automated Retrieval System (TEKTRAN)

    This is a chapter for a book titled “Livestock Epigenetics” edited by Dr. Hasan Khatib and published by Wiley-Blackwell. This chapter is focused on the research development in our laboratory in the area of interaction of nutrients and genomic phonotype in bovine cells. Briefly, the Research on nutri...

  15. Linkage mapping bovine EST-based SNP

    PubMed Central

    Snelling, Warren M; Casas, Eduardo; Stone, Roger T; Keele, John W; Harhay, Gregory P; Bennett, Gary L; Smith, Timothy PL

    2005-01-01

    Background Existing linkage maps of the bovine genome primarily contain anonymous microsatellite markers. These maps have proved valuable for mapping quantitative trait loci (QTL) to broad regions of the genome, but more closely spaced markers are needed to fine-map QTL, and markers associated with genes and annotated sequence are needed to identify genes and sequence variation that may explain QTL. Results Bovine expressed sequence tag (EST) and bacterial artificial chromosome (BAC)sequence data were used to develop 918 single nucleotide polymorphism (SNP) markers to map genes on the bovine linkage map. DNA of sires from the MARC reference population was used to detect SNPs, and progeny and mates of heterozygous sires were genotyped. Chromosome assignments for 861 SNPs were determined by twopoint analysis, and positions for 735 SNPs were established by multipoint analyses. Linkage maps of bovine autosomes with these SNPs represent 4585 markers in 2475 positions spanning 3058 cM . Markers include 3612 microsatellites, 913 SNPs and 60 other markers. Mean separation between marker positions is 1.2 cM. New SNP markers appear in 511 positions, with mean separation of 4.7 cM. Multi-allelic markers, mostly microsatellites, had a mean (maximum) of 216 (366) informative meioses, and a mean 3-lod confidence interval of 3.6 cM Bi-allelic markers, including SNP and other marker types, had a mean (maximum) of 55 (191) informative meioses, and were placed within a mean 8.5 cM 3-lod confidence interval. Homologous human sequences were identified for 1159 markers, including 582 newly developed and mapped SNP. Conclusion Addition of these EST- and BAC-based SNPs to the bovine linkage map not only increases marker density, but provides connections to gene-rich physical maps, including annotated human sequence. The map provides a resource for fine-mapping quantitative trait loci and identification of positional candidate genes, and can be integrated with other data to guide and

  16. Molecular definition of bovine argininosuccinate synthetase deficiency.

    PubMed Central

    Dennis, J A; Healy, P J; Beaudet, A L; O'Brien, W E

    1989-01-01

    Citrullinemia is an inborn error of metabolism due to deficiency of the urea cycle enzyme, argininosuccinate synthetase [L-citrulline:L-aspartate ligase (AMP-forming), EC 6.3.4.5]. The disease was first described in humans but was recently reported in dairy cattle in Australia. Here we report the nucleotide sequence of the normal bovine cDNA for argininosuccinate synthetase and the mutation present in animals with citrullinemia. Analysis of DNA from affected animals by Southern blotting did not readily identify the mutation in the bovine gene. RNA (Northern) blotting revealed a major reduction in the steady-state amount of mRNA in the liver of affected animals to less than 5% of controls. The bovine cDNA was cloned and sequenced and revealed 96% identity with the deduced human sequence at the amino acid level. Starting with mutant bovine liver, the mRNA was reverse-transcribed; the cDNA product was amplified with the polymerase chain reaction, cloned, and sequenced. The sequence revealed a C----T transition converting arginine-86 (CGA) to a nonsense codon (TGA). A second C----T transition represented a polymorphism in proline-175 (CCC----CCT). The mutation and the polymorphism were confirmed by amplification of genomic DNA and demonstration with restriction endonuclease enzymes of both the loss of an Ava II site in DNA from mutant animals at codon 86 and the presence or absence of a Dde I site at codon 175. The loss of the Ava II site can be used for rapid, economical, nonradioactive detection of heterozygotes for bovine citrullinemia. Images PMID:2813370

  17. A physical map of the bovine genome

    PubMed Central

    Snelling, Warren M; Chiu, Readman; Schein, Jacqueline E; Hobbs, Matthew; Abbey, Colette A; Adelson, David L; Aerts, Jan; Bennett, Gary L; Bosdet, Ian E; Boussaha, Mekki; Brauning, Rudiger; Caetano, Alexandre R; Costa, Marcos M; Crawford, Allan M; Dalrymple, Brian P; Eggen, André; Everts-van der Wind, Annelie; Floriot, Sandrine; Gautier, Mathieu; Gill, Clare A; Green, Ronnie D; Holt, Robert; Jann, Oliver; Jones, Steven JM; Kappes, Steven M; Keele, John W; de Jong, Pieter J; Larkin, Denis M; Lewin, Harris A; McEwan, John C; McKay, Stephanie; Marra, Marco A; Mathewson, Carrie A; Matukumalli, Lakshmi K; Moore, Stephen S; Murdoch, Brenda; Nicholas, Frank W; Osoegawa, Kazutoyo; Roy, Alice; Salih, Hanni; Schibler, Laurent; Schnabel, Robert D; Silveri, Licia; Skow, Loren C; Smith, Timothy PL; Sonstegard, Tad S; Taylor, Jeremy F; Tellam, Ross; Van Tassell, Curtis P; Williams, John L; Womack, James E; Wye, Natasja H; Yang, George; Zhao, Shaying

    2007-01-01

    Background Cattle are important agriculturally and relevant as a model organism. Previously described genetic and radiation hybrid (RH) maps of the bovine genome have been used to identify genomic regions and genes affecting specific traits. Application of these maps to identify influential genetic polymorphisms will be enhanced by integration with each other and with bacterial artificial chromosome (BAC) libraries. The BAC libraries and clone maps are essential for the hybrid clone-by-clone/whole-genome shotgun sequencing approach taken by the bovine genome sequencing project. Results A bovine BAC map was constructed with HindIII restriction digest fragments of 290,797 BAC clones from animals of three different breeds. Comparative mapping of 422,522 BAC end sequences assisted with BAC map ordering and assembly. Genotypes and pedigree from two genetic maps and marker scores from three whole-genome RH panels were consolidated on a 17,254-marker composite map. Sequence similarity allowed integrating the BAC and composite maps with the bovine draft assembly (Btau3.1), establishing a comprehensive resource describing the bovine genome. Agreement between the marker and BAC maps and the draft assembly is high, although discrepancies exist. The composite and BAC maps are more similar than either is to the draft assembly. Conclusion Further refinement of the maps and greater integration into the genome assembly process may contribute to a high quality assembly. The maps provide resources to associate phenotypic variation with underlying genomic variation, and are crucial resources for understanding the biology underpinning this important ruminant species so closely associated with humans. PMID:17697342

  18. Bovine Rhinitis Viruses Are Common in U.S. Cattle with Bovine Respiratory Disease

    PubMed Central

    Hause, Ben M.; Collin, Emily A.; Anderson, Joe; Hesse, Richard A.; Anderson, Gary

    2015-01-01

    Bovine rhinitis viruses (BRV) are established etiological agents of bovine respiratory disease complex however little research into their epidemiology and ecology has been published for several decades. In the U.S., only bovine rhinitis A virus 1 (BRAV1) has been identified while bovine rhinitis A virus 2 (BRAV2) and bovine rhinitis B virus (BRBV) were previously only identified in England and Japan, respectively. Metagenomic sequencing of a nasal swab from a bovine respiratory disease (BRD) diagnostic submission from Kansas identified contigs with approximately 90% nucleotide similarity to BRAV2 and BRBV. A combination of de novo and templated assemblies using reference genomes yielded near complete BRAV2 and BRBV genomes. The near complete genome of bovine rhinitis A virus 1 (BRAV1) was also determined from a historical isolate to enable further molecular epidemiological studies. A 5’-nuclease reverse transcription PCR assay targeting the 3D polymerase gene was designed and used to screen 204 archived BRD clinical specimens. Thirteen (6.4%) were positive. Metagenomic sequencing of six positive samples identified mixed BRAV1/BRAV2, BRAV1/BRBV and BRAV2/BRBV infections for five samples. One sample showed infection only with BRAV1. Seroprevalence studies using a cell culture adapted BRBV found immunofluorescence assay-reactive antibodies were common in the herds analyzed. Altogether, these results demonstrate that BRV infections are common in cattle with respiratory disease and that BRAV1, BRAV2 and BRBV co-circulate in U.S. cattle and have high similarity to viruses isolated more than 30 years ago from diverse locations. PMID:25789939

  19. Bovine rhinitis viruses are common in U.S. cattle with bovine respiratory disease.

    PubMed

    Hause, Ben M; Collin, Emily A; Anderson, Joe; Hesse, Richard A; Anderson, Gary

    2015-01-01

    Bovine rhinitis viruses (BRV) are established etiological agents of bovine respiratory disease complex however little research into their epidemiology and ecology has been published for several decades. In the U.S., only bovine rhinitis A virus 1 (BRAV1) has been identified while bovine rhinitis A virus 2 (BRAV2) and bovine rhinitis B virus (BRBV) were previously only identified in England and Japan, respectively. Metagenomic sequencing of a nasal swab from a bovine respiratory disease (BRD) diagnostic submission from Kansas identified contigs with approximately 90% nucleotide similarity to BRAV2 and BRBV. A combination of de novo and templated assemblies using reference genomes yielded near complete BRAV2 and BRBV genomes. The near complete genome of bovine rhinitis A virus 1 (BRAV1) was also determined from a historical isolate to enable further molecular epidemiological studies. A 5'-nuclease reverse transcription PCR assay targeting the 3D polymerase gene was designed and used to screen 204 archived BRD clinical specimens. Thirteen (6.4%) were positive. Metagenomic sequencing of six positive samples identified mixed BRAV1/BRAV2, BRAV1/BRBV and BRAV2/BRBV infections for five samples. One sample showed infection only with BRAV1. Seroprevalence studies using a cell culture adapted BRBV found immunofluorescence assay-reactive antibodies were common in the herds analyzed. Altogether, these results demonstrate that BRV infections are common in cattle with respiratory disease and that BRAV1, BRAV2 and BRBV co-circulate in U.S. cattle and have high similarity to viruses isolated more than 30 years ago from diverse locations. PMID:25789939

  20. Linkage Disequilibrium Estimation of Chinese Beef Simmental Cattle Using High-density SNP Panels

    PubMed Central

    Zhu, M.; Zhu, B.; Wang, Y. H.; Wu, Y.; Xu, L.; Guo, L. P.; Yuan, Z. R.; Zhang, L. P.; Gao, X.; Gao, H. J.; Xu, S. Z.; Li, J. Y.

    2013-01-01

    Linkage disequilibrium (LD) plays an important role in genomic selection and mapping quantitative trait loci (QTL). In this study, the pattern of LD and effective population size (Ne) were investigated in Chinese beef Simmental cattle. A total of 640 bulls were genotyped with IlluminaBovinSNP50BeadChip and IlluminaBovinHDBeadChip. We estimated LD for each autosomal chromosome at the distance between two random SNPs of <0 to 25 kb, 25 to 50 kb, 50 to 100 kb, 100 to 500 kb, 0.5 to 1 Mb, 1 to 5 Mb and 5 to 10 Mb. The mean values of r2 were 0.30, 0.16 and 0.08, when the separation between SNPs ranged from 0 to 25 kb to 50 to 100 kb and then to 0.5 to 1 Mb, respectively. The LD estimates decreased as the distance increased in SNP pairs, and increased with the increase of minor allelic frequency (MAF) and with the decrease of sample sizes. Estimates of effective population size for Chinese beef Simmental cattle decreased in the past generations and Ne was 73 at five generations ago. PMID:25049849

  1. Genome-wide association analysis identifies loci for left-sided displacement of the abomasum in German Holstein cattle.

    PubMed

    Mömke, S; Sickinger, M; Lichtner, P; Doll, K; Rehage, J; Distl, O

    2013-06-01

    Left-sided displacement of the abomasum (LDA) is one of the most common disorders of the digestive system in many dairy breeds and particularly in Holstein dairy cows. We performed a genome-wide association study for 854 German Holstein cows, including 225 cases and 629 controls. All cows were genotyped using the Illumina Bovine SNP50 BeadChip (Illumina Inc., San Diego, CA). After quality control of genotypes, a total of 36,226 informative single nucleotide polymorphisms (SNP) were left for analysis. We used a mixed linear model approach for a genome-wide association study of LDA. In total, 36 SNP located on 17 bovine (Bos taurus) chromosomes (BTA) showed associations with LDA at nominal -log10P-values >3.0. Two of these SNP, located on BTA11 at 46.70 Mb and BTA20 at 16.67 Mb, showed genome-wide significant associations with LDA at -log10P-values >4.6. Pathway analyses indicated genes involved in calcium metabolism and insulin-dependent diabetes mellitus to be factors in the pathogenesis of LDA in German Holstein cows. PMID:23548285

  2. Identification of Novel Ghanaian G8P[6] Human-Bovine Reassortant Rotavirus Strain by Next Generation Sequencing

    PubMed Central

    Dennis, Francis E.; Fujii, Yoshiki; Haga, Kei; Damanka, Susan; Lartey, Belinda; Agbemabiese, Chantal A.; Ohta, Nobuo; Armah, George E.; Katayama, Kazuhiko

    2014-01-01

    Group A rotaviruses (RVAs) are the most important etiological agent of acute gastroenteritis in children <5 years of age worldwide. The monovalent rotavirus vaccine Rotarix was introduced into the national Expanded Programme on Immunization (EPI) in Ghana in May 2012. However, there is a paucity of genetic and phylogenetic data on the complete genomes of human RVAs in circulation pre-vaccine introduction. The common bovine rotavirus VP7 genotype G8 has been sporadically detected in Ghanaian children, usually in combination with the VP4 genotype P[6]. To investigate the genomic constellations and phylogeny of RVA strains in circulation prior to vaccine introduction, the full genomes of two unusual G8P[6] strains, GH018-08 and GH019-08, detected during burden of disease surveillance, were characterized by Illumina MiSeq sequencing. The Ghanaian isolates, GH018-08 and GH019-08, exhibited the unusual, previously unreported genotype constellation G8-P[6]-I2-R2-C2-M2-A2-N2-T2-E2-H3. Phylogenetic analyses confirmed that 10 out of the 11 genes of GH018-08 and GH019-08 were identical/nearly identical, with significant variation detected only in their VP1 genes, and clearly established the occurrence of multiple independent interspecies transmission and reassortment events between co-circulating bovine/ovine/caprine rotaviruses and human DS-1-like RVA strains. These findings highlight the contribution of reassortment and interspecies transmission events to the high rotavirus diversity in this region of Africa, and justify the need for simultaneous monitoring of animal and human rotavirus strains. PMID:24971993

  3. Ontogeny of the Bovine Immune Response 1

    PubMed Central

    Schultz, R. D.; Dunne, H. W.; Heist, C. E.

    1973-01-01

    The ontogenesis of the bovine immune response was studied in three embryos (<40 days) and 106 fetuses of various ages. In the absence of overt antigenic stimulation, fetuses had lymphoid development of the thymus at 42 days of gestation, the spleen was structurally present at 55 days, and certain peripheral lymph nodes were present at 60 days. Mesenteric lymph nodes were structurally present by 100 days of gestation, and lymphoid tissue of the gastrointestinal tract, particularly the lower ileum, was observed in histologic sections of a 175-day fetus with a bacterial infection. Pyroninophilic cells, plasma cells, and germinal centers were present in lymph node sections of antigenically stimulated fetuses. Lymphoid tissue developed more rapidly in fetuses with bacteria, viral antigens, or apparent maternal red-blood-cell antigens than in the normal fetus. Thymic and splenic indices reached maximal values in the 205- to 220-day fetal age group. Immunoglobulin M (IgM)-containing cells were first observed, by immunofluorescence, in a single fetus at 59 days of gestation. Immunoglobulin G (IgG)-containing cells were observed at 145 days of gestation in one fetus with a bacterial and viral infection. IgM-containing cells were observed in 36 fetuses and IgM and IgG cells were present in seven fetuses. Spleen, lymph nodes, thymus, bone marrow, and liver of one fetus from a dam with lymphosarcoma had immunoglobulin-containing cells. Hemal lymph nodes, blood (buffy coat), Peyer patches, and heart and lung sections from fetuses with immunoglobulin-containing cells in spleen or lymph node did not have immunoglobulin-containing cells. Antigens of the virus of bovine virus diarrhea-mucosal disease (BVD) were detected in one fetus, and antigens of infectious bovine rhinotracheitis (IBR) virus were detected in three fetuses; however, viruses were not isolated in primary bovine embryonic kidney cells. Two of the three fetuses with IBR virus antigens had neutralizing serum antibody

  4. Cloning, sequence, and expression of bovine interleukin 2.

    PubMed Central

    Cerretti, D P; McKereghan, K; Larsen, A; Cantrell, M A; Anderson, D; Gillis, S; Cosman, D; Baker, P E

    1986-01-01

    Interleukin 2 (IL-2) cDNA clones have been isolated from both human and murine sources. We report here the isolation of a cDNA clone encoding bovine IL-2. This was accomplished by screening a cDNA library constructed from lectin-stimulated bovine lymph node cells, using a human IL-2 probe. Bovine IL-2 is composed of 155 amino acids and has a predicted molecular weight of 19,555. Alignment of the amino acid sequence with human IL-2 indicates that mature bovine IL-2 is composed of 135 amino acids and has a predicted molecular weight of 15,452. It has an amino acid homology of 65% with human IL-2 and 50% with murine IL-2. Bovine IL-2 is unique among IL-2 homologs in that it has a single N-linked glycosylation site. Biologically active bovine IL-2 was synthesized in an Escherichia coli expression system. Images PMID:3517854

  5. Whole genome detection of rotavirus mixed infections in human, porcine and bovine samples co-infected with various rotavirus strains collected from sub-Saharan Africa.

    PubMed

    Nyaga, Martin M; Jere, Khuzwayo C; Esona, Mathew D; Seheri, Mapaseka L; Stucker, Karla M; Halpin, Rebecca A; Akopov, Asmik; Stockwell, Timothy B; Peenze, Ina; Diop, Amadou; Ndiaye, Kader; Boula, Angeline; Maphalala, Gugu; Berejena, Chipo; Mwenda, Jason M; Steele, A Duncan; Wentworth, David E; Mphahlele, M Jeffrey

    2015-04-01

    Group A rotaviruses (RVA) are among the main global causes of severe diarrhea in children under the age of 5years. Strain diversity, mixed infections and untypeable RVA strains are frequently reported in Africa. We analysed rotavirus-positive human stool samples (n=13) obtained from hospitalised children under the age of 5years who presented with acute gastroenteritis at sentinel hospital sites in six African countries, as well as bovine and porcine stool samples (n=1 each), to gain insights into rotavirus diversity and evolution. Polyacrylamide gel electrophoresis (PAGE) analysis and genotyping with G-(VP7) and P-specific (VP4) typing primers suggested that 13 of the 15 samples contained more than 11 segments and/or mixed G/P genotypes. Full-length amplicons for each segment were generated using RVA-specific primers and sequenced using the Ion Torrent and/or Illumina MiSeq next-generation sequencing platforms. Sequencing detected at least one segment in each sample for which duplicate sequences, often having distinct genotypes, existed. This supported and extended the PAGE and RT-PCR genotyping findings that suggested these samples were collected from individuals that had mixed rotavirus infections. The study reports the first porcine (MRC-DPRU1567) and bovine (MRC-DPRU3010) mixed infections. We also report a unique genome segment 9 (VP7), whose G9 genotype belongs to lineage VI and clusters with porcine reference strains. Previously, African G9 strains have all been in lineage III. Furthermore, additional RVA segments isolated from humans have a clear evolutionary relationship with porcine, bovine and ovine rotavirus sequences, indicating relatively recent interspecies transmission and reassortment. Thus, multiple RVA strains from sub-Saharan Africa are infecting mammalian hosts with unpredictable variations in their gene segment combinations. Whole-genome sequence analyses of mixed RVA strains underscore the considerable diversity of rotavirus sequences and

  6. Whole genome detection of rotavirus mixed infections in human, porcine and bovine samples co-infected with various rotavirus strains collected from sub-Saharan Africa

    PubMed Central

    Nyaga, Martin M.; Jere, Khuzwayo C.; Esona, Mathew D.; Seheri, Mapaseka L.; Stucker, Karla M.; Halpin, Rebecca A.; Akopov, Asmik; Stockwell, Timothy B.; Peenze, Ina; Diop, Amadou; Ndiaye, Kader; Boula, Angeline; Maphalala, Gugu; Berejena, Chipo; Mwenda, Jason M.; Steele, A. Duncan; Wentworth, David E.; Mphahlele, M. Jeffrey

    2015-01-01

    Group A rotaviruses (RVA) are among the main global causes of severe diarrhea in children under the age of 5 years. Strain diversity, mixed infections and untypeable RVA strains are frequently reported in Africa. We analysed rotavirus-positive human stool samples (n=13) obtained from hospitalised children under the age of 5 years who presented with acute gastroenteritis at sentinel hospital sites in six African countries, as well as bovine and porcine stool samples (n=1 each), to gain insights into rotavirus diversity and evolution. Polyacrylamide gel electrophoresis (PAGE) analysis and genotyping with G- (VP7) and P-specific (VP4) typing primers suggested that 13 of the 15 samples contained more than 11 segments and/or mixed G/P genotypes. Full-length amplicons for each segment were generated using RVA-specific primers and sequenced using the Ion Torrent and/or Illumina MiSeq next-generation sequencing platforms. Sequencing detected at least one segment in each sample for which duplicate sequences, often having distinct genotypes, existed. This supported and extended the PAGE and RT-PCR genotyping findings that suggested these samples were collected from individuals that had mixed rotavirus infections. The study reports the first porcine (MRC-DPRU1567) and bovine (MRC-DPRU3010) mixed infections. We also report a unique genome segment 9 (VP7), whose G9 genotype belongs to lineage VI and clusters with porcine reference strains. Previously, African G9 strains have all been in lineage III. Furthermore, additional RVA segments isolated from humans have a clear evolutionary relationship with porcine, bovine and ovine rotavirus sequences, indicating relatively recent interspecies transmission and reassortment. Thus, multiple RVA strains from sub-Saharan Africa are infecting mammalian hosts with unpredictable variations in their gene segment combinations. Whole-genome sequence analyses of mixed RVA strains underscore the considerable diversity of rotavirus sequences and

  7. Killing of Brucella abortus by bovine serum.

    PubMed Central

    Corbeil, L B; Blau, K; Inzana, T J; Nielsen, K H; Jacobson, R H; Corbeil, R R; Winter, A J

    1988-01-01

    Studies of the serum bactericidal system in bovine brucellosis were undertaken to investigate the role of the humoral immune response in protection of cattle against the facultative intracellular parasite Brucella abortus. Fresh sera from normal control cattle, infected cattle, and cattle immunized with B. abortus cell envelopes were collected before treatment and during the course of immunization or infection. Normal fresh bovine serum or fresh agammaglobulinemic serum from colostrum-deprived calves was effective in killing smooth virulent B. abortus 2308, but rough strains RB51 (a rough mutant of strain 2308) and 45/20 were much more sensitive to serum. The difference in susceptibility to serum was shown to be correlated with differences in lipopolysaccharide chemotype, with the more resistant strain 2308 having O polysaccharide and the more susceptible strains 45/20 and RB51 lacking O side chains. By treatment of fresh serum with MgCl2 and EGTA [ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid] killing was shown to occur via the classical pathway of complement activation. When antibody to B. abortus was present, killing of strain RB51 increased but killing of smooth strain 2308 decreased. The earliest antibody response in serum from infected animals did not interfere with killing. When affinity-purified bovine immunoglobulins specific for B. abortus smooth lipopolysaccharide were added to fresh normal bovine serum, immunoglobulin G1 (IgG1) and IgG2 isotypes blocked killing but IgM and IgA isotypes did not. Thus, it appears that serum from previously unexposed animals or animals early during infection can kill smooth B. abortus, an appropriate defense mechanism before the organism becomes intracellular. At later stages of infection, blocking antibodies predominate. Images PMID:3141287

  8. Random laser action in bovine semen

    NASA Astrophysics Data System (ADS)

    Smuk, Andrei; Lazaro, Edgar; Olson, Leif P.; Lawandy, N. M.

    2011-03-01

    Experiments using bovine semen reveal that the addition of a high-gain water soluble dye results in random laser action when excited by a Q-switched, frequency doubled, Nd:Yag laser. The data shows that the linewidth collapse of the emission is correlated to the sperm count of the individual samples, potentially making this a rapid, low sample volume approach to count determination.

  9. Unraveling bovin phylogeny: accomplishments and challenges

    PubMed Central

    2010-01-01

    The phylogenetic systematics of bovin species forms a common basis for studies at multiple scales, from the level of domestication in populations to major cladogenesis. The main big-picture accomplishments of this productive field, including two recent works, one in BMC Genomics, are reviewed with an eye for some of the limitations and challenges impeding progress. See Research article http://www.biomedcentral.com/1471-2164/10/177 PMID:20525112

  10. PHYSICOCHEMICAL CHARACTERIZATION OF LYOPHILIZED BOVINE BONE GRAFTS

    PubMed Central

    Galia, Carlos Roberto; Lourenço, André Luis; Rosito, Ricardo; Souza Macedo, Carlos Alberto; Camargo, Lourdes Maria Araujo Quaresma

    2015-01-01

    To evaluate the physicochemical characteristics of lyophilized bovine grafts manufactured on a semi-industrial scale (Orthogen; Baumer S/A*) in accordance with a protocol previously developed by the authors. Methods: The lyophilized bovine bone grafts were characterized by means of scanning electron microscopy (SEM), energy dispersive spectroscopy (EDS), X-ray diffractometry (XRD), thermogravimetric (TG) analysis, differential exploratory scanning calorimetry (DSC) and Fourier-transform infrared (FT-IR) spectroscopy. Results: Ca was the main component (60%) found in the samples, followed by P (28%) and O (5%). The mean (sd) pore size was 316 μm (146.7), ranging from 91.2 to 497.8 μm, and 333.5 μm (304.8), ranging from 87.2 to 963.9 μm, at 50x and 150x magnification, respectively. The hydroxyapatite peaks were at 26°C and 32°C, and mass losses were observed between 250°C and 640°C, corresponding to organic material and water. Two temperature transitions (45.67°C and 91.89°C) showed denaturation of type 1 collagen and dehydration of hydroxyapatite. Conclusion: The physicochemical assessment of lyophilized bovine bone grafts in accordance with the protocol developed at semi-industrial scale confirmed that this product presents excellent biocompatibility, with characteristics similar to natural bone. PMID:27027036

  11. Oxytocin binding sites in bovine mammary tissue

    SciTech Connect

    Zhao, Xin.

    1989-01-01

    Oxytocin binding sites were identified and characterized in bovine mammary tissue. ({sup 3}H)-oxytocin binding reached equilibrium by 50 min at 20{degree}C and by 8 hr at 4{degree}C. The half-time of displacement at 20{degree}C was approximately 1 hr. Thyrotropin releasing hormone, adrenocorticotropin, angiotensin I, angiotensin II, pentagastrin, bradykinin, xenopsin and L-valyl-histidyl-L-leucyl-L-threonyl-L-prolyl-L-valyl-L-glutamyl-L-lysine were not competitive. In the presence of 10 nM LiCl, addition of oxytocin to dispersed bovine mammary cells, in which phosphatidylinositol was pre-labelled, caused a time and dose-dependent increase in radioactive inositiol monophosphate incorporation. The possibility that there are distinct vasopressin receptors in bovine mammary tissue was investigated. ({sup 3}H)-vasopressin binding reached equilibrium by 40 min at 20{degree}. The half-time of displacement at 20{degree}C was approximately 1 hr. The ability of the peptides to inhibit ({sup 3}H)-vasopressin binding was: (Thr{sup 4},Gly{sup 7})-oxytocin > Arg{sup 8}-vasopressin > (lys{sup 8})-vasopressin > (Deamino{sup 1},D-arg{sup 8})-vasopressin > oxytocin > d (CH{sub 2}){sub 5}Tyr(Me)AVP.

  12. Bovine Postparturient Hemoglobinuria: A Review of the Literature

    PubMed Central

    Macwilliams, P. S.; Searcy, G. P.; Bellamy, J. E. C.

    1982-01-01

    Predisposing causes of bovine post-parturient hemoglobinuria are summarized along with suspected pathogeneses, clinical signs, laboratory findings, clinical management and early experimentation. PMID:17422195

  13. Approaches to biosecurity in bovine embryo transfer programs.

    PubMed

    Givens, M D; Marley, S D

    2008-01-01

    Although transfer of bovine embryos is much less likely to result in transmission of pathogens than transport of postnatal cattle, the epidemiologic risk associated with bovine embryo transfer merits examination. Much research has validated the efficacy of internationally approved processing protocols to render bovine in vivo-derived embryos free of specified pathogens. The purpose of this review is to summarize current sanitary recommendations for bovine embryo transfer, while emphasizing recent research to develop and validate novel approaches to biosecurity. Continued research will enable the development and validation of novel embryo treatments and culture reagents to minimize requirements for testing of embryo or oocyte donors, and testing of embryo recipients. PMID:18028999

  14. The bovine patella as a model of early osteoarthritis

    PubMed Central

    Hargrave-Thomas, E J; Thambyah, A; McGlashan, S R; Broom, N D

    2013-01-01

    The bovine patella model has been used extensively for studying important structure–function aspects of articular cartilage, including its degeneration. However, the degeneration seen in this model has, to our knowledge, never been adequately compared with human osteoarthritis (OA). In this study, bovine patellae displaying normal to severely degenerate states were compared with human tissue displaying intact cartilage to severe OA. Comparisons of normal and OA features were made with histological scoring, morphometric measurements, and qualitative observations. Differential interference contrast microscopy was used to image early OA changes in the articular cartilage matrix and to investigate whether this method provided comparable quality of visualisation of key structural features with standard histology. The intact bovine cartilage was found to be similar to healthy human cartilage and the degenerate bovine cartilage resembled the human OA tissues with regard to structural disruption, cellularity changes, and staining loss. The extent of degeneration in the bovine tissues matched the mild to moderate range of human OA tissues; however, no bovine samples exhibited late-stage OA. Additionally, in both bovine and human tissues, cartilage degeneration was accompanied by calcified cartilage thickening, tidemark duplication, and the advancement of the cement line by protrusions of bony spicules into the calcified cartilage. This comparison of degeneration in the bovine and human tissues suggests a common pathway for the progression of OA and thus the bovine patella is proposed to be an appropriate model for investigating the structural changes associated with early OA. PMID:24111904

  15. Bovine besnoitiosis emerging in Central-Eastern Europe, Hungary

    PubMed Central

    2014-01-01

    Background Besnoitia besnoiti, the cause of bovine besnoitiosis, is a cyst-forming coccidian parasite that has recently been shown to be spreading in several Western and Southern European countries. Findings Clinical cases of bovine besnoitiosis were confirmed for the first time in Hungary, by histological, serological and PCR analyses. Conclusions This is the first report of autochthonous bovine besnoitiosis in Central-Eastern Europe. The emergence of bovine besnoitiosis in this region represents a further example, when human activity (i.e. cattle trading) is the main factor involved in the geographical spread of an infectious disease. PMID:24410743

  16. Lactic Acid Bacteria Isolated from Bovine Mammary Microbiota: Potential Allies against Bovine Mastitis

    PubMed Central

    Saraoui, Taous; Rault, Lucie; Germon, Pierre; Gonzalez-Moreno, Candelaria; Nader-Macias, Fatima M. E.; Baud, Damien; François, Patrice; Chuat, Victoria; Chain, Florian; Langella, Philippe; Nicoli, Jacques; Le Loir, Yves; Even, Sergine

    2015-01-01

    Bovine mastitis is a costly disease in dairy cattle worldwide. As of yet, the control of bovine mastitis is mostly based on prevention by thorough hygienic procedures during milking. Additional strategies include vaccination and utilization of antibiotics. Despite these measures, mastitis is not fully under control, thus prompting the need for alternative strategies. The goal of this study was to isolate autochthonous lactic acid bacteria (LAB) from bovine mammary microbiota that exhibit beneficial properties that could be used for mastitis prevention and/or treatment. Sampling of the teat canal led to the isolation of 165 isolates, among which a selection of ten non-redundant LAB strains belonging to the genera Lactobacillus and Lactococcus were further characterized with regard to several properties: surface properties (hydrophobicity, autoaggregation); inhibition potential of three main mastitis pathogens, Staphylococcus aureus, Escherichia coli and Streptococcus uberis; colonization capacities of bovine mammary epithelial cells (bMEC); and immunomodulation properties. Three strains, Lactobacillus brevis 1595 and 1597 and Lactobacillus plantarum 1610, showed high colonization capacities and a medium surface hydrophobicity. These strains are good candidates to compete with pathogens for mammary gland colonization. Moreover, nine strains exhibited anti-inflammatory properties, as illustrated by the lower IL-8 secretion by E. coli-stimulated bMEC in the presence of these LAB. Full genome sequencing of five candidate strains allowed to check for undesirable genetic elements such as antibiotic resistance genes and to identify potential bacterial determinants involved in the beneficial properties. This large screening of beneficial properties while checking for undesirable genetic markers allowed the selection of promising candidate LAB strains from bovine mammary microbiota for the prevention and/or treatment of bovine mastitis. PMID:26713450

  17. Diagnosis and Control of Viral Diseases of Reproductive Importance: Infectious Bovine Rhinotracheitis and Bovine Viral Diarrhea.

    PubMed

    Newcomer, Benjamin W; Givens, Daniel

    2016-07-01

    Both bovine viral diarrhea virus and bovine herpesvirus 1 can have significant negative reproductive impacts on cattle health. Vaccination is the primary control method for the viral pathogens in US cattle herds. Polyvalent, modified-live vaccines are recommended to provide optimal protection against various viral field strains. Of particular importance to bovine viral diarrhea control is the limitation of contact of pregnant cattle with potential viral reservoirs during the critical first 125 days of gestation. PMID:27140298

  18. Evaluation of DNA extraction kits and phylogenetic diversity of the porcine gastrointestinal tract based on Illumina sequencing of two hypervariable regions.

    PubMed

    Burbach, Katharina; Seifert, Jana; Pieper, Dietmar H; Camarinha-Silva, Amélia

    2016-02-01

    A robust DNA extraction method is important to identify the majority of microorganisms present in environmental microbial communities and to enable a consistent comparison between different studies. Here, 15 manual and four automated commercial DNA extraction kits were evaluated for their efficiency to extract DNA from porcine feces and ileal digesta samples. DNA yield, integrity, and purity varied among the different methods. Terminal restriction fragment length polymorphism (T-RFLP) and Illumina amplicon sequencing were used to characterize the diversity and composition of the microbial communities. We also compared phylogenetic profiles of two regions of the 16S rRNA gene, one of the most used region (V1-2) and the V5-6 region. A high correlation between community structures obtained by analyzing both regions was observed at genus and family level for ileum digesta and feces. Based on our findings, we want to recommend the FastDNA(™) SPIN Kit for Soil (MP Biomedical) as a suitable kit for the analyses of porcine gastrointestinal tract samples. PMID:26541370

  19. Performance Comparison of Illumina and Ion Torrent Next-Generation Sequencing Platforms for 16S rRNA-Based Bacterial Community Profiling

    PubMed Central

    Kawashima, Toana; Rosenthal, Christopher; Hoogestraat, Daniel R.; Cummings, Lisa A.; Sengupta, Dhruba J.; Harkins, Timothy T.; Cookson, Brad T.

    2014-01-01

    High-throughput sequencing of the taxonomically informative 16S rRNA gene provides a powerful approach for exploring microbial diversity. Here we compare the performances of two common “benchtop” sequencing platforms, Illumina MiSeq and Ion Torrent Personal Genome Machine (PGM), for bacterial community profiling by 16S rRNA (V1-V2) amplicon sequencing. We benchmarked performance by using a 20-organism mock bacterial community and a collection of primary human specimens. We observed comparatively higher error rates with the Ion Torrent platform and report a pattern of premature sequence truncation specific to semiconductor sequencing. Read truncation was dependent on both the directionality of sequencing and the target species, resulting in organism-specific biases in community profiles. We found that these sequencing artifacts could be minimized by using bidirectional amplicon sequencing and an optimized flow order on the Ion Torrent platform. Results of bacterial community profiling performed on the mock community and a collection of 18 human-derived microbiological specimens were generally in good agreement for both platforms; however, in some cases, results differed significantly. Disparities could be attributed to the failure to generate full-length reads for particular organisms on the Ion Torrent platform, organism-dependent differences in sequence error rates affecting classification of certain species, or some combination of these factors. This study demonstrates the potential for differential bias in bacterial community profiles resulting from the choice of sequencing platform alone. PMID:25261520

  20. SSR_pipeline--computer software for the identification of microsatellite sequences from paired-end Illumina high-throughput DNA sequence data

    USGS Publications Warehouse

    Miller, Mark P.; Knaus, Brian J.; Mullins, Thomas D.; Haig, Susan M.

    2013-01-01

    SSR_pipeline is a flexible set of programs designed to efficiently identify simple sequence repeats (SSRs; for example, microsatellites) from paired-end high-throughput Illumina DNA sequencing data. The program suite contains three analysis modules along with a fourth control module that can be used to automate analyses of large volumes of data. The modules are used to (1) identify the subset of paired-end sequences that pass quality standards, (2) align paired-end reads into a single composite DNA sequence, and (3) identify sequences that possess microsatellites conforming to user specified parameters. Each of the three separate analysis modules also can be used independently to provide greater flexibility or to work with FASTQ or FASTA files generated from other sequencing platforms (Roche 454, Ion Torrent, etc). All modules are implemented in the Python programming language and can therefore be used from nearly any computer operating system (Linux, Macintosh, Windows). The program suite relies on a compiled Python extension module to perform paired-end alignments. Instructions for compiling the extension from source code are provided in the documentation. Users who do not have Python installed on their computers or who do not have the ability to compile software also may choose to download packaged executable files. These files include all Python scripts, a copy of the compiled extension module, and a minimal installation of Python in a single binary executable. See program documentation for more information.

  1. Isolation and Characterization of Microsatellite DNA Markers in the Deep-Sea Amphipod Paralicella tenuipes by Illumina MiSeq Sequencing.

    PubMed

    Ritchie, Heather; Jamieson, Alan J; Piertney, Stuart B

    2016-07-01

    Here, we describe the development of 16 polymorphic microsatellite markers using an Illumina MiSeq sequencing approach in the deep-sea amphipod Paralicella tenuipes A total of 25 577 844 DNA sequences were filtered for microsatellite motifs of which 197 873 sequences were identified. From these sequences, 64 had sufficient flanking regions for primer design and 16 of these loci were polymorphic. Between 5 and 30 alleles were detected per locus, with an average of 13.63 alleles per locus, across a total of 120 individuals from 5 separate deep sea trenches from the Pacific Ocean. For the 16 loci, observed and expected heterozygosity values ranged from 0.116 to 0.414 and 0.422 to 0.820, respectively, with one locus displaying significant deviation from Hardy-Weinberg equilibrium. The microsatellite loci that have been isolated and described here are the first molecular markers developed for deep sea amphipods and will be invaluable for elucidating the genetic population structure and the extent of connectivity between deep ocean trenches. PMID:27012615

  2. Transcriptome analysis of colored calla lily (Zantedeschia rehmannii Engl.) by Illumina sequencing: de novo assembly, annotation and EST-SSR marker development

    PubMed Central

    Cui, Binbin; Zhang, Qixiang; Xiong, Min; Wang, Xian

    2016-01-01

    Colored calla lily is the short name for the species or hybrids in section Aestivae of genus Zantedeschia. It is currently one of the most popular flower plants in the world due to its beautiful flower spathe and long postharvest life. However, little genomic information and few molecular markers are available for its genetic improvement. Here, de novo transcriptome sequencing was performed to produce large transcript sequences for Z. rehmannii cv. ‘Rehmannii’ using an Illumina HiSeq 2000 instrument. More than 59.9 million cDNA sequence reads were obtained and assembled into 39,298 unigenes with an average length of 1,038 bp. Among these, 21,077 unigenes showed significant similarity to protein sequences in the non-redundant protein database (Nr) and in the Swiss-Prot, Gene Ontology (GO), Cluster of Orthologous Group (COG) and Kyoto Encyclopedia of Genes and Genomes (KEGG) databases. Moreover, a total of 117 unique transcripts were then defined that might regulate the flower spathe development of colored calla lily. Additionally, 9,933 simple sequence repeats (SSRs) and 7,162 single nucleotide polymorphisms (SNPs) were identified as putative molecular markers. High-quality primers for 200 SSR loci were designed and selected, of which 58 amplified reproducible amplicons were polymorphic among 21 accessions of colored calla lily. The sequence information and molecular markers in the present study will provide valuable resources for genetic diversity analysis, germplasm characterization and marker-assisted selection in the genus Zantedeschia.

  3. De novo Transcriptome Generation and Annotation for Two Korean Endemic Land Snails, Aegista chejuensis and Aegista quelpartensis, Using Illumina Paired-End Sequencing Technology

    PubMed Central

    Kang, Se Won; Patnaik, Bharat Bhusan; Hwang, Hee-Ju; Park, So Young; Wang, Tae Hun; Park, Eun Bi; Chung, Jong Min; Song, Dae Kwon; Patnaik, Hongray Howrelia; Lee, Jae Bong; Kim, Changmu; Kim, Soonok; Park, Hong Seog; Lee, Jun Sang; Han, Yeon Soo; Lee, Yong Seok

    2016-01-01

    Aegista chejuensis and Aegista quelpartensis (Family-Bradybaenidae) are endemic to Korea, and are considered vulnerable due to declines in their population. The limited genetic resources for these species restricts the ability to prioritize conservation efforts. We sequenced the transcriptomes of these species using Illumina paired-end technology. Approximately 257 and 240 million reads were obtained and assembled into 198,531 and 230,497 unigenes for A. chejuensis and A. quelpartensis, respectively. The average and N50 unigene lengths were 735.4 and 1073 bp, respectively, for A. chejuensis, and 705.6 and 1001 bp, respectively, for A. quelpartensis. In total, 68,484 (34.5%) and 77,745 (33.73%) unigenes for A. chejuensis and A. quelpartensis, respectively, were annotated to databases. Gene Ontology terms were assigned to 23,778 (11.98%) and 26,396 (11.45) unigenes, for A. chejuensis and A. quelpartensis, respectively, while 5050 and 5838 unigenes were mapped to 117 and 124 pathways in the Kyoto Encyclopedia of Genes and Genomes database. In addition, we identified and annotated 9542 and 10,395 putative simple sequence repeats (SSRs) in unigenes from A. chejuensis and A. quelpartensis, respectively. We designed a list of PCR primers flanking the putative SSR regions. These microsatellites may be utilized for future phylogenetics and conservation initiatives. PMID:26999110

  4. Characterization of the Kenaf (Hibiscus cannabinus) Global Transcriptome Using Illumina Paired-End Sequencing and Development of EST-SSR Markers.

    PubMed

    Li, Hui; Li, Defang; Chen, Anguo; Tang, Huijuan; Li, Jianjun; Huang, Siqi

    2016-01-01

    Kenaf (Hibiscus cannabinus L.) is an economically important natural fiber crop grown worldwide. However, only 20 expressed tag sequences (ESTs) for kenaf are available in public databases. The aim of this study was to develop large-scale simple sequence repeat (SSR) markers to lay a solid foundation for the construction of genetic linkage maps and marker-assisted breeding in kenaf. We used Illumina paired-end sequencing technology to generate new EST-simple sequences and MISA software to mine SSR markers. We identified 71,318 unigenes with an average length of 1143 nt and annotated these unigenes using four different protein databases. Overall, 9324 complementary pairs were designated as EST-SSR markers, and their quality was validated using 100 randomly selected SSR markers. In total, 72 primer pairs reproducibly amplified target amplicons, and 61 of these primer pairs detected significant polymorphism among 28 kenaf accessions. Thus, in this study, we have developed large-scale SSR markers for kenaf, and this new resource will facilitate construction of genetic linkage maps, investigation of fiber growth and development in kenaf, and also be of value to novel gene discovery and functional genomic studies. PMID:26960153

  5. Characterization of the Kenaf (Hibiscus cannabinus) Global Transcriptome Using Illumina Paired-End Sequencing and Development of EST-SSR Markers

    PubMed Central

    Li, Hui; Li, Defang; Chen, Anguo; Tang, Huijuan; Li, Jianjun; Huang, Siqi

    2016-01-01

    Kenaf (Hibiscus cannabinus L.) is an economically important natural fiber crop grown worldwide. However, only 20 expressed tag sequences (ESTs) for kenaf are available in public databases. The aim of this study was to develop large-scale simple sequence repeat (SSR) markers to lay a solid foundation for the construction of genetic linkage maps and marker-assisted breeding in kenaf. We used Illumina paired-end sequencing technology to generate new EST-simple sequences and MISA software to mine SSR markers. We identified 71,318 unigenes with an average length of 1143 nt and annotated these unigenes using four different protein databases. Overall, 9324 complementary pairs were designated as EST-SSR markers, and their quality was validated using 100 randomly selected SSR markers. In total, 72 primer pairs reproducibly amplified target amplicons, and 61 of these primer pairs detected significant polymorphism among 28 kenaf accessions. Thus, in this study, we have developed large-scale SSR markers for kenaf, and this new resource will facilitate construction of genetic linkage maps, investigation of fiber growth and development in kenaf, and also be of value to novel gene discovery and functional genomic studies. PMID:26960153

  6. Illumina MiSeq sequencing reveals the key microorganisms involved in partial nitritation followed by simultaneous sludge fermentation, denitrification and anammox process.

    PubMed

    Wang, Bo; Peng, Yongzhen; Guo, Yuanyuan; Zhao, Mengyue; Wang, Shuying

    2016-05-01

    A combined process including a partial nitritation SBR (PN-SBR) followed by a simultaneous sludge fermentation, denitrification and anammox reactor (SFDA) was established to treat low C/N domestic wastewater in this study. An average nitrite accumulation rate of 97.8% and total nitrogen of 9.4mg/L in the effluent was achieved during 140days' operation. The underlying mechanisms were investigated by using Illumina MiSeq sequencing to analyze the microbial community structures in the PN-SBR and SFDA. Results showed that the predominant bacterial phylum was Proteobacteria in the external waste activated sludge (WAS, added to the SFDA) and SFDA while Bacteroidetes in the PN-SBR. Further study indicated that in the PN-SBR, the dominant nitrobacteria, Nitrosomonas genus, facilitated nitritation and little nitrate was generated in the PN-SBR effluent. In the SFDA, the co-existence of functional microorganisms Thauera, Candidatus Anammoximicrobium and Pseudomonas were found to contribute to simultaneous sludge fermentation, denitrification and anammox. PMID:26874440

  7. Illumina sequencing-based analysis of free-living bacterial community dynamics during an Akashiwo sanguine bloom in Xiamen sea, China

    PubMed Central

    Yang, Caiyun; Li, Yi; Zhou, Benjamin; Zhou, Yanyan; Zheng, Wei; Tian, Yun; Van Nostrand, Joy D.; Wu, Liyou; He, Zhili; Zhou, Jizhong; Zheng, Tianling

    2015-01-01

    Although phytoplankton are the major source of marine dissolved organic matter (DOM), their blooms are a global problem that can greatly affect marine ecological systems, especially free-living bacteria, which are the primary DOM degraders. In this study, we analyzed free-living bacterial communities from Xiamen sea during an Akashiwo sanguine bloom using Illumina MiSeq sequencing of 16S rRNA gene amplicons. The bloom was probably stimulated by low salinity and ended after abatement of eutrophication pollution. A total of 658,446 sequence reads and 11,807 OTUs were obtained in both bloom and control samples with Alpha-proteobacteria and Gamma-proteobacteria being the predominant classes detected. The bloom decreased bacterial diversity, increased species evenness, and significantly changed the bacterial community structure. Bacterial communities within the bloom were more homogeneous than those within the control area. The bacteria stimulated by this bloom included the SAR86 and SAR116 clades and the AEGEAN-169 marine group, but a few were suppressed. In addition, many bacteria known to be associated with phytoplankton were detected only in the bloom samples. This study revealed the great influence of an A. sanguinea bloom on free-living bacterial communities, and provided new insights into the relationship between bacteria and A. sanguinea in marine ecosystems. PMID:25684124

  8. De novo transcriptome analysis of an imminent biofuel crop, Camelina sativa L. using Illumina GAIIX sequencing platform and identification of SSR markers.

    PubMed

    Mudalkar, Shalini; Golla, Ramesh; Ghatty, Sreenivas; Reddy, Attipalli Ramachandra

    2014-01-01

    Camelina sativa L. is an emerging biofuel crop with potential applications in industry, medicine, cosmetics and human nutrition. The crop is unexploited owing to very limited availability of transcriptome and genomic data. In order to analyse the various metabolic pathways, we performed de novo assembly of the transcriptome on Illumina GAIIX platform with paired end sequencing for obtaining short reads. The sequencing output generated a FastQ file size of 2.97 GB with 10.83 million reads having a maximum read length of 101 nucleotides. The number of contigs generated was 53,854 with maximum and minimum lengths of 10,086 and 200 nucleotides respectively. These trancripts were annotated using BLAST search against the Aracyc, Swiss-Prot, TrEMBL, gene ontology and clusters of orthologous groups (KOG) databases. The genes involved in lipid metabolism were studied and the transcription factors were identified. Sequence similarity studies of Camelina with the other related organisms indicated the close relatedness of Camelina with Arabidopsis. In addition, bioinformatics analysis revealed the presence of a total of 19,379 simple sequence repeats. This is the first report on Camelina sativa L., where the transcriptome of the entire plant, including seedlings, seed, root, leaves and stem was done. Our data established an excellent resource for gene discovery and provide useful information for functional and comparative genomic studies in this promising biofuel crop. PMID:24002439

  9. Transcriptome Profiling and Identification of Transcription Factors in Ramie (Boehmeria nivea L. Gaud) in Response to PEG Treatment, Using Illumina Paired-End Sequencing Technology

    PubMed Central

    An, Xia; Chen, Jie; Zhang, Jingyu; Liao, Yiwen; Dai, Lunjin; Wang, Bo; Liu, Lijun; Peng, Dingxiang

    2015-01-01

    Ramie (Boehmeria nivea L. Gaud), commonly known as China grass, is a perennial bast fiber plant of the Urticaceae. In China, ramie farming, industry, and trade provide income for about five million people. Drought stress severely affects ramie stem growth and causes a dramatic decrease in ramie fiber production. There is a need to enhance ramie’s tolerance to drought stress. However, the drought stress regulatory mechanism in ramie remains unknown. Water stress imposed by polyethylene glycol (PEG) is a common and convenient method to evaluate plant drought tolerance. In this study, transcriptome analysis of cDNA collections from ramie subjected to PEG treatment was conducted using Illumina paired-end sequencing, which generated 170 million raw sequence reads. Between leaves and roots subjected to 24 (L2 and R2) and 72 (L3 and R3) h of PEG treatment, 16,798 genes were differentially expressed (9281 in leaves and 8627 in roots). Among these, 25 transcription factors (TFs) from the AP2 (3), MYB (6), NAC (9), zinc finger (5), and bZIP (2) families were considered to be associated with drought stress. The identified TFs could be used to further investigate drought adaptation in ramie. PMID:25658800

  10. Illumina MiSeq sequencing reveals long-term impacts of single-walled carbon nanotubes on microbial communities of wastewater treatment systems.

    PubMed

    Qu, Yuanyuan; Zhang, Xuwang; Shen, Wenli; Ma, Qiao; You, Shengnan; Pei, Xiaofang; Li, Shuzhen; Ma, Fang; Zhou, Jiti

    2016-07-01

    In this study, phenol wastewater treatment systems treated with different concentrations of single-walled carbon nanotubes (SWCNTs) (0-3.5g/L) were exposed to phenol and carbon nanotubes (CNTs) shock loadings to investigate the long-term impacts of SWCNTs on microbial communities. Phenol removal remained high efficiency (>98%) in SWCNTs-treated groups but decreased in non-treated group (85.1±1.9%) when exposed to high concentration of phenol (500mg/L). However, secondary dosing of SWCNTs in SWCNTs-treated groups would decrease the phenol removal efficiency. Illumina MiSeq sequencing revealed that the diversity, richness and structure of microbial communities were shifted under phenol shock loading, especially under high phenol concentration, but not under CNTs shock loading. In response to phenol and CNTs shock loadings, Rudaea, Burkholderia, Sphingomonas, Acinetobacter, Methylocystis and Thauera became dominant genera, which should be involved in phenol removal. These results suggested that a proper amount of SWCNTs might have positive effects on phenol wastewater treatment systems. PMID:27017131

  11. Characterization of Global Transcriptome Using Illumina Paired-End Sequencing and Development of EST-SSR Markers in Two Species of Gynostemma (Cucurbitaceae).

    PubMed

    Zhao, Yue-Mei; Zhou, Tao; Li, Zhong-Hu; Zhao, Gui-Fang

    2015-01-01

    Gynostemma pentaphyllum is an important medicinal herb of the Cucurbitaceae family, but limited genomic data have hindered genetic studies. In this study, transcriptomes of two closely-related Gynostemma species, Gynostemma cardiospermum and G. pentaphyllum, were sequenced using Illumina paired-end sequencing technology. A total of 71,607 nonredundant unigenes were assembled. Of these unigenes, 60.45% (43,288) were annotated based on sequence similarity search with known proteins. A total of 11,059 unigenes were identified in the Kyoto Encyclopedia of Genes and Genomes Pathway (KEGG) database. A total of 3891 simple sequence repeats (SSRs) were detected in 3526 nonredundant unigenes, 2596 primer pairs were designed and 360 of them were randomly selected for validation. Of these, 268 primer pairs yielded clear products among six G. pentaphyllum samples. Thirty polymorphic SSR markers were used to test polymorphism and transferability in Gynostemma. Finally, 15 SSR makers that amplified in all 12 Gynostemma species were used to assess genetic diversity. Our results generated a comprehensive sequence resource for Gynostemma research. PMID:26633323

  12. The utility of diversity profiling using Illumina 18S rRNA gene amplicon deep sequencing to detect and discriminate Toxoplasma gondii among the cyst-forming coccidia.

    PubMed

    Cooper, Madalyn K; Phalen, David N; Donahoe, Shannon L; Rose, Karrie; Šlapeta, Jan

    2016-01-30

    Next-generation sequencing (NGS) has the capacity to screen a single DNA sample and detect pathogen DNA from thousands of host DNA sequence reads, making it a versatile and informative tool for investigation of pathogens in diseased animals. The technique is effective and labor saving in the initial identification of pathogens, and will complement conventional diagnostic tests to associate the candidate pathogen with a disease process. In this report, we investigated the utility of the diversity profiling NGS approach using Illumina small subunit ribosomal RNA (18S rRNA) gene amplicon deep sequencing to detect Toxoplasma gondii in previously confirmed cases of toxoplasmosis. We then tested the diagnostic approach with species-specific PCR genotyping, histopathology and immunohistochemistry of toxoplasmosis in a Risso's dolphin (Grampus griseus) to systematically characterise the disease and associate causality. We show that the Euk7A/Euk570R primer set targeting the V1-V3 hypervariable region of the 18S rRNA gene can be used as a species-specific assay for cyst-forming coccidia and discriminate T. gondii. Overall, the approach is cost-effective and improves diagnostic decision support by narrowing the differential diagnosis list with more certainty than was previously possible. Furthermore, it supplements the limitations of cryptic protozoan morphology and surpasses the need for species-specific PCR primer combinations. PMID:26801593

  13. Development and Applications of a Bovine 50,000 SNP Chip

    Technology Transfer Automated Retrieval System (TEKTRAN)

    To develop an Illumina iSelect high density single nucleotide polymorphism (SNP) assay for cattle, the collaborative iBMC (Illumina, USDA ARS Beltsville, University of Missouri, USDA ARS Clay Center) Consortium first performed a de novo SNP discovery project in which genomic reduced representation l...

  14. Bovine viral diarrhea virus infection alters global transcription profiles in bovine endothelial cells

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bovine viral diarrhea viruses (BVDV) are significant pathogens of cattle worldwide. These viruses exist in both non-cytopathic and cytopathic biotypes. Non-cytopathic BVDV can establish persistent lifelong infections in cattle and are a frequent contaminant of biological reagents such as cell cultur...

  15. Bovine central memory T cells are highly proliferative in response to bovine tuberculosis infection

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Long-term (i.e., 14 days) cultured IFN-gamma ELISPOT assays measure central memory T cell (Tcm) responses in both humans and cattle. With bovine tuberculosis, vaccine-elicited long-term IFN-gamma ELISPOT responses correlate with protection. In other species, Tcm’s pose low activation threshold and a...

  16. Description and analysis of the Bovine Gene Atlas - An extensive compendium of bovine transcript profiles

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The Bovine Gene Atlas (BGA) is a compendium of over 7.2 million unique 20-base transcript tags profiled from 81 tissues acquired from the cow “L1 Dominette 01449” (L1D), her male fetus, her 255-day-old heifer calf, and her father. The BGA tags were generated on a next-generation massively parallel ...

  17. Bovine central memory T cells are highly proliferative in response to bovine tuberculosis infection

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Long-term (i.e., 14 days) cultured IFN-gamma responses of peripheral blood mononuclear cells are used as a correlate of T cell central memory (Tcm) responses in both humans and cattle. With bovine tuberculosis, vaccine-elicited long-term IFN-gamma ELISPOT assays are a correlate of protection. Recent...

  18. Bovine Genetic Diversity Revealed By mtDNA Sequence Variation

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Mitochondrial DNA single nucleotide polymorphism (SNP) data were used to determine genetic distance, nucleotide diversity, construction of haplotypes, estimation of information contents, and phylogenic relationships in bovine HapMap breeds. The Bovine International HapMap panel consists of 720 anima...

  19. Prevalence, transmission and impact of bovine leukosis in Michigan dairies

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bovine leukosis, caused by infection with the retrovirus bovine leukemia virus (BLV), has been characterized as a contagious, but practically benign disease of the immune system. National Animal Health Monitoring Surveys in 1996 and 2007 indicate complacency has resulted in high prevalence of infect...

  20. Identification of highly active flocculant proteins in bovine blood

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bovine blood is an excellent flocculating agent, faster acting and as effective on a mass basis as polyacrylamide, the most widely utilized polymeric flocculant. To determine the molecular basis of flocculation activity, whole bovine blood (BB) and BB plasma were fractionated by size exclusion chro...

  1. 9 CFR 113.68 - Pasteurella Haemolytica Vaccine, Bovine.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 9 Animals and Animal Products 1 2013-01-01 2013-01-01 false Pasteurella Haemolytica Vaccine... REQUIREMENTS Live Bacterial Vaccines § 113.68 Pasteurella Haemolytica Vaccine, Bovine. Pasteurella Haemolytica Vaccine, Bovine, shall be prepared as a desiccated live culture bacterial vaccine of an avirulent...

  2. 9 CFR 113.68 - Pasteurella Haemolytica Vaccine, Bovine.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 9 Animals and Animal Products 1 2012-01-01 2012-01-01 false Pasteurella Haemolytica Vaccine... REQUIREMENTS Live Bacterial Vaccines § 113.68 Pasteurella Haemolytica Vaccine, Bovine. Pasteurella Haemolytica Vaccine, Bovine, shall be prepared as a desiccated live culture bacterial vaccine of an avirulent...

  3. 9 CFR 113.68 - Pasteurella Haemolytica Vaccine, Bovine.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 9 Animals and Animal Products 1 2011-01-01 2011-01-01 false Pasteurella Haemolytica Vaccine... REQUIREMENTS Live Bacterial Vaccines § 113.68 Pasteurella Haemolytica Vaccine, Bovine. Pasteurella Haemolytica Vaccine, Bovine, shall be prepared as a desiccated live culture bacterial vaccine of an avirulent...

  4. 9 CFR 113.68 - Pasteurella Haemolytica Vaccine, Bovine.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 9 Animals and Animal Products 1 2014-01-01 2014-01-01 false Pasteurella Haemolytica Vaccine... REQUIREMENTS Live Bacterial Vaccines § 113.68 Pasteurella Haemolytica Vaccine, Bovine. Pasteurella Haemolytica Vaccine, Bovine, shall be prepared as a desiccated live culture bacterial vaccine of an avirulent...

  5. Bovine viral diarrhea virus modulations of monocyte derived macrophages

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bovine viral diarrhea virus (BVDV) is a single stranded, positive sense RNA virus and is the causative agent of bovine viral diarrhea (BVD). Disease can range from persistently infected (PI) animals displaying no clinical symptoms of disease to an acute, severe disease. Presently, limited studies ha...

  6. Bovine tuberculosis: Immune response and vaccine efficacy studies

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bovine tuberculosis (TB) is a re-emerging disease of cattle within the United States, primarily due to importation of infected cattle from Mexico and the emergence of a wildlife reservoir (white-tailed deer) in Michigan. While the mainstay of bovine TB control has been abattoir inspection plus targe...

  7. Diagnosis of enzootic bovine leucosis in single and pooled samples.

    PubMed

    Hoff-Jørgensen, R

    1990-12-01

    Diagnosis of enzootic bovine leucosis is based on detection of antibodies against bovine leukemia virus, BLV. Some ELISA modifications have proved sensitive enough for use in the examination of pooled blood samples from slaughterhouses, milk and pooled milk samples. Suggestions for the standardisation of different ELISA modifications using a common reference serum are presented. PMID:1966753

  8. Bovine Viral Diarrhea Virus Modulation of Monocyte Derived Macrophage

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bovine Viral Diarrhea Virus (BVDV) is a single stranded, positive sense virus of the Flaviviridae family and is the causative agent of the disease known as Bovine Viral Diarrhea (BVD). Disease can range from persistently infected (PI) animals displaying no clinical symptoms of disease to an acute, s...

  9. Polymorphisms and haplotype structure of bovine PRND (doppel) and PRNT

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bovine spongiform encephalopathy (BSE) is a fatal neurological disorder characterized by abnormal deposits of a protease-resistant isoform of the prion protein. In previous studies, we have characterized linkage disequilibrium (LD) and haplotype networks within the bovine prion gene (PRNP). Other ...

  10. 21 CFR 184.1034 - Catalase (bovine liver).

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... with 5 U.S.C. 552(a) and 1 CFR part 51. Copies are available from the National Academy Press, 2101... 21 Food and Drugs 3 2011-04-01 2011-04-01 false Catalase (bovine liver). 184.1034 Section 184.1034... Listing of Specific Substances Affirmed as GRAS § 184.1034 Catalase (bovine liver). (a) Catalase...

  11. 9 CFR 113.311 - Bovine Virus Diarrhea Vaccine.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... serum dilution in a varying serum-constant virus neutralization test with less than 500 TCID50 of bovine...-neutralization tests from each of the vaccinates. The test virus shall be less than 500 TCID50 of bovine virus... as prescribed in paragraph (c)(3) of this section, show neutralization in all tubes of the...

  12. 9 CFR 113.311 - Bovine Virus Diarrhea Vaccine.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... serum dilution in a varying serum-constant virus neutralization test with less than 500 TCID50 of bovine...-neutralization tests from each of the vaccinates. The test virus shall be less than 500 TCID50 of bovine virus... as prescribed in paragraph (c)(3) of this section, show neutralization in all tubes of the...

  13. 9 CFR 113.311 - Bovine Virus Diarrhea Vaccine.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... serum dilution in a varying serum-constant virus neutralization test with less than 500 TCID50 of bovine...-neutralization tests from each of the vaccinates. The test virus shall be less than 500 TCID50 of bovine virus... as prescribed in paragraph (c)(3) of this section, show neutralization in all tubes of the...

  14. 9 CFR 113.311 - Bovine Virus Diarrhea Vaccine.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... serum dilution in a varying serum-constant virus neutralization test with less than 500 TCID50 of bovine...-neutralization tests from each of the vaccinates. The test virus shall be less than 500 TCID50 of bovine virus... as prescribed in paragraph (c)(3) of this section, show neutralization in all tubes of the...

  15. 9 CFR 113.311 - Bovine Virus Diarrhea Vaccine.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... serum dilution in a varying serum-constant virus neutralization test with less than 500 TCID50 of bovine...-neutralization tests from each of the vaccinates. The test virus shall be less than 500 TCID50 of bovine virus... as prescribed in paragraph (c)(3) of this section, show neutralization in all tubes of the...

  16. Bovine Papillomavirus Type 13 DNA in Equine Sarcoids

    PubMed Central

    Lunardi, Michele; de Alcântara, Brígida Kussumoto; Otonel, Rodrigo Alejandro Arellano; Rodrigues, Wagner Borges; Alfieri, Alice Fernandes

    2013-01-01

    Equine sarcoids are locally aggressive fibroblastic neoplasms considered to be the most common skin tumors of horses worldwide. Bovine papillomavirus types 1 and 2 have typically been associated with sarcoids in equids. Investigations aiming to identify papillomavirus strains, aside from bovine papillomaviruses 1 and 2, which might be associated with sarcoid lesions, have been lacking. The aim of this article is to report the identification of a third bovine papillomavirus type, bovine papillomavirus 13, associated with equine sarcoids. Six sarcoid lesions were collected from diverse anatomical sites on two horses from southern Brazil. To detect a broad spectrum of papillomavirus strains, eight degenerate primer pairs designed to detect conserved regions on the L1 and E1 genes were tested on the DNA samples. Direct sequencing was then performed on the obtained amplicons, and sequence identities were compared with sequences from all bovine papillomavirus types. The FAP59/FAP64, MY09/MY11, and AR-E1F2/AR-E1R4 sequences generated from the sarcoids were shown to present 99 to 100% identity with bovine papillomavirus 13, a new bovine papillomavirus type previously described in cattle. The results from this study suggest that there is a need to identify bovine papillomavirus type 13 and other papillomavirus strains that might be associated with sarcoids in diverse geographical areas; such investigations might establish the frequency of occurrence of this viral type in these common tumors of equids. PMID:23637294

  17. Comparative molecular analysis of ovine and bovine Streptococcus uberis isolates.

    PubMed

    Gilchrist, T L; Smith, D G E; Fitzpatrick, J L; Zadoks, R N; Fontaine, M C

    2013-02-01

    Streptococcus uberis causes clinical and subclinical mastitis in cattle and sheep, but it is unknown whether the composition of Strep. uberis populations differs between host species. To address this, we characterized a collection of bovine and ovine Strep. uberis isolates with shared geographical and temporal origins by means of an expanded multilocus sequence typing scheme. Among 14 ovine and 35 bovine isolates, 35 allelic profiles were detected. Each allelic profile was associated with a single host species and all but one were new to the multilocus sequence typing database. The median number of new alleles per isolate was higher for ovine isolates than for bovine isolates. None of the ovine isolates belonged to the global clonal complexes 5 or 143, which are commonly associated with bovine mastitis and which have a wide geographical distribution. Ovine isolates also differed from bovine isolates in carriage of plasminogen activator genes, with significantly higher prevalence of pauB in ovine isolates. Isolates that were negative for yqiL, one of the targets of multilocus sequence typing, were found among ovine and bovine isolates and were not associated with a specific sequence type or global clonal complex. One bovine isolate carried a gapC allele that was probably acquired through lateral gene transfer, most likely from Streptococcus salivarius. We conclude that ovine isolates are distinct from bovine isolates of Strep. uberis, and that recombination between isolates from different host species or bacterial species could contribute to changes in virulence gene profiles with relevance for vaccine development. PMID:23200465

  18. Discovery of a Bovine Enterovirus in Alpaca

    PubMed Central

    McClenahan, Shasta D.; Scherba, Gail; Borst, Luke; Fredrickson, Richard L.; Krause, Philip R.; Uhlenhaut, Christine

    2013-01-01

    A cytopathic virus was isolated using Madin-Darby bovine kidney (MDBK) cells from lung tissue of alpaca that died of a severe respiratory infection. To identify the virus, the infected cell culture supernatant was enriched for virus particles and a generic, PCR-based method was used to amplify potential viral sequences. Genomic sequence data of the alpaca isolate was obtained and compared with sequences of known viruses. The new alpaca virus sequence was most similar to recently designated Enterovirus species F, previously bovine enterovirus (BEVs), viruses that are globally prevalent in cattle, although they appear not to cause significant disease. Because bovine enteroviruses have not been previously reported in U.S. alpaca, we suspect that this type of infection is fairly rare, and in this case appeared not to spread beyond the original outbreak. The capsid sequence of the detected virus had greatest homology to Enterovirus F type 1 (indicating that the virus should be considered a member of serotype 1), but the virus had greater homology in 2A protease sequence to type 3, suggesting that it may have been a recombinant. Identifying pathogens that infect a new host species for the first time can be challenging. As the disease in a new host species may be quite different from that in the original or natural host, the pathogen may not be suspected based on the clinical presentation, delaying diagnosis. Although this virus replicated in MDBK cells, existing standard culture and molecular methods could not identify it. In this case, a highly sensitive generic PCR-based pathogen-detection method was used to identify this pathogen. PMID:23950875

  19. Bovine and equine peritubular and intertubular dentin.

    PubMed

    Stock, S R; Deymier-Black, A C; Veis, A; Telser, A; Lux, E; Cai, Z

    2014-09-01

    Dentin contains 1-2μm diameter tubules extending from the pulp cavity to near the junction with enamel. Peritubular dentin (PTD) borders the tubule lumens and is surrounded by intertubular dentin (ITD). Differences in PTD and ITD composition and microstructure remain poorly understood. Here, a (∼200nm)(2), 10.1keV synchrotron X-ray beam maps X-ray fluorescence and X-ray diffraction simultaneously around tubules in 15-30μm thick bovine and equine specimens. Increased Ca fluorescence surrounding tubule lumens confirms that PTD is present, and the relative intensities in PTD and ITD correspond to carbonated apatite (cAp) volume fraction of ∼0.8 in PTD vs. 0.65 assumed for ITD. In the PTD near the lumen edges, Zn intensity is strongly peaked, corresponding to a Zn content of ∼0.9mgg(-1) for an assumed concentration of ∼0.4mgg(-1) for ITD. In the equine specimen, the Zn K-edge position indicates that Zn(2+) is present, similar to bovine dentin (Deymier-Black et al., 2013), and the above edge structure is consistent with spectra from macromolecules related to biomineralization. Transmission X-ray diffraction shows only cAp, and the 00.2 diffraction peak (Miller-Bravais indices) width is constant from ITD to the lumen edge. The cAp 00.2 average preferred orientation is axisymmetric (about the tubule axis) in both bovine and equine dentin, and the axisymmetric preferred orientation continues from ITD through the PTD to the tubule lumen. These data indicate that cAp structure does not vary from PTD to ITD. PMID:24911530

  20. Rosette nanotubes inhibit bovine neutrophil chemotaxis

    PubMed Central

    Le, Minh Hong Anh; Suri, Sarabjeet Singh; Rakotondradany, Felaniaina; Fenniri, Hicham; Singh, Baljit

    2010-01-01

    Migration of activated neutrophils that have prolonged lifespan into inflamed organs is an important component of host defense but also contributes to tissue damage and mortality. In this report, we used biologically-inspired RGD-tagged rosette nanotubes (RNT) to inhibit neutrophil chemotaxis. We hypothesize that RGD-RNT will block neutrophil migration through inhibition of MAPK. In this report, RNT conjugated to lysine (K–RNT) and arginine-glycine-aspartic acid-serine-lysine (RGDSK-RNT) were co-assembled in a molar ratio of 95/5. The effect of the resulting composite RNT (RGDSK/K–RNT) on neutrophil chemotaxis, cell signaling and apoptosis was then investigated. Exposure to RGDSK/K–RNT reduced bovine neutrophil migration when compared to the non-treated group (p < 0.001). Similar effect was seen following treatment with ERK1/2 or p38 MAPK inhibitors. Phosphorylation of the ERK1/2 and p38 MAPK was inhibited at 5 min by RGDSK/K–RNT (p < 0.05). The RGDSD/K-RNT did not affect the migration of neutrophils pre-treated with αvβ3 integrin antibody suggesting that both bind to the same receptor. RGDSK/K–RNT did not induce apoptosis in bovine neutrophils, which was suppressed by pre-exposing them to LPS (p < 0.001). We conclude that RGDSK/K–RNT inhibit phosphorylation of ERK1/2 and p38 MAPK and inhibit chemotaxis of bovine neutrophils. PMID:20663476

  1. Bovine neonatal pancytopenia in calves in Poland.

    PubMed

    Jaśkowski, J M; Polak, M P; Marszałek, A; Nowak, K; Frackowiak, H; Olechnowicz, J

    2012-01-01

    Bovine neonatal pancytopenia (BNP), a newly emerged syndrome of discussed etiology in calves, has been diagnosed since 2006. Here we describe first cases of BNP in Poland. Between September 2008 and April 2011, 62 cases of BNP were diagnosed in dairy calves. Bleeding skin lesions were mostly pronounced in summer and early autumn. Severe thrombocytopenia was observed in all sick animals. All calves came from herds vaccinated against BVDV infection with PregSure BVD vaccine (Pfizer). Substitution of colostrum from dams of BNP positive calves with colostrum from dams from herds free of BNP was the only effective measure to avoid new cases in affected herds. PMID:23214380

  2. Bovine viral diarrhea virus: global status.

    PubMed

    Ridpath, Julia F

    2010-03-01

    Despite the success of regional bovine viral diarrhea viruses (BVDV) eradication programs, infections remain a source of economic loss for producers. The wide variation among BVDV results in differences in genotype, biotype, virulence, and types of infections. BVDV infect a range of domestic and wild ruminants. Clinical presentation varies depending on strain of virus, species of host, immune status of host, reproductive status of host, age of host, and concurrent infections. Recent advances in BVDV research and diagnostics have led to the development of regional eradication/control programs, the most efficacious of which focus on biosecurity, surveillance, and control. PMID:20117546

  3. Transcriptional organization of bovine papillomavirus type 1.

    PubMed

    Engel, L W; Heilman, C A; Howley, P M

    1983-09-01

    Multiple bovine papillomavirus type 1 (BPV-1)-specific polyadenylated RNA species in a BPV-1-infected bovine fibropapilloma were identified and mapped. All of the RNA species were transcribed from the same DNA strand of the BPV-1 genome. Five RNA species previously identified in BPV-1-transformed mouse cells were also present in the bovine fibropapilloma. These five species measured 1,050, 1,150, 1,700, 3,800, and 4,050 bases, mapped within the 69% transforming segment of the BPV-1 genome, and shared a 3' coterminus at 0.53 map units (m.u.). The 5' ends of the bodies of these distinct transcripts were located at ca. 0.03, 0.09, 0.34, 0.39, and 0.41 m.u. Additional polyadenylated RNA species not present in BPV-1-transformed mouse cells were specific for the BPV-1-infected bovine fibropapilloma and measured 1,700, 3,700, 3,800, 6,700, and 8,000 bases. These wart-specific species shared a 3' coterminus at 0.90 m.u. The 5' termini of the bodies of the 1,700- and 3,800-base species mapped at 0.71 and 0.42 m.u., respectively. Exonuclease VII analysis failed to reveal any internal splicing in these two species; however, the presence of small remote 5' leader sequences could not be ruled out. The 3,700-base species hybridized to DNA fragments from the 69% transforming segment as well as from the 31% nontransforming segment of the BPV-1 genome; however, this species was not precisely mapped. The 5' termini of the two largest RNA species (6,700 and 8,000 bases in size) were located at ca. 0.01 and 0.90 m.u., respectively. Since the 5' ends of these mapped adjacent to a TATAAA sequence which could possibly serve as an element of a transcriptional promoter, it is possible that one or both of these species represent nonspliced precursor RNA molecules. PMID:6137574

  4. Evaluation of power of the Illumina HumanOmni5M-4v1 BeadChip to detect risk variants for human complex diseases.

    PubMed

    Xing, Chuanhua; Huang, Jie; Hsu, Yi-Hsiang; DeStefano, Anita L; Heard-Costa, Nancy L; Wolf, Philip A; Seshadri, Sudha; Kiel, Douglas P; Cupples, L Adrienne; Dupuis, Josée

    2016-07-01

    Although emerging sequencing technologies can characterize all genetic variants, the cost is still high. Illumina released the HumanOmni5M-4v1 (Omni5) genotype array with ~4.3M assayed SNPs, a much denser array compared with other available arrays. The Omni5 balances both cost and array density. In this article, we illustrate the power of Omni5 to detect genetic associations. The Omni5 includes variants with a wide range of minor allele frequencies down to <1%. The theoretical power calculation examples indicate the increased power of the Omni5 array compared with other arrays with lower density when evaluating associations with some known loci, although there are exceptions. We further evaluate the genetic associations between known loci and several quantitative traits in the Framingham Heart Study: femoral neck bone mineral density, lumbar spine bone mineral density and hippocampal volume. Finally, we search genome wide for novel associations using the Omni5 genotypes. We compare our association results from Affymetrix 500K+MIPS 50K arrays and two imputed data sets: (1) HapMap Phase II and (2) 1000 Genomes reference panel. We observed increased evidence for genotype-phenotype associations with smaller P-values for selected known loci using the Omni5 genotypes. With limited sample sizes, we identify novel variants with genome-wide significant P-values. Our observations support the notion that dense genotyping using the Omni5 can be powerful in detecting novel associated variants. Comparison with imputed data with higher density also suggests that imputation helps but cannot replace genotyping, especially when imputation quality is low. PMID:26577045

  5. High-Resolution Genotyping of Wild Barley Introgression Lines and Fine-Mapping of the Threshability Locus thresh-1 Using the Illumina GoldenGate Assay

    PubMed Central

    Schmalenbach, Inga; March, Timothy J.; Bringezu, Thomas; Waugh, Robbie; Pillen, Klaus

    2011-01-01

    Genetically well-characterized mapping populations are a key tool for rapid and precise localization of quantitative trait loci (QTL) and subsequent identification of the underlying genes. In this study, a set of 73 introgression lines (S42ILs) originating from a cross between the spring barley cultivar Scarlett (Hordeum vulgare ssp. vulgare) and the wild barley accession ISR42-8 (H. v. ssp. spontaneum) was subjected to high-resolution genotyping with an Illumina 1536-SNP array. The array enabled a precise localization of the wild barley introgressions in the elite barley background. Based on 636 informative SNPs, the S42IL set represents 87.3% of the wild barley genome, where each line contains on average 3.3% of the donor genome. Furthermore, segregating high-resolution mapping populations (S42IL-HRs) were developed for 70 S42ILs in order to facilitate QTL fine-mapping and cloning. As a case study, we used the developed genetic resources to rapidly identify and fine-map the novel locus thresh-1 on chromosome 1H that controls grain threshability. Here, the recessive wild barley allele confers a difficult to thresh phenotype, suggesting that thresh-1 played an important role during barley domestication. Using a S42IL-HR population, thresh-1 was fine-mapped within a 4.3cM interval that was predicted to contain candidate genes involved in regulation of plant cell wall composition. The set of wild barley introgression lines and derived high-resolution populations are ideal tools to speed up the process of mapping and further dissecting QTL, which ultimately clears the way for isolating the genes behind QTL effects. PMID:22384330

  6. Assessment of a 16S rRNA amplicon Illumina sequencing procedure for studying the microbiome of a symbiont-rich aphid genus.

    PubMed

    Jousselin, E; Clamens, A-L; Galan, M; Bernard, M; Maman, S; Gschloessl, B; Duport, G; Meseguer, A S; Calevro, F; Coeur d'acier, A

    2016-05-01

    The bacterial communities inhabiting arthropods are generally dominated by a few endosymbionts that play an important role in the ecology of their hosts. Rather than comparing bacterial species richness across samples, ecological studies on arthropod endosymbionts often seek to identify the main bacterial strains associated with each specimen studied. The filtering out of contaminants from the results and the accurate taxonomic assignment of sequences are therefore crucial in arthropod microbiome studies. We aimed here to validate an Illumina 16S rRNA gene sequencing protocol and analytical pipeline for investigating endosymbiotic bacteria associated with aphids. Using replicate DNA samples from 12 species (Aphididae: Lachninae, Cinara) and several controls, we removed individual sequences not meeting a minimum threshold number of reads in each sample and carried out taxonomic assignment for the remaining sequences. With this approach, we show that (i) contaminants accounted for a negligible proportion of the bacteria identified in our samples; (ii) the taxonomic composition of our samples and the relative abundance of reads assigned to a taxon were very similar across PCR and DNA replicates for each aphid sample; in particular, bacterial DNA concentration had no impact on the results. Furthermore, by analysing the distribution of unique sequences across samples rather than aggregating them into operational taxonomic units (OTUs), we gained insight into the specificity of endosymbionts for their hosts. Our results confirm that Serratia symbiotica is often present in Cinara species, in addition to the primary symbiont, Buchnera aphidicola. Furthermore, our findings reveal new symbiotic associations with Erwinia- and Sodalis-related bacteria. We conclude with suggestions for generating and analysing 16S rRNA gene sequences for arthropod-endosymbiont studies. PMID:26458227

  7. Illumina MiSeq Sequencing Reveals Diverse Microbial Communities of Activated Sludge Systems Stimulated by Different Aromatics for Indigo Biosynthesis from Indole

    PubMed Central

    Zhang, Xuwang; Qu, Yuanyuan; Ma, Qiao; Zhang, Zhaojing; Li, Duanxing; Wang, Jingwei; Shen, Wenli; Shen, E; Zhou, Jiti

    2015-01-01

    Indole, as a typical N-heteroaromatic compound existed in coking wastewater, can be used for bio-indigo production. The microbial production of indigo from indole has been widely reported during the last decades using culture-dependent methods, but few studies have been carried out by microbial communities. Herein, three activated sludge systems stimulated by different aromatics, i.e. naphthalene plus indole (G1), phenol plus indole (G2) and indole only (G3), were constructed for indigo production from indole. During the operation, G1 produced the highest indigo yield in the early stage, but it switched to G3 in the late stage. Based on LC-MS analysis, indigo was the major product in G1 and G3, while the purple product 2-(7-oxo-1H-indol-6(7H)-ylidene) indolin-3-one was dominant in G2. Illumina MiSeq sequencing of 16S rRNA gene amplicons was applied to analyze the microbial community structure and composition. Detrended correspondence analysis (DCA) and dissimilarity tests showed that the overall community structures of three groups changed significantly during the operation (P<0.05). Nevertheless, the bacteria assigned to phylum Proteobacteria, family Comamonadaceae, and genera Diaphorobacter, Comamonas and Aquamicrobium were commonly shared dominant populations. Pearson correlations were calculated to discern the relationship between microbial communities and indigo yields. The typical indigo-producing populations Comamonas and Pseudomonas showed no positive correlations with indigo yields, while there emerged many other genera that exhibited positive relationships, such as Aquamicrobium, Truepera and Pusillimonas, which had not been reported for indigo production previously. The present study should provide new insights into indigo bio-production by microbial communities from indole. PMID:25928424

  8. Development and assessment of an optimized next-generation DNA sequencing approach for the mtgenome using the Illumina MiSeq

    PubMed Central

    McElhoe, Jennifer A.; Holland, Mitchell M.; Makova, Kateryna D.; Su, Marcia Shu-Wei; Paul, Ian M.; Baker, Christine H.; Faith, Seth A.; Young, Brian

    2015-01-01

    The development of molecular tools to detect and report mitochondrial DNA (mtDNA) heteroplasmy will increase the discrimination potential of the testing method when applied to forensic cases. The inherent limitations of the current state-of-the-art, Sanger-based sequencing, including constrictions in speed, throughput, and resolution, have hindered progress in this area. With the advent of next-generation sequencing (NGS) approaches, it is now possible to clearly identify heteroplasmic variants, and at a much lower level than previously possible. However, in order to bring these approaches into forensic laboratories and subsequently as accepted scientific information in a court of law, validated methods will be required to produce and analyze NGS data. We report here on the development of an optimized approach to NGS analysis for the mtDNA genome (mtgenome) using the Illumina MiSeq instrument. This optimized protocol allows for the production of more than 5 gigabases of mtDNA sequence per run, sufficient for detection and reliable reporting of minor heteroplasmic variants down to approximately 0.5–1.0% when multiplexing twelve samples. Depending on sample throughput needs, sequence coverage rates can be set at various levels, but were optimized here for at least 5,000 reads. In addition, analysis parameters are provided for a commercially available software package that identify the highest quality sequencing reads and effectively filter out sequencing-based noise. With this method it will be possible to measure the rates of low-level heteroplasmy across the mtgenome, evaluate the transmission of heteroplasmy between the generations of maternal lineages, and assess the drift of variant sequences between different tissue types within an individual. PMID:25051226

  9. De novo assembly and characterization of global transcriptome of coconut palm (Cocos nucifera L.) embryogenic calli using Illumina paired-end sequencing.

    PubMed

    Rajesh, M K; Fayas, T P; Naganeeswaran, S; Rachana, K E; Bhavyashree, U; Sajini, K K; Karun, Anitha

    2016-05-01

    Production and supply of quality planting material is significant to coconut cultivation but is one of the major constraints in coconut productivity. Rapid multiplication of coconut through in vitro techniques, therefore, is of paramount importance. Although somatic embryogenesis in coconut is a promising technique that will allow for the mass production of high quality palms, coconut is highly recalcitrant to in vitro culture. In order to overcome the bottlenecks in coconut somatic embryogenesis and to develop a repeatable protocol, it is imperative to understand, identify, and characterize molecular events involved in coconut somatic embryogenesis pathway. Transcriptome analysis (RNA-Seq) of coconut embryogenic calli, derived from plumular explants of West Coast Tall cultivar, was undertaken on an Illumina HiSeq 2000 platform. After de novo transcriptome assembly and functional annotation, we have obtained 40,367 transcripts which showed significant BLASTx matches with similarity greater than 40 % and E value of ≤10(-5). Fourteen genes known to be involved in somatic embryogenesis were identified. Quantitative real-time PCR (qRT-PCR) analyses of these 14 genes were carried in six developmental stages. The result showed that CLV was upregulated in the initial stage of callogenesis. Transcripts GLP, GST, PKL, WUS, and WRKY were expressed more in somatic embryo stage. The expression of SERK, MAPK, AP2, SAUR, ECP, AGP, LEA, and ANT were higher in the embryogenic callus stage compared to initial culture and somatic embryo stages. This study provides the first insights into the gene expression patterns during somatic embryogenesis in coconut. PMID:26210639

  10. Concordance and reproducibility of a next generation mtGenome sequencing method for high-quality samples using the Illumina MiSeq.

    PubMed

    Peck, Michelle A; Brandhagen, Michael D; Marshall, Charla; Diegoli, Toni M; Irwin, Jodi A; Sturk-Andreaggi, Kimberly

    2016-09-01

    Sanger-type sequencing (STS) of mitochondrial DNA (mtDNA), specifically the control region (CR), is routinely employed in forensics in human identification and missing persons scenarios. Yet next-generation sequencing (NGS) has the potential to overcome some of the major limitations of STS processing, permitting reasonable paths forward for full mitochondrial genome (mtGenome) sequencing, while also offering higher-throughput and higher sensitivity capabilities. To establish the accuracy and reproducibility of NGS for the development of mtDNA data, 90 DNA extracts that were previously used to generate forensic quality full mtGenomes using STS were sequenced using Nextera XT library preparation and the Illumina MiSeq. Using the same amplicon product, replicate library sets were generated and sequenced at different laboratories, and analysis was performed in replicate using the CLC Genomics Workbench. Both sequencing sets resulted in 99.998% of positions with greater than 10X coverage when 96 samples (including controls) were multiplexed. Overall, 99.9996% concordance was observed between the NGS data and the STS data for the full mtGenome. The only "discordant" calls involved low level point heteroplasmies, with the differences resulting from stochastic variation and/or the increased sensitivity of NGS. Higher sensitivity also allowed for the detection of a mixed sample previously not detected with STS. Additionally, variant calls were reproducible between sequencing sets and between software analysis versions with the variant frequency only differing by 0.23% and 0.01%, respectively. Further validation studies and specialized software functionality tailored to forensic practice should facilitate the incorporation of NGS processing into standard casework applications. The data herein comprise the largest, and likely most thoroughly examined, complete mtGenome STS-NGS concordance dataset available. PMID:27368088

  11. Illumina-based analysis of the rhizosphere microbial communities associated with healthy and wilted Lanzhou lily (Lilium davidii var. unicolor) plants grown in the field.

    PubMed

    Shang, Qianhan; Yang, Guo; Wang, Yun; Wu, Xiukun; Zhao, Xia; Hao, Haiting; Li, Yuyao; Xie, Zhongkui; Zhang, Yubao; Wang, Ruoyu

    2016-06-01

    Lanzhou lily (Liliumdavidii var. unicolor) is the best edible lily as well as a traditional medicinal plant in China. The microbes associated with plant roots play crucial roles in plant growth and health. However, little is known about the differences of rhizosphere microbes between healthy and wilted Lanzhou lily (Lilium davidii var. unicolor) plants. The objective of this study was to compare the rhizosphere microbial community and functional diversity of healthy and wilted plants, and to identify potential biocontrol agents with significant effect. Paired end Illumina Mi-Seq sequencing of 16S rRNA and ITS gene amplicons was employed to study the bacterial and fungal communities in the rhizosphere soil of Lanzhou lily plants. BIOLOG technology was adopted to investigate the microbial functional diversity. Our results indicated that there were major differences in the rhizosphere microbial composition and functional diversity of wilted samples compared with healthy samples. Healthy Lanzhou lily plants exhibited lower rhizosphere-associated bacterial diversity than diseased plants, whereas fungi exhibited the opposite trend. The dominant phyla in both the healthy and wilted samples were Proteobacteria and Ascomycota, i.e., 34.45 and 64.01 %, respectively. The microbial functional diversity was suppressed in wilted soil samples. Besides Fusarium, the higher relative abundances of Rhizoctonia, Verticillium, Penicillium, and Ilyonectria (Neonectria) in the wilted samples suggest they may pathogenetic root rot fungi. The high relative abundances of Bacillus in Firmicutes in healthy samples may have significant roles as biological control agents against soilborne pathogens. This is the first study to find evidence of major differences between the microbial communities in the rhizospheric soil of healthy and wilted Lanzhou lily, which may be linked to the health status of plants. PMID:27116961

  12. Characterization of Intestinal Microbiomes of Hirschsprung's Disease Patients with or without Enterocolitis Using Illumina-MiSeq High-Throughput Sequencing.

    PubMed

    Li, Yuqing; Poroyko, Valeriy; Yan, Zhilong; Pan, Liya; Feng, Yi; Zhao, Peihua; Xie, Zhoulonglong; Hong, Li

    2016-01-01

    Hirschsprung-associated enterocolitis (HAEC) is a life-threatening complication of Hirschsprung's disease (HD). Although the pathological mechanisms are still unclear, studies have shown that HAEC has a close relationship with the disturbance of intestinal microbiota. This study aimed to investigate the characteristics of the intestinal microbiome of HD patients with or without enterocolitis. During routine or emergency surgery, we collected 35 intestinal content samples from five patients with HAEC and eight HD patients, including three HD patients with a history of enterocolitis who were in a HAEC remission (HAEC-R) phase. Using Illumina-MiSeq high-throughput sequencing, we sequenced the V4 region of bacterial 16S rRNA, and operational taxonomic units (OTUs) were defined by 97% sequence similarity. Principal coordinate analysis (PCoA) of weighted UniFrac distances was performed to evaluate the diversity of each intestinal microbiome sample. The microbiota differed significantly between the HD patients (characterized by the prevalence of Bacteroidetes) and HAEC patients (characterized by the prevalence of Proteobacteria), while the microbiota of the HAEC-R patients was more similar to that of the HAEC patients. We also observed that the specimens from different intestinal sites of each HD patient differed significantly, while the specimens from different intestinal sites of each HAEC and HAEC-R patient were more similar. In conclusion, the microbiome pattern of the HAEC-R patients was more similar to that of the HAEC patients than to that of the HD patients. The HD patients had a relatively distinct, more stable community than the HAEC and HAEC-R patients, suggesting that enterocolitis may either be caused by or result in a disruption of the patient's uniquely adapted intestinal flora. The intestinal microbiota associated with enterocolitis may persist following symptom resolution and can be implicated in the symptom recurrence. PMID:27603009

  13. The first metagenome of activated sludge from full-scale anaerobic/anoxic/oxic (A2O) nitrogen and phosphorus removal reactor using Illumina sequencing.

    PubMed

    Tian, Mei; Zhao, Fangqing; Shen, Xin; Chu, Kahou; Wang, Jinfeng; Chen, Shuai; Guo, Yan; Liu, Hanhu

    2015-09-01

    The anaerobic/anoxic/oxic (A2O) process is globally one of the widely used biological sewage treatment processes. This is the first report of a metagenomic analysis using Illumina sequencing of full-scale A2O sludge from a municipal sewage treatment plant. With more than 530,000 clean reads from different taxa and metabolic categories, the metagenome results allow us to gain insight into the functioning of the biological community of the A2O sludge. There are 51 phyla and nearly 900 genera identified from the A2O activated sludge ecosystem. Proteobacteria, Bacteroidetes, Nitrospirae and Chloroflexi are predominant phyla in the activated sludge, suggesting that these organisms play key roles in the biodegradation processes in the A2O sewage treatment system. Nitrospira, Thauera, Dechloromonas and Ignavibacterium, which have abilities to metabolize nitrogen and aromatic compounds, are most prevalent genera. The percent of nitrogen and phosphorus metabolism in the A2O sludge is 2.72% and 1.48%, respectively. In the current A2O sludge, the proportion of Candidatus Accumulibacter is 1.37%, which is several times more than that reported in a recent study of A2O sludge. Among the four processes of nitrogen metabolism, denitrification related genes had the highest number of sequences (76.74%), followed by ammonification (15.77%), nitrogen fixation (3.88%) and nitrification (3.61%). In phylum Planctomycetes, four genera (Planctomyces, Pirellula, Gemmata and Singulisphaera) are included in the top 30 abundant genera, suggesting the key role of ANAMMOX in nitrogen metabolism in the A2O sludge. PMID:26354707

  14. Markers on Bovine Chromosome 20 Associated with Carcass Quality and Composition Traits and Incidence of Contracting Infectious Bovine Keratoconjunctivitis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The objective of this study was to evaluate single nucleotide polymorphisms (SNP) on bovine chromosome 20 to fine map a previously identified QTL associated with the incidence of infectious bovine keratoconjunctivitis (IBK). Crossbred steers (GPE7; n = 539) derived from sires of 7 Bos taurus breeds...

  15. Markers on bovine chromosome 20 associated with carcass quality and composition traits and incidence of contracting infectious bovine keratoconjunctivitis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The objective of this study was to use single nucleotide polymorphisms (SNP) on bovine chromosome 20 to fine map a previously identified QTL associated with the incidence of infectious bovine keratoconjunctivitis (IBK). Crossbred steers (GPE7; n = 539) derived from sires of 7 Bos Taurus breeds and h...

  16. Single Nucleotide Polymorphism Discovery in Bovine Pituitary Gland Using RNA-Seq Technology.

    PubMed

    Pareek, Chandra Shekhar; Smoczyński, Rafał; Kadarmideen, Haja N; Dziuba, Piotr; Błaszczyk, Paweł; Sikora, Marcin; Walendzik, Paulina; Grzybowski, Tomasz; Pierzchała, Mariusz; Horbańczuk, Jarosław; Szostak, Agnieszka; Ogluszka, Magdalena; Zwierzchowski, Lech; Czarnik, Urszula; Fraser, Leyland; Sobiech, Przemysław; Wąsowicz, Krzysztof; Gelfand, Brian; Feng, Yaping; Kumar, Dibyendu

    2016-01-01

    Examination of bovine pituitary gland transcriptome by strand-specific RNA-seq allows detection of putative single nucleotide polymorphisms (SNPs) within potential candidate genes (CGs) or QTLs regions as well as to understand the genomics variations that contribute to economic trait. Here we report a breed-specific model to successfully perform the detection of SNPs in the pituitary gland of young growing bulls representing Polish Holstein-Friesian (HF), Polish Red, and Hereford breeds at three developmental ages viz., six months, nine months, and twelve months. A total of 18 bovine pituitary gland polyA transcriptome libraries were prepared and sequenced using the Illumina NextSeq 500 platform. Sequenced FastQ databases of all 18 young bulls were submitted to NCBI-SRA database with NCBI-SRA accession numbers SRS1296732. For the investigated young bulls, a total of 113,882,3098 raw paired-end reads with a length of 156 bases were obtained, resulting in an approximately 63 million paired-end reads per library. Breed-wise, a total of 515.38, 215.39, and 408.04 million paired-end reads were obtained for Polish HF, Polish Red, and Hereford breeds, respectively. Burrows-Wheeler Aligner (BWA) read alignments showed 93.04%, 94.39%, and 83.46% of the mapped sequencing reads were properly paired to the Polish HF, Polish Red, and Hereford breeds, respectively. Constructed breed-specific SNP-db of three cattle breeds yielded at 13,775,885 SNPs. On an average 765,326 breed-specific SNPs per young bull were identified. Using two stringent filtering parameters, i.e., a minimum 10 SNP reads per base with an accuracy ≥ 90% and a minimum 10 SNP reads per base with an accuracy = 100%, SNP-db records were trimmed to construct a highly reliable SNP-db. This resulted in a reduction of 95,7% and 96,4% cut-off mark of constructed raw SNP-db. Finally, SNP discoveries using RNA-Seq data were validated by KASP™ SNP genotyping assay. The comprehensive QTLs/CGs analysis of 76 QTLs

  17. Recovery of Native Genetic Background in Admixed Populations Using Haplotypes, Phenotypes, and Pedigree Information – Using Cika Cattle as a Case Breed

    PubMed Central

    Simčič, Mojca; Smetko, Anamarija; Sölkner, Johann; Seichter, Doris; Gorjanc, Gregor; Kompan, Dragomir; Medugorac, Ivica

    2015-01-01

    The aim of this study was to obtain unbiased estimates of the diversity parameters, the population history, and the degree of admixture in Cika cattle which represents the local admixed breeds at risk of extinction undergoing challenging conservation programs. Genetic analyses were performed on the genome-wide Single Nucleotide Polymorphism (SNP) Illumina Bovine SNP50 array data of 76 Cika animals and 531 animals from 14 reference populations. To obtain unbiased estimates we used short haplotypes spanning four markers instead of single SNPs to avoid an ascertainment bias of the BovineSNP50 array. Genome-wide haplotypes combined with partial pedigree and type trait classification show the potential to improve identification of purebred animals with a low degree of admixture. Phylogenetic analyses demonstrated unique genetic identity of Cika animals. Genetic distance matrix presented by rooted Neighbour-Net suggested long and broad phylogenetic connection between Cika and Pinzgauer. Unsupervised clustering performed by the admixture analysis and two-dimensional presentation of the genetic distances between individuals also suggest Cika is a distinct breed despite being similar in appearance to Pinzgauer. Animals identified as the most purebred could be used as a nucleus for a recovery of the native genetic background in the current admixed population. The results show that local well-adapted strains, which have never been intensively managed and differentiated into specific breeds, exhibit large haplotype diversity. They suggest a conservation and recovery approach that does not rely exclusively on the search for the original native genetic background but rather on the identification and removal of common introgressed haplotypes would be more powerful. Successful implementation of such an approach should be based on combining phenotype, pedigree, and genome-wide haplotype data of the breed of interest and a spectrum of reference breeds which potentially have had

  18. Recovery of native genetic background in admixed populations using haplotypes, phenotypes, and pedigree information--using Cika cattle as a case breed.

    PubMed

    Simčič, Mojca; Smetko, Anamarija; Sölkner, Johann; Seichter, Doris; Gorjanc, Gregor; Kompan, Dragomir; Medugorac, Ivica

    2015-01-01

    The aim of this study was to obtain unbiased estimates of the diversity parameters, the population history, and the degree of admixture in Cika cattle which represents the local admixed breeds at risk of extinction undergoing challenging conservation programs. Genetic analyses were performed on the genome-wide Single Nucleotide Polymorphism (SNP) Illumina Bovine SNP50 array data of 76 Cika animals and 531 animals from 14 reference populations. To obtain unbiased estimates we used short haplotypes spanning four markers instead of single SNPs to avoid an ascertainment bias of the BovineSNP50 array. Genome-wide haplotypes combined with partial pedigree and type trait classification show the potential to improve identification of purebred animals with a low degree of admixture. Phylogenetic analyses demonstrated unique genetic identity of Cika animals. Genetic distance matrix presented by rooted Neighbour-Net suggested long and broad phylogenetic connection between Cika and Pinzgauer. Unsupervised clustering performed by the admixture analysis and two-dimensional presentation of the genetic distances between individuals also suggest Cika is a distinct breed despite being similar in appearance to Pinzgauer. Animals identified as the most purebred could be used as a nucleus for a recovery of the native genetic background in the current admixed population. The results show that local well-adapted strains, which have never been intensively managed and differentiated into specific breeds, exhibit large haplotype diversity. They suggest a conservation and recovery approach that does not rely exclusively on the search for the original native genetic background but rather on the identification and removal of common introgressed haplotypes would be more powerful. Successful implementation of such an approach should be based on combining phenotype, pedigree, and genome-wide haplotype data of the breed of interest and a spectrum of reference breeds which potentially have had

  19. Toxicology of a bovine paraplegic syndrome.

    PubMed

    Sevcik, C; Brito, J C; D'Suze, G; Domínguez-Bello, M G; Lovera, M; Mijares, A J; Bónoli, S

    1993-12-01

    A clinical entity named 'bovine paraplegic syndrome' ('síndrome parapléjico de los bovinos') has spread alarmingly in the cattle-growing areas of the central and eastern plains of Venezuela. It is estimated that four million cattle are bred in the area where the disease occurs. The mortality ranges from 5 to 25% of the animals at risk, mostly pregnant or lactating cows. The principal characteristic of the bovine paraplegic syndrome is ventral or sternal decubitus, in animals that make vain efforts to stand when stimulated. The diagnosis is established when all other possible causes (e.g. paralytic rabies, botulism and blood parasites such as Anaplasma marginal, Babesia bovis, B. bigemina, and Trypanosoma vivax) have been ruled out clinically and by laboratory tests. Death always occurs, usually after a few days, and there is no known treatment. In this work, we describe results that show the presence of a toxin in the cattle suffering from, or liable to suffer from the syndrome. The toxin is produced by ruminal bacteria. In squid giant axons under voltage clamp conditions, the toxin blocks the sodium current. We detected the toxin analytically by absorbance measurements at 340 nm after reacting with picrylsulfonic acid. We obtained a good separation of the toxin with isocratic high pressure liquid chromatography, using 40% methanol in water on phenylborasil columns. PMID:8146871

  20. [Serological study of bovine leptospirosis in Mexico].

    PubMed

    Moles Cervantes, Luis Pedro; Cisneros Puebla, Miguel Angel; Rosas, Dolores Gavaldón; Serranía, Nora Rojas; Torres Barranca, Jorge Isaac

    2002-01-01

    The results of 4 043 bovine sera samples from various Mexican regions, which were sent to a diagnosis lab, were analyzed. The method was the agglutination technique, taking the dilution rate 1:1000 or higher as positive. The analysis revealed 31,1% of seroprevalence and the most frequent serovarietes were hardjo (strain H 89 isolated in Mexico), wolffi and tarassovi. There is coincidence with early data obtained in Mexico on a 34% of prevalence found in a similar study performed in 1994, and with the scientific literature from other countries. The former study also indicated that tarassovi and wolffi were the most common leptospira, so there is coincidence with the figures in the reviewed literature. It was concluded that there was no significant variation in the prevalence rate between the 1994 study and the present one; therefore, it is recommended that this study be promoted so as to increase the bovine vaccination and achieve a reduction in leptospirosis in Mexico. PMID:15846936

  1. Tensile strength of bovine trabecular bone.

    PubMed

    Kaplan, S J; Hayes, W C; Stone, J L; Beaupré, G S

    1985-01-01

    Data on the tensile and compressive properties of trabecular bone are needed to define input parameters and failure criteria for modeling total joint replacements. To help resolve differences in reports comparing tensile and compressive properties of trabecular bone, we have developed new methods, based on porous foam technology, for tensile testing of fresh/frozen trabecular bone specimens. Using bovine trabecular bone from an isotropic region from the proximal humerus as a model material, we measured ultimate strengths in tension and compression for two groups of 24 specimens each. The average ultimate strength in tension was 7.6 +/- 2.2 (95% C.I.) MPa and in compression was 12.4 +/- 3.2 MPa. This difference was statistically significant (p = 0.013) and was not related to density differences between the test groups (p = 0.28). Strength was related by a power-law function of the local apparent density, but, even accounting for density influences, isotropic bovine trabecular bone exhibits significantly lower strengths in tension than in compression. PMID:4077868

  2. Affinity purification of aprotinin from bovine lung.

    PubMed

    Xin, Yu; Liu, Lanhua; Chen, Beizhan; Zhang, Ling; Tong, Yanjun

    2015-05-01

    An affinity protocol for the purification of aprotinin from bovine lung was developed. To simulate the structure of sucrose octasulfate, a natural specific probe for aprotinin, the affinity ligand was composed of an acidic head and a hydrophobic stick, and was then linked with Sepharose. The sorbent was then subjected to adsorption analysis with pure aprotinin. The purification process consisted of one step of affinity chromatography and another step of ultrafiltration. Then purified aprotinin was subjected to sodium dodecyl sulfate polyacrylamide gel electrophoresis, trypsin inhibitor activity, gel-filtration, and thin-layer chromatography analysis. As calculated, the theoretical maximum adsorption (Qmax ) of the affinity sorbent was 25,476.0 ± 184.8 kallikrein inactivator unit/g wet gel; the dissociation constant of the complex "immobilized ligand-aprotinin" (Kd ) was 4.6 ± 0.1 kallikrein inactivator unit/mL. After the affinity separation of bovine lung aprotinin, reducing sodium dodecyl sulfate polyacrylamide gel electrophoresis analysis and gel-filtration chromatography revealed that the protein was a single polypeptide, and the purities were ∼ 97 and 100%, respectively; the purified peptide was also confirmed with aprotinin standard by gel-filtration chromatography and thin-layer chromatography. After the whole purification process, protein, and bioactivity recoveries were 2.2 and 92.6%, respectively; and the specific activity was up to 15,907.1 ± 10.2 kallikrein inactivator unit/mg. PMID:25677462

  3. Epidemiological Modeling of Bovine Brucellosis in India

    PubMed Central

    Kang, Gloria J.; Gunaseelan, L.; Abbas, Kaja M.

    2015-01-01

    The study objective is to develop an epidemiological model of brucellosis transmission dynamics among cattle in India and to estimate the impact of different prevention and control strategies. The prevention and control strategies are test-and-slaughter, transmission rate reduction, and mass vaccination. We developed a mathematical model based on the susceptible-infectious-recovered epidemic model to simulate brucellosis transmission dynamics, calibrated to the endemically stable levels of bovine brucellosis prevalence of cattle in India. We analyzed the epidemiological benefit of different rates of reduced transmission and vaccination. Test-and-slaughter is an effective strategy for elimination and eradication of brucellosis, but socio-cultural constraints forbid culling of cattle in India. Reducing transmission rates lowered the endemically stable levels of brucellosis prevalence correspondingly. One-time vaccination lowered prevalence initially but increased with influx of new susceptible births. While this epidemiological model is a basic representation of brucellosis transmission dynamics in India and constrained by limitations in surveillance data, this study illustrates the comparative epidemiological impact of different bovine brucellosis prevention and control strategies. PMID:26280026

  4. Tensile strength and durability of bovine dentin.

    PubMed

    Inoue, Toshiko; Miyazaki, Takashi; Nishimura, Fumio

    2007-05-01

    This study investigated the effects of thermal cycling on the tensile strength of dentin. Bovine dentin were divided into 10 groups, which were then subjected to various conditions: intact after preparation, thereby serving as a control; heating in boiling water for 45 minutes; 10,000 thermal cycles in water; 10,000 thermal cycles in PBS; storage in water at 5, 23, or 55 degrees C for two weeks; and storage in PBS at 5, 23, or 55 degrees C for two weeks. Subsequently, bovine dentin were trimmed into dumbbell-shaped specimens and the tensile test performed in distilled water at 37 degrees C. Mean tensile strengths were compared statistically by one-way ANOVA and Fisher's PLTD test (p<0.05). Fracture surfaces were observed by scanning electron microscopy, and reliability of the results was analyzed with Weibull distribution. Tensile strength did not significantly change after thermal cycling or storage in water and PBS at all temperatures tested (71.2-77.0 MPa) but decreased after treatment with boiling water (65.5 MPa). PMID:17694743

  5. PCR detection of bovine herpesviruses from nonbovine ruminants in Hungary.

    PubMed

    Kálmán, Dóra; Egyed, László

    2005-07-01

    Polymerase chain reaction (PCR) was used to test six different nonbovine ruminant species for five bovine herpesviruses including infectious bovine rhinotracheitis virus (BoHV-1), bovine herpes mammillitis virus (BoHV-2), Movar-type herpesvirus (BoHV-4), bovine herpesvirus type 5 (BoHV-5), and alcelaphine herpesvirus 1 (AlHV-1). Species tested included 56 roe deer (Capreolus capreolus), 66 red deer (Cervus elaphus), 20 fallow deer (Dama dama), 16 mouflon (Ovis musimon), 34 domestic sheep, and 44 domestic goats, which were sampled in Hungary in 2003. Tracheal and popliteal lymph nodes collected from these animals were tested for the presence of the five bovine herpesviruses using three nested (two of which were duplex) PCR assays. Three bovine herpesviruses (BoHV-1, -2, and -4) were detected, whereas no evidence of AlHV-1 or BoHV-5 was observed. Prevalence of BoHV-1 ranged from 12% to 47%, and PCR-positive results were observed in all species tested. BoHV-2 was detected from roe deer, red deer, fallow deer, mouflon, and domestic sheep, and the prevalence in these species ranged from 3% to 50%. BoHV-4 was detected in all species, with prevalence ranging from 12% to 69%. Sequenced PCR products were 99-100% identical to bovine herpesviral sequences deposited in the GenBank. PMID:16244057

  6. Characterization of the Nanog 5'-flanking Region in Bovine.

    PubMed

    Choi, Don-Ho; Kim, Duk-Jung; Song, Ki-Duk; Park, Hwan-Hee; Ko, Tae Hyun; Pyao, Yuliya; Chung, Ku-Min; Cha, Seok Ho; Sin, Young-Su; Kim, Nam-Hyung; Lee, Woon-Kyu

    2016-10-01

    Bovine embryonic stem cells have potential for use in research, such as transgenic cattle generation and the study of developmental gene regulation. The Nanog may play a critical role in maintenance of the undifferentiated state of embryonic stem cells in the bovine, as in murine and human. Nevertheless, efforts to study the bovine Nanog for pluripotency-maintaining factors have been insufficient. In this study, in order to understand the mechanisms of transcriptional regulation of the bovine Nanog, the 5'-flanking region of the Nanog was isolated from ear cells of Hanwoo. Results of transient transfection using a luciferase reporter gene under the control of serially deleted 5'-flanking sequences revealed that the -134 to -19 region contained the positive regulatory sequences for the transcription of the bovine Nanog. Results from mutagenesis studies demonstrated that the Sp1-binding site that is located in the proximal promoter region plays an important role in transcriptional activity of the bovine Nanog promoter. The electrophoretic mobility shift assay with the Sp1 specific antibody confirmed the specific binding of Sp1 transcription factor to this site. In addition, significant inhibition of Nanog promoter activity by the Sp1 mutant was observed in murine embryonic stem cells. Furthermore, chromatin-immunoprecipitation assay with the Sp1 specific antibody confirmed the specific binding of Sp1 transcription factor to this site. These results suggest that Sp1 is an essential regulatory factor for bovine Nanog transcriptional activity. PMID:27165025

  7. MicroRNA Expression Profile in Bovine Granulosa Cells of Preovulatory Dominant and Subordinate Follicles during the Late Follicular Phase of the Estrous Cycle

    PubMed Central

    Gebremedhn, Samuel; Salilew-Wondim, Dessie; Ahmad, Ijaz; Sahadevan, Sudeep; Hossain, Md Munir; Hoelker, Michael; Rings, Franca; Neuhoff, Christiane; Tholen, Ernst; Looft, Christian; Schellander, Karl; Tesfaye, Dawit

    2015-01-01

    In bovine, ovarian follicles grow in a wave-like fashion with commonly 2 or 3 follicular waves emerging per estrous cycle. The dominant follicle of the follicular wave which coincides with the LH-surge becomes ovulatory, leaving the subordinate follicles to undergo atresia. These physiological processes are controlled by timely and spatially expressed genes and gene products, which in turn are regulated by post-transcriptional regulators. MicroRNAs, a class of short non-coding RNA molecules, are one of the important posttranscriptional regulators of genes associated with various cellular processes. Here we investigated the expression pattern of miRNAs in granulosa cells of bovine preovulatory dominant and subordinate follicles during the late follicular phase of bovine estrous cycle using Illumina miRNA deep sequencing. In addition to 11 putative novel miRNAs, a total of 315 and 323 known miRNAs were detected in preovulatory dominant and subordinate follicles, respectively. Moreover, in comparison with the subordinate follicles, a total of 64 miRNAs were found to be differentially expressed in preovulatory dominant follicles, of which 34 miRNAs including the miR-132 and miR-183 clusters were significantly enriched, and 30 miRNAs including the miR-17-92 cluster, bta-miR-409a and bta-miR-378 were significantly down regulated in preovulatory dominant follicles. In-silico pathway analysis revealed that canonical pathways related to oncogenesis, cell adhesion, cell proliferation, apoptosis and metabolism were significantly enriched by the predicted target genes of differentially expressed miRNAs. Furthermore, Luciferase reporter assay analysis showed that one of the differentially regulated miRNAs, the miR-183 cluster miRNAs, were validated to target the 3´-UTR of FOXO1 gene. Moreover FOXO1 was highly enriched in granulosa cells of subordinate follicles in comparison with the preovulatory dominant follicles demonstrating reciprocal expression pattern with miR-183

  8. Epidemiology, pathology, immunology and diagnosis of bovine farcy: a review.

    PubMed

    Hamid, Mohamed E

    2012-06-01

    Bovine farcy (which is caused by Mycobacterium farcinogenes and Mycobacterium senegalense) is a chronic suppurative granulomatous inflammation of the skin and lymphatics of cattle and is seen mostly in sub-Saharan Africa. It is not yet certain whether Nocardia farcinica causes cutaneous nocardiosis (farcy) in animals that mimics bovine farcy. Epidemiological data have steadily reported finding bovine farcy in adult cattle of the transhumance pastoralist tribes of the Sahel and the Sudanian savannah zones. M. farcinogenes and or M. senegalense do not affect other domestic or non-domestic animals; it is not known whether these bacteria are zoonotic. The disease--once widespread in many regions--has disappeared from some countries historically known to have it. Reports of bovine farcy prevalence seem to be linked to the existence of survey initiatives by governments and diagnostic capabilities in each country. Farcy causes economic loss due to damaged hides and also is a public-health burden (because the lymphadenitis due to farcy resembles the lesions of bovine tuberculosis in carcasses and the meat is considered inappropriate for human consumption). The current literature is deficient in establishing definitely the prevalence, transmission patterns, and risk factors of bovine farcy. Ixodid ticks transmit other skin diseases (such as dermatophilosis) and might play a role in bovine farcy (given the similarity in the bio-physiology and geographic distribution of the disease). In addition, the tick-resistance of cattle breeds such as the N'Dama, Fulani or the Nilotic might explain their resistance to bovine farcy. Apart from the judicious use of conventional smear-and-culture methods, few diagnostic tests have been developed; the molecular and serological tests have not been evaluated for reproducibility and accuracy. This review points out aspects of bovine farcy that need further research and updates available data on the prevalence, distribution, risk factors

  9. Mannheimia haemolytica leukotoxin binds cyclophilin D on bovine neutrophil mitochondria.

    PubMed

    Aulik, Nicole A; Hellenbrand, Katrina M; Kisiela, Dagmara; Czuprynski, Charles J

    2011-01-01

    Mannheimia haemolytica is an important member of the bovine respiratory disease (BRD) complex that causes fibrinous and necrotizing pleuropneumonia in cattle. BRD is characterized by abundant neutrophil infiltration into the alveoli and fibrin deposition. The most important virulence factor of M. haemolytica is its leukotoxin. Previous research in our laboratory has shown that the leukotoxin is able to enter into and traffic to the mitochondria of a bovine lymphoblastoid cell line (BL-3). In this study, we evaluated the ability of LKT to be internalized and travel to mitochondria in bovine neutrophils. We demonstrate that LKT binds bovine neutrophil mitochondria and co-immunoprecipitates with TOM22 and TOM40, which are members of the translocase of the outer mitochondrial (TOM) membrane family. Upon entry into mitochondria, LKT co-immunoprecipitates with cyclophilin D, a member of the mitochondria permeability transition pore. Unlike BL-3 cells, bovine neutrophil mitochondria are not protected against LKT by the membrane-stabilizing agent cyclosporin A, nor were bovine neutrophil mitochondria protected by the permeability transition pore antagonist bongkrekic acid. In addition, we found that bovine neutrophil cyclophilin D is significantly smaller than that found in BL-3 cells. Bovine neutrophils were protected against LKT by protein transfection of an anti-cyclophilin D antibody directed at the C-terminal amino acids, but not an antibody against the first 50 N-terminal amino acids. In contrast, BL-3 cells were protected by antibodies against either the C-terminus or N-terminus of cyclophilin. These data confirm that LKT binds to bovine neutrophil mitochondria, but indicate there are distinctions between neutrophil and BL-3 mitochondria that might reflect differences in cyclophilin D. PMID:21220005

  10. Genome analysis of an atypical bovine pestivirus from fetal bovine serum.

    PubMed

    Gao, Shandian; Du, Junzheng; Tian, Zhancheng; Xing, Shanshan; Chang, Huiyun; Liu, Guangyuan; Luo, Jianxun; Yin, Hong

    2016-08-01

    We report the complete genome sequence of a bovine pestivirus LVRI/cont-1 originated from a commercial batch of fetal bovine serum. Its complete genome consists of 12,282 nucleotides (nt), which contain an open reading frame (ORF) of 11,700 bp flanked by 5' and 3' untranslated regions (383 and 199 bp). The size of the 5'UTR and the individual protein coding region of LVRI/cont-1 are identical to those of the reference virus Th/04_KhonKaen, but it has a deletion of the first 56 nt in the 3'UTR. Alignment of the complete nucleotide sequence and phylogenetic analysis indicate that this viral isolate belongs to atypical pestiviruses. PMID:27052733

  11. Survey on vertical infection of bovine viral diarrhea virus from fetal bovine sera in the field

    PubMed Central

    NAGAYAMA, Kumiko; OGUMA, Keisuke; SENTSUI, Hiroshi

    2015-01-01

    Bovine viral diarrhea virus (BVDV) isolation and antibody survey were performed using 2,758 fetal bovine sera (FBS) collected from slaughterhouses in New Zealand, Australia and the Dominican Republic, and then sent to Japan to manufacture commercial serum for cell culture use. FBS in the Dominican Republic were pooled for each several individuals, and those collected in other countries were separated according to each individual and subjected to the tests. BVDV was isolated from 25 (0.91%) FBS, and the BVDV antibody was detected in 44 (1.60%) FBS. The survey on 139 sets of paired sera of a dam and her fetus revealed that neither the BVDV antibody nor BVDV was detected in all FBS from BVDV antibody-positive dams. PMID:26074250

  12. Immunological release of histamine from bovine leucocytes. Unusual adrenergic modulation.

    PubMed Central

    Holroyde, M C; Eyre, P

    1976-01-01

    Sensitized bovine granulocytes release histamine when exposed to specific antigen. In comparison with in vitro systems in several other species, the modulation of this release by adrenergic agents is unique. Beta-Adrenoceptor stimulation potentiates (rather than inhibits), whereas alpha-adrenoceptor stimulation inhibits (rather than potentiates) histamine release. Adrenaline, which is generally considered to be a physiological antagonist of the anaphylactic reaction, potentiated histamine release in this study. Dopamine, which is present in high concentration in bovine mast cells, was without effect. The results are discussed in terms of the possible role of granulocytes in bovine hypersensitivity. PMID:60282

  13. Transcriptome Analysis of Androgenic Gland for Discovery of Novel Genes from the Oriental River Prawn, Macrobrachium nipponense, Using Illumina Hiseq 2000

    PubMed Central

    Jin, Shubo; Fu, Hongtuo; Zhou, Qiao; Sun, Shengming; Jiang, Sufei; Xiong, Yiwei; Gong, Yongsheng; Qiao, Hui; Zhang, Wenyi

    2013-01-01

    Background The oriental river prawn, Macrobrachium nipponense, is an important aquaculture species in China, even in whole of Asia. The androgenic gland produces hormones that play crucial roles in sexual differentiation to maleness. This study is the first de novo M. nipponense transcriptome analysis using cDNA prepared from mRNA isolated from the androgenic gland. Illumina/Solexa was used for sequencing. Methodology and Principal Finding The total volume of RNA sample was more than 5 ug. We generated 70,853,361 high quality reads after eliminating adapter sequences and filtering out low-quality reads. A total of 78,408 isosequences were obtained by clustering and assembly of the clean reads, producing 57,619 non-redundant transcripts with an average length of 1244.19 bp. In total 70,702 isosequences were matched to the Nr database, additional analyses were performed by GO (33,203), KEGG (17,868), and COG analyses (13,817), identifying the potential genes and their functions. A total of 47 sex-determination related gene families were identified from the M. nipponense androgenic gland transcriptome based on the functional annotation of non-redundant transcripts and comparisons with the published literature. Furthermore, a total of 40 candidate novel genes were found, that may contribute to sex-determination based on their extremely high expression levels in the androgenic compared to other sex glands,. Further, 437 SSRs and 65,535 high-confidence SNPs were identified in this EST dataset from which 14 EST-SSR markers have been isolated. Conclusion Our study provides new sequence information for M. nipponense, which will be the basis for further genetic studies on decapods crustaceans. More importantly, this study dramatically improves understanding of sex-determination mechanisms, and advances sex-determination research in all crustacean species. The huge number of potential SSR and SNP markers isolated from the transcriptome may shed the lights on research in many

  14. De Novo Transcriptome Sequencing of the Octopus vulgaris Hemocytes Using Illumina RNA-Seq Technology: Response to the Infection by the Gastrointestinal Parasite Aggregata octopiana

    PubMed Central

    Castellanos-Martínez, Sheila; Arteta, David; Catarino, Susana; Gestal, Camino

    2014-01-01

    Background Octopus vulgaris is a highly valuable species of great commercial interest and excellent candidate for aquaculture diversification; however, the octopus’ well-being is impaired by pathogens, of which the gastrointestinal coccidian parasite Aggregata octopiana is one of the most important. The knowledge of the molecular mechanisms of the immune response in cephalopods, especially in octopus is scarce. The transcriptome of the hemocytes of O. vulgaris was de novo sequenced using the high-throughput paired-end Illumina technology to identify genes involved in immune defense and to understand the molecular basis of octopus tolerance/resistance to coccidiosis. Results A bi-directional mRNA library was constructed from hemocytes of two groups of octopus according to the infection by A. octopiana, sick octopus, suffering coccidiosis, and healthy octopus, and reads were de novo assembled together. The differential expression of transcripts was analysed using the general assembly as a reference for mapping the reads from each condition. After sequencing, a total of 75,571,280 high quality reads were obtained from the sick octopus group and 74,731,646 from the healthy group. The general transcriptome of the O. vulgaris hemocytes was assembled in 254,506 contigs. A total of 48,225 contigs were successfully identified, and 538 transcripts exhibited differential expression between groups of infection. The general transcriptome revealed genes involved in pathways like NF-kB, TLR and Complement. Differential expression of TLR-2, PGRP, C1q and PRDX genes due to infection was validated using RT-qPCR. In sick octopuses, only TLR-2 was up-regulated in hemocytes, but all of them were up-regulated in caecum and gills. Conclusion The transcriptome reported here de novo establishes the first molecular clues to understand how the octopus immune system works and interacts with a highly pathogenic coccidian. The data provided here will contribute to identification of biomarkers

  15. The role of melanin pathways in extremotolerance and virulence of Fonsecaea revealed by de novo assembly transcriptomics using illumina paired-end sequencing.

    PubMed

    Li, X Q; Guo, B L; Cai, W Y; Zhang, J M; Huang, H Q; Zhan, P; Xi, L Y; Vicente, V A; Stielow, B; Sun, J F; de Hoog, G S

    2016-01-01

    Melanisation has been considered to be an important virulence factor of Fonsecaea monophora. However, the biosynthetic mechanisms of melanisation remain unknown. We therefore used next generation sequencing technology to investigate the transcriptome and digital gene expression data, which are valuable resources to better understand the molecular and biological mechanisms regulating melanisation in F. monophora. We performed de novo transcriptome assembly and digital gene expression (DGE) profiling analyses of parent (CBS 122845) and albino (CBS 125194) strains using the Illumina RNA-seq system. A total of 17 352 annotated unigenes were found by BLAST search of NR, Swiss-Prot, Gene Ontology, Clusters of Orthologous Groups and Kyoto Encyclopedia of Genes and Genomes (KEGG) (E-value <1e‒5). A total of 2 283 unigenes were judged to be the differentially expressed between the two genotypes. We identified most of the genes coding for key enzymes involved in melanin biosynthesis pathways, including polyketide synthase (pks), multicopper oxidase (mco), laccase, tyrosinase and homogentisate 1,2-dioxygenase (hmgA). DEG analysis showed extensive down-regulation of key genes in the DHN pathway, while up-regulation was noted in the DOPA pathway of the albino mutant. The transcript levels of partial genes were confirmed by real time RT-PCR, while the crucial role of key enzymes was confirmed by either inhibitor or substrate tests in vitro. Meanwhile, numbers of genes involved in light sensing, cell wall synthesis, morphology and environmental stress were identified in the transcriptome of F. monophora. In addition, 3 353 SSRs (Simple Sequence Repeats) markers were identified from 21 600 consensus sequences. Blocking of the DNH pathway is the most likely reason of melanin deficiency in the albino strain, while the production of pheomelanin and pyomelanin were probably regulated by unknown transcription factors on upstream of both pathways. Most of genes involved in

  16. Illumina MiSeq Phylogenetic Amplicon Sequencing Shows a Large Reduction of an Uncharacterised Succinivibrionaceae and an Increase of the Methanobrevibacter gottschalkii Clade in Feed Restricted Cattle

    PubMed Central

    McCabe, Matthew Sean; Cormican, Paul; Keogh, Kate; O’Connor, Aaron; O’Hara, Eoin; Palladino, Rafael Alejandro; Kenny, David Anthony; Waters, Sinéad Mary

    2015-01-01

    Periodic feed restriction is used in cattle production to reduce feed costs. When normal feed levels are resumed, cattle catch up to a normal weight by an acceleration of normal growth rate, known as compensatory growth, which is not yet fully understood. Illumina Miseq Phylogenetic marker amplicon sequencing of DNA extracted from rumen contents of 55 bulls showed that restriction of feed (70% concentrate, 30% grass silage) for 125 days, to levels that caused a 60% reduction of growth rate, resulted in a large increase of relative abundance of Methanobrevibacter gottschalkii clade (designated as OTU-M7), and a large reduction of an uncharacterised Succinivibrionaceae species (designated as OTU-S3004). There was a strong negative Spearman correlation (ρ = -0.72, P = <1x10-20) between relative abundances of OTU-3004 and OTU-M7 in the liquid rumen fraction. There was also a significant increase in acetate:propionate ratio (A:P) in feed restricted animals that showed a negative Spearman correlation (ρ = -0.69, P = <1x10-20) with the relative abundance of OTU-S3004 in the rumen liquid fraction but not the solid fraction, and a strong positive Spearman correlation with OTU-M7 in the rumen liquid (ρ = 0.74, P = <1x10-20) and solid (ρ = 0.69, P = <1x10-20) fractions. Reduced A:P ratios in the rumen are associated with increased feed efficiency and reduced production of methane which has a global warming potential (GWP 100 years) of 28. Succinivibrionaceae growth in the rumen was previously suggested to reduce methane emissions as some members of this family utilise hydrogen, which is also utilised by methanogens for methanogenesis, to generate succinate which is converted to propionate. Relative abundance of OTU-S3004 showed a positive Spearman correlation with propionate (ρ = 0.41, P = <0.01) but not acetate in the liquid rumen fraction. PMID:26226343

  17. SNP Discovery by Illumina-Based Transcriptome Sequencing of the Olive and the Genetic Characterization of Turkish Olive Genotypes Revealed by AFLP, SSR and SNP Markers

    PubMed Central

    Kaya, Hilal Betul; Cetin, Oznur; Kaya, Hulya; Sahin, Mustafa; Sefer, Filiz; Kahraman, Abdullah; Tanyolac, Bahattin

    2013-01-01

    Background The olive tree (Olea europaea L.) is a diploid (2n = 2x = 46) outcrossing species mainly grown in the Mediterranean area, where it is the most important oil-producing crop. Because of its economic, cultural and ecological importance, various DNA markers have been used in the olive to characterize and elucidate homonyms, synonyms and unknown accessions. However, a comprehensive characterization and a full sequence of its transcriptome are unavailable, leading to the importance of an efficient large-scale single nucleotide polymorphism (SNP) discovery in olive. The objectives of this study were (1) to discover olive SNPs using next-generation sequencing and to identify SNP primers for cultivar identification and (2) to characterize 96 olive genotypes originating from different regions of Turkey. Methodology/Principal Findings Next-generation sequencing technology was used with five distinct olive genotypes and generated cDNA, producing 126,542,413 reads using an Illumina Genome Analyzer IIx. Following quality and size trimming, the high-quality reads were assembled into 22,052 contigs with an average length of 1,321 bases and 45 singletons. The SNPs were filtered and 2,987 high-quality putative SNP primers were identified. The assembled sequences and singletons were subjected to BLAST similarity searches and annotated with a Gene Ontology identifier. To identify the 96 olive genotypes, these SNP primers were applied to the genotypes in combination with amplified fragment length polymorphism (AFLP) and simple sequence repeats (SSR) markers. Conclusions/Significance This study marks the highest number of SNP markers discovered to date from olive genotypes using transcriptome sequencing. The developed SNP markers will provide a useful source for molecular genetic studies, such as genetic diversity and characterization, high density quantitative trait locus (QTL) analysis, association mapping and map-based gene cloning in the olive. High levels of

  18. Seroprevalence of bovine immunodeficiency virus and bovine leukemia virus in draught animals in Cambodia.

    PubMed

    Meas, S; Ohashi, K; Tum, S; Chhin, M; Te, K; Miura, K; Sugimoto, C; Onuma, M

    2000-07-01

    Since bovine immunodeficiency virus (BIV), known as bovine lentivirus, has been detected in dairy and beef cattle in various countries around the world, a prevalence study of antibodies to BIV and bovine leukemia virus (BLV) was conducted in draught animals in five provinces in Cambodia, where protozoan parasite infections were suspected in some animals. To clarify the status of draught animals including Haryana, Brahman, mixed-breed, local breed cattle and muscle water buffaloes, a total of 544 cattle and 42 buffaloes were tested, and 26.3 and 16.7%, respectively, were found positive for anti-BIV p26 antibodies determined by Western blotting. There were 5.3% positive for anti-BLV antibodies detected by immunodiffusion test among the cattle, but no reactors among buffaloes and no dual infection for both BIV and BLV was determined in this study. Peripheral blood mononuclear cells from BIV-seropositive cattle were found to have BIV-provirus DNA, as detected by polymerase chain reaction and subsequent Southern blot hybridization. This is the first evidence for the presence of BIV and BLV infections in draught animals in tropical countries such as Cambodia. This wide distribution of BIV suggests its association with problems in animal health as reported worldwide, and that a primary BIV infection can predispose death of affected animals by other aggressive pathogens or stresses. PMID:10945301

  19. Bovine achondrogenesis: evidence for defective chondrocyte differentiation.

    PubMed

    Horton, W A; Jayo, M J; Leipold, H W; Machado, M A; Campbell, D; Ahmed, S

    1987-01-01

    A survey study of growth cartilage abnormalities in bovine bone dysplasias revealed that a disorder in Holstein cattle called bulldog calf closely resembles human achondrogenesis Type II. Substantial amounts of Type I collagen and other non Type II collagens were detected in the bulldog cartilage which was comprised primarily of extensive vascular canals and cells having the characteristics of hypertrophic and degenerative chondrocytes normally found in the growth plate. It is proposed that chondrocytes throughout the bulldog growth cartilage prematurely differentiate into hypertrophic cells that degenerate and predispose the cartilage to vascular invasion and the formation of cartilage canals. The presence of these canals probably accounts for most of the observed collagen abnormalities. PMID:3606909

  20. Tear and decohesion of bovine pericardial tissue.

    PubMed

    Tobaruela, Almudena; Elices, Manuel; Bourges, Jean Yves; Rojo, Francisco Javier; Atienza, José Miguel; Guinea, Gustavo

    2016-10-01

    The aim of this study was to evaluate quantitatively the fracture-by tear and delamination-of bovine pericardium tissues which are usually employed for the manufacture of bioprosthetic valves. A large number of samples (77) were tested in root-to-apex and circumferential directions, according to a standardised tear test (ASTM D 1938). Before performing the tear test, some samples were subjected to 1000 cycles of fatigue to a maximum stress of 3MPa. Fracture toughness of tearing and delamination were computed by following a simple fracture model. The study showed significantly lower values of delamination toughness compared with tear delamination. Moreover, tear forces were different in each test direction, revealing a clear orthotropic behaviour. All these results, as well as the testing procedure, could be of value for future research in the physiological function of pericardium tissues and clinical applications. PMID:27337412

  1. Diagnosis and Control of Bovine Neosporosis.

    PubMed

    McAllister, Milton M

    2016-07-01

    Neosporosis is one of the most common and widespread causes of bovine abortion. The causative parasite is transmitted in at least two ways, horizontally from canids, and by endogenous transmission within maternal lines of infected cattle. The prevalence of neosporosis is higher in the dairy industry than in the beef industry because of risk factors associated with intensive feeding. There are no vaccines, but logical management options are discussed that can lower the risk of abortion outbreaks and gradually reduce the prevalence of infection within herds. Steps should be taken to prevent total mixed rations from becoming contaminated by canine feces. If a herd has a high rate of infection that is associated with abortions in heifers, then the rate of reduction of infection prevalence can be speeded by only selecting seronegative replacement heifers to enter the breeding herd. Elimination of all infected cattle is not a recommended goal. PMID:27161392

  2. Properties of intracellular bovine papillomavirus chromatin.

    PubMed Central

    Rösl, F; Waldeck, W; Zentgraf, H; Sauer, G

    1986-01-01

    Episomal nucleoprotein complexes of bovine papillomavirus type 1 (BPV-1) in transformed cells were exposed to DNase I treatment to localize hypersensitive regions. Such regions, which are indicative for gene expression, were found within the noncoding part of the genome, coinciding with the origin of replication and the 5' ends of most of the early mRNAs. However, there were also regions of hypersensitivity within the structural genes. These intragenic perturbations of the chromatin structure coincide with regulatory sequences at the DNA level. One of these regions maps in close proximity to a Z-DNA antibody-binding site which is located near the putative BPV-1 enhancer sequence. Images PMID:3009863

  3. Somatic Cell Counts in Bovine Milk

    PubMed Central

    Dohoo, I. R.; Meek, A. H.

    1982-01-01

    Factors which influence somatic cell counts in bovine milk are reviewed and guidelines for their interpretation are presented. It is suggested that the thresholds of 300 000 and 250 000 cells/mL be used to identify infected quarters and cows respectively. However, it is stressed that somatic cell counts are general indicators of udder health which are subject to the influence of many factors. Therefore the evaluation of several successive counts is preferable to the interpretation of an individual count. Relationships between somatic cell counts and both milk production and milk composition are discussed. Subclinical mastitis reduces milk quality and decreases yield although the relationship between production loss and somatic cell count requires clarification. Finally the availability of somatic cell counting programs in Canada is presented. PMID:17422127

  4. Diminishing returns in bovine tuberculosis control.

    PubMed

    Hone, J

    2013-07-01

    Mycobacterium bovis causes bovine tuberculosis (bTB) in many mammals including cattle, deer and brushtail possum. The aim of this study was to estimate the strength of association, using model selection (AICc) regression analyses, between the proportion of cattle and farmed deer herds with bTB in New Zealand and annual costs of TB control, namely disease control in livestock, in wildlife or in a combination of the two. There was more support for curved (concave up) than linear models which related the proportion of cattle and farmed deer herds with bTB to the annual control costs. The curved, concave-up, best-fitting relationships showed diminishing returns with no positive asymptote and implied TB eradication is feasible in New Zealand. PMID:23632097

  5. Lipidome signatures in early bovine embryo development.

    PubMed

    Sudano, Mateus J; Rascado, Tatiana D S; Tata, Alessandra; Belaz, Katia R A; Santos, Vanessa G; Valente, Roniele S; Mesquita, Fernando S; Ferreira, Christina R; Araújo, João P; Eberlin, Marcos N; Landim-Alvarenga, Fernanda D C

    2016-07-15

    Mammalian preimplantation embryonic development is a complex, conserved, and well-orchestrated process involving dynamic molecular and structural changes. Understanding membrane lipid profile fluctuation during this crucial period is fundamental to address mechanisms governing embryogenesis. Therefore, the aim of the present work was to perform a comprehensive assessment of stage-specific lipid profiles during early bovine embryonic development and associate with the mRNA abundance of lipid metabolism-related genes (ACSL3, ELOVL5, and ELOVL6) and with the amount of cytoplasmic lipid droplets. Immature oocytes were recovered from slaughterhouse-derived ovaries, two-cell embryos, and eight- to 16-cell embryos, morula, and blastocysts that were in vitro produced under different environmental conditions. Lipid droplets content and mRNA transcript levels for ACSL3, ELOVL5, and ELOVL6, monitored by lipid staining and quantitative polymerase chain reaction, respectively, increased at morula followed by a decrease at blastocyst stage. Relative mRNA abundance changes of ACSL3 were closely related to cytoplasmic lipid droplet accumulation. Characteristic dynamic changes of phospholipid profiles were observed during early embryo development and related to unsaturation level, acyl chain length, and class composition. ELOVL5 and ELOVL6 mRNA levels were suggestive of overexpression of membrane phospholipids containing elongated fatty acids with 16, 18, and 20 carbons. In addition, putative biomarkers of key events of embryogenesis, embryo lipid accumulation, and elongation were identified. This study provides a comprehensive description of stage-specific lipidome signatures and proposes a mechanism to explain its potential relationship with the fluctuation of both cytoplasmic lipid droplets content and mRNA levels of lipid metabolism-related genes during early bovine embryo development. PMID:27107972

  6. Nonenzymatic glycosylation of bovine myelin basic protein

    SciTech Connect

    Hitz, J.B.

    1987-01-01

    In the CNS myelin sheath the nonenzymatic glycosylation reaction (at the early stage of the Amadori product) occurs only with the myelin basic protein and not with the other myelin proteins. This was observed in isolated bovine myelin by in vitro incubation with (/sup 14/C)-galactose and (/sup 14/C)-glucose. The respective in-vitro incorporation rates for purified bovine myelin basic protein with D-galactose, D-glucose and D-mannose were 7.2, 2.4 and 2.4 mmoles/mole myelin basic protein per day at 37/sup 0/C. A more rapid, HPLC method was devised and characterized to specifically analyze for the Amadori product. The HPLC method was correlated to the (/sup 14/C)-sugar incorporation method for myelin basic protein under a set of standard reaction conditions using (/sup 14/C)-glucose and (/sup 14/C)-mannose with HPLC values at 1/6 and 1/5 of the (/sup 14/C)-sugar incorporation method. A novel myelin basic protein purification step has been developed that yields a relativity proteolytic free preparation that is easy to work with, being totally soluble at a neutral pH. Nine new spots appear for a trypsinized glycosylated MBP in the paper peptide map of which eight correspond to positions of the (/sup 3/H)-labeled Amadori product in affinity isolated peptides. These studies provide a general characterization of and a structural basis for investigations on nonenzymatically glycosylated MBP as well as identifying MBP as the only nonenzymatically glycosylated protein in the CNS myelin sheath which may accumulate during aging, diabetes, and demyelinating diseases in general.

  7. Tylosin susceptibility of Staphylococci from bovine mastitis.

    PubMed

    Entorf, Monika; Feßler, Andrea T; Kadlec, Kristina; Kaspar, Heike; Mankertz, Joachim; Peters, Thomas; Schwarz, Stefan

    2014-07-16

    Although the 16-membered macrolide tylosin is commonly used for the treatment of bovine mastitis, little information is currently available about the susceptibility of mastitis pathogens to tylosin. In the present study, 112 Staphylococcus aureus and 110 coagulase-negative Staphylococcus (CoNS) spp. isolates from cases of bovine mastitis were tested by broth microdilution and agar disk diffusion with 30 μg tylosin disks. Susceptibility to erythromycin was tested by broth microdilution and disk diffusion using 15 μg disks. Both test populations showed bimodal distributions of minimal inhibitory concentrations (MICs) and zone diameters with eleven S. aureus and eight CoNS isolates showing tylosin MICs of ≥ 256 μg/ml and no zones of growth inhibition around the tylosin 30 μg disks. All 19 isolates with tylosin MICs of ≥ 256 μg/ml were also resistant to erythromycin. For six additional erythromycin-resistant isolates, tylosin MICs of 1-8 μg/ml were observed. One S. aureus and two CoNS isolates showed inducible macrolide resistance. PCR analysis of the 25 erythromycin-resistant staphylococcal isolates identified the resistance genes erm(A), erm(B), erm(C), erm(T), mph(C) and msr(A) alone or in different combinations. An excellent correlation between the results of the different tylosin susceptibility tests (broth microdilution versus disk diffusion) was seen for S. aureus and CoNS isolates. Since tylosin does not induce the expression of the aforementioned erm genes, isolates with an inducible resistance phenotype may - if only tylosin is tested - be falsely classified as tylosin-susceptible. Thus, erythromycin should be tested in parallel and tylosin should only be used for the treatment of infections caused by erythromycin-susceptible staphylococci. PMID:24461550

  8. Detection and characterization of viruses as field and vaccine strains in feedlot cattle with bovine respiratory disease

    Technology Transfer Automated Retrieval System (TEKTRAN)

    This study investigated viruses in bovine respiratory disease (BRD) cases in feedlots, including bovine herpesvirus-1 (BoHV-1), bovine viral diarrhea virus (BVDV), bovine respiratory syncytial virus (BRSV), bovine coronaviruses (BoCV) and parainfluenza-3 virus (PI3V). Nasal swabs were collected fro...

  9. MRSA endocarditis of bovine Contegra valved conduit: a case report

    PubMed Central

    2009-01-01

    Background Different techniques are used for the right ventricular outflow tract reconstruction, including homo- or porcine xenografts, which have several limitations. Contegra, a bovine jugular vein graft, is an interesting alternative to overcome these limitations. It consists of a bovine jugular vein with a naturally integrated valve in it. Isolated pulmonary valve endocarditis is extremely rare. Case presentation We report the case of a 20 years old male patient with acute endocarditis of bovine Contegra valved conduit, four years after right ventricular outflow tract reconstruction and atrial septal defect correction, associated with acute glomerulonephritis, renal failure and severe anemia, secondary to methicillin-resistant Staphylococcus aureus infection (MRSA). Conclusion We present a complex patient with acute endocarditis of bovine Contegra valved conduit. We believe that the presentation of this case should encourage the researchers for the discussing of the implantation of this conduit and the prevention of endocarditis in these patients. PMID:19146664

  10. Cartilage (Bovine and Shark) (PDQ®)—Patient Version

    Cancer.gov

    Expert-reviewed information summary about the use of bovine and shark cartilage as a treatment for people with cancer. Note: The information in this summary is no longer being updated and is provided for reference purposes only.

  11. Cartilage (Bovine and Shark) (PDQ®)—Health Professional Version

    Cancer.gov

    Expert-reviewed information summary about the use of bovine and shark cartilage as a treatment for people with cancer. Note: The information in this summary is no longer being updated and is provided for reference purposes only.

  12. Association of a bovine prion gene haplotype with atypical BSE

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Background: Atypical bovine spongiform encephalopathies (BSEs) are recently recognized prion diseases of cattle. Atypical BSEs are rare; approximately 30 cases have been identified worldwide. We tested prion gene (PRNP) haplotypes for an association with atypical BSE. Methodology/Principal Findin...

  13. Immune suppression in calves with bovine immunodeficiency virus.

    PubMed Central

    Zhang, S; Wood, C; Xue, W; Krukenberg, S M; Chen, Q; Minocha, H C

    1997-01-01

    The present study was designed to evaluate the effect of bovine immunodeficiency virus (BIV) infection on immune functions and possible interactions between BIV and other bovine viruses in calves. Ten calves were inoculated intravenously with BIV, and five served as controls. An increased lymphocyte proliferation to BIV gag protein was demonstrated 2 to 6 weeks after BIV inoculation (P < 0.05). Lymphocyte subset differentiation revealed a decreased CD4/CD8 ratio (P < 0.05) during weeks 2 to 7, suggesting a possible immune dysfunction in BIV-infected calves. When the calves were inoculated with bovine herpesvirus type 1 (BHV-1), the antibody response to BHV-1 in BIV-infected calves was delayed and the antibody titers were significantly lower (P < 0.05). Injection of bovine viral diarrhea virus vaccine also elicited a lower neutralizing antibody response in BIV-infected calves. The results indicated that immune suppression occurred in BIV-infected calves. PMID:9067663

  14. Molecular and biological aspects of the bovine immunodeficiency virus.

    PubMed

    Corredor, Andrea G; St-Louis, Marie-Claude; Archambault, Denis

    2010-01-01

    The bovine immunodeficiency virus (BIV) was isolated in 1969 from a cow, R-29, with a wasting syndrome suggesting bovine leucosis. The virus, first designated bovine visna-like virus, remained unstudied until HIV was discovered in 1983. Then, it was demonstrated in 1987 that the bovine R-29 isolate was a lentivirus with striking similarity to the human immunodeficiency virus (HIV). Moreover, BIV has the most complex genomic structure among all identified lentiviruses shown by several regulatory/accessory genes encoding proteins, some of which are involved in the regulation of virus gene expression. This manuscript aims to review biological and molecular aspects of BIV, with emphasis on regulatory/accessory viral genes/proteins which are involved in virus expression. PMID:20210777

  15. 9 CFR 113.310 - Bovine Rhinotracheitis Vaccine.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ...-neutralization tests from each of the vaccinates. The test virus shall be 100 to 500 TCID50 bovine...-injection serum samples tested as prescribed in paragraph (c)(3) of this section show neutralization in...

  16. 9 CFR 113.310 - Bovine Rhinotracheitis Vaccine.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ...-neutralization tests from each of the vaccinates. The test virus shall be 100 to 500 TCID50 bovine...-injection serum samples tested as prescribed in paragraph (c)(3) of this section show neutralization in...

  17. 9 CFR 113.310 - Bovine Rhinotracheitis Vaccine.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ...-neutralization tests from each of the vaccinates. The test virus shall be 100 to 500 TCID50 bovine...-injection serum samples tested as prescribed in paragraph (c)(3) of this section show neutralization in...

  18. 9 CFR 113.310 - Bovine Rhinotracheitis Vaccine.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ...-neutralization tests from each of the vaccinates. The test virus shall be 100 to 500 TCID50 bovine...-injection serum samples tested as prescribed in paragraph (c)(3) of this section show neutralization in...

  19. 9 CFR 113.310 - Bovine Rhinotracheitis Vaccine.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ...-neutralization tests from each of the vaccinates. The test virus shall be 100 to 500 TCID50 bovine...-injection serum samples tested as prescribed in paragraph (c)(3) of this section show neutralization in...

  20. BTA2 and BTA26 are linked with bovine respiratory disease and associated with persistent infection of bovine viral diarrhea virus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bovine viral diarrhea virus is a pathogen associated with bovine respiratory disease (BRD). BRD causes 28% of all cattle deaths and an annual U.S. loss over $692 million. The objective of this study was to refine the linkage of BRD and association of bovine viral diarrhea-persistent infection (BVD-P...

  1. Fine mapping of loci on BTA2 and BTA26 associated with bovine viral diarrhea persistent infection and linked with bovine respiratory disease in cattle

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bovine respiratory disease (BRD) is considered to be the most costly infectious disease in the cattle industry. Bovine viral diarrhea virus (BVDV) is one of the pathogens involved with the BRD complex of disease. Bovine viral diarrhea virus infection also negatively impacts cow reproduction and calf...

  2. Isolation and Transfection of Primary Culture Bovine Retinal Pericytes.

    PubMed

    Primo, Vincent A; Arboleda-Velasquez, Joseph F

    2016-01-01

    This protocol describes an enzymatic approach for isolating homogeneous cultures of pericytes from retinas of bovine source. In summary, retinas are dissected, washed, digested, filtered, cultured in specific media to select for pericytes, and finally expanded for a low passage culture of about 14 million bovine retinal pericytes (BRP) within 4-6 weeks. This protocol also describes a liposomal-based technique for transfection of BRPs. PMID:27172949

  3. On-Farm Use of Ultrasonography for Bovine Respiratory Disease.

    PubMed

    Ollivett, Theresa L; Buczinski, Sébastien

    2016-03-01

    Thoracic ultrasonography (TUS) in young cattle has recently gained momentum as an accurate and practical tool for identifying the lung lesions associated with bovine respiratory disease. As cattle producers increasingly seek input from their veterinarians on respiratory health issues, bovine practitioners should consider adding TUS to their practice models. This article discusses the relevant literature regarding TUS in young cattle, current acceptable techniques, and practical on-farm applications. PMID:26922110

  4. Bovine coronavirus antibody titers at weaning negatively correlate with incidence of bovine respiratory disease in the feed yard

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bovine respiratory disease complex (BRDC) is a multifactorial disease caused by complex interactions among viral and bacterial pathogens, stressful management practices and host genetic variability. Although vaccines and antibiotic treatments are readily available to prevent and treat infection caus...

  5. Stability of Bovine viral diarrhea virus 1 nucleic acid in fetal bovine samples stored under different conditions

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Infection of pregnant cattle with bovine viral diarrhea viruses can result in reproductive disease that includes fetal reabsorption, mummification, abortion, still births, congenital defects affecting structural, neural, reproductive and immune systems and the birth of calves persistently infected w...

  6. Effects of candidate gene polymorphisms on the detailed fatty acids profile determined by gas chromatography in bovine milk.

    PubMed

    Pegolo, S; Cecchinato, A; Mele, M; Conte, G; Schiavon, S; Bittante, G

    2016-06-01

    Association analyses between candidate genes and bovine milk fatty acids can improve our understanding of genetic variation in milk fatty acid profiles and reveal potential opportunities to tailor milk fat composition through selection strategies. In this work, we investigated the association of 51 single nucleotide polymorphisms (SNP) selected from 37 candidate genes using a functional and positional approach, with 47 fatty acids, 9 fatty acid groups, and 5 Δ(9)-desaturation indices in milk samples from Brown Swiss cows. Individual milk samples were collected from 1,158 Italian Brown Swiss cows, and gas chromatography was used to obtain detailed milk fatty acid compositions. A GoldenGate assay system (Illumina, San Diego, CA) was used to perform genotype 96 selected SNP located in 54 genes across 22 chromosomes. In total, 51 polymorphic SNP in 37 candidate genes were retained for the association analysis. A Bayesian linear animal model was used to estimate the contribution of each SNP. A total of 129 tests indicated relevant additive effects between a given SNP and a single fatty acid trait; 38 SNP belonging to 30 genes were relevant for a total of 57 fatty acid traits. Most of the studied fatty acid traits (~81%) were relevantly associated with multiple SNP. Relevantly associated SNP were mainly found in genes related to fat metabolism, linked to or contained in previously identified quantitative trait loci for fat yield or content, or associated with genes previously identified in association analyses with milk fatty acid profiles in other cow breeds. The most representative candidate genes were LEP, PRL, STAT5A, CCL3, ACACA, GHR, ADRB2, LPIN1, STAT1, FABP4, and CSN2. In particular, relevant associations with SNP located on bovine chromosome 19 (BTA19) were found. Two candidate genes on BTA19 (CCL3 and ACACA) were relevantly associated with de novo short- and medium-chain fatty acids, likely explaining the high heritability values found for these fatty acids

  7. Increased bovine Tim-3 and its ligand expressions during bovine leukemia virus infection

    PubMed Central

    2012-01-01

    The immunoinhibitory receptor T cell immunoglobulin domain and mucin domain-3 (Tim-3) and its ligand, galectin-9 (Gal-9), are involved in the immune evasion mechanisms for several pathogens causing chronic infections. However, there is no report concerning the role of Tim-3 in diseases of domestic animals. In this study, cDNA encoding for bovine Tim-3 and Gal-9 were cloned and sequenced, and their expression and role in immune reactivation were analyzed in bovine leukemia virus (BLV)-infected cattle. Predicted amino acid sequences of Tim-3 and Gal-9 shared high homologies with human and mouse homologues. Functional domains, including tyrosine kinase phosphorylation motif in the intracellular domain of Tim-3 were highly conserved among cattle and other species. Quantitative real-time PCR analysis showed that bovine Tim-3 mRNA is mainly expressed in T cells such as CD4+ and CD8+ cells, while Gal-9 mRNA is mainly expressed in monocyte and T cells. Tim-3 mRNA expression in CD4+ and CD8+ cells was upregulated during disease progression of BLV infection. Interestingly, expression levels for Tim-3 and Gal-9 correlated positively with viral load in infected cattle. Furthermore, Tim-3 expression level closely correlated with up-regulation of IL-10 in infected cattle. The expression of IFN-γ and IL-2 mRNA was upregulated when PBMC from BLV-infected cattle were cultured with Cos-7 cells expressing Tim-3 to inhibit the Tim-3/Gal-9 pathway. Moreover, combined blockade of the Tim-3/Gal-9 and PD-1/PD-L1 pathways significantly promoted IFN-γ mRNA expression compared with blockade of the PD-1/PD-L1 pathway alone. These results suggest that Tim-3 is involved in the suppression of T cell function during BLV infection. PMID:22621175

  8. Malignant transformation of hamster cells following infection with bovine herpesvirus (infectious bovine rhinotracheitis virus.

    PubMed

    Michalski, F; Hsiung, G D

    1975-03-01

    Hamster embryo cells, following infection with IBR virus, showed malignant transformation. Hamsters of all ages, inbred or random bred, inoculated with two of the transformed cell lines developed solid tumors. Preliminary characterization of the tumors induced by one of the cell lines has indicated undifferentiated sarcomas. Viral specific antigen was detected in about 5% of the transformed cells and 10% of primary tumor cells in culture. Viral specific antibody was detected in the serum of tumor-bearing hamsters by the indirect immunofluorescent method, but no neutralizing antibodies were found. Infectious virus has not been recovered from either the transformed or tumor cells by cocultivation with bovine embryonic kidney cells. PMID:165538

  9. Cerebral Candidal Abscess and Bovine Viral Diarrhoea Virus Infection in an Aborted Bovine Fetus.

    PubMed

    Vilander, A C; Niles, G A; Frank, C B

    2016-01-01

    Candida species are opportunistic fungi associated with immunosuppression and are the most commonly isolated fungal pathogens from the human central nervous system. Invasive candidiasis is reported uncommonly in animals and there have only been two reports of candidal infection of the brain. This report presents a case of a cerebral candidal abscess in an aborted late-term calf co-infected with bovine viral diarrhoea virus. Candida etchellsii, a species not previously identified as pathogenic, was identified as the causative agent by polymerase chain reaction. PMID:26895887

  10. Sequence and structural implications of a bovine corneal keratan sulfate proteoglycan core protein. Protein 37B represents bovine lumican and proteins 37A and 25 are unique

    NASA Technical Reports Server (NTRS)

    Funderburgh, J. L.; Funderburgh, M. L.; Brown, S. J.; Vergnes, J. P.; Hassell, J. R.; Mann, M. M.; Conrad, G. W.; Spooner, B. S. (Principal Investigator)

    1993-01-01

    Amino acid sequence from tryptic peptides of three different bovine corneal keratan sulfate proteoglycan (KSPG) core proteins (designated 37A, 37B, and 25) showed similarities to the sequence of a chicken KSPG core protein lumican. Bovine lumican cDNA was isolated from a bovine corneal expression library by screening with chicken lumican cDNA. The bovine cDNA codes for a 342-amino acid protein, M(r) 38,712, containing amino acid sequences identified in the 37B KSPG core protein. The bovine lumican is 68% identical to chicken lumican, with an 83% identity excluding the N-terminal 40 amino acids. Location of 6 cysteine and 4 consensus N-glycosylation sites in the bovine sequence were identical to those in chicken lumican. Bovine lumican had about 50% identity to bovine fibromodulin and 20% identity to bovine decorin and biglycan. About two-thirds of the lumican protein consists of a series of 10 amino acid leucine-rich repeats that occur in regions of calculated high beta-hydrophobic moment, suggesting that the leucine-rich repeats contribute to beta-sheet formation in these proteins. Sequences obtained from 37A and 25 core proteins were absent in bovine lumican, thus predicting a unique primary structure and separate mRNA for each of the three bovine KSPG core proteins.

  11. Preparation of Monoclonal Antibodies Against Bovine Haptoglobin

    PubMed Central

    Wang, Caihong; Gu, Cheng; Guo, Donghua; Gao, Jing; Li, Chunqiu; Liu, Na; Geng, Yufei; Su, Mingjun; Wang, Xinyu

    2014-01-01

    Female, 8-week-old BALB/c mice were immunized with purified recombinant proteins of the predicted immunodominant region of bovine haptoglobin (pirBoHp). Two monoclonal antibodies (MAbs), named 1B3 and 6D6, were prepared by conventional B lymphocyte hybridoma technique. Titers of ascitic fluid and cell culture supernatant of MAb 1B3 were 1:9.6×108 and 1:8.2×104, respectively, and that of MAb 6D6 were 1:4.4×105 and 1:1.0×104, respectively. The subtype of MAbs 1B3 and 6D6 was IgG1κ. In Western blot analysis, MAbs 1B3 and 6D6 could recognize the α-chain of native BoHp from plasma of dairy cows. These data indicated that MAbs 1B3 and 6D6 have a potential use for developing diagnostic reagents of BoHp. PMID:25358005

  12. Femtosecond laser ablation of bovine cortical bone

    NASA Astrophysics Data System (ADS)

    Cangueiro, Liliana T.; Vilar, Rui; Botelho do Rego, Ana M.; Muralha, Vania S. F.

    2012-12-01

    We study the surface topographical, structural, and compositional modifications induced in bovine cortical bone by femtosecond laser ablation. The tests are performed in air, with a Yb:KYW chirped-pulse-regenerative amplification laser system (500 fs, 1030 nm) at fluences ranging from 0.55 to 2.24 J/cm2. The ablation process is monitored by acoustic emission measurements. The topography of the laser-treated surfaces is studied by scanning electron microscopy, and their constitution is characterized by glancing incidence x-ray diffraction, x-ray photoelectron spectroscopy, Fourier transform infrared spectroscopy, and micro-Raman spectroscopy. The results show that femtosecond laser ablation allows removing bone without melting, carbonization, or cracking. The structure and composition of the remaining tissue are essentially preserved, the only constitutional changes observed being a reduction of the organic material content and a partial recrystallization of hydroxyapatite in the most superficial region of samples. The results suggest that, within this fluence range, ablation occurs by a combination of thermal and electrostatic mechanisms, with the first type of mechanism predominating at lower fluences. The associated thermal effects explain the constitutional changes observed. We show that femtosecond lasers are a promising tool for delicate orthopaedic surgeries, where small amounts of bone must be cut with negligible damage, thus minimizing surgical trauma.

  13. The release of bradykinin in bovine mastitis.

    PubMed

    Eshraghi, H R; Zeitlin, I J; Fitzpatrick, J L; Ternent, H; Logue, D

    1999-01-01

    The kinin peptides are released during inflammation and are amongst the most potent known mediators of vasodilatation, pain and oedema. Despite early reports of the presence of kinins in milk, no previous study has investigated the role of the kinin system in bovine mastitis. The present study indicated that mastitis was accompanied by raised levels of bradykinin (BK) in milk and the increased levels of BK correlated with the severity of mastitis. Raised BK levels in mastitic milk were not dependent on the presence of inflammatory cells, nor were they secondary to changes in blood levels of BK. In milk from sub-clinically inflamed quarters, BK was raised in those milks where Staphylococcus aureus (S. aureus) was isolated but not in those milks where no pathogen was isolated. Increasing S. aureus artificially, also caused an increase in the milk BK. Increases in milk BK were not restricted only to the mastitic quarters of the udder. In udders in which mastitis was detected in one or more quarters, BK increases were also detected in the apparently uninvolved quarters. PMID:10328527

  14. Recombinant viral vaccines for enzootic bovine leucosis.

    PubMed

    Daniel, R C; Gatei, M H; Good, M F; Boyle, D B; Lavin, M F

    1993-10-01

    Recently published studies on the development and use of recombinant vaccinia virus (VV) vaccines incorporating either the complete envelope (env) gene or only a fragment of the env gene consisting of the coding sequence for the env glycoprotein 51 (gp51) and part of gp30 of the bovine leukaemia virus (BLV) are described. It has been reported that vaccination of sheep with recombinant VV vaccines containing the complete env gene appears to protect sheep against challenge infection with BLV. The evidence for this protection is based on the lack of persistence of high titres of anti-gp51 antibodies compared with unvaccinated BLV infected controls, on the enhanced CD4 proliferative responses to specific BLV gp51 synthetic peptides in the vaccinated sheep, and on the inability to detect BLV pro-virus by polymerase chain reaction in the vaccinated sheep after 4 months following challenge infection compared with continual detection in unvaccinated sheep over a 16 month trial period. It has been suggested that cell-mediated immune responses may be an important aspect of protective immunity against BLV infection and it has been reported that large tracts of amino acid sequences within the env and pol genes are highly conserved in different isolates from different countries which is of importance in designing peptide derived vaccines. PMID:8270269

  15. [Enzootic bovine leukosis--a retrovirus disease].

    PubMed

    Straub, O C

    1988-01-01

    After a short historical introduction and a description of some of the properties of the bovine leucosis (leukemia) virus (BLV) the natural pathways of transmission, for which most likely intact infected cells are a prerequisite, are described and the role of possible vectors mentioned. The iatrogenic transmission by needles, dehorning and perhaps even rectal examinations is discussed. Semen, embryo transfer, saliva, urine, faeces and breath do obviously not play any role in contrast to nasal secretions containing viable cells. It is pointed out that it is not inevitable to eliminate all seropositive cattle from the premises during an eradication programme provided some additional tests are carried out: determination of the white blood picture, the p 24 status and perhaps even proof of antigen production in short-term buffy coat cultures. Offspring of seropositive cattle requires some special attention and careful testing at certain intervals. The report according to which BVD/MD virus-infected cattle are superinfected with BLV leading to rather difficult situations is cited. PMID:2851883

  16. Bovine leukocyte adhesion deficiency (BLAD): a review.

    PubMed

    Nagahata, Hajime

    2004-12-01

    Bovine leukocyte adhesion deficiency (BLAD) in Holstein cattle is an autosomal recessive congenital disease characterized by recurrent bacterial infections, delayed wound healing and stunted growth, and is also associated with persistent marked neutrophilia. The molecular basis of BLAD is a single point mutation (adenine to guanine) at position 383 of the CD18 gene, which caused an aspartic acid to glycine substitution at amino acid 128 (D128G) in the adhesion molecule CD18. Neutrophils from BLAD cattle have impaired expression of the beta2 integrin (CD11a,b,c/CD18) of the leukocyte adhesion molecule. Abnormalities in a wide spectrum of adherence dependent functions of leukocytes have been fully characterized. Cattle affected with BLAD have severe ulcers on oral mucous membranes, severe periodontitis, loss of teeth, chronic pneumonia and recurrent or chronic diarrhea. Affected cattle die at an early age due to the infectious complications. Holstein bulls, including carrier sires that had a mutant BLAD gene in heterozygote were controlled from dairy cattle for a decade. The control of BLAD in Holstein cattle by publishing the genotypes and avoiding the mating between BLAD carriers was found to be successful. This paper provides an overview of the genetic disease BLAD with reference to the disease in Holstein cattle. PMID:15644595

  17. Bioelectrical impedance analysis of bovine milk fat

    NASA Astrophysics Data System (ADS)

    Veiga, E. A.; Bertemes-Filho, P.

    2012-12-01

    Three samples of 250ml at home temperature of 20°C were obtained from whole, low fat and fat free bovine UHT milk. They were analysed by measuring both impedance spectra and dc conductivity in order to establish the relationship between samples related to fat content. An impedance measuring system was developed, which is based on digital oscilloscope, a current source and a FPGA. Data was measured by the oscilloscope in the frequency 1 kHz to 100 kHz. It was showed that there is approximately 7.9% difference in the conductivity between whole and low fat milk whereas 15.9% between low fat and free fat one. The change of fatness in the milk can be significantly sensed by both impedance spectra measurements and dc conductivity. This result might be useful for detecting fat content of milk in a very simple way and also may help the development of sensors for measuring milk quality, as for example the detection of mastitis.

  18. Membrane associated phospholipase C from bovine brain

    SciTech Connect

    Lee, K.; Ryu, S.H.; Suh, P.; Choi, W.C.; Rhee, S.G.

    1987-05-01

    Cytosolic fractions of bovine brain contain 2 immunologically distinct phosphoinositide-specific phospholipase (PLC), PLC-I and PLC-II, whose MW are 150,000 and 145,000 respectively, under a denaturing condition. Monoclonal antibodies were derived against each form and specific radioimmunoassays were developed. Distribution of PLC-I and PLC-II in cytosolic and particulate fractions was measured using the radioimmunoassay. More than 90% of PLC-II was found in the cytosolic fraction, while the anti-PLC-I antibody cross-reacting protein was distributed nearly equally between the soluble fraction and the 2 M KCl extract of particulate fraction. The PLC enzyme in the particulate fraction was purified to homogeneity, yielding 2 proteins of 140 KDa and 150 KDa when analyzed on SDS-PAGE. Neither of the 2 enzymes cross-reacted with anti-PLC-II antibodies, but both could be immunoblotted by all 4 different anti-PLC-I antibodies. This suggests that the 140 KDa PLC was derived from the 150 KDa form. The 150 Kda form from particulate fraction was indistinguishable from the cytosolic PLC-I when their mixture was analyzed on SDS-PAGE. In addition, the elution profile of tryptic peptides derived from the 150 KDa particulate form was identical to that of cytosolic PLC-I. This result indicates that PLC-I is reversibly associated to membranes.

  19. Complexes of dendrimers with bovine serum albumin.

    PubMed

    Mandeville, J S; Tajmir-Riahi, H A

    2010-02-01

    We report the complexation of bovine serum albumin (BSA) with several dendrimers of different compositions mPEG-PAMAM (G3), mPEG-PAMAM (G4), and PAMAM (G4) at physiological conditions using constant protein concentration and various dendrimer contents. FTIR, CD, and fluorescence spectroscopic methods were used to analyze polymer binding mode, the binding constant, and the effects of dendrimer complexation on BSA stability and conformation. Structural analysis showed that dendrimers bind BSA via hydrophilic and hydrophobic interactions with a number of bound polymers (n): 1.30 for mPEG-PAMAM-G3, 1.30 for mPEG-PAMAM-G4, and 1.0 for PAMAM-G4. The polymer-BSA binding constants were K(mPEG-G3) = 5.0 (+/-0.8) x 10(3) M(-1), K(mPEG-G4) = 1.0 (+/-0.3) x 10(4) M(-1), and K(PAMAM-G4) = 1.1 (+/-0.4) x 10(4) M(-1). Dendrimer binding altered BSA conformation with a major reduction of alpha-helix and an increase in random coil and turn structures, indicating a partial protein unfolding. PMID:20085247

  20. Spatial and temporal epidemiology of bovine trichomoniasis and bovine genital campylobacteriosis in La Pampa province (Argentina).

    PubMed

    Molina, L; Perea, J; Meglia, G; Angón, E; García, A

    2013-07-01

    The venereal diseases bovine trichomoniasis (BT) and bovine genital campylobacteriosis (BCG) cause economic losses in endemic areas like La Pampa province in Argentina, where beef cattle are usually managed extensively. This study used data compiled under a Provincial Programme for the Control and Eradication of BT and BGC (PCE) to determine the spatio-temporal distribution of these diseases and identify spatial clusters. The study population comprised 29,178 non-virgin bulls drawn from 3766 herds, tested for BT and BGC in 2010. Preputial smegma samples were cultured for BT detection, while BGC was diagnosed by direct immunofluorescence testing of these samples. Campylobacter fetus infection was detected in 1.5% of bulls and 2.3% of herds, and Tritrichomonas foetus infection was found in 1.1% of bulls and 5.1% of herds. The proportion of positive tests was highest in February for BT, while in April it was highest for BCG, and was inversely related to the number of tests, which was greatest during the breeding season (spring). An elliptical spatial cluster of high risk for BGC and a circular cluster for BT were both identified in the south of La Pampa province, which could not be explained by cattle herd density. The spatial and temporal patterns identified in this study provide baseline data for monitoring the success of BT and BGC control activities in La Pampa. PMID:23518247

  1. Bovine leucosis virus contamination of a vaccine produced in vivo against bovine babesiosis and anaplasmosis.

    PubMed

    Rogers, R J; Dimmock, C K; de Vos, A J; Rodwell, B J

    1988-09-01

    Contamination of a batch of tick fever (babesiosis and anaplasmosis) vaccine with bovine leucosis virus (BLV) was detected when a herd, in the final stages of an enzootic bovine leucosis (EBL) accreditation program, developed a large number of seropositive cattle following use of tick fever vaccine. Investigations incriminated a single calf used to produce Anaplasma centrale vaccine from which 13,959 doses were distributed. The failure of this calf to give a positive agar gel immunodiffusion (AGID) test before use was not fully explained. A total of 22,627 cattle from 111 herds receiving contaminated vaccine was tested to validate claims for compensation. Results showed infection rates of 62% and 51.8% in vaccinated dairy and beef cattle, respectively, compared with 6.1% and 1.5% in non-vaccinated cattle in the same herds. The results also indicated that infection did not spread from vaccinated to non-vaccinated in-contact cattle. Heavy reliance is now placed on purchase of calves for vaccine production from EBL accredited-free herds and on transmission tests from the calves to sheep to prevent a recurrence of contamination. The need for a BLV antigen detection test, with the sensitivity of the sheep transmission test but simpler and faster to perform, is evident. PMID:2847702

  2. Fraction of bovine leukemia virus-infected dairy cattle developing enzootic bovine leukosis.

    PubMed

    Tsutsui, Toshiyuki; Kobayashi, Sota; Hayama, Yoko; Yamamoto, Takehisa

    2016-02-01

    Enzootic bovine leucosis (EBL) is a transmissible disease caused by the bovine leukemia virus that is prevalent in cattle herds in many countries. Only a small fraction of infected animals develops clinical symptoms, such as malignant lymphosarcoma, after a long incubation period. In the present study, we aimed to determine the fraction of EBL-infected dairy cattle that develop lymphosarcoma and the length of the incubation period before clinical symptoms emerge. These parameters were determined by a mathematical modeling approach based on the maximum-likelihood estimation method, using the results of a nationwide serological survey of prevalence in cattle and passive surveillance records. The best-fit distribution to estimate the disease incubation period was determined to be the Weibull distribution, with a median and average incubation period of 7.0 years. The fraction of infected animals developing clinical disease was estimated to be 1.4% with a 95% confidence interval of 1.2-1.6%. The parameters estimated here contribute to an examination of efficient control strategies making quantitative evaluation available. PMID:26754928

  3. Cellular Requirements for Bovine Immunodeficiency Virus Vif-Mediated Inactivation of Bovine APOBEC3 Proteins

    PubMed Central

    Zhang, Wenyan; Wang, Hong; Li, Zhaolong; Liu, Xin; Liu, Guanchen; Harris, Reuben S.

    2014-01-01

    ABSTRACT Human immunodeficiency virus type 1 (HIV-1) and simian immunodeficiency virus (SIV) viral infectivity factor (Vif) form a CRL5 E3 ubiquitin ligase complex to suppress virus restriction by host APOBEC3 (A3) proteins. The primate lentiviral Vif complex is composed of the unique cofactor core binding factor β (CBF-β) and canonical ligase components Cullin 5 (CUL5), Elongin B/C (ELOB/C), and RBX2. However, the mechanism by which the Vif protein of the related lentivirus bovine immunodeficiency virus (BIV) overcomes its host A3 proteins is less clear. In this study, we show that BIV Vif interacts with Cullin 2 (CUL2), ELOB/C, and RBX1, but not with CBF-β or CUL5, to form a CRL2 E3 ubiquitin ligase and degrade the restrictive bovine A3 proteins (A3Z2Z3 and A3Z3). RNA interference-mediated knockdown of ELOB or CUL2 inhibited BIV Vif-mediated degradation of these A3 proteins, whereas knockdown of CUL5 or CBF-β did not. BIV Vif with mutations in the BC box (Vif SLQ-AAA) or putative VHL box (Vif YI-AA), which cannot interact with ELOB/C or CUL2, respectively, lost the ability to counteract bovine A3 proteins. Moreover, CUL2 and UBE2M dominant negative mutants competitively inhibited the BIV Vif-mediated degradation mechanism. Thus, although the general strategy for inhibiting A3 proteins is conserved between HIV-1/SIV and BIV, the precise mechanisms can differ substantially, with only the HIV-1/SIV Vif proteins requiring CBF-β as a cofactor, HIV-1/SIV Vif using CUL5-RBX2, and BIV Vif using CUL2-RBX1. IMPORTANCE Primate lentivirus HIV-1 and SIV Vif proteins form a ubiquitin ligase complex to target host antiviral APOBEC3 proteins for degradation. However, the mechanism by which the nonprimate lentivirus BIV Vif inhibits bovine APOBEC3 proteins is unclear. In the present study, we determined the mechanism for BIV Vif-mediated degradation of bovine APOBEC3 proteins and found that it differs from the mechanism of HIV-1/SIV Vif by being CBF-β independent and

  4. FTO gene variants are associated with growth and carcass traits in cattle.

    PubMed

    Jevsinek Skok, D; Kunej, T; Kovac, M; Malovrh, S; Potocnik, K; Petric, N; Zgur, S; Dovc, P; Horvat, S

    2016-04-01

    An important aim in animal breeding is the improvement of growth and meat quality traits. Previous studies have demonstrated that genetic variants in the fat mass and obesity associated (FTO) gene have a relatively large effect on human obesity as well as on body composition in rodents and, more recently, in livestock. Here, we examined the effects of the FTO gene variants on growth and carcass traits in the Slovenian population of Simmental (SS) and Brown (SB) cattle. To validate and identify new polymorphisms, we used sequencing, PCR-RFLP analysis and TaqMan assays in the SS breed and FTO gene variants data from the Illumina BovineSNP50 v1 array for the SB breed. Sequencing of the eight samples of progeny-tested SS sires detected 108 single nucleotide polymorphisms (SNPs) in the bovine FTO gene. Statistical analyses between growth and carcass traits and 34 FTO polymorphisms revealed significant association of FTO variants with lean meat percentage in both breeds. Additionally, FTO SNPs analyzed in SS cattle were associated with fat percentage, bone weight and live weight at slaughter. The FTO gene can thus be regarded as a candidate gene for the marker-assisted selection programs in our and possibly other populations of cattle. Future studies in cattle might reveal novel roles for the FTO gene in shaping carcass traits in livestock species as well as body composition control in other mammals. PMID:26708680

  5. Food Safety: Bovine Spongiform Encephalopathy (Mad Cow Disease).

    PubMed

    Acheson, David W. K.

    2002-01-01

    Bovine spongiform encephalopathy is just one of a group of diseases known as transmissible spongiform encephalopathies. Only recently has it become recognized that transmissible spongiform encephalopathies are likely due to proteins known as prions. Although it has been recognized that transmissible spongiform encephalopathies may readily spread within species, the recent observations that bovine spongiform encephalopathy in cattle may have originated from another transmissible spongiform encephalopathy in sheep, known as scrapie, is cause for concern. Further, bovine spongiform encephalopathy has now been strongly linked with a universally fatal human neurologic disease known as new variant Creutzfeldt-Jakob disease. Currently the only approach to preventing bovine spongiform encephalopathy, and subsequent new variant Creutzfeldt-Jakob disease in humans, from ingestion of bovine spongiform encephalopathy-infected material is to avoid consumption of contaminated food. Little can be done to treat food that will destroy prions and leave a palatable product. At this stage we are continuing to learn about transmissible spongiform encephalopathies and their implications on human health. This is an ever-changing situation and has an unpredictable element in terms of the extent of the current outbreaks in England and other parts of Europe. PMID:11984426

  6. Epidemiology of Bovine Mastitis in Cows of Dharwad District

    PubMed Central

    Kurjogi, Mahantesh M.; Kaliwal, Basappa B.

    2014-01-01

    Bovine mastitis is very common in cows of both developed and developing countries. The prevalence of clinical and subclinical mastitis (SCM) varies from region to region. Hence, the present study was carried out to determine the prevalence of mastitis using three diagnostic tests by considering different risk factors like age, lactation, breed, season, quarters, and herd. The results showed that surf field mastitis test (SFMT) is the most sensitive test for diagnosis of bovine mastitis, the older age and cows with later part of lactation period were more prone to bovine mastitis, and exotic breeds like Holstein freshen (HF) were more susceptible to bovine mastitis. The highest incidence of mastitis was recorded in monsoon season. The prevalence of subclinical and clinical mastitis was more in single and two quarters, respectively, and the rate of bovine mastitis was more in unorganized herds. The study concluded that SCM is directly associated with age, lactation period, and environmental factors of the cow and clinical mastitis is more associated with breed of the cow and environmental conditions. PMID:27382623

  7. Structure of an Enteric Pathogen, Bovine Parvovirus

    PubMed Central

    Kailasan, Shweta; Halder, Sujata; Gurda, Brittney; Bladek, Heather; Chipman, Paul R.; McKenna, Robert; Brown, Kevin

    2014-01-01

    ABSTRACT Bovine parvovirus (BPV), the causative agent of respiratory and gastrointestinal disease in cows, is the type member of the Bocaparvovirus genus of the Parvoviridae family. Toward efforts to obtain a template for the development of vaccines and small-molecule inhibitors for this pathogen, the structure of the BPV capsid, assembled from the major capsid viral protein 2 (VP2), was determined using X-ray crystallography as well as cryo-electron microscopy and three-dimensional image reconstruction (cryo-reconstruction) to 3.2- and 8.8-Å resolutions, respectively. The VP2 region ordered in the crystal structure, from residues 39 to 536, conserves the parvoviral eight-stranded jellyroll motif and an αA helix. The BPV capsid displays common parvovirus features: a channel at and depressions surrounding the 5-fold axes and protrusions surrounding the 3-fold axes. However, rather than a depression centered at the 2-fold axes, a raised surface loop divides this feature in BPV. Additional observed density in the capsid interior in the cryo-reconstructed map, compared to the crystal structure, is interpreted as 10 additional N-terminal residues, residues 29 to 38, that radially extend the channel under the 5-fold axis, as observed for human bocavirus 1 (HBoV1). Surface loops of various lengths and conformations extend from the core jellyroll motif of VP2. These loops confer the unique surface topology of the BPV capsid, making it strikingly different from HBoV1 as well as the type members of other Parvovirinae genera for which structures have been determined. For the type members, regions structurally analogous to those decorating the BPV capsid surface serve as determinants of receptor recognition, tissue and host tropism, pathogenicity, and antigenicity. IMPORTANCE Bovine parvovirus (BPV), identified in the 1960s in diarrheic calves, is the type member of the Bocaparvovirus genus of the nonenveloped, single-stranded DNA (ssDNA) Parvoviridae family. The recent

  8. A Major Gene for Bovine Ovulation Rate

    PubMed Central

    Kirkpatrick, Brian W.; Morris, Chris A.

    2015-01-01

    Half-sib daughters sired by a bull believed to be a carrier of a major gene for high ovulation rate were evaluated for ovulation rate and genotyped in an effort to both test the hypothesis of segregation of a major gene and to map the gene’s location. A total of 131 daughters were produced over four consecutive years at a University of Wisconsin-Madison research farm. All were evaluated for ovulation rate over an average of four estrous cycles using transrectal ultrasonography. The sire and all daughters were genotyped using a 3K SNP chip and the genotype and phenotype data were used in a linkage analysis. Subsequently, daughters recombinant within the QTL region and the sire were genotyped successively with 50K and 777K SNP chips to refine the location of the causative polymorphism. Positional candidate genes within the fine-mapped region were examined for polymorphism by Sanger sequencing of PCR amplicons encompassing coding and 5’ and 3’ flanking regions of the genes. Sire DNA was used as template in the PCR reactions. Strong evidence of a major gene for ovulation rate was observed (p<1x10-28) with the gene localized to bovine chromosome 10. Fine-mapping subsequently reduced the location to a 1.2 Mb region between 13.6 and 14.8 Mb on chromosome 10. The location identified does not correspond to that for any previously identified major gene for ovulation rate. This region contains three candidate genes, SMAD3, SMAD6 and IQCH. While candidate gene screening failed to identify the causative polymorphism, three polymorphisms were identified that can be used as a haplotype to track inheritance of the high ovulation rate allele in descendants of the carrier sire. PMID:26046917

  9. Quantitative Risk Assessment of Bovine Spongiform Encephalopathy

    NASA Astrophysics Data System (ADS)

    Tsutsui, Toshiyuki; Kasuga, Fumiko

    Bovine spongiform encephalopathy (BSE) is a progressive neurological disease of cattle affecting the central nervous system and was first diagnosed in the United Kingdom (UK) in 1986 (Wells et al., 1987). This disease is one of the transmissible spongiform encephalopathy (TSE) which includes Creutzfeldt-Jakob disease (CJD) in humans and scrapie in sheep. The causative agent of TSE is considered to be an abnormal form of prion protein. However, the details of its pathogenic mechanism have not been fully identified. Scrapie, which causes neurological symptoms in sheep and goats, has existed in the UK for 200 years (Hoinville, 1996) and spread across the rest of the world in the 1900s (Detwiler & Baylis, 2003). There has been no report so far that scrapie can be transmitted to humans. Initially, BSE was also considered as a disease affecting only animals. However, a variant type of Creutzfeldt-Jakob disease (vCJD) was first reported in the UK, and exposure to a BSE agent was suspected (Collinge, Sidle, Meads, Ironside, & Hill, 1996). vCJD is clinically and pathologically different from the sporadic type of CJD, and age at clinical onset of vCJD is younger than sporadic type (Will et al., 1996). Since the UK government announced the possible association between BSE and vCJD in 1996, BSE has become a huge public health concern all over the world. Of particular concern about vCJD, the fatal disease in younger age, distorted consumer confidence in beef safety, and as a result reduced beef consumption has been seen in many BSE-affected countries.

  10. Morphological classification of bovine ovarian follicles.

    PubMed

    Rodgers, R J; Irving-Rodgers, H F

    2010-02-01

    Follicle classification is an important aid to the understanding of follicular development and atresia. Some bovine primordial follicles have the classical primordial shape, but ellipsoidal shaped follicles with some cuboidal granulosa cells at the poles are far more common. Preantral follicles have one of two basal lamina phenotypes, either a single aligned layer or one with additional layers. In antral follicles <5 mm diameter, half of the healthy follicles have columnar shaped basal granulosa cells and additional layers of basal lamina, which appear as loops in cross section ('loopy'). The remainder have aligned single-layered follicular basal laminas with rounded basal cells, and contain better quality oocytes than the loopy/columnar follicles. In sizes >5 mm, only aligned/rounded phenotypes are present. Dominant and subordinate follicles can be identified by ultrasound and/or histological examination of pairs of ovaries. Atretic follicles <5 mm are either basal atretic or antral atretic, named on the basis of the location in the membrana granulosa where cells die first. Basal atretic follicles have considerable biological differences to antral atretic follicles. In follicles >5 mm, only antral atresia is observed. The concentrations of follicular fluid steroid hormones can be used to classify atresia and distinguish some of the different types of atresia; however, this method is unlikely to identify follicles early in atresia, and hence misclassify them as healthy. Other biochemical and histological methods can be used, but since cell death is a part of normal homoeostatis, deciding when a follicle has entered atresia remains somewhat subjective. PMID:19786400

  11. Dynamics of perinatal bovine leukemia virus infection

    PubMed Central

    2014-01-01

    Background Bovine leukemia virus (BLV) is highly endemic in many countries, including Argentina. As prevention of the spread from infected animals is of primary importance in breaking the cycle of BLV transmission, it is important to know the pathophysiology of BLV infection in young animals, as they are the main source of animal movement. In this work, we determined the proviral load and antibody titers of infected newborn calves from birth to first parturition (36 months). Results All calves under study were born to infected dams with high proviral load (PVL) in blood and high antibody titers and detectable provirus in the colostrum. The PVL for five out of seven calves was low at birth. All animals reached PVLs of more than 1% infected peripheral blood mononuclear cells (PBMCs), three at 3 months, one at 6 months, and one at 12 months. High PVLs persisted until the end of the study, and, in two animals, exceeded one BLV copy per cell. Two other calves maintained a high PVL from birth until the end of the study. Antibody titers were 32 or higher in the first sample from six out of seven calves. These decayed at 3–6 months to 16 or lower, and then increased again after this point. Conclusions Calves infected during the first week of life could play an active role in early propagation of BLV to susceptible animals, since their PVL raised up during the first 12 months and persist as high for years. Early elimination could help to prevent transmission to young susceptible animals and to their own offspring. To our knowledge, this is the first study of the kinetics of BLV proviral load and antibody titers in newborn infected calves. PMID:24708791

  12. Prion biology relevant to bovine spongiform encephalopathy.

    PubMed

    Novakofski, J; Brewer, M S; Mateus-Pinilla, N; Killefer, J; McCusker, R H

    2005-06-01

    Bovine spongiform encephalopathy (BSE) and chronic wasting disease (CWD) of deer and elk are a threat to agriculture and natural resources, as well as a human health concern. Both diseases are transmissible spongiform encephalopathies (TSE), or prion diseases, caused by autocatalytic conversion of endogenously encoded prion protein (PrP) to an abnormal, neurotoxic conformation designated PrPsc. Most mammalian species are susceptible to TSE, which, despite a range of species-linked names, is caused by a single highly conserved protein, with no apparent normal function. In the simplest sense, TSE transmission can occur because PrPsc is resistant to both endogenous and environmental proteinases, although many details remain unclear. Questions about the transmission of TSE are central to practical issues such as livestock testing, access to international livestock markets, and wildlife management strategies, as well as intangible issues such as consumer confidence in the safety of the meat supply. The majority of BSE cases seem to have been transmitted by feed containing meat and bone meal from infected animals. In the United Kingdom, there was a dramatic decrease in BSE cases after neural tissue and, later, all ruminant tissues were banned from ruminant feed. However, probably because of heightened awareness and widespread testing, there is growing evidence that new variants of BSE are arising "spontaneously," suggesting ongoing surveillance will continue to find infected animals. Interspecies transmission is inefficient and depends on exposure, sequence homology, TSE donor strain, genetic polymorphism of the host, and architecture of the visceral nerves if exposure is by an oral route. Considering the low probability of interspecies transmission, the low efficiency of oral transmission, and the low prion levels in nonnervous tissues, consumption of conventional animal products represents minimal risk. However, detection of rare events is challenging, and TSE

  13. Development of a rapid and sensitive polymerase chain reaction assay for detection of bovine herpesvirus type 1 in bovine semen.

    PubMed Central

    van Engelenburg, F A; Maes, R K; van Oirschot, J T; Rijsewijk, F A

    1993-01-01

    We developed a polymerase chain reaction (PCR) assay to detect bovine herpesvirus type 1 (BHV-1) in bovine semen. Since bovine semen contains components that inhibit PCR amplification, a protocol was developed to purify BHV-1 DNA from bovine semen. To identify failures of PCR amplification, we used an internal control template that was coamplified by the same PCR primers. When separated fractions of BHV-1-contaminated semen were analyzed by the PCR, we found that more than 90% of the BHV-1 DNA was present in a pooled fraction consisting of seminal fluid, nonsperm cells, and virus adsorbed to spermatozoa. By using this fraction, three to five molecules of BHV-1 DNA in 50 microliters of bovine semen could be detected. A pilot study to compare this PCR assay with the routinely used virus isolation method showed that this PCR assay is 2- to 100-fold more sensitive. In addition, the results of the PCR assay are available in 1 day, whereas the virus isolation method takes 7 days. Therefore, the PCR assay may be a good alternative to the virus isolation method. Images PMID:8308103

  14. Dynamic compressive properties of bovine knee layered tissue

    NASA Astrophysics Data System (ADS)

    Nishida, Masahiro; Hino, Yuki; Todo, Mitsugu

    2015-09-01

    In Japan, the most common articular disease is knee osteoarthritis. Among many treatment methodologies, tissue engineering and regenerative medicine have recently received a lot of attention. In this field, cells and scaffolds are important, both ex vivo and in vivo. From the viewpoint of effective treatment, in addition to histological features, the compatibility of mechanical properties is also important. In this study, the dynamic and static compressive properties of bovine articular cartilage-cancellous bone layered tissue were measured using a universal testing machine and a split Hopkinson pressure bar method. The compressive behaviors of bovine articular cartilage-cancellous bone layered tissue were examined. The effects of strain rate on the maximum stress and the slope of stress-strain curves of the bovine articular cartilage-cancellous bone layered tissue were discussed.

  15. Immunogenicity of a modified-live virus vaccine against bovine viral diarrhea virus types 1 and 2, infectious bovine rhinotracheitis virus, bovine parainfluenza-3 virus, and bovine respiratory syncytial virus when administered intranasally in young calves.

    PubMed

    Xue, Wenzhi; Ellis, John; Mattick, Debra; Smith, Linda; Brady, Ryan; Trigo, Emilio

    2010-05-14

    The immunogenicity of an intranasally-administered modified-live virus (MLV) vaccine in 3-8 day old calves was evaluated against bovine viral diarrhea virus (BVDV) types 1 and 2, infectious bovine rhinotracheitis (IBR) virus, parainfluenza-3 (PI-3) virus and bovine respiratory syncytial virus (BRSV). Calves were intranasally vaccinated with a single dose of a multivalent MLV vaccine and were challenged with one of the respective viruses three to four weeks post-vaccination in five separate studies. There was significant sparing of diseases in calves intranasally vaccinated with the MLV vaccine, as indicated by significantly fewer clinical signs, lower rectal temperatures, reduced viral shedding, greater white blood cell and platelet counts, and less severe pulmonary lesions than control animals. This was the first MLV combination vaccine to demonstrate efficacy against BVDV types 1 and 2, IBR, PI-3 and BRSV in calves 3-8 days of age. PMID:20381643

  16. Molecular cloning and expression vector construction of bovine TRIM28.

    PubMed

    Ma, X; Zhai, Z C; Zhang, M L; Song, B H; Zhu, Y R; Yang, S B; Dong, X Q; Su, L Y; Wang, C F; Ma, H X; Luan, W M

    2016-01-01

    The bovine TRIM28 gene was amplified from ovary tissue by using RT-PCR. The TRIM28 gene was inserted into the eukaryotic expression vector pIRES2-EGFP and transfected into bovine fetal fibroblasts by using Lipofectamine 3000. TRIM28 mRNA and protein were detected by fluorescence microscope and western blotting. The results showed that the full length of TRIM28 was cloned and pIRES2-EGFP-TRIM28 was constructed successfully. EGFP expression was observed, and the pIRES2-EGFP-TRIM28 transfected group expressed more TRIM28 protein than that by the pIRES2-EGFP group. The TIMR28 gene has been successfully transferred into bovine fetal fibroblasts. PMID:27420979

  17. Isolation and characterization of four antibacterial peptides from bovine hemoglobin.

    PubMed

    Nedjar-Arroume, Naima; Dubois-Delval, Véronique; Miloudi, Khalil; Daoud, Rachid; Krier, François; Kouach, Mostafa; Briand, Gilbert; Guillochon, Didier

    2006-09-01

    Peptic digestion of bovine hemoglobin at low degree of hydrolysis yields several intermediate peptide fractions after separation by reversed phase HPLC exhibiting antibacterial activity against Micrococcus luteus A270, Listeria innocua, Escherichia coli, and Salmonella enteritidis. From these fractions, four new antibacterial peptides were isolated and analyzed by ESI-MS/MS. Three of these peptides correspond to fragments of the alpha-chain of bovine hemoglobin: alpha107-141, alpha137-141, and alpha133-141, and one peptide to the beta-chain: beta126-145. The minimum inhibitory concentrations (MIC) of these peptides towards the four strains and their hemolytic activity towards bovine erythrocytes were determined. PMID:16730859

  18. Bundling of bovine and brine shrimp (Artemia) microtubules in vitro.

    PubMed

    MacRae, T H

    1984-06-01

    Cell-free extracts from embryos of the brine shrimp (Artemia) induced bundling of bovine microtubules assembled in the presence of glycerol and Mg++. Sedimentation of microtubules through sucrose cushions and subsequent electrophoresis revealed that bundling occurred independently of accessory proteins tightly bound to the microtubules. Bovine microtubules containing microtubule-associated proteins (MAPS) or assembled with taxol did not bundle. The unusual polymerization properties of homogeneous Artemia tubulin, bundling in the absence of added factors and the small number of microtubules assembled in crude embryo extracts upon addition of taxol precluded a complete comparative study of Artemia and bovine microtubule bundling. Interesting properties of the in vitro assembly of Artemia microtubules were, however, elaborated and putative Artemia MAPs were observed as a consequence of the work with brine shrimp embryos. PMID:6744423

  19. Control of Bovine Mastitis: Old and Recent Therapeutic Approaches.

    PubMed

    Gomes, Fernanda; Henriques, Mariana

    2016-04-01

    Mastitis is defined as the inflammatory response resulting of the infection of the udder tissue and it is reported in numerous species, namely in domestic dairy animals. This pathology is the most frequent disease of dairy cattle and can be potentially fatal. Mastitis is an economically important pathology associated with reduced milk production, changes in milk composition and quality, being considered one of the most costly to dairy industry. Therefore, the majority of research in the field has focused on control of bovine mastitis and many efforts are being made for the development of new and effective anti-mastitis drugs. Antibiotic treatment is an established component of mastitis control programs; however, the continuous search for new therapeutic alternatives, effective in the control and treatment of bovine mastitis, is urgent. This review will provide an overview of some conventional and emerging approaches in the management of bovine mastitis' infections. PMID:26687332

  20. Current ante-mortem techniques for diagnosis of bovine tuberculosis.

    PubMed

    Bezos, Javier; Casal, Carmen; Romero, Beatriz; Schroeder, Bjoern; Hardegger, Roland; Raeber, Alex J; López, Lissette; Rueda, Paloma; Domínguez, Lucas

    2014-10-01

    Bovine tuberculosis (TB), mainly caused by Mycobacterium bovis, is a zoonotic disease with implications for Public Health and having an economic impact due to decreased production and limitations to the trade. Bovine TB is subjected to official eradication campaigns mainly based on a test and slaughter policy using diagnostic assays based on the cell-mediated immune response as the intradermal tuberculin test and the gamma-interferon (IFN-γ) assay. Moreover, several diagnostic assays based on the detection of specific antibodies (Abs) have been developed in the last few years with the aim of complementing the current diagnostic techniques in the near future. This review provides an overview of the current ante-mortem diagnostic tools for diagnosis of bovine TB regarding historical background, methodologies and sensitivity (Se) and specificity (Sp) obtained in previous studies under different epidemiological situations. PMID:24768355

  1. Expression of β-Defensin Genes in Bovine Alveolar Macrophages

    PubMed Central

    Ryan, Lisa K.; Rhodes, Janice; Bhat, Meenakshi; Diamond, Gill

    1998-01-01

    Bovine alveolar macrophages (BAM) were examined for the expression of β-defensins and to determine whether their expression could be upregulated by bacterial lipopolysaccharide (LPS), as observed with β-defensins expressed in bovine tracheal epithelial cells. Four β-defensins were expressed constitutively in BAM, with bovine neutrophil β-defensin (BNBD)-4 and BNBD-5 being the most predominant. This is the first evidence of β-defensin gene expression in a mature myeloid cell. LPS had no effect on β-defensin expression in BAM, even though tumor necrosis factor alpha (TNF-α) production was induced. Nonbacterial inflammatory particles had little effect on β-defensin gene expression or TNF-α production in BAM. We hypothesize that constitutively expressed β-defensins of alveolar macrophages may have a role in lung host defense. PMID:9453661

  2. Sequence analysis of a bovine rhinovirus type 1 strain RS3x

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bovine rhinoviruses, known to cause clinical and subclinical upper respiratory infections in bovines worldwide, include three serotypes. Bovine rhinovirus (BRV) 1, 2 and 3 were originally classified as tentative members of the genus Rhinovirus (family Picornaviridae), however, in 2008 this genus was...

  3. Identification of genetic variation and putative regulatory regions in bovine CARD15

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Mutations in caspase recruitment domain 15 (CARD15) are associated with susceptibility to Crohn's disease and Blau Syndrome. We performed comparative analyses of the bovine, murine, and human CARD15 transcripts to elucidate functionality of bovine CARD15 and examine its potential role in bovine dise...

  4. 70 FR 18251 - Bovine Spongiform Encephalopathy; Minimal-Risk Regions and Importation of Commodities; Finding of...

    Code of Federal Regulations, 2014 CFR

    2005-04-08

    ... Agriculture Animal and Plant Health Inspection Service 9 CFR Part 93, et al. Bovine Spongiform Encephalopathy... Implementing Procedures (7 CFR part 372). Bovine Spongiform Encephalopathy; Minimal-Risk Regions and... pointed to the four confirmed cases of bovine spongiform encephalopathy (BSE) in cows of Canadian...

  5. Construction and validation of a Bovine Innate Immune Microarray

    PubMed Central

    Donaldson, Laurelea; Vuocolo, Tony; Gray, Christian; Strandberg, Ylva; Reverter, Antonio; McWilliam, Sean; Wang, YongHong; Byrne, Keren; Tellam, Ross

    2005-01-01

    Background Microarray transcript profiling has the potential to illuminate the molecular processes that are involved in the responses of cattle to disease challenges. This knowledge may allow the development of strategies that exploit these genes to enhance resistance to disease in an individual or animal population. Results The Bovine Innate Immune Microarray developed in this study consists of 1480 characterised genes identified by literature searches, 31 positive and negative control elements and 5376 cDNAs derived from subtracted and normalised libraries. The cDNA libraries were produced from 'challenged' bovine epithelial and leukocyte cells. The microarray was found to have a limit of detection of 1 pg/μg of total RNA and a mean slide-to-slide correlation co-efficient of 0.88. The profiles of differentially expressed genes from Concanavalin A (ConA) stimulated bovine peripheral blood lymphocytes were determined. Three distinct profiles highlighted 19 genes that were rapidly up-regulated within 30 minutes and returned to basal levels by 24 h; 76 genes that were up-regulated between 2–8 hours and sustained high levels of expression until 24 h and 10 genes that were down-regulated. Quantitative real-time RT-PCR on selected genes was used to confirm the results from the microarray analysis. The results indicate that there is a dynamic process involving gene activation and regulatory mechanisms re-establishing homeostasis in the ConA activated lymphocytes. The Bovine Innate Immune Microarray was also used to determine the cross-species hybridisation capabilities of an ovine PBL sample. Conclusion The Bovine Innate Immune Microarray has been developed which contains a set of well-characterised genes and anonymous cDNAs from a number of different bovine cell types. The microarray can be used to determine the gene expression profiles underlying innate immune responses in cattle and sheep. PMID:16176586

  6. Predictive qualitative risk model of bovine rabies occurrence in Brazil.

    PubMed

    Braga, Guilherme Basseto; Grisi-Filho, José Henrique Hildebrand; Leite, Bruno Meireles; de Sena, Elaine Fátima; Dias, Ricardo Augusto

    2014-03-01

    Bovine rabies remains endemic in Brazil and despite control efforts, the disease still spreads insidiously. The main vector is the hematophagous bat, Desmodus rotundus. The present work aimed to create a predictive qualitative model of the occurrence of bovine rabies in each municipality in 25 of the 27 Brazilian States. The risk of rabies transmission from bats to bovine was estimated using decision-tree models of receptivity and vulnerability. Questionnaires, which covered a number of questions related to the surveillance of possible risk factors, such as bovine rabies outbreaks in the previous year, the presence of bat roosts, bat rabies positivity and environmental changes, were sent to the local veterinary units of each State. The bovine density and geomorphologic features were obtained from national databases and geographic information systems. Of the 433 municipalities presenting bovine rabies outbreaks in 2010, 178 (41.1%) were classified by the model as high risk, 212 (49.0%) were classified as moderate risk, 25 (5.8%) were classified as low risk, whereas the risk was undetermined in 18 municipalities (4.1%). An ROC curve was built to determine if the risk evaluated by the model could adequately discriminate between municipalities with and without rabies occurrence in future years. The risk estimator for the year 2011 was classified as moderately accurate. In the future, these models could allow the targeting of rabies control efforts, with the adoption of control measures directed to the higher risk locations and the optimization of the field veterinary staff deployment throughout the country. Additionally, efforts must be made to encourage continuous surveillance of risk factors. PMID:24433635

  7. The use of fetal bovine serum: ethical or scientific problem?

    PubMed

    Jochems, Carlo E A; van der Valk, Jan B F; Stafleu, Frans R; Baumans, Vera

    2002-01-01

    Fetal bovine serum (FBS) is a common component of animal cell culture media. It is harvested from bovine fetuses taken from pregnant cows during slaughter. FBS is commonly harvested by means of a cardiac puncture without any form of anaesthesia. Fetuses are probably exposed to pain and/or discomfort, so the current practice of fetal blood harvesting is inhumane. Apart from moral concerns, several scientific and technical problems exist with regard to the use of FBS in cell culture. Efforts should be made to reduce the use of FBS or, preferably, to replace it with synthetic alternatives. PMID:11971757

  8. [Culture and control of cells producing bovine leukemia virus].

    PubMed

    Granátová, M

    1987-10-01

    In the field surveys of the occurrence of enzootic bovine leucosis caused by the bovine leucosis virus (BLV), the identification of positive animals is based on the detection of specific antiviral antibodies by serological methods. The reliability of these tests (particularly their sensitivity and specificity) depends on the quality of the virus antigen. The preparation of the antigen is based on the cultivation of BLV virus in cultures of the FLS cell line. A modified procedure of preparing the BLV antigen in the FLS cell culture is described, along with the control of its production by the immunoperoxidase test. PMID:2827363

  9. Bovine Viral Diarrhea Virus-Associated Disease in Feedlot Cattle.

    PubMed

    Larson, Robert L

    2015-11-01

    Bovine viral diarrhea virus (BVDv) is associated with bovine respiratory disease complex and other diseases of feedlot cattle. Although occasionally a primary pathogen, BVDv's impact on cattle health is through the immunosuppressive effects of the virus and its synergism with other pathogens. The simple presence or absence of BVDv does not result in consistent health outcomes because BVDv is only one of many risk factors that contribute to disease syndromes. Current interventions have limitations and the optimum strategy for their uses to limit the health, production, and economic costs associated with BVDv have to be carefully considered for optimum cost-effectiveness. PMID:26210765

  10. Accuracy of predicting genomic breeding values for residual feed intake in Angus and Charolais beef cattle.

    PubMed

    Chen, L; Schenkel, F; Vinsky, M; Crews, D H; Li, C

    2013-10-01

    In beef cattle, phenotypic data that are difficult and/or costly to measure, such as feed efficiency, and DNA marker genotypes are usually available on a small number of animals of different breeds or populations. To achieve a maximal accuracy of genomic prediction using the phenotype and genotype data, strategies for forming a training population to predict genomic breeding values (GEBV) of the selection candidates need to be evaluated. In this study, we examined the accuracy of predicting GEBV for residual feed intake (RFI) based on 522 Angus and 395 Charolais steers genotyped on SNP with the Illumina Bovine SNP50 Beadchip for 3 training population forming strategies: within breed, across breed, and by pooling data from the 2 breeds (i.e., combined). Two other scenarios with the training and validation data split by birth year and by sire family within a breed were also investigated to assess the impact of genetic relationships on the accuracy of genomic prediction. Three statistical methods including the best linear unbiased prediction with the relationship matrix defined based on the pedigree (PBLUP), based on the SNP genotypes (GBLUP), and a Bayesian method (BayesB) were used to predict the GEBV. The results showed that the accuracy of the GEBV prediction was the highest when the prediction was within breed and when the validation population had greater genetic relationships with the training population, with a maximum of 0.58 for Angus and 0.64 for Charolais. The within-breed prediction accuracies dropped to 0.29 and 0.38, respectively, when the validation populations had a minimal pedigree link with the training population. When the training population of a different breed was used to predict the GEBV of the validation population, that is, across-breed genomic prediction, the accuracies were further reduced to 0.10 to 0.22, depending on the prediction method used. Pooling data from the 2 breeds to form the training population resulted in accuracies increased

  11. Design and Construction of Chimeric VP8-S2 Antigen for Bovine Rotavirus and Bovine Coronavirus

    PubMed Central

    Nasiri, Khadijeh; Nassiri, Mohammadreza; Tahmoorespur, Mojtaba; Haghparast, Alireza; Zibaee, Saeed

    2016-01-01

    Purpose: Bovine Rotavirus and Bovine Coronavirus are the most important causes of diarrhea in newborn calves and in some other species such as pigs and sheep. Rotavirus VP8 subunit is the major determinant of the viral infectivity and neutralization. Spike glycoprotein of coronavirus is responsible for induction of neutralizing antibody response. Methods: In the present study, several prediction programs were used to predict B and T-cells epitopes, secondary and tertiary structures, antigenicity ability and enzymatic degradation sites. Finally, a chimeric antigen was designed using computational techniques. The chimeric VP8-S2 antigen was constructed. It was cloned and sub-cloned into pGH and pET32a(+) expression vector. The recombinant pET32a(+)-VP8-S2 vector was transferred into E.oli BL21CodonPlus (DE3) as expression host. The recombinant VP8-S2 protein was purified by Ni-NTA chromatography column. Results: The results of colony PCR, enzyme digestion and sequencing showed that the VP8-S2 chimeric antigen has been successfully cloned and sub-cloned into pGH and pET32a(+).The results showed that E.coli was able to express VP8-S2 protein appropriately. This protein was expressed by induction of IPTG at concentration of 1mM and it was confirmed by Ni–NTA column, dot-blotting analysis and SDS-PAGE electrophoresis. Conclusion: The results of this study showed that E.coli can be used as an appropriate host to produce the recombinant VP8-S2 protein. This recombinant protein may be suitable to investigate to produce immunoglobulin, recombinant vaccine and diagnostic kit in future studies after it passes biological activity tests in vivo in animal model and or other suitable procedure. PMID:27123423

  12. Microarray chip based identification of a mixed infection of bovine herpesvirus 1 and bovine viral diarrhea 2 from Indian cattle.

    PubMed

    Ratta, Barkha; Yadav, Brijesh Singh; Pokhriyal, Mayank; Saxena, Meeta; Sharma, Bhaskar

    2014-01-01

    Bovine herpesvirus 1 (BHV1) and bovine viral diarrhea virus 2 (BVD2) are endemic in India although no mixed infection with these viruses has been reported from India. We report first mixed infection of these viruses in cattle during routine screening with a microarray chip. 62 of the 69 probes of BHV1 and 42 of the 57 BVD2 probes in the chip gave positive signals for the virus. The virus infections were subsequently confirmed by RT-PCR. We also discuss the implications of these findings. PMID:24026447

  13. Delayed-onset enzootic bovine leukosis possibly caused by superinfection with bovine leukemia virus mutated in the pol gene.

    PubMed

    Watanabe, Tadaaki; Inoue, Emi; Mori, Hiroshi; Osawa, Yoshiaki; Okazaki, Katsunori

    2015-08-01

    Bovine leukemia virus (BLV) is the causative agent of enzootic bovine leucosis (EBL), to which animals are most susceptible at 4-8 years of age. In this study, we examined tumor cells associated with EBL in an 18-year-old cow to reveal that the cells carried at least two different copies of the virus, one of which was predicted to encode a reverse transcriptase (RT) lacking ribonuclease H activity and no integrase. Such a deficient enzyme may exhibit a dominant negative effect on the wild-type RT and cause insufficient viral replication, resulting in delayed tumor development in this cow. PMID:26025155

  14. Bovine spongiform encephalopathy in Sweden: an H-type variant

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bovine spongiform encephalopathy (BSE) had never been detected in Sweden until 2006, when the active surveillance identified a case in a 12-year-old cow. The case was an unusual form since several molecular features of the protease-resistant prion protein (PrP**res) were different from classical BSE...

  15. Activation of bovine oocytes penetrated after germinal vesicle breakdown.

    PubMed

    Abeydeera, L R; Okuda, K; Niwa, K

    1994-11-01

    The present study was designed to examine the ability of bovine oocytes, after germinal vesicle breakdown (GVBD), to be activated by sperm penetration and the sequence of sperm nuclear transformation. Bovine oocytes cultured for 8 h in maturation medium (tissue culture medium TCM-199 containing 10% fetal calf serum) were inseminated in Brackett and Oliphant's medium supplemented with bovine serum albumin (10 mg/ml), caffeine (5 mM) and heparin (10 micrograms/ml). When oocytes were transferred to the maturation medium 8 h after insemination and additionally cultured for 5-40 h at 39 degrees C in 5% CO2 in air, 71-76% of oocytes were penetrated and polyspermy (67-75%) was common. The proportions of penetrated oocytes that were activated significantly increased with the lapse of the additional culture time, reaching 88% and 87% by 25 and 40 h after additional culture, respectively. When compared with unpenetrated oocytes, significantly higher proportions of penetrated oocytes reached metaphase II or beyond 15 and 25 h after additional culture. After penetration, sperm nuclei were transformed into metaphase chromosomes and then to telophase chromosomes before the formation of male pronuclei. These results provide evidence that bovine oocytes acquire the ability to respond to sperm-mediated activation soon after GVBD. PMID:8665157

  16. Bovine colostrum against gut inflammatory lesions in preterm pigs

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bovine colostrum is rich in bioactive factors and may prevent necrotizing enterocolitis (NEC) in pre-term neonates. We hypothesized that both native and sterilized, heat-treated colostrum protect neonates against NEC following preterm birth and formula feeding. Further, we aimed to investigate if pr...

  17. Association of a bovine prion gene haplotype with atypical BSE

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Transmissible spongiform encephalopathies (TSEs), also known as prion diseases, are a class of fatal neurodegenerative disorders that occur in humans, ruminants, cats, and mink. Three distinct TSEs afflict cattle: classical bovine spongiform encephalopathy (BSE), atypical H-type BSE, and atypical ...

  18. 21 CFR 184.1034 - Catalase (bovine liver).

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... with 5 U.S.C. 552(a) and 1 CFR part 51. Copies are available from the National Academy Press, 2101... liver) (CAS Reg. No. 81457-95-6) is an enzyme preparation obtained from extracts of bovine liver. It is a partially purified liquid or powder. Its characterizing enzyme activity is catalase (EC...

  19. 21 CFR 184.1034 - Catalase (bovine liver).

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... with 5 U.S.C. 552(a) and 1 CFR part 51. Copies are available from the National Academy Press, 2101... liver) (CAS Reg. No. 81457-95-6) is an enzyme preparation obtained from extracts of bovine liver. It is a partially purified liquid or powder. Its characterizing enzyme activity is catalase (EC...

  20. 21 CFR 184.1034 - Catalase (bovine liver).

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... with 5 U.S.C. 552(a) and 1 CFR part 51. Copies are available from the National Academy Press, 2101... liver) (CAS Reg. No. 81457-95-6) is an enzyme preparation obtained from extracts of bovine liver. It is a partially purified liquid or powder. Its characterizing enzyme activity is catalase (EC...

  1. 21 CFR 184.1034 - Catalase (bovine liver).

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    .... 110, which is incorporated by reference in accordance with 5 U.S.C. 552(a) and 1 CFR part 51. Copies... enzyme preparation obtained from extracts of bovine liver. It is a partially purified liquid or powder. Its characterizing enzyme activity is catalase (EC 1.11.1.6). (b) The ingredient meets the...

  2. Bovine plasma proteins increase virulence of Haemophilus somnus in mice.

    PubMed

    Geertsema, Roger S; Kimball, Richard A; Corbeil, Lynette B

    2007-01-01

    The role of bovine serum or plasma proteins in Haemophilus somnus virulence was investigated in a mouse model of septicemia. An increase in virulence was detected when the organism was pre-incubated for 5 min and inoculated with fetal calf serum. When purified bovine serum or plasma proteins were pre-incubated with H. somnus before inoculating into mice, transferrin was found to increase virulence. Bovine lactoferrin was also noted to increase virulence, but to a lesser extent and had a delayed time course when compared with transferrin. Using an ELISA assay, an increased amount of H. somnus whole cells and culture supernatant bound to bovine transferrin when the organism was grown in iron-restricted media. Lactoferrin also bound to H. somnus, but binding was not affected by growth in iron-restricted media and it was eliminated with 2M NaCl, which reversed charge mediated binding. Transferrin, but not lactoferrin, supported growth of H. somnus on iron-depleted agar based media using a disk assay. Therefore, lactoferrin increased virulence by an undetermined mechanism whereas transferrin increased virulence of H. somnus by binding to iron-regulated outer-membrane proteins (IROMPs) and providing iron to the pathogen. PMID:17125964

  3. Genomics of bovine respiratory disease complex at USMARC

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Selection for genetic resistance/resilience bovine respiratory disease complex (BRDC) would significantly increase the efficiency of beef production in the U.S. through decreased treatment costs, productivity, and death loss. Unfortunately, selection for resistance to BRDC is challenging to implemen...

  4. Production of Bioactive Recombinant Bovine Chymosin in Tobacco Plants

    PubMed Central

    Wei, Zheng-Yi; Zhang, Yu-Ying; Wang, Yun-Peng; Fan, Ming-Xia; Zhong, Xiao-Fang; Xu, Nuo; Lin, Feng; Xing, Shao-Chen

    2016-01-01

    Chymosin (also known as rennin) plays an essential role in the coagulation of milk in the cheese industry. Chymosin is traditionally extracted from the rumen of calves and is of high cost. Here, we present an alternative method to producing bovine chymosin from transgenic tobacco plants. The CYM gene, which encodes a preprochymosin from bovine, was introduced into the tobacco nuclear genome under control of the viral 35S cauliflower mosaic promoter. The integration and transcription of the foreign gene were confirmed with Southern blotting and reverse transcription PCR (RT-PCR) analyses, respectively. Immunoblotting analyses were performed to demonstrate expression of chymosin, and the expression level was quantified by enzyme-linked immunosorbent assay (ELISA). The results indicated recombinant bovine chymosin was successfully expressed at an average level of 83.5 ng/g fresh weight, which is 0.52% of the total soluble protein. The tobacco-derived chymosin exhibited similar native milk coagulation bioactivity as the commercial product extracted from bovine rumen. PMID:27136529

  5. Bovine viral diarrhea virus: characteristics of the virus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    This paper reviews the history of research on bovine viral diarrhea viruses (BVDV) from their discovery in the 1940's to the design of current BVDV eradication programs. The physiochemical characteristics of BVDV are discussed and well as classification of BVDV into biotypes and genotypes. The trans...

  6. Pathogen reduction in minimally managed composting of bovine manure

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Persistence of pathogenic bacteria such as E. coli O157:H7, Salmonella spp., and Listeria monocytogenes in bovine feces and contaminated soils is an important risk factor in perpetuating the initial infection as well as re-infection of cattle and dissemination of pathogens throughout agricultural la...

  7. Genetic relationships of the Portuguese Lidia bovine populations

    PubMed Central

    Correia, P; Baron, E; da Silva, J. M; Cortés, O

    2014-01-01

    To clarify the genetic relationships among the Lidia breed lineages and two main Portuguese Lidia bovine populations, Casta Portuguesa and Brava dos Açores, 24 autosomal microsatellites were analyzed in 120 samples. Brava dos Açores showed the highest observed and expected heterozygosity (0.73 and 0.70, respectively) while Casta Portuguesa showed the lowest observed and expected heterozygosity (0.51 and 0.50, respectively). The results of this study were compared with the previous microsatellites data from the main Lidia bovine lineages. Casta Portuguesa was the most genetically isolated Lidia bovine population as revealed by the average FST genetic distance value with respect to the other lineages (32%). All the populations of Portuguese Lidia had negative FIS values. The Neighbour-joining dendrogram grouped Casta Portuguesa in the same branch with Miura, which was supported by the STRUCTURE software. The results evidenced low levels of genetic diversity and high levels of genetic differentiation in Casta Portuguesa and high levels of genetic diversity in Brava dos Açores populations, probably due to the crossbreeding of different bovine lineages at origin, and genetic flow among herds. PMID:27175132

  8. Bovine colostrum immunoglobulin concentrate for cryptosporidiosis in AIDS.

    PubMed Central

    Shield, J; Melville, C; Novelli, V; Anderson, G; Scheimberg, I; Gibb, D; Milla, P

    1993-01-01

    Lactobin-R is a commercial hyperimmune bovine colostrum with potent anticryptosporidial activity. It was administered to a 4 year old child with AIDS and severe diarrhoea associated with cryptosporidiosis. There was significant clinical improvement in the diarrhoea and permanent elimination of the parasite from the gut as assessed through serial jejunal biopsy and stool specimens. Images PMID:8259880

  9. Control of declared origin of bovine serum, a pilot study

    NASA Astrophysics Data System (ADS)

    Horacek, M.; Papesch, W.

    2009-04-01

    Bovine serum is the essential culture medium for cell cultures. Therefore it is highly demanded and the quality of the serum, e.g.: absence of bacteria, viruses certain antibodies, etc.., are important criteria. as some cattle diseases are endemic in certain regions, the origin of bovine serum is an important quality measure for its value. Thus the need to control the declared origins is present. Bovine serum was measured for d2H, d13C, d15N and d34S of proteine (dry residue) and d2H and d18O of the serum water. The hydrogen and oxygen are mainly depending by the isotopic composition of the water ingested by the cattle, and thus usually influenced by the isotopic signal of the precipitation. The carbon isotope signal is reflecting the diet of the cattle, whether it mainly feed on C3- or C4-plants. The nitrogen and sulphur isotope ratio is transferred from the ground/soil into the plant material and into the animal tissue, with some offset for nitrogen and without any significant offset for sulphur. Bovine serum samples from Canada, USA, Mexico, Brazil, Australia and New Zealand have been analysed. Due to the variations in the environmental conditions in different countries and regions which influence the isotope signatures of the serum samples it is possible to discriminate samples of different origin. Main discriminating parameters are d2H and d18O, d13C and d34S.

  10. Is there a genetic solution to bovine respiratory disease complex?

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bovine respiratory disease complex (BRDC) is a complex multi-factor disease, which increases costs and reduces revenue from feedlot cattle. Multiple stressors and pathogens (viral and bacterial) have been implicated in the etiology of BRDC, therefore multiple approaches will be needed to evaluate a...

  11. Typical and atypical cases of bovine spongiform encephalopathy

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bovine spongiform encephalopathy (BSE) is a transmissible spongiform encephalopathy of cattle, first detected in 1986 in the United Kingdom and subsequently in other countries. It is the most likely cause of variant Creutzfeldt-Jakob disease (vCJD) in humans, but the origin of BSE has not been eluci...

  12. Control of Bovine Viral Diarrhea Virus in Ruminants

    Technology Transfer Automated Retrieval System (TEKTRAN)

    This document is a consensus statement, produced at the request of the American College of Veterinary Internal Medicine that reflects the opinion of an expert panel regarding the prevalence and host range, clinical manifestations, and the potential for ultimate eradication of bovine viral diarrhea v...

  13. Effect of Ergot Alkaloids on Bovine Foregut Vasculature

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Ergot alkaloids induce vasoconstriction of bovine foregut vasculature. Ergovaline induced the greatest response in ruminal artery while ergovaline and ergotamine induced the greatest response in ruminal vein. Lysergic acid did not stimulate a contractile response in either the ruminal artery or vein...

  14. Effects of Ergot Alkaloids on Bovine Sperm Motility In Vitro

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Ergot alkaloids are synthesized by endophyte-infected (Neotyphodium coenophialum) tall fescue (Lolium arundinaceum (Schreb.) S.J. Darbyshire). Our objective was to determine direct effects of ergot alkaloids (ergotamine, dihydroergotamine and ergonovine) on the motility of bovine spermatozoa in vit...

  15. Endocrine and exocrine function of the bovine testis. Chapter 2

    Technology Transfer Automated Retrieval System (TEKTRAN)

    This chapter is devoted to the endocrine and exocrine function of the normal bovine male testes. The discussion begins with a historical review of the literature dating back to Aristotle’s (300 BC) initial description of the anatomy of the mammalian testes. The first microscopic examination of the t...

  16. [Bovine leukocyte adhesion deficiency: clinical picture and differential diagnosis].

    PubMed

    Lienau, A; Stöber, M; Kehrli, M E; Tammen, I; Schwenger, B; Kuczka, A; Pohlenz, J

    1994-10-01

    The pathological clinical and laboratory findings obtained in 50 calves and young cattle affected with Bovine Leukocyte Adhesion Deficiency are compared with those found in 114 calves and young cattle showing marked neutrophil leukocytosis of other origin (age: < 2 years; leukocyte count: > 30,000 per microl; percentage of lymphocytes: < 55%). PMID:7851303

  17. Bovine coronaviruses from the respiratory tract: Antigenic and genetic diversity

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bovine corona viruses (BoCV) isolated from respiratory tract, nasal swab and broncho alveolar washing fluid samples were evaluated for genetic and antigenic differences. These BoCV from the respiratory tract of healthy and clinically ill cattle with BRD signs were compared to reference and vaccine ...

  18. Salmonella prevalence in bovine lymph nodes differs among feedyards

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Lymphatic tissue, specifically lymph nodes, is commonly incorporated into ground beef products as a component of lean trimmings. Salmonella and other pathogenic bacteria have been identified in bovine lymph nodes. Although Salmonella prevalence has been examined among lymph nodes within an animal,...

  19. Clinical bovine viral diarrhoea virus infection in Jordan.

    PubMed

    Abutarbush, S M; Alqawasmeh, D M

    2010-12-01

    A 1-year-old Holstein Friesian heifer was presented for anorexia and acute diarrhoea. The heifer was born and raised at the farm. Bovine viral diarrhoea virus (BVDV) infection was diagnosed using clinical signs and RT-PCR. Clinical BVDV infection has never been reported in Jordan. PMID:21117287

  20. Full Genome Sequence of a Bovine Enterovirus Isolated in China

    PubMed Central

    Peng, Xiao-wei; Dong, Hao; Wu, Qing-min

    2014-01-01

    We report the full genome sequence of an isolate of bovine enterovirus type B from China. The virus (BEV-BJ001) was isolated from Beijing, China, from fecal swabs of cattle suffering from severe diarrhea. This genome sequence will give useful insight for future molecular epidemiological studies in China. PMID:24970832

  1. Production of Bioactive Recombinant Bovine Chymosin in Tobacco Plants.

    PubMed

    Wei, Zheng-Yi; Zhang, Yu-Ying; Wang, Yun-Peng; Fan, Ming-Xia; Zhong, Xiao-Fang; Xu, Nuo; Lin, Feng; Xing, Shao-Chen

    2016-01-01

    Chymosin (also known as rennin) plays an essential role in the coagulation of milk in the cheese industry. Chymosin is traditionally extracted from the rumen of calves and is of high cost. Here, we present an alternative method to producing bovine chymosin from transgenic tobacco plants. The CYM gene, which encodes a preprochymosin from bovine, was introduced into the tobacco nuclear genome under control of the viral 35S cauliflower mosaic promoter. The integration and transcription of the foreign gene were confirmed with Southern blotting and reverse transcription PCR (RT-PCR) analyses, respectively. Immunoblotting analyses were performed to demonstrate expression of chymosin, and the expression level was quantified by enzyme-linked immunosorbent assay (ELISA). The results indicated recombinant bovine chymosin was successfully expressed at an average level of 83.5 ng/g fresh weight, which is 0.52% of the total soluble protein. The tobacco-derived chymosin exhibited similar native milk coagulation bioactivity as the commercial product extracted from bovine rumen. PMID:27136529

  2. Identification, localization, and sequencing of fetal bovine VASA homolog.

    PubMed

    Bartholomew, Rachel A; Parks, John E

    2007-10-01

    The vasa gene, first described in Drosophila, is purported to be important in germ cell development. Vasa is present across several invertebrate and vertebrate taxa, including frogs, fish, chickens, and humans. Vasa, a DEAD (asparagine-glutamine-alanine-asparagine) box protein shown to function as an RNA helicase in vitro, has not been investigated previously in fetal stage cattle. Total RNA was extracted from bovine fetal gonads obtained at 35-55 days, 55-80 days, and 80-120 days of gestation to amplify a 296 bp reverse transcription polymerase chain reaction (RT-PCR) product using primers for human vasa. The complete coding sequence of bovine vasa was cloned with 5' and 3' random amplification of cDNA ends polymerase chain reaction (RACE-PCR) and subsequently identified as bovine vasa homolog (BVH). Northern blot analysis revealed that among the tissues examined (gonad, liver, heart, brain, and femur), the vasa gene was expressed in the gonad. This localization, the conserved pattern of gene expression, and the gene sequence suggests that BVH plays a role in bovine germ cell development as proposed for other mammalian species. PMID:17150314

  3. Binding of Pasteurella haemolytica leukotoxin to bovine leukocytes.

    PubMed Central

    Brown, J F; Leite, F; Czuprynski, C J

    1997-01-01

    Pasteurella haemolytica is the principal bacterial pathogen in the bovine respiratory disease complex. This organism produces an exotoxin (referred to as leukotoxin) during logarithmic-phase growth that is a potent leukocyte-modulating agent. At low concentrations, it activates neutrophils and mononuclear phagocytes to release inflammatory mediators, while at the same time making these cells destined to undergo apoptotic cell death. At higher concentrations, the toxin causes rapid swelling and loss of cell viability. In this study, we demonstrated that toxin binding can be directly evaluated by flow cytometry with biologically active biotinylated leukotoxin. Leukotoxin binding was blocked by the addition of a neutralizing anti-leukotoxin monoclonal antibody and was not detected when bovine leukocytes were incubated with culture filtrates from a mutant strain of P. haemolytica that does not produce biologically active leukotoxin. In addition, treatment of bovine leukocytes with protease K eliminated subsequent binding of leukotoxin, suggesting that there is a protein on the leukocyte surface that is either a leukotoxin binding site or is required for stabilization of leukotoxin binding. We did not detect binding of biotinylated leukotoxin to porcine or human leukocytes, which have been reported previously to be resistant to the lytic effects of the leukotoxin. These findings suggest that there may be a specific binding site for P. haemolytica leukotoxin on bovine but not on porcine or human leukocytes and that it might be involved in the activation and lytic activities of the leukotoxin. PMID:9284143

  4. On the search for markers of tick resistance in bovines

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Genetic differences in susceptibility to ticks (Rhipicephalus (Boophilus) microplus) are considerable in bovines. Here, mapping, association and gene expression approaches were employed to further advance our understanding of the molecular basis of tick resistance. A B. taurus x B. indicus F2 popul...

  5. Investigation of bovine serum albumin glycation by THz spectroscopy

    NASA Astrophysics Data System (ADS)

    Cherkasova, Olga P.; Nazarov, Maxim M.; Shkurinov, Alexander P.

    2016-04-01

    Protein glycation is accelerated under hyperglycemic conditions resulting to loss in the structure and biological functions of proteins. The transmission THz spectroscopy has been used for measuring of bovine serum albumin glycation dynamics. It was found that amplitude of albumin THz absorption depends on type of sugars and incubation time.

  6. Bovine tuberculosis vaccine research: historical perspectives and recent advances

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The emergence of wildlife reservoirs of Mycobacterium bovis infection in cattle as well as increased inter-regional trade with associated spread of M. bovis has led to renewed interest in the use of vaccines for the control of bovine tuberculosis (TB). Field efficacy trials performed in the early 20...

  7. Analysis of copy number variation in the bovine genome

    Technology Transfer Automated Retrieval System (TEKTRAN)

    We initiated a systematic study of the copy number variation (CNV) within the Bovine HapMap cattle population using array comparative genomic hybridization (array CGH). Oligonucleotide CGH arrays were designed and fabricated to provide a genome-wide coverage with an average interval of 6 kb using t...

  8. Bacterial expression and purification of recombinant bovine Fab fragments.

    PubMed

    O'Brien, Philippa M; Maxwell, Gavin; Campo, M Saveria

    2002-02-01

    We have previously described a recombinant phagemid expression vector, pComBov, designed for the production of native sequence bovine monoclonal antibodies (mAb) generated by antibody phage display. Bovine mAb Fab fragments isolated from libraries constructed using pComBov in Escherichia coli strain XL1-Blue, which is routinely used for antibodies expressed on the surface of phage, were expressed at very low yields. Therefore, a study was undertaken to determine optimal growth conditions for maximal expression of bovine Fab fragments in E. coli. By varying the E. coli strain, and the temperature and length of the culture growth, we were able to substantially increase the yield of soluble Fab fragments. A high yield of Fab fragments was found in the culture growth medium, which enabled us to devise a rapid and simple single-step method for the purification of native (nondenatured) Fabs based on immobilized metal affinity chromatography against a six-histidine amino acid carboxyl-terminal extension of the heavy-chain constant region. Using these methods we were able to express and purify antigen-specific bovine Fab fragments from E. coli. PMID:11812221

  9. Interferon Gamma Assay for the Diagnosis of Bovine Tuberculosis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Contact Irene Schiller Prionics AG Wagistrasse 27A CH-8952 Schlieren Switzerland irene.schiller@prionics.com Introduction Bovine tuberculosis (bTB), a zoonotic disease with a major economic impact, continues to be a significant problem with a global perspective and increasing prevalence in vario...

  10. 9 CFR 113.69 - Pasteurella Multocida Vaccine, Bovine.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 9 Animals and Animal Products 1 2010-01-01 2010-01-01 false Pasteurella Multocida Vaccine, Bovine. 113.69 Section 113.69 Animals and Animal Products ANIMAL AND PLANT HEALTH INSPECTION SERVICE, DEPARTMENT OF AGRICULTURE VIRUSES, SERUMS, TOXINS, AND ANALOGOUS PRODUCTS; ORGANISMS AND VECTORS STANDARD REQUIREMENTS Live Bacterial Vaccines §...

  11. 9 CFR 113.68 - Pasteurella Haemolytica Vaccine, Bovine.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 9 Animals and Animal Products 1 2010-01-01 2010-01-01 false Pasteurella Haemolytica Vaccine, Bovine. 113.68 Section 113.68 Animals and Animal Products ANIMAL AND PLANT HEALTH INSPECTION SERVICE, DEPARTMENT OF AGRICULTURE VIRUSES, SERUMS, TOXINS, AND ANALOGOUS PRODUCTS; ORGANISMS AND VECTORS STANDARD REQUIREMENTS Live Bacterial Vaccines §...

  12. 9 CFR 113.69 - Pasteurella Multocida Vaccine, Bovine.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 9 Animals and Animal Products 1 2014-01-01 2014-01-01 false Pasteurella Multocida Vaccine, Bovine. 113.69 Section 113.69 Animals and Animal Products ANIMAL AND PLANT HEALTH INSPECTION SERVICE, DEPARTMENT OF AGRICULTURE VIRUSES, SERUMS, TOXINS, AND ANALOGOUS PRODUCTS; ORGANISMS AND VECTORS STANDARD REQUIREMENTS Live Bacterial Vaccines §...

  13. Porcine brain natriuretic peptide receptor in bovine adrenal cortex

    SciTech Connect

    Higuchi, K.; Hashiguchi, T.; Ohashi, M.; Takayanagi, R.; Haji, M.; Matsuo, H.; Nawata, H.

    1989-01-01

    The action of porcine brain natriuretic peptide (pBNP) on the steroidogenesis was investigated in cultured bovine adrenocortical cells. Porcine BNP induced a significant dose-dependent inhibition of both ACTH- and A II-stimulated aldosterone secretion. 10/sup /minus/8/M and 10/sup /minus/7/M pBNP also significantly inhibited ACTH-stimulated cortisol and dehydroepiandrosterone (DHEA) secretions. Binding studies of (/sup 125/I)-pBNP to bovine adrenocortical membrane fractions showed that adrenal cortex had high-affinity and low-capacity pBNP binding sites, with a dissociation constant (Kd) of 1.70 x 10/sup /minus/10/M and a maximal binding capacity (Bmax) of 19.9 fmol/mg protein. Finally, the 135 Kd radioactive band was specially visualized in the affinity labeling of bovine adrenal cortex with disuccinimidyl suberate (DSS). These results suggest that pBNP may have receptor-mediated suppressive actions on bovine adrenal steroidogenesis, similar to that in atrial natriuretic peptide (ANP).

  14. Adherence of Tritrichomonas foetus to bovine vaginal epithelial cells.

    PubMed Central

    Corbeil, L B; Hodgson, J L; Jones, D W; Corbeil, R R; Widders, P R; Stephens, L R

    1989-01-01

    Adherence of Tritrichomonas foetus to bovine vaginal epithelial cells (VECs) in vitro was investigated with fresh washed bovine VECs and log-phase cultures of T. foetus. Observation under phase-contrast microscopy showed that T. foetus usually adhered first by the posterior flagellum and later by the body. Significantly more keratinized squamous epithelial cells were detected with attached parasites than nonkeratinized round epithelial cells. The optimal pH range for attachment was 6.0 to 7.5, with peak attachment at pH 6.5 for squamous VECs. Surface-reactive bovine antiserum to T. foetus prevented adherence to bovine squamous VECs. Inhibition of adherence occurred at nonagglutinating, nonimmobilizing serum dilutions. Antiserum fractions enriched for immunoglobulin G1 inhibited adherence, but fractions enriched for immunoglobulin G2 did not. The inhibitory antiserum was specific for several medium- to high-molecular-weight membrane antigens as detected in Western blots (immunoblots). The ability of surface-reactive antibodies to prevent adherence and to agglutinate and immobilize T. foetus indicates that they may be protective. Images PMID:2471692

  15. Bovine copy number variation and its implication in animal health

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Recently it has become apparent that previously unappreciated genomic structural variation, including copy number variations (CNV), contributes significantly to individual health and disease in humans and rodents. As a complement to the bovine HapMap project, we initiated a systematic study of the C...

  16. Hedgehog signaling pathway in small bovine ovarian follicles

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The hedgehog signaling pathway is involved in the regulation of cell proliferation, differentiation, and turnover in a variety of mammalian embryonic and adult tissues including bovine ovarian granulosa and theca cells. Binding of hedgehog to the patch receptor derepresses smoothened resulting in t...

  17. Killing of juvenile Fasciola hepatica by purified bovine eosinophil proteins.

    PubMed Central

    Duffus, W P; Thorne, K; Oliver, R

    1980-01-01

    Eosinophils were isolated from the mammary gland of Fasciola hepatica-infected cattle by intramammary infusion with a crude extract from adult F. hepatica. Up to 5 x 10(9) eosinophils with a purity of over 90% could be obtained from a single quarter of the gland. The major contaminating cells were monocytes which reached their peak several days following the eosinophil peak. Two major proteins were isolated from bovine eosinophil granules, a high molecular weight peroxidase-active protein and a smaller molecular weight predominantly basic protein. This smaller protein was thought to be the bovine equivalent of guinea-pig and human major basic protein (MBP), although it possessed an unusually high concentration of cysteine. The bovine MBP had a profound effect on juvenile F. hepatica in vitro causing damage and death at concentrations down to 1 x 10(-6) M. The damage was detected by a 51Cr release assay and/or a viability assay involving microscopical examination of the flukes. Other cations, especially protamine sulphate, were also shown to kill flukes, although both lysozyme, found in neutrophils, and the peroxidase-positive peak from bovine eosinophils were unable to mediate any detectable damage. Images Fig. 4 PMID:7438542

  18. Phenotypic and Functional Heterogeneity of Bovine Blood Monocytes

    PubMed Central

    Hussen, Jamal; Düvel, Anna; Sandra, Olivier; Smith, David; Sheldon, Iain Martin; Zieger, Peter; Schuberth, Hans-Joachim

    2013-01-01

    Murine and human peripheral blood monocytes are heterogeneous in size, granularity, nuclear morphology, phenotype and function. Whether and how bovine blood monocytes follow this pattern was analyzed in this study. Flow cytometrically, classical monocytes (cM) CD14+ CD16−, intermediate monocytes (intM) CD14+ CD16+ and nonclassical monocytes (ncM) CD14+ CD16+ were identified, with cM being the predominant subset (89%). cM showed a significant lower expression of CD172a, intM expressed the highest level of MHC class II molecules, and ncM were low positive for CD163. Compared to cM and intM, ncM showed a significantly reduced phagocytosis capacity, a significantly reduced generation of reactive oxygen species, and reduced mRNA expression of CXCL8, CXCL1 and IL-1β after LPS stimulation. Based on IL-1β secretion after LPS/ATP stimulation, the inflammasome could be activated in cM and intM, but not in ncM. IFNγ increased the expression of CD16 selectively on cM and induced a shift from cM into intM in vitro. In summary, bovine CD172a-positive mononuclear cells define three monocyte subsets with distinct phenotypic and functional differences. Bovine cM and intM share homologies with their human counterparts, whereas bovine ncM are not inflammatory monocytes. PMID:23967219

  19. Bovine lactoferrin interacts with cable pili of Burkholderia cenocepacia.

    PubMed

    Ammendolia, Maria Grazia; Bertuccini, Lucia; Iosi, Francesca; Minelli, Fabio; Berlutti, Francesca; Valenti, Piera; Superti, Fabiana

    2010-06-01

    In this study we evaluated the ability of lactoferrin, the most abundant antimicrobial protein in airway secretions, to bind the surface structures of a Burkholderia strain cystic fibrosis-isolated. Burkholderia cenocepacia is a gram-negative bacterium involved as respiratory pathogen in cystic fibrosis patient infections. This bacterium possesses filamentous structures, named cable pili that have been proposed as virulence factors because of their ability to bind to respiratory epithelia and mucin. Previously, we demonstrated that bovine lactoferrin was able to influence the efficiency of invasion of different iron-regulated morphological forms of B. cenocepacia. Bovine lactoferrin showed to efficiently inhibit invasion of alveolar epithelial cells by free-living bacteria or iron-induced aggregates or biofilm. Results of the present study demonstrate that bovine lactoferrin is also able to specifically bind to B. cenocepacia cells and show that cable pili are involved in this interaction. The attachment of bovine lactoferrin to pili led to a reduced binding of bacterial cells to mucin. Since cable pili are implicated in mediating the bacterial interactions with mucin and epithelial cells, lactoferrin binding to these structures could play an important role in neutralizing bacterial infection in cystic fibrosis patients. PMID:20364433

  20. Illumina Amplicon Sequencing of 16S rRNA Tag Reveals Bacterial Community Development in the Rhizosphere of Apple Nurseries at a Replant Disease Site and a New Planting Site

    PubMed Central

    Sun, Jian; Zhang, Qiang; Zhou, Jia; Wei, Qinping

    2014-01-01

    We used a next-generation, Illumina-based sequencing approach to characterize the bacterial community development of apple rhizosphere soil in a replant site (RePlant) and a new planting site (NewPlant) in Beijing. Dwarfing apple nurseries of ‘Fuji’/SH6/Pingyitiancha trees were planted in the spring of 2013. Before planting, soil from the apple rhizosphere of the replant site (ReSoil) and from the new planting site (NewSoil) was sampled for analysis on the Illumina MiSeq platform. In late September, the rhizosphere soil from both sites was resampled (RePlant and NewPlant). More than 16,000 valid reads were obtained for each replicate, and the community was composed of five dominant groups (Proteobacteria, Acidobacteria, Bacteroidetes, Gemmatimonadetes and Actinobacteria). The bacterial diversity decreased after apple planting. Principal component analyses revealed that the rhizosphere samples were significantly different among treatments. Apple nursery planting showed a large impact on the soil bacterial community, and the community development was significantly different between the replanted and newly planted soils. Verrucomicrobia were less abundant in RePlant soil, while Pseudomonas and Lysobacter were increased in RePlant compared with ReSoil and NewPlant. Both RePlant and ReSoil showed relatively higher invertase and cellulase activities than NewPlant and NewSoil, but only NewPlant soil showed higher urease activity, and this soil also had the higher plant growth. Our experimental results suggest that planting apple nurseries has a significant impact on soil bacterial community development at both replant and new planting sites, and planting on new site resulted in significantly higher soil urease activity and a different bacterial community composition. PMID:25360786

  1. Mining biomass-degrading genes through Illumina-based de novo sequencing and metagenomic analysis of free-living bacteria in the gut of the lower termite Coptotermes gestroi harvested in Vietnam.

    PubMed

    Do, Thi Huyen; Nguyen, Thi Thao; Nguyen, Thanh Ngoc; Le, Quynh Giang; Nguyen, Cuong; Kimura, Keitarou; Truong, Nam Hai

    2014-12-01

    The 5.6 Gb metagenome of free-living microbial flora in the gut of the lower termite Coptotermes gestroi, harvested in Vietnam, was sequenced using Illumina technology. Genes related to biomass degradation were mined for a better understanding of biomass digestion in the termite gut and to identify lignocellulolytic enzymes applicable to biofuel production. The sequencing generated 5.4 Gb of useful reads, containing 125,431 ORFs spanning 78,271,365 bp, 80% of which was derived from bacteria. The 12 most abundant bacterial orders were Spirochaetales, Lactobacillales, Bacteroidales, Clostridiales, Enterobacteriales, Pseudomonades, Synergistales, Desulfovibrionales, Xanthomonadales, Burkholderiales, Bacillales, and Actinomycetales, and 1460 species were estimated. Of more than 12,000 ORFs with predicted functions related to carbohydrate metabolism, 587 encoding hydrolytic enzymes for cellulose, hemicellulose, and pectin were identified. Among them, 316 ORFs were related to cellulose degradation, and included β-glucosidases, 6-phospho-β-glucosidases, licheninases, glucan endo-1,3-β-D-glucosidases, endoglucanases, cellulose 1,4-β-cellobiosidases, glucan 1,3-β-glucosidases, and cellobiose phosphorylases. In addition, 259 ORFs were related to hemicellulose degradation, encoding endo-1,4-β-xylanases, α-galactosidases, α-N-arabinofuranosidases, xylan 1,4-β-xylosidases, arabinan endo-1,5-α-L-arabinosidases, endo-1,4-β-mannanases, and α-glucuronidases. Twelve ORFs encoding pectinesterases and pectate lyases were also obtained. To our knowledge, this is the first successful application of Illumina-based de novo sequencing for the analysis of a free-living bacterial community in the gut of a lower termite C. gestroi and for mining genes related to lignocellulose degradation from the gut bacteria. PMID:24928651

  2. Prevalence of antibodies to infectious bovine rhinotracheitis, parainfluenza 3, bovine respiratory syncytial, and bovine viral diarrhea viruses in cattle in Saskatchewan and Alberta

    PubMed Central

    Durham, Peter J.K.; Hassard, Lori E.

    1990-01-01

    A total of 1745 healthy cattle from 295 farms in Saskatchewan and Alberta was tested by ELISA for antibodies to four viruses. Antibodies to infectious bovine rhinotracheitis (IBR) virus were found in 37.8% of sera (59.5% of properties), to parainfluenza 3 (PI3) virus in 93.9% of sera (99.7% of properties), to bovine respiratory syncytial (BRS) virus in 78.5% of sera (86.6% of properties), and to bovine viral diarrhea (BVD) virus in 40.6% of sera (66.7% of properties) The prevalence of PI3 viral antibodies among Saskatchewan cattle was not affected by district of origin, breed, sex, age, or vaccination practices, though BRS viral antibodies appeared less frequent in young, male, and unvaccinated animals. Antibodies to IBR and BVD viruses were less prevalent in the Prince Albert/Tisdale districts and in young, male, and unvaccinated animals, but were more common in Holstein cattle. Antibodies to IBR virus appeared less frequent in Herefords. Antibodies were more prevalent in cattle which had been vaccinated against IBR, BRS, and BVD virus infections. The relatively small number of cattle sampled from Alberta had a similar prevalence of antibodies to PI3 and BRS viruses to that seen in cattle in Saskatchewan, though IBR and BVD prevalence rates were lower. PMID:17423704

  3. Vaccenic acid metabolism in the liver of rat and bovine.

    PubMed

    Gruffat, Dominique; De La Torre, Anne; Chardigny, Jean-Michel; Durand, Denys; Loreau, Olivier; Bauchart, Dominique

    2005-03-01

    Hepatic metabolism of vaccenic acid (VA), especially its conversion into CLA, was studied in the bovine (ruminant species that synthesizes CLA) and in the rat (model for non-ruminant) by using the in vitro technique of liver explants. Liver tissue samples were collected from fed animals (5 male Wistar rats and 5 Charolais steers) and incubated at 37 degrees C for 17 h under an atmosphere of 95% O2/5% CO2 in medium supplemented with 0.75 mM of FA mixture and with 55 microM [1-14C]VA. VA uptake was about sixfold lower in bovine than in rat liver slices (P< 0.01). For both species, VA that was oxidized to partial oxidation products represented about 20% of VA incorporated by cells. The chemical structure of VA was not modified in bovine liver cells, whereas in rat liver cells, 3.2% of VA was converted into 16:0 and only 0.33% into CLA. The extent of esterification of VA was similar for both animal species (70-80% of incorporated VA). Secretion of VA as part of VLDL particles was very low and similar in rat and bovine liver (around 0.07% of incorporated VA). In conclusion, characteristics of the hepatic metabolism of VA were similar for rat and bovine animals, the liver not being involved in tissue VA conversion into CLA in spite of its high capacity for FA desaturation especially in the rat. This indicates that endogenous synthesis of CLA should take place exclusively in peripheral tissues. PMID:15957256

  4. Propionate induces the bovine cytosolic phosphoenolpyruvate carboxykinase promoter activity.

    PubMed

    Zhang, Qian; Koser, Stephanie L; Donkin, Shawn S

    2016-08-01

    Cytosolic phosphoenolpyruvate carboxykinase (PCK1) is a critical enzyme within the metabolic networks for gluconeogenesis, hepatic energy metabolism, and tricarboxylic acid cycle function, and is controlled by several transcription factors including hepatic nuclear factor 4α (HNF4α). The primary objective of the present study was to determine whether propionate regulates bovine PCK1 transcription. The second objective was to determine the action of cyclic AMP (cAMP), glucocorticoids, and insulin, hormonal cues known to modulate glucose metabolism, on bovine PCK1 transcriptional activity. The proximal promoter of the bovine PCK1 gene was ligated to a Firefly luciferase reporter and transfected into H4IIE hepatoma cells. Cells were exposed to treatments for 23 h and luciferase activity was determined in cell lysates. Activity of the PCK1 promoter was linearly induced by propionate, and maximally increased 7-fold with 2.5 mM propionate, which was not muted by 100 nM insulin. Activity of the PCK1 promoter was increased 1-fold by either 1.0 mM cAMP or 5.0µM dexamethasone, and 2.2-fold by their combination. Induction by cAMP and dexamethasone was repressed 50% by 100 nM insulin. Propionate, cAMP, and dexamethasone acted synergistically to induce the PCK1 promoter activity. Propionate-responsive regions, identified by 5' deletion analysis, were located between -1,238 and -409 bp and between -85 and +221 bp. Deletions of the core sequences of the 2 putative HNF4α sites decreased the responsiveness to propionate by approximately 40%. These data indicate that propionate regulates its own metabolism through transcriptional stimulation of the bovine PCK1 gene. This induction is mediated, in part, by the 2 putative HNF4α binding sites in the bovine PCK1 promoter. PMID:27289145

  5. Effect of triiodothyronine on developmental competence of bovine oocytes.

    PubMed

    Costa, N N; Cordeiro, M S; Silva, T V G; Sastre, D; Santana, P P B; Sá, A L A; Sampaio, R V; Santos, S S D; Adona, P R; Miranda, M S; Ohashi, O M

    2013-09-01

    Developmental competence of in vitro-matured bovine oocytes is a limiting factor in production of embryos in vitro. Several studies have suggested a potential positive effect of thyroid hormones on cultured oocytes and/or their supporting cells. Therefore, the aim of the present study was to ascertain whether medium supplementation with triiodothyronine (T3) improved subsequent developmental competence of in vitro-matured bovine oocytes. For this purpose, we first documented (using reverse transcription PCR) that whereas bovine cumulus cells expressed both thyroid hormone receptor (TR)-α and TRβ, immature bovine oocytes expressed TRα only. Thereafter, to test the effects of TH on developmental competence, abattoir-derived oocytes were matured in vitro in a medium containing 0, 25, 50, or 100 nM T3 and subjected to in vitro fertilization. Embryo quality was evaluated by assessing cleavage and blastocyst rates, morphological quality, development kinetics, and total cell number on Day 8 of culture. Notably, addition of 50 or 100 nM T3 to the in vitro maturation medium increased (P < 0.05) the rate of hatched blastocysts on the eighth day of culture, as compared with other groups (62.4 ± 11.7, 53.1 ± 16.3, and 32.4 ± 5.3, respectively). Next, the relative expression levels of genes related to embryo quality POU-domain transcription factor (POU5F1) and glucose transporter-1 (GLUT 1) were compared between in vivo- and in vitro-produced blastocysts. On the basis of the previous experiments, IVP embryos originating from oocytes that were matured in vitro in the presence or absence of 50 nM T3 were evaluated. The treatment had no effect (P > 0.05) on gene expression. We concluded that supplementation of bovine oocyte in vitro maturation medium with T3 may have a beneficial effect on the kinetics of embryo development. PMID:23683691

  6. Global Genetic Profiles of Gene Network Disruption in Bovine Peripheral Blood Mononuclear Cells Induced Bovine Leukemia Virus (BLV) Infection

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Efficient nutrient assimilation into useful animal-derived products is the ultimate requirement for successful animal production. Infection in young growing animals can decrease energy and nutrient use required for growth rate by redirection of nutrients to support immune defense processes. Bovine l...

  7. Markers on bovine chromosome 20 associated with fat related traits and incidence of contracting bovine respiratory disease

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The objective of this study was to assess the association of single nucleotide polymorphism (SNP) markers with incidence of bovine respiratory disease (BRD) and fat related traits. Steers from the Cycle 7 of the Germplasm Evaluation Program (GPE7; n=565) were used. The GPE7 population comprises anim...

  8. Evaluation of endocrine and immune disruption of infectious bovine rhinotracheitis virus vaccinated steers challenged with infectious bovine rhinotracheitis virus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The objective of this study was to evaluate the endocrine and immune responses of steers challenged with infectious bovine rhinotracheitis virus (IBRV). For the study, twelve crossbred beef steers weighing approximately 228.82 kg were fitted with indwelling rectal temperature monitoring devices and ...

  9. Bovine leukosis. V. Epidemiological study of bovine C-type virus by the use of the complement fixation test.

    PubMed Central

    Tabel, H; Chander, S; Van Der Maaten, M J; Miller, J M

    1976-01-01

    Persistent levels of serum antibodies to bovine C-type virus were demonstrated by the complement fixation test in cattle of a leukosis herd during an observation period of one and one half years. Using the same method, no antibodies were detected in a control herd. PMID:187302

  10. The synergistic necrohemorrhagic action of Clostridium perfringens perfringolysin and alpha toxin in the bovine intestine and against bovine endothelial cells

    PubMed Central

    2013-01-01

    Bovine necrohemorrhagic enteritis is a major cause of mortality in veal calves. Clostridium perfringens is considered as the causative agent, but there has been controversy on the toxins responsible for the disease. Recently, it has been demonstrated that a variety of C. perfringens type A strains can induce necrohemorrhagic lesions in a calf intestinal loop assay. These results put forward alpha toxin and perfringolysin as potential causative toxins, since both are produced by all C. perfringens type A strains. The importance of perfringolysin in the pathogenesis of bovine necrohemorrhagic enteritis has not been studied before. Therefore, the objective of the current study was to evaluate the role of perfringolysin in the development of necrohemorrhagic enteritis lesions in calves and its synergism with alpha toxin. A perfringolysin-deficient mutant, an alpha toxin-deficient mutant and a perfringolysin alpha toxin double mutant were less able to induce necrosis in a calf intestinal loop assay as compared to the wild-type strain. Only complementation with both toxins could restore the activity to that of the wild-type. In addition, perfringolysin and alpha toxin had a synergistic cytotoxic effect on bovine endothelial cells. This endothelial cell damage potentially explains why capillary hemorrhages are an initial step in the development of bovine necrohemorrhagic enteritis. Taken together, our results show that perfringolysin acts synergistically with alpha toxin in the development of necrohemorrhagic enteritis in a calf intestinal loop model and we hypothesize that both toxins act by targeting the endothelial cells. PMID:23782465

  11. Effects of Preinfection With Bovine Viral Diarrhea Virus on Immune Cells From the Lungs of Calves Inoculated With Bovine Herpesvirus 1.1.

    PubMed

    Risalde, M A; Molina, V; Sánchez-Cordón, P J; Romero-Palomo, F; Pedrera, M; Gómez-Villamandos, J C

    2015-07-01

    The aim of this work was to study the interstitial aggregates of immune cells observed in pulmonary parenchyma of calves preinfected with bovine viral diarrhea virus and challenged later with bovine herpesvirus 1. In addition, the intent of this research was to clarify the role of bovine viral diarrhea virus in local cell-mediated immunity and potentially in predisposing animals to bovine respiratory disease complex. Twelve Friesian calves, aged 8 to 9 months, were inoculated with noncytopathic bovine viral diarrhea virus genotype 1. Ten were subsequently challenged with bovine herpesvirus 1 and euthanized at 1, 2, 4, 7, or 14 days postinoculation. The other 2 calves were euthanized prior to the second inoculation. Another cohort of 10 calves was inoculated only with bovine herpesvirus 1 and then were euthanized at the same time points. Two calves were not inoculated with any agent and were used as negative controls. Pulmonary lesions were evaluated in all animals, while quantitative and biosynthetic changes in immune cells were concurrently examined immunohistochemically to compare coinfected calves and calves challenged only with bovine herpesvirus 1. Calves preinfected with bovine viral diarrhea virus demonstrated moderate respiratory clinical signs and histopathologic evidence of interstitial pneumonia with aggregates of mononuclear cells, which predominated at 4 days postinoculation. Furthermore, this group of animals was noted to have a suppression of interleukin-10 and associated alterations in the Th1-driven cytokine response in the lungs, as well as inhibition of the response of CD8+ and CD4+ T lymphocytes against bovine herpesvirus 1. These findings suggest that bovine viral diarrhea virus preinfection could affect the regulation of the immune response as modulated by regulatory T cells, as well as impair local cell-mediated immunity to secondary respiratory pathogens. PMID:25322747

  12. Eight-year results of site retention of anorganic bovine bone and anorganic bovine matrix.

    PubMed

    Degidi, Marco; Perrotti, Vittoria; Piattelli, Adriano; Iezzi, Giovanna

    2013-12-01

    The long-term fate of some biomaterials is still unknown, and the reports present in the literature are not conclusive as to whether these biomaterials are resorbed over time or not. Different reports can be found with regard to the resorption behavior of anorganic bovine bone (ABB). The aim of the present study was to provide a comparative histological and histomorphometrical evaluation, in the same patient, of 2 specimens retrieved from a sinus augmented with ABB and with anorganic bovine matrix added to a cell-binding peptide (PepGen P-15), respectively, after a healing period of 6 months and after 8 years of implant loading, to evaluate the resorption of both biomaterials. A unilateral sinus augmentation procedure with ABB (50%) and with PepGen P-15 (50%) was performed in a 54-year-old male patient. Two titanium dental implants with a sandblasted and acid-etched surface were inserted after 6 months. During this procedure, 2 tissue cores were retrieved from the sinus with a trephine, before implant insertion. After an additional 6 months, a fixed prosthetic restoration was fabricated. One of these implants, after a loading period of 8 years, fractured in the coronal portion and was removed. Both specimens, one retrieved after a 6-month healing period and the other after an 8-year loading period, were treated to obtain thin ground sections. In the 6-month specimen, the histomorphometry showed that the percentage of newly formed bone was 27.2% ± 3.6%, marrow spaces 35.6% ± 2.3%, residual ABB particles 25.1% ± 1.2%, and residual PepGen P-15 particles 12.1% ± 2.2%. In the 8-year specimen, the histomorphometry showed that the percentage of newly formed bone was 51.4% ± 4.8%, marrow spaces 40% ± 7.1%, residual ABB particles 6.2% ± 0.7%, and residual PepGen P-15 particles 2.4% ± 0.5%. Both biomaterials underwent significant resorption over the course of this study. PMID:22103882

  13. High resolution radiation hybrid maps of bovine chromosomes 19 and 29: comparison with the bovine genome sequence assembly

    PubMed Central

    Prasad, Aparna; Schiex, Thomas; McKay, Stephanie; Murdoch, Brenda; Wang, Zhiquan; Womack, James E; Stothard, Paul; Moore, Stephen S

    2007-01-01

    Background High resolution radiation hybrid (RH) maps can facilitate genome sequence assembly by correctly ordering genes and genetic markers along chromosomes. The objective of the present study was to generate high resolution RH maps of bovine chromosomes 19 (BTA19) and 29 (BTA29), and compare them with the current 7.1X bovine genome sequence assembly (bovine build 3.1). We have chosen BTA19 and 29 as candidate chromosomes for mapping, since many Quantitative Trait Loci (QTL) for the traits of carcass merit and residual feed intake have been identified on these chromosomes. Results We have constructed high resolution maps of BTA19 and BTA29 consisting of 555 and 253 Single Nucleotide Polymorphism (SNP) markers respectively using a 12,000 rad whole genome RH panel. With these markers, the RH map of BTA19 and BTA29 extended to 4591.4 cR and 2884.1 cR in length respectively. When aligned with the current bovine build 3.1, the order of markers on the RH map for BTA19 and 29 showed inconsistencies with respect to the genome assembly. Maps of both the chromosomes show that there is a significant internal rearrangement of the markers involving displacement, inversion and flips within the scaffolds with some scaffolds being misplaced in the genome assembly. We also constructed cattle-human comparative maps of these chromosomes which showed an overall agreement with the comparative maps published previously. However, minor discrepancies in the orientation of few homologous synteny blocks were observed. Conclusion The high resolution maps of BTA19 (average 1 locus/139 kb) and BTA29 (average 1 locus/208 kb) presented in this study suggest that by the incorporation of RH mapping information, the current bovine genome sequence assembly can be significantly improved. Furthermore, these maps can serve as a potential resource for fine mapping QTL and identification of causative mutations underlying QTL for economically important traits. PMID:17784962

  14. Detection and Characterisation of Lactobacillus spp. in the Bovine Uterus and Their Influence on Bovine Endometrial Epithelial Cells In Vitro

    PubMed Central

    Gärtner, Martina A.; Bondzio, Angelika; Braun, Nicole; Jung, Markus; Einspanier, Ralf; Gabler, Christoph

    2015-01-01

    Bacterial infections and inflammation of the uterus are common in dairy cattle after parturition. In particular, pathogenic bacteria that cause endometritis have been the focus of research in cattle reproduction in the last ten years. The aim of the present study was to identify commensal lactobacilli in the bovine uterus and to examine their influence on the synthesis of pro-inflammatory factors in bovine endometrial epithelial cells in vitro. Lactobacillus species were isolated from healthy bovine uteri and further characterised. Bovine endometrial epithelial cells in the second passage (n = 5 animals) were co-cultured with the autochthonous isolates L. buchneri, L. ruminis and L. amylovorus as well as with a commercially available L. vaginalis in different multiplicities of infection (MOI = 1, 5 and 10, respectively). Endometrial epithelial cells cultured without bacteria served as controls. At distinct points in time (2, 4 and 6 h) total RNA was extracted from co-cultured epithelial cells and subjected to reverse transcription quantitative PCR of pro-inflammatory factors. Furthermore, the release of such factors by co-cultured epithelial cells was measured by ELISA or EIA after 24 and 48 h. L. ruminis and L. amylovorus induced increased interleukin (IL) IL1A, IL6, IL8 and prostaglandin-endoperoxide synthase 2 mRNA levels and the release of IL8 and prostaglandin F2α in endometrial epithelial cells compared with control cells. In contrast, L. buchneri did not significantly influence the expression and release of these factors. Toll-like receptors 2 and 6 transcripts were found unchanged in co-cultured and untreated epithelial cells in vitro. However, endometrial epithelial cells of each animal showed individual differences in the response to bacterial load. These results suggest that Lactobacillus species are present in the bovine uterus, revealing immunomodulatory properties. PMID:25803719

  15. Detection and characterisation of Lactobacillus spp. in the bovine uterus and their influence on bovine endometrial epithelial cells in vitro.

    PubMed

    Gärtner, Martina A; Bondzio, Angelika; Braun, Nicole; Jung, Markus; Einspanier, Ralf; Gabler, Christoph

    2015-01-01

    Bacterial infections and inflammation of the uterus are common in dairy cattle after parturition. In particular, pathogenic bacteria that cause endometritis have been the focus of research in cattle reproduction in the last ten years. The aim of the present study was to identify commensal lactobacilli in the bovine uterus and to examine their influence on the synthesis of pro-inflammatory factors in bovine endometrial epithelial cells in vitro. Lactobacillus species were isolated from healthy bovine uteri and further characterised. Bovine endometrial epithelial cells in the second passage (n = 5 animals) were co-cultured with the autochthonous isolates L. buchneri, L. ruminis and L. amylovorus as well as with a commercially available L. vaginalis in different multiplicities of infection (MOI = 1, 5 and 10, respectively). Endometrial epithelial cells cultured without bacteria served as controls. At distinct points in time (2, 4 and 6 h) total RNA was extracted from co-cultured epithelial cells and subjected to reverse transcription quantitative PCR of pro-inflammatory factors. Furthermore, the release of such factors by co-cultured epithelial cells was measured by ELISA or EIA after 24 and 48 h. L. ruminis and L. amylovorus induced increased interleukin (IL) IL1A, IL6, IL8 and prostaglandin-endoperoxide synthase 2 mRNA levels and the release of IL8 and prostaglandin F(2α) in endometrial epithelial cells compared with control cells. In contrast, L. buchneri did not significantly influence the expression and release of these factors. Toll-like receptors 2 and 6 transcripts were found unchanged in co-cultured and untreated epithelial cells in vitro. However, endometrial epithelial cells of each animal showed individual differences in the response to bacterial load. These results suggest that Lactobacillus species are present in the bovine uterus, revealing immunomodulatory properties. PMID:25803719

  16. Performance Assessment PCR-Based Assays Targeting Bacteroidales Genetic Markers of Bovine Fecal Pollution▿

    PubMed Central

    Shanks, Orin C.; White, Karen; Kelty, Catherine A.; Hayes, Sam; Sivaganesan, Mano; Jenkins, Michael; Varma, Manju; Haugland, Richard A.

    2010-01-01

    There are numerous PCR-based assays available to characterize bovine fecal pollution in ambient waters. The determination of which approaches are most suitable for field applications can be difficult because each assay targets a different gene, in many cases from different microorganisms, leading to variation in assay performance. We describe a performance evaluation of seven end-point PCR and real-time quantitative PCR (qPCR) assays reported to be associated with either ruminant or bovine feces. Each assay was tested against a reference collection of DNA extracts from 247 individual bovine fecal samples representing 11 different populations and 175 fecal DNA extracts from 24 different animal species. Bovine-associated genetic markers were broadly distributed among individual bovine samples ranging from 39 to 93%. Specificity levels of the assays spanned 47.4% to 100%. End-point PCR sensitivity also varied between assays and among different bovine populations. For qPCR assays, the abundance of each host-associated genetic marker was measured within each bovine population and compared to results of a qPCR assay targeting 16S rRNA gene sequences from Bacteroidales. Experiments indicate large discrepancies in the performance of bovine-associated assays across different bovine populations. Variability in assay performance between host populations suggests that the use of bovine microbial source-tracking applications will require a priori characterization at each watershed of interest. PMID:20061457

  17. Subcellular Localization and Polymorphism of Bovine FABP4 in Bovine Intramuscular Adipocytes.

    PubMed

    Yonekura, Shinichi; Hirota, Shohei; Miyazaki, Honami; Tokutake, Yukako

    2016-01-01

    Fatty acid binding protein 4 (FABP4) I74 V, a gene polymorphism associated with unsaturated fatty acid contents, was discovered in Japanese Black cattle. Individuals with FABP4 I/I genotype contain a significantly high level of palmitoleic acid compared to those with FABP4 V/V genotype. It remains unknown how the FABP4 polymorphism leads to different palmitoleic acid contents. We overexpressed FABP4 of different genotypes in bovine intramuscular preadipocytes and examined whether the intracellular localization of FABP4 and the expression levels of lipid metabolism-related genes were different among cells expressing different genotypes. Nuclear localization was observed for the FABP4 V/V, while the FABP4 I/I almost did not. The cells expressing FABP4 of different genotypes were comparable in terms of the expression levels of genes involved in lipid metabolism. FABP4 I/I was localized in most of the lipid droplets 4 days after differentiation induction, whereas approximately 25% lipid droplet co-localized with FABP4 in cells expressing FABP4 V/V. The lipid droplet size increased when palmitoleic acid was added compared to the size observed when palmitic acid was added. These results suggest that lipid droplet enlargement caused by palmitoleic acid and genotype-dependent differences in the fatty acid transporting capacity underlie variations in palmitoleic acid content among FABP4 polymorphisms. PMID:26913550

  18. Whole genome analysis of Japanese bovine toroviruses reveals natural recombination between porcine and bovine toroviruses.

    PubMed

    Ito, Mika; Tsuchiaka, Shinobu; Naoi, Yuki; Otomaru, Konosuke; Sato, Mitsuo; Masuda, Tsuneyuki; Haga, Kei; Oka, Tomoichiro; Yamasato, Hiroshi; Omatsu, Tsutomu; Sugimura, Satoshi; Aoki, Hiroshi; Furuya, Tetsuya; Katayama, Yukie; Oba, Mami; Shirai, Junsuke; Katayama, Kazuhiko; Mizutani, Tetsuya; Nagai, Makoto

    2016-03-01

    Bovine toroviruses (BToVs), belong to the subfamily Toroviridae within the family Coronaviridae, and are pathogens, causing enteric disease in cattle. In Japan, BToVs are distributed throughout the country and cause gastrointestinal infection of calves and cows. In the present study, complete genome sequences of two Japanese BToVs and partial genome sequences of two Japanese BToVs and one porcine torovirus (PToV) from distant regions in Japan were determined and genetic analyses were performed. Pairwise nucleotide comparison and phylogenetic analyses revealed that Japanese BToVs shared high identity with each other and showed high similarities with BToV Breda1 strain in S, M, and HE coding regions. Japanese BToVs showed high similarities with porcine toroviruses in ORF1a, ORF1b, and N coding regions and the 5' and 3' untranslated regions, suggestive of a natural recombination event. Recombination analyses mapped the putative recombinant breakpoints to the 3' ends of the ORF1b and HE regions. These findings suggest that the interspecies recombinant nature of Japanese BToVs resulted in a closer relationship between BToV Breda1 and PToVs. PMID:26708248

  19. THE ABILITY OF MYCOBACTERIUM AVIUM SUBSP. PARATUBERCULOSIS TO ENTER BOVINE EPITHELIAL CELLS IS INFLUENCED BY PREEXPOSURE TO A HYPEROSMOLAR ENVIRONMENT AND INTRACELLULAR PASSAGE IN BOVINE MAMMARY EPITHELIAL CELLS

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Mycobacterium avium subsp. paratuberculosis is the cause of Johne’s disease in cattle and other ruminants. M. avium subsp. paratuberculosis infection of the bovine host is not well understood; however, it is assumed that crossing the bovine intestinal mucosa is important in order for M. avium subsp...

  20. Loci on Bos taurus chromosome 2 and Bos taurus chromosome 26 are linked with bovine respiratory disease and associated with persistent infection of bovine viral diarrhea virus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The objective of this study was to identify loci linked with bovine respiratory disease (BRD) and subsequently to determine if these same loci were associated with bovine viral diarrhea virus persistent infection (BVD-PI) in BVD-PI calves or their dams. A genome-wide linkage study using 312 microsa...

  1. Bovine Staphylococcus aureus: diagnostic properties of specific media.

    PubMed

    Graber, H U; Pfister, S; Burgener, P; Boss, R; Meylan, M; Hummerjohann, J

    2013-08-01

    As accurate discrimination between Staphylococcus (S.) aureus and NSA (non-S. aureus staphylococci) involved in bovine mastitis is essential in terms of clinical prognosis and outcome, the aim of this study was to reevaluate the classical bacteriological procedures to identify these agents. Various media and the coagulase tube test were investigated using 116 strains of S. aureus and 115 of NSA, all isolated from cows with spontaneous intramammary infections (IMI). Furthermore, 25 NSA reference strains were analyzed. The study demonstrated that a few media were appropriate for differentiating S. aureus from NSA, provided that the staphylococci were isolated from bovine IMI. Evaluation of hemolysis further revealed that double or incomplete hemolysis are specific for S. aureus and are, therefore, a decisive diagnostic criterion. For strains showing complete hemolysis, maximal discrimination between S. aureus and NSA was observed by subculturing them on CHROMagar Staph. aureus. PMID:23548479

  2. Thioltransferase activity of bovine lens glutathione S-transferase.

    PubMed Central

    Dal Monte, M; Cecconi, I; Buono, F; Vilardo, P G; Del Corso, A; Mura, U

    1998-01-01

    A Mu-class glutathione S-transferase purified to electrophoretic homogeneity from bovine lens displayed thioltransferase activity, catalysing the transthiolation reaction between GSH and hydroxyethyldisulphide. The thiol-transfer reaction is composed of two steps, the formation of GSSG occurring through the generation of an intermediate mixed disulphide between GSH and the target disulphide. Unlike glutaredoxin, which is only able to catalyse the second step of the transthiolation process, glutathioneS-transferase catalyses both steps of the reaction. Data are presented showing that bovine lens glutathione S-transferase and rat liver glutaredoxin, which was used as a thioltransferase enzyme model, can operate in synergy to catalyse the GSH-dependent reduction of hydroxyethyldisulphide. PMID:9693102

  3. Application of Functional Genomics for Bovine Respiratory Disease Diagnostics

    PubMed Central

    Rai, Aswathy N.; Epperson, William B.; Nanduri, Bindu

    2015-01-01

    Bovine respiratory disease (BRD) is the most common economically important disease affecting cattle. For developing accurate diagnostics that can predict disease susceptibility/resistance and stratification, it is necessary to identify the molecular mechanisms that underlie BRD. To study the complex interactions among the bovine host and the multitude of viral and bacterial pathogens, as well as the environmental factors associated with BRD etiology, genome-scale high-throughput functional genomics methods such as microarrays, RNA-seq, and proteomics are helpful. In this review, we summarize the progress made in our understanding of BRD using functional genomics approaches. We also discuss some of the available bioinformatics resources for analyzing high-throughput data, in the context of biological pathways and molecular interactions. Although resources for studying host response to infection are avail-able, the corresponding information is lacking for majority of BRD pathogens, impeding progress in identifying diagnostic signatures for BRD using functional genomics approaches. PMID:26526746

  4. Critical role of cellular cholesterol in bovine rotavirus infection

    PubMed Central

    2014-01-01

    Background Bovine rotavirus (BRV) is a non-enveloped dsRNA virus that cause neonatal calf diarrhea. Lipid rafts are cholesterol-enrich membrane mircodomains that play a vital role in many cellular processes. In this study, the effect of cellular cholesterol depletion on infection of MA-104 cells with bovine rotavirus was investigated. Results We demonstrated that cholesterol depletion of the plasma membrane by MβCD had no effect on BRV binding to cells but significantly impaired BRV entry in a dose-dependent manner and the effect was partially reversed by addition of exogenous cholesterol, suggesting the reduction of BRV infection by MβCD was specifically due to cholesterol depletion. Cholesterol depletion after virus entry did not reduce BRV replication, whereas affected virus assembly. Conclusions Taken together, our results demonstrate that cell membrane cholesterol is essential to BRV infectivity. PMID:24884772

  5. Epidemiology and control of bovine herpesvirus 1 infection in Europe.

    PubMed

    Raaperi, Kerli; Orro, Toomas; Viltrop, Arvo

    2014-09-01

    Bovine herpesvirus 1 (BHV-1) causes infectious bovine rhinotracheitis (IBR), infectious pustular vulvovaginitis, abortion and balanoposthitis, as well as neurological and systemic disease in cattle. The virus is endemic in cattle populations worldwide although in Europe six countries and several regions in other countries have achieved 'IBR-free' status by implementing control measures. According to European Union (EU) directives, all member states must comply with specific requirements related to BHV-1 infection status in semen and embryos. The requirement that 'IBR-free' states restrict the importation of cattle from endemically infected regions has motivated several European countries to instigate disease eradication programmes. Despite such control measures within the EU, outbreaks of IBR persist in 'IBR-free' states contiguous with infected countries. This review presents a summary of recent research on the epidemiology of BHV-1, highlights the control measures and surveillance systems in place, and discusses the challenges facing eradication schemes. PMID:24954868

  6. Phylogenetic analysis and characterization of Korean bovine viral diarrhea viruses.

    PubMed

    Oem, Jae-Ku; Hyun, Bang-Hun; Cha, Sang-Ho; Lee, Kyoung-Ki; Kim, Seong-Hee; Kim, Hye-Ryoung; Park, Choi-Kyu; Joo, Yi-Seok

    2009-11-18

    Thirty-six bovine viral disease viruses (BVDVs) were identified in bovine feces (n=16), brains (n=2), and aborted fetuses (n=18) in Korea. To reveal the genetic diversity and characteristics of these Korean strains, the sequences of their 5'-untranslated regions (5'-UTRs) were determined and then compared with published reference sequences. Neighbor-joining phylogenetic analysis revealed that most of the Korean viruses were of the BVDV subtypes 1a (n=17) or 2a (n=17). The remaining strains were of subtypes 1b (n=1) and 1n (n=1). This analysis indicates that the 1a and 2a BVDV subtypes are predominant and widespread in Korea. In addition, the prevalence of BVDV-2 was markedly higher in aborted fetuses than in other samples and was more often associated with reproductive problems and significant mortality in cattle. PMID:19589650

  7. Long range restriction analysis of the bovine casein genes.

    PubMed Central

    Ferretti, L; Leone, P; Sgaramella, V

    1990-01-01

    Pulsed field gel electrophoresis (PFGE) was used to analyse the organization of the bovine alpha s1, alpha s2, beta and kappa casein genes. High molecular weight DNA was prepared from fibroblasts and lymphocytes embedded in agarose and was digested with the restriction endonucleases Clal, Sall, Smal, Xhol. The digestion products were separated by PFGE, transfered to nitrocellulose filters and hybridized to probes corresponding to the cDNAs of the four bovine casein genes. The casein genes were demonstrated to be physically linked within a region of 300 kb, represented by two adjacent Xhol fragments in fibroblasts and by a single fragment in lymphocytes. A restriction map of the casein locus was derived and the order of the genes was shown to be kappa, alpha s2, beta, alpha s1. Images PMID:2263448

  8. In vitro digestibility and immunoreactivity of bovine milk proteins.

    PubMed

    Do, Andrew B; Williams, Kristina; Toomer, Ondulla T

    2016-01-01

    Current models of digestibility solely utilize pepsin stability to assess the safety of allergenic food proteins. However, in vivo complete protein digestion requires acid denaturation and pepsin, trypsin, and/or chymotrypsin cleavage. This study aimed to identify the immunoreactivity and allergenicity of stable bovine milk proteins, using an improved digestibility model to simulate physiological gastric and intestinal conditions in vitro. Gel electrophoresis and immunoblot analysis were used to determine protein stability and immunoreactivity, respectively. Immunoreactivity of bovine milk proteins, β-lactoglobulin (β-LG) and casein (CN) was greatly diminished with gastric simulation (0-60 min), but some proteins were stable and immunoreactive with simulated intestinal digestive conditions (0-60 min). This study demonstrates the need for improved digestibility models for more accurate assessment of the behavior of food allergens in vivo. PMID:26213013

  9. Bovine Leukemia Virus DNA in Human Breast Tissue

    PubMed Central

    Shen, Hua Min; Jensen, Hanne M.; Choi, K. Yeon; Sun, Dejun; Nuovo, Gerard

    2014-01-01

    Bovine leukemia virus (BLV), a deltaretrovirus, causes B-cell leukemia/lymphoma in cattle and is prevalent in herds globally. A previous finding of antibodies against BLV in humans led us to examine the possibility of human infection with BLV. We focused on breast tissue because, in cattle, BLV DNA and protein have been found to be more abundant in mammary epithelium than in lymphocytes. In human breast tissue specimens, we identified BLV DNA by using nested liquid-phase PCR and DNA sequencing. Variations from the bovine reference sequence were infrequent and limited to base substitutions. In situ PCR and immunohistochemical testing localized BLV to the secretory epithelium of the breast. Our finding of BLV in human tissues indicates a risk for the acquisition and proliferation of this virus in humans. Further research is needed to determine whether BLV may play a direct role in human disease. PMID:24750974

  10. Application of Functional Genomics for Bovine Respiratory Disease Diagnostics.

    PubMed

    Rai, Aswathy N; Epperson, William B; Nanduri, Bindu

    2015-01-01

    Bovine respiratory disease (BRD) is the most common economically important disease affecting cattle. For developing accurate diagnostics that can predict disease susceptibility/resistance and stratification, it is necessary to identify the molecular mechanisms that underlie BRD. To study the complex interactions among the bovine host and the multitude of viral and bacterial pathogens, as well as the environmental factors associated with BRD etiology, genome-scale high-throughput functional genomics methods such as microarrays, RNA-seq, and proteomics are helpful. In this review, we summarize the progress made in our understanding of BRD using functional genomics approaches. We also discuss some of the available bioinformatics resources for analyzing high-throughput data, in the context of biological pathways and molecular interactions. Although resources for studying host response to infection are avail-able, the corresponding information is lacking for majority of BRD pathogens, impeding progress in identifying diagnostic signatures for BRD using functional genomics approaches. PMID:26526746

  11. Bioelectrical impedance analysis for bovine milk: Preliminary results

    NASA Astrophysics Data System (ADS)

    Bertemes-Filho, P.; Valicheski, R.; Pereira, R. M.; Paterno, A. S.

    2010-04-01

    This work reports the investigation and analysis of bovine milk quality by using biological impedance measurements using electrical impedance spectroscopy (EIS). The samples were distinguished by a first chemical analysis using Fourier transform midinfrared spectroscopy (FTIR) and flow citometry. A set of milk samples (100ml each) obtained from 17 different cows in lactation with and without mastitis were analyzed with the proposed technique using EIS. The samples were adulterated by adding distilled water and hydrogen peroxide in a controlled manner. FTIR spectroscopy and flow cytometry were performed, and impedance measurements were made in a frequency range from 500Hz up to 1MHz with an implemented EIS system. The system's phase shift was compensated by measuring saline solutions. It was possible to show that the results obtained with the Bioelectrical Impedance Analysis (BIA) technique may detect changes in the milk caused by mastitis and the presence of water and hydrogen peroxide in the bovine milk.

  12. Bovine Spongiform Encephalopathy Infectivity in Greater Kudu (Tragelaphus strepsiceros)

    PubMed Central

    Kirkwood, James K.; Dawson, Michael; Spencer, Yvonne I.; Green, Robert B.; Wells, Gerald A.H.

    2004-01-01

    Of all the species exposed naturally to the bovine spongiform encephalopathy (BSE) agent, the greater kudu (Tragelaphus strepsiceros), a nondomesticated bovine from Africa, appears to be the most susceptible to the disease. We present the results of mouse bioassay studies to show that, contrary to findings in cattle with BSE in which the tissue distribution of infectivity is the most limited recorded for any of the transmissible spongiform encephalopathies (TSE), infectivity in greater kudu with BSE is distributed in as wide a range of tissues as occurs in any TSE. BSE agent was also detected in skin, conjunctiva, and salivary gland, tissues in which infectivity has not previously been reported in any naturally occurring TSE. The distribution of infectivity in greater kudu with BSE suggests possible routes for transmission of the disease and highlights the need for further research into the distribution of TSE infectious agents in other host species. PMID:15207051

  13. Antimicrobial susceptibilities of Mycoplasma isolated from bovine mastitis in Japan.

    PubMed

    Kawai, Kazuhiro; Higuchi, Hidetoshi; Iwano, Hidetomo; Iwakuma, Akihiro; Onda, Ken; Sato, Reiichiro; Hayashi, Tomohito; Nagahata, Hajime; Oshida, Toshio

    2014-01-01

    Mycoplasma spp. are highly contagious pathogens and intramammary Mycoplasma infection is a serious issue for the dairy industry. As there is no effective vaccine for Mycoplasma infection, control depends on good husbandry and chemo-antibiotic therapy. In this study, antimicrobial susceptibility of Mycoplasma strains recently isolated from cases of bovine mastitis in Japan was evaluated by minimum inhibitory concentration (MIC). All Mycoplasma bovis strains were sensitive to pirlimycin, danofloxacin and enrofloxacin, but not kanamycin, oxytetracycline, tilmicosin or tylosin. M. californicum and M. bovigenitalium strains were sensitive to pirlimycin, danofloxacin, enrofloxacin, oxytetracycline, tilmicosin and tylosin, but not to kanamycin. This is the first report to describe the MIC of major antimicrobial agents for Mycoplasma species isolated from bovine mastitis in Japan. PMID:24261609

  14. Vaccination against δ-retroviruses: the bovine leukemia virus paradigm.

    PubMed

    Gutiérrez, Gerónimo; Rodríguez, Sabrina M; de Brogniez, Alix; Gillet, Nicolas; Golime, Ramarao; Burny, Arsène; Jaworski, Juan-Pablo; Alvarez, Irene; Vagnoni, Lucas; Trono, Karina; Willems, Luc

    2014-06-01

    Bovine leukemia virus (BLV) and human T-lymphotropic virus type 1 (HTLV-1) are closely related d-retroviruses that induce hematological diseases. HTLV-1 infects about 15 million people worldwide, mainly in subtropical areas. HTLV-1 induces a wide spectrum of diseases (e.g., HTLV-associated myelopathy/tropical spastic paraparesis) and leukemia/lymphoma (adult T-cell leukemia). Bovine leukemia virus is a major pathogen of cattle, causing important economic losses due to a reduction in production, export limitations and lymphoma-associated death. In the absence of satisfactory treatment for these diseases and besides the prevention of transmission, the best option to reduce the prevalence of d-retroviruses is vaccination. Here, we provide an overview of the different vaccination strategies in the BLV model and outline key parameters required for vaccine efficacy. PMID:24956179

  15. Bovine tuberculosis and badgers in Britain: relevance of the past.

    PubMed

    Atkins, P J; Robinson, P A

    2013-07-01

    The European badger (Meles meles) has been identified as a wildlife reservoir of bovine tuberculosis and a source of transmission to cattle in Britain and Ireland. Both behavioural ecology and statistical ecological modelling have indicated the long-term persistence of the disease in some badger communities, and this is postulated to account for the high incidence of bovine tuberculosis in cattle across large tracts of England and Wales. This paper questions this consensus by using historical cartographic evidence to show that tuberculosis in cattle had a very different spatial distribution before 1960 to the present day. Since few of the badgers collected in road traffic accidents between 1972 and 1990 had tuberculosis in counties such as Cheshire, where the disease had until shortly before that been rife in the cattle population, the role of badgers as reservoirs in spreading disease in similar counties outside the south-west of England has to be questioned. PMID:23347609

  16. One-step multiplex real time RT-PCR for the detection of bovine respiratory syncytial virus, bovine herpesvirus 1 and bovine parainfluenza virus 3

    PubMed Central

    2012-01-01

    Background Detection of respiratory viruses in veterinary species has traditionally relied on virus detection by isolation or immunofluorescence and/or detection of circulating antibody using ELISA or serum neutralising antibody tests. Multiplex real time PCR is increasingly used to diagnose respiratory viruses in humans and has proved to be superior to traditional methods. Bovine respiratory disease (BRD) is one of the most common causes of morbidity and mortality in housed cattle and virus infections can play a major role. We describe here a one step multiplex reverse transcriptase quantitative polymerase chain reaction (mRT-qPCR) to detect the viruses commonly implicated in BRD. Results A mRT-qPCR assay was developed and optimised for the simultaneous detection of bovine respiratory syncytial virus (BRSV), bovine herpes virus type 1 (BoHV-1) and bovine parainfluenza virus type 3 (BPI3 i & ii) nucleic acids in clinical samples from cattle. The assay targets the highly conserved glycoprotein B gene of BoHV-1, nucleocapsid gene of BRSV and nucleoprotein gene of BPI3. This mRT-qPCR assay was assessed for sensitivity, specificity and repeatability using in vitro transcribed RNA and recent field isolates. For clinical validation, 541 samples from clinically affected animals were tested and mRT-qPCR result compared to those obtained by conventional testing using virus isolation (VI) and/or indirect fluorescent antibody test (IFAT). Conclusions The mRT-qPCR assay was rapid, highly repeatable, specific and had a sensitivity of 97% in detecting 102 copies of BRSV, BoHV-1 and BPI3 i & ii. This is the first mRT-qPCR developed to detect the three primary viral agents of BRD and the first multiplex designed using locked nucleic acid (LNA), minor groove binding (MGB) and TaqMan probes in one reaction mix. This test was more sensitive than both VI and IFAT and can replace the aforesaid methods for virus detection during outbreaks of BRD. PMID:22455597

  17. Molecular and immunodiagnostic investigations on bovine neosporosis in Switzerland.

    PubMed

    Gottstein, B; Hentrich, B; Wyss, R; Thür, B; Busato, A; Stärk, K D; Müller, N

    1998-04-01

    Neospora caninum has gained considerable attention through its role in the aetiology of bovine abortion. Due to its close phylogenetic relationship with Toxoplasma gondii, respective unequivocal differential diagnosis deserves special consideration. In order to evaluate the diagnostic performance of molecular and immunodiagnostic techniques and to provide insights into the epidemiological significance of bovine neosporosis in Switzerland, we conducted a study on 83 cases of bovine abortion: of these, 24 (29%) foetal brains were positive by Neospora-PCR, six of these foetuses were simultaneously seropositive in Neospora-IFAT and/or somatic antigen-ELISA. Conversely, four (5%) foetal brains were considered positive by Toxoplasma-PCR, two of which were also seropositive in the Toxoplasma-P30-ELISA and/or direct agglutination test. The seroprevalence in 1689 cattle sera obtained from 113 diary farms was 11.5% (95% confidence interval: 9.2-13.8) by Neospora-somatic antigen-ELISA were and 10.7% (95% confidence interval: 8.3-12.6) by Toxoplasma-P30-ELISA. From the same samples, 1.1%, less than statistically expected, were positive in both ELISA. Within selected groups of cow-calf farms, the seroprevalence determined using the Neospora-somatic antigen-ELISA was 14% (95% confidence interval 5.0-23.0) for dams and 15% (95% confidence interval: 3.0-28.0) for offspring calves. Seroprevalences determined by Toxoplasma-P30-ELISA were 8% (95% confidence interval: 4.0-12.0) for dams and 3% (95% confidence interval: 0.3-6.0) for calves. None of the sera gave a positive reaction in both ELISA. Our data indicated that prenatal neosporosis appears as an important cause of bovine abortion in Switzerland. PMID:9602392

  18. Reaction of goats to infection with infectious bovine rhinotracheitis virus.

    PubMed

    Wafula, J S; Mushi, E Z; Wamwayi, H

    1985-07-01

    Intranasal exposure of goats to infectious bovine rhinotracheitis virus resulted in mild respiratory disease and virus reisolation from nasal secretions. No disease was produced in goats exposed to the same virus by the genital or ocular routes. There was serological evidence of contact transmission of infection from infected goats to cattle. Virus recrudescence was not detected in goats treated with dexamethasone two months after virus inoculation. PMID:2994191

  19. Successful vitrification of bovine blastocysts on paper container.

    PubMed

    Kim, Y M; Uhm, S J; Gupta, M K; Yang, J S; Lim, J-G; Das, Z C; Heo, Y T; Chung, H-J; Kong, I-K; Kim, N-H; Lee, H T; Ko, D H

    2012-09-15

    Cryopreservation of bovine embryos can be performed by a variety of methods with variable degree of success. Here, we report a new, easy to perform, simple, inexpensive, and successful method for vitrification of bovine blastocysts. In vitro produced bovine blastocysts were exposed to vitrification solution (5.5 m ethylene glycol, 10% serum and 1% sucrose) in one single step for 20 s, loaded on a paper container prepared from commonly available non-slippery, absorbent writing paper, and then were directly plunged into liquid nitrogen for storage. Vitrified blastocysts were warmed by serial rinsing in 0.5, 0.25 and 0.125 m sucrose solution for 1 min each. Results showed that one step exposure of bovine blastocysts to cryoprotective agents was sufficient to achieve successful cryopreservation. Under these conditions, more than 95% of blastocysts survived the vitrification-warming on paper containers which was significantly higher than those obtained from other containers, such as electron microscope (EM) grid (78.1%), open pulled straw (OPS; 80.2%), cryoloop (76.2%) or plastic straw (73.9%). Embryo transfer of blastocysts vitrified-warmed on paper container resulted in successful conception (19.3%) and full-term live birth of offspring (12.3%) which were lower (P < 0.05) than those obtained from non-vitrified blastocysts (38.0 and 32.7%) but were comparable (P > 0.05) to those obtained from blastocysts vitrified-warmed on EM grid (23.3 and 14.2%). Our results, therefore, suggest that paper may be an inexpensive and useful container for the cryopreservation of animal embryos. PMID:22763071

  20. Binding of several benzodiazepines to bovine serum albumin: Fluorescence study

    NASA Astrophysics Data System (ADS)

    Machicote, Roberta G.; Pacheco, María E.; Bruzzone, Liliana

    2010-10-01

    The interactions of lorazepam, oxazepam and bromazepam with bovine serum albumin (BSA) were studied by fluorescence spectrometry. The Stern-Volmer quenching constants and corresponding thermodynamic parameters Δ H, Δ G and Δ S were calculated. The binding constants and the number of binding sites were also investigated. The distances between the donor (BSA) and the acceptors (benzodiazepines) were obtained according to fluorescence resonance energy transfer and conformational changes of BSA were observed from synchronous fluorescence spectra.