Reference genomes of channel catfish and blue catfish reveal multiple pericentric chromosome inversions.

BACKGROUND: Channel catfish and blue catfish are the most important aquacultured species in the USA. The species do not readily intermate naturally but F1 hybrids can be produced through artificial spawning. F1 hybrids produced by mating channel catfish female with blue catfish male exhibit heterosis and provide an ideal system to study reproductive isolation and hybrid vigor. The purpose of the study was to generate high-quality chromosome level reference genome sequences and to determine their genomic similarities and differences. RESULTS: We present high-quality reference genome sequences for both channel catfish and blue catfish, containing only 67 and 139 total gaps, respectively. We also report three pericentric chromosome inversions between the two genomes, as evidenced by long reads across the inversion junctions from distinct individuals, genetic linkage mapping, and PCR amplicons across the inversion junctions. Recombination rates within the inversional segments, detected as double crossovers, are extremely low among backcross progenies (progenies of channel catfish female × F1 hybrid male), suggesting that the pericentric inversions interrupt postzygotic recombination or survival of recombinants. Identification of channel catfish- and blue catfish-specific genes, along with expansions of immunoglobulin genes and centromeric Xba elements, provides insights into genomic hallmarks of these species. CONCLUSIONS: We generated high-quality reference genome sequences for both blue catfish and channel catfish and identified major chromosomal inversions on chromosomes 6, 11, and 24. These perimetric inversions were validated by additional sequencing analysis, genetic linkage mapping, and PCR analysis across the inversion junctions. The reference genome sequences, as well as the contrasted chromosomal architecture should provide guidance for the interspecific breeding programs.

New tools to screen wild peanut species for aflatoxin accumulation and genetic fingerprinting.

BACKGROUND: Aflatoxin contamination in peanut seeds is still a serious problem for the industry and human health. No stable aflatoxin resistant cultivars have yet been produced, and given the narrow genetic background of cultivated peanuts, wild species became an important source of genetic diversity. Wild peanut seeds, however, are not abundant, thus, an effective method of screening for aflatoxin accumulation using minimal seeds is highly desirable. In addition, keeping record of genetic fingerprinting of each accession would be very useful for breeding programs and for the identification of accessions within germplasm collections. RESULTS: In this study, we report a method of screening for aflatoxin accumulation that is applicable to the small-size seeds of wild peanuts, increases the reliability by testing seed viability, and records the genetic fingerprinting of the samples. Aflatoxin levels observed among 20 wild peanut species varied from zero to 19000 ng.g-1 and 155 ng.g-1 of aflatoxin B1 and B2, respectively. We report the screening of 373 molecular markers, including 288 novel SSRs, tested on 20 wild peanut species. Multivariate analysis by Neighbor-Joining, Principal Component Analysis and 3D-Principal Coordinate Analysis using 134 (36 %) transferable markers, in general grouped the samples according to their reported genomes. The best 88 markers, those with high fluorescence, good scorability and transferability, are reported with BLAST results. High quality markers (total 98) that discriminated genomes are reported. A high quality marker with UPIC score 16 (16 out of 20 species discriminated) had significant hits on BLAST2GO to a pentatricopeptide-repeat protein, another marker with score 5 had hits on UDP-D-apiose synthase, and a third one with score 12 had BLASTn hits on La-RP 1B protein. Together, these three markers discriminated all 20 species tested. CONCLUSIONS: This study provides a reliable method to screen wild species of peanut for aflatoxin resistance using minimal seeds. In addition we report 288 new SSRs for peanut, and a cost-effective combination of markers sufficient to discriminate all 20 species tested. These tools can be used for the systematic search of aflatoxin resistant germplasm keeping record of the genetic fingerprinting of the accessions tested for breeding purpose.

Genome Sequences of Three Strains of Aspergillus flavus for the Biological Control of Aflatoxin.

Aflatoxin is a carcinogenic contaminant of many commodities that are infected by Aspergillus flavus Nonaflatoxigenic strains of A. flavus have been utilized as biological control agents. Here, we report the genome sequences from three biocontrol strains. This information will be useful in developing markers for postrelease monitoring of these fungi.

Naturally occurring high oleic acid cottonseed oil: identification and functional analysis of a mutant allele of Gossypium barbadense fatty acid desaturase-2.

MAIN CONCLUSION: Some naturally occurring cotton accessions contain commercially attractive seed oil fatty acid profiles. The likely causal factor for a high-oleate trait in pima cotton ( Gossypium barbadense ) accession GB-713 is described here. Vegetable oils are broadly used in the manufacture of many human and animal nutritional products, and in various industrial applications. Along with other well-known edible plant oils from soybean, corn, and canola, cottonseed oil is a valuable commodity. Cottonseed oil is a co-product derived from the processing of cottonseed fiber. In the past, it was used extensively in a variety of food applications. However, cottonseed oil has lost market share in recent years due to less than optimal ratios of the constituent fatty acids found in either traditional or partially hydrogenated oil. Increased awareness of the negative health consequences of dietary trans-fats, along with the public wariness associated with genetically modified organisms has created high demand for naturally occurring oil with high monounsaturate/polyunsaturate ratios. Here, we report the discovery of multiple exotic accessions of pima cotton that contain elevated seed oil oleate content. The genome of one such accession was sequenced, and a mutant candidate fatty acid desaturase-2 (FAD2-1D) gene was identified. The mutant protein produced significantly less linoleic acid in infiltrated Arabidopsis leaf assays, compared to a repaired version of the same enzyme. Identification of this gene provides a valuable resource. Development of markers associated with this mutant locus will be very useful in efforts to breed the high-oleate trait into agronomic fiber accessions of upland cotton.

Colistin Resistance mcr-1-Gene-Bearing Escherichia coli Strain from the United States.

Transmissible colistin resistance in the form of an mcr-1-gene-bearing plasmid has been recently reported in Enterobacteriaceae in several parts of the world. We report here the completed genome sequence of an Escherichia coli strain isolated from swine in the United States that carried the mcr-1 gene on an IncI2-type plasmid.

The channel catfish genome sequence provides insights into the evolution of scale formation in teleosts.

Catfish represent 12% of teleost or 6.3% of all vertebrate species, and are of enormous economic value. Here we report a high-quality reference genome sequence of channel catfish (Ictalurus punctatus), the major aquaculture species in the US. The reference genome sequence was validated by genetic mapping of 54,000 SNPs, and annotated with 26,661 predicted protein-coding genes. Through comparative analysis of genomes and transcriptomes of scaled and scaleless fish and scale regeneration experiments, we address the genomic basis for the most striking physical characteristic of catfish, the evolutionary loss of scales and provide evidence that lack of secretory calcium-binding phosphoproteins accounts for the evolutionary loss of scales in catfish. The channel catfish reference genome sequence, along with two additional genome sequences and transcriptomes of scaled catfishes, provide crucial resources for evolutionary and biological studies. This work also demonstrates the power of comparative subtraction of candidate genes for traits of structural significance.

Development of a Large Set of Microsatellite Markers in Zapote Mamey (Pouteria sapota (Jacq.) H.E. Moore & Stearn) and Their Potential Use in the Study of the Species.

Pouteria sapota is known for its edible fruits that contain unique carotenoids, as well as for its fungitoxic, anti-inflammatory and anti-oxidant activity. However, its genetics is mostly unknown, including aspects about its genetic diversity and domestication process. We did high-throughput sequencing of microsatellite-enriched libraries of P. sapota, generated 5223 contig DNA sequences, 1.8 Mbp, developed 368 microsatellites markers and tested them on 29 individuals from 10 populations (seven wild, three cultivated) from Mexico, its putative domestication center. Gene ontology BLAST analysis of the DNA sequences containing microsatellites showed potential association to physiological functions. Genetic diversity was slightly higher in cultivated than in the wild gene pool (HE = 0.41 and HE = 0.35, respectively), although modified Garza-Williamson Index and Bottleneck software showed evidence for a reduction in genetic diversity for the cultivated one. Neighbor Joining, 3D Principal Coordinates Analysis and assignment tests grouped most individuals according to their geographic origin but no clear separation was observed between wild or cultivated gene pools due to, perhaps, the existence of several admixed populations. The developed microsatellites have a great potential in genetic population and domestication studies of P. sapota but additional sampling will be necessary to better understand how the domestication process has impacted the genetic diversity of this fruit crop.

Metagenomic analysis of the turkey gut RNA virus community.

Viral enteric disease is an ongoing economic burden to poultry producers worldwide, and despite considerable research, no single virus has emerged as a likely causative agent and target for prevention and control efforts. Historically, electron microscopy has been used to identify suspect viruses, with many small, round viruses eluding classification based solely on morphology. National and regional surveys using molecular diagnostics have revealed that suspect viruses continuously circulate in United States poultry, with many viruses appearing concomitantly and in healthy birds. High-throughput nucleic acid pyrosequencing is a powerful diagnostic technology capable of determining the full genomic repertoire present in a complex environmental sample. We utilized the Roche/454 Life Sciences GS-FLX platform to compile an RNA virus metagenome from turkey flocks experiencing enteric disease. This approach yielded numerous sequences homologous to viruses in the BLAST nr protein database, many of which have not been described in turkeys. Our analysis of this turkey gut RNA metagenome focuses in particular on the turkey-origin members of the Picornavirales, the Caliciviridae, and the turkey Picobirnaviruses.

UPIC: Perl scripts to determine the number of SSR markers to run.

UNLABELLED: We introduce here the concept of Unique Pattern Informative Combinations (UPIC), a decision tool for the cost-effective design of DNA fingerprinting/genotyping experiments using simple-sequence/tandem repeat (SSR/STR) markers. After the first screening of SSR-markers tested on a subset of DNA samples, the user can apply UPIC to find marker combinations that maximize the genetic information obtained by a minimum or desirable number of markers. This allows a cost-effective planning of future experiments. We have developed Perl scripts to calculate all possible subset combinations of SSR markers, and determine based on unique patterns or alleles, which combinations can discriminate among all DNA samples included in a test. This makes UPIC an essential tool for optimizing resources when working with microsatellites. An example using real data from eight markers and 12 genotypes shows that UPIC detected groups of as few as three markers sufficient to discriminate all 12- DNA samples. Should markers for future experiments be chosen based only on polymorphism-information content (PIC), the necessary number of markers for discrimination of all samples cannot be determined. We also show that choosing markers using UPIC, an informative combination of four markers can provide similar information as using a combination of six markers (23 vs. 25 patterns, respectively), granting a more efficient planning of experiments. Perl scripts with documentation are also included to calculate the percentage of heterozygous loci on the DNA samples tested and to calculate three PIC values depending on the type of fertilization and allele frequency of the organism. AVAILABILITY: Perl scripts are freely available for download from http://www.ars.usda.gov/msa/jwdsrc/gbru.

An examination of antibacterial and antifungal properties of constituents described in traditional Ulster cures and remedies.

Traditional herbal cures and remedies have played an important historical role in the treatment of a variety of illnesses and diseases in Northern Ireland for the last three hundred years. Recently, these have been reviewed in the publication by Linda Ballard from the Ulster Folk and Transport Museum at Cultra, Co. Down, which details the variety of local plants used and for what purpose. From this publication and another related publication, we note the description of several plant species that consistently appear in traditional cures and remedies, particularly used to treat infections and infectious diseases. Unfortunately, although these plants have strong associations with the local historical evidence base, there are very limited and mainly no formal publications in the medical/scientific evidence base, examining their scientific background and clinical efficacy.

Development of a physical map of the soybean pathogen Fusarium virguliforme based on synteny with Fusarium graminearum genomic DNA.

BACKGROUND: Reference genome sequences within the major taxa can be used to assist the development of genomic tools for related organisms. A major constraint in the use of these sequenced and annotated genomes is divergent evolution. Divergence of organisms from a common ancestor may have occurred millions of years ago, leading to apparently un-related and un-syntenic genomes when sequence alignment is attempted. RESULTS: A series of programs were written to prepare 36 Mbp of Fusarium graminearum sequence in 19 scaffolds as a reference genome. Exactly 4,152 Bacterial artificial chromosome (BAC) end sequences from 2,178 large-insert Fusarium virguliforme clones were tested against this sequence. A total of 94 maps of F. graminearum sequence scaffolds, annotated exonic fragments and associated F. virguliforme sequences resulted. CONCLUSION: Developed here was a technique that allowed the comparison of genomes based on small, 15 bp regions of shared identity. The main power of this method lay in its ability to align diverged sequences. This work is unique in that discontinuous sequences were used for the analysis and information not readily apparent, such as match direction, are presented. The 94 maps and JAVA programs are freely available on the Web and by request.

High-throughput detection of glutathione s-transferase polymorphic alleles in a pediatric cancer population.

Polymorphisms of glutathione S-transferase (GST) enzymes have been correlated with altered risk of several cancers, as well as altered response and toxicity from cancer chemotherapy. We report a low cost, highly reproducible and specific PCR-based high-throughput assay for genotyping different GSTs designed for use in large clinical trials. In comparison to an alternative genotyping method (single nucleotide extension), the sensitivity and specificity of the high throughput assay was shown to be 92 and 97%, respectively, depending on the source of genomic DNA. Using the high-throughput assay, we demonstrate by multivariate analysis an increased risk of acute lymphoblastic leukemia, glial brain tumors, and osteosarcoma for patients carrying nonnull alleles of GSTM1 and/or GSTT1.

Strategies for genotyping: Effectiveness of tailing primers to increase accuracy in short tandem repeat determinations.

A problem associated with automated analysis of fluorescently labeled fragments separated by slab gel or capillary electrophoresis is the doublet peak formed when Taq DNA Polymerase adds a nontemplated nucleotide (generally an adenosine) to the 3' end of the product.This nontemplated addition (plus A) is primarily dependent on the 5' sequence of the reverse primer and, to a lesser extent, polymerase chain reaction (PCR) conditions. Primers may amplify the true product, the plus A product or a doublet product comprised of both. When using markers based on dinucleotide repeats, this single base pair difference can make binning and accurate automated analysis problematic. To drive the PCR reaction consistently to the plus A product, the sequence of the nonfluorescent primer used in amplification can be modified by adding a 5' tail favoring the nontemplated addition. The present study, conducted by the Fragment Analysis Research Group (FARG) of the Association of Biomolecular Resource Facilities, provided researchers with an opportunity to compare normal products amplified with a dinucleotide marker to products amplified with the same primer to which a 5' tail designed to promote the plus A product had been added. The study also included a sample amplified with a tetranucleotide repeat marker for comparison. The results from this study were returned to the FARG for comprehensive analysis and are reported here.