Relationships among pest flour beetles of the genus Tribolium (Tenebrionidae) inferred from multiple molecular markers

Model species often provide initial hypotheses and tools for studies of development, genetics and molecular evolution in closely related species. Flour beetles of the genus Tribolium Macleay (1825) are one group with potential for such comparative studies. Tribolium castaneum (Herbst 1797) is an increasingly useful developmental genetic system. The convenience with which congeneric and other species of tenebrionid flour beetles can be reared in the laboratory makes this group attractive for comparative studies on a small phylogenetic scale. Here we present the results of phylogenetic analyses of relationships among the major pest species of Tribolium based on two mitochondrial and three nuclear markers (cytochrome oxidase 1, 16S ribosomal DNA, wingless, 28S ribosomal DNA and histone H3). The utility of partitioning the dataset in a manner informed by biological structure and function is demonstrated by comparing various partitioning strategies. In parsimony and partitioned Bayesian analyses of the combined dataset, the castaneum and confusum species groups are supported as monophyletic and as each other's closest relatives. However, a sister group relationship between this clade and Tribolium brevicornis (Leconte 1859) is not supported. The inferred phylogeny provides an evolutionary framework for comparative studies using flour beetles.     

畜禽遗传资源系统保存的理论与模拟实验研究──Ⅱ．畜禽遗传资源系统保存的模拟实验

赤拟谷盗和计算机模拟实验结果,验证了畜禽遗传资源系统保存理论的可行性、合理性,及其数学模型的生物学意义。同时它还表明,系统保存群体应保持一定大小的群体有效含量,控制群体间的基因流动,并根据保存基因（或性状）的群体遗传特征,确定适宜的选择方法和交配制度。     

The development of PCR-based markers for molecular sex identification in the model insect species Tribolium castaneum

The red flour beetle, Tribolium castaneum (Herbst) (Coleoptera: Tenebrionidae), is a major pest of stored grain and cereal crops. It is also an important model species in genetic, ecological, and evolutionary research. The majority of its genome was recently sequenced and published. However, the genomic sequence of the small Y-chromosome is still undetermined, which hinders the development of molecular sex identification methods. Traditional methods for sexing adult forms of Tribolium beetles are troublesome. Therefore, a method for molecular sex identification in the red flour beetle was developed. One sex-linked randomly amplified polymorphic DNA marker was converted into a sequence-characterized amplified region (SCAR). The SCAR was aligned with the T. castaneum reference whole-genome sequence and fully matched a fragment of a single contig of unknown genomic location. The novelty of the method is that the fragment consists of shorter DNA fragments that are also present at other locations around the genome, but the particular order of these fragments within the sequenced region appeared to be Y-specific and this property was utilized for marker development. A set of three primers for multiplex PCR reaction was designed resulting in amplification of different length Y-specific and not-Y-specific (control) DNA fragments in a single PCR, which allows to distinguish males from females. The primers successfully sexed pre-sexed pupae and adult beetles from six laboratory strains, showing that the order of the repeated fragments is conserved in the species and is not strain-specific.     

Maxillopedia is the Tribolium ortholog of proboscipedia

SUMMARY Null mutations in the Drosophila melanogaster homeotic gene proboscipedia ( pb ) cause transformation of the adult labial palps to legs. The similar phenotype produced by mutations in the Tribolium castaneum homeotic complex (HOMC) gene maxillopedia ( mxp ) has led to suggestions that the two genes may be orthologous. We have cloned the Tribolium ortholog of pb , which predicts a protein with a homeodomain identical to that of Drosophila Pb. The two proteins also share several additional regions of identity, including an N-box, a motif unique to Pb orthologs. We have identified a frameshift mutation within Tribolium pb associated with an mxp null mutation, demonstrating that Tribolium pb corresponds to the mxp genetic locus. Thus, we will refer to the cloned gene as mxp . In addition, we have begun to construct a molecular map of the Tribolium HOMC. Two overlapping BAC clones which span the mxp locus also include the Tribolium labial ortholog ( Tclabial   ) and part of Tczerknüllt , indicating that the order of these genes in the HOMC is conserved between Drosophila and Tribolium.     

Molecular genetic manipulation of the red flour beetle: Genome organization and cloning of a ribosomal protein gene

We have assessed the potential for molecular genetic analysis of the red flour beetle, Tribolium castaneum, to augment the high resolution genetic analysis demonstrated for this organism by Beeman et al. (Dev. Biol.33, 196–209, 1989). The genome size, 0.21 pg, the long period interspersion pattern of repetitive elements and a low methyl-cytosine content indicate molecular studies should be straight-forward. We constructed a genomic library and isolated clones containing RPS14, a small, highly conserved, near single copy ribosomal protein gene. We found three RPS14 clones in a screen of four genome equivalents, suggesting our library is representative of the genome. We also demonstrated the use of an RFLP, identified by the RPS14 clone, in genetic mapping.     

Genetics of gene transfer between species

Bacteria transfer genetic information to members of at least three of the five biological kingdoms. Gene transfer between species may play the same role as sex between members of a single species, providing genetic diversity and material for repair of genomic damage.     

Integrated analysis of genome-wide genetic and epigenetic association data for identification of disease mechanisms

Many human diseases are multifactorial, involving multiple genetic and environmental factors impacting on one or more biological pathways. Much of the environmental effect is believed to be mediated through epigenetic changes. Although many genome-wide genetic and epigenetic association studies have been conducted for different diseases and traits, it is still far from clear to what extent the genomic loci and biological pathways identified in the genetic and epigenetic studies are shared. There is also a lack of statistical tools to assess these important aspects of disease mechanisms. In the present study, we describe a protocol for the integrated analysis of genome-wide genetic and epigenetic data based on permutation of a sum statistic for the combined effects in a locus or pathway. The method was then applied to published type 1 diabetes (T1D) genome-wide- and epigenome-wide-association studies data to identify genomic loci and biological pathways that are associated with T1D genetically and epigenetically. Through combined analysis, novel loci and pathways were also identified, which could add to our understanding of disease mechanisms of T1D as well as complex diseases in general.     

Metamorphosis and adult development of the mushroom bodies of the red flour beetle, Tribolium castaneum

The insect mushroom bodies play important roles in a number of higher processing functions such as sensory integration, higher level olfactory processing, and spatial and associative learning and memory. These functions have been established through studies in a handful of tractable model systems, of which only the fruit fly Drosophila melanogaster has been readily amenable to genetic manipulations. The red flour beetle Tribolium castaneum has a sequenced genome and has been subject to the development of molecular tools for the ready manipulation of gene expression; however, little is known about the development and organization of the mushroom bodies of this insect. The present account bridges this gap by demonstrating that the organization of the Tribolium mushroom bodies is strikingly like that of the fruit fly, with the significant exception that the timeline of neurogenesis is shifted so that the last population of Kenyon cells is born entirely after adult eclosion. Tribolium Kenyon cells are generated by two large neuroblasts per hemisphere and segregate into an early-born delta lobe subpopulation followed by clear homologs of the Drosophila gamma, alpha'/beta' and alpha/beta lobe subpopulations, with the larval-born cohorts undergoing dendritic reorganization during metamorphosis. BrdU labeling and immunohistochemical staining also reveal that a proportion of individual Tribolium have variable numbers of mushroom body neuroblasts. If heritable, this variation represents a unique opportunity for further studies of the genetic control of brain region size through the control of neuroblast number and cell cycle dynamics.     

The 3Cs provide a novel concept of bacterial species: messages from the genome as illustrated by Salmonella

A key issue troubling bacterial taxonomy and systematics is the lack of a biological species definition. Criteria to be used for defining bacterial species on genetic and biological bases should be able to reveal clear-cut boundaries among clusters of bacteria. To date, DNA–DNA re-association assays and ribosomal RNA sequence comparison have been useful in determining relative evolutionary distances among bacteria but the data are continuous and thus cannot define bacterial clusters as taxonomic units to be called species. Using Salmonella as models, we have looked for definite genetic and biologic uniqueness of clusters of bacteria. Based on our findings that each Salmonella lineage has a unique genome structure shared by strains of the same lineage but not overlapping with strains of other Salmonella lineages, we conclude that this is a result of genetic isolation following divergence of the bacteria. We propose that there should be genetic boundaries between different species of bacteria at the genomic level, which awaits further genomic information for validation.     

The genomes and transposable elements in plants: are they friends or foes?

Transposable elements (TEs) are mobile genetic elements that are present in prokaryotes and eukaryotes. The ubiquity and abundance of these self-replicating entities, bereft of cellular function, had earned them the label of ‘genomic parasites’. However, the status of TEs has been revised, with ample genomic and biological evidence now portraying them as “genomic gold”. They are perceived as a major participant in the evolution of species. This review addresses the classification of TEs as well as their role and significance in the evolution of genomes, genetic diversity, gene regulation, and exaptation of contemporary species of the plant and animal kingdoms.     

Identification of microsatellite markers in the red flour beetle,Tribolium castaneum

We isolated and characterized microsatellite markers for the red flour beetle Tribolium castaneum, an important model species for studies in various areas of evolutionary biology and ecology. A microsatellite‐enriched genomic library was constructed and screened with single stranded oligonucleotide probes [(CCT)₁₇, (AAT)₁₇ and (CAG)₁₇]. Forty‐five primer pairs were designed of which 19 pairs produced successful amplification. Polymorphism screening involved beetles from five beetle strains and revealed 15 polymorphic and four monomorphic markers. The development of polymorphic microsatellite markers will facilitate future ecological and genetic studies involving T. castaneum beetles.     

Efficient transformation of the beetle Tribolium castaneum using the Minos transposable element: quantitative and qualitative analysis of genomic integration events

Genetic transformation in insects holds great promise as a tool for genetic manipulation in species of particular scientific, economic, or medical interest. A number of transposable elements have been tested recently as potential vectors for transformation in a range of insects. Minos is one of the most promising elements because it appears to be active in diverse species and has the capacity to carry large inserts. We report here the use of the Minos element as a transformation vector in the red flour beetle Tribolium castaneum (Coleoptera), an important species for comparative developmental and pest management studies. Transgenic G(1) beetles were recovered from 32.4% of fertile G(0)'s injected with a plasmid carrying a 3xP3-EGFP-marked transposon and in vitro synthesized mRNA encoding the Minos transposase. This transformation efficiency is 2.8-fold higher than that observed when using a plasmid helper. Molecular and genetic analyses show that several independent insertions can be recovered from a single injected parent, but that the majority of transformed individuals carry single Minos insertions. These results establish Minos as one of the most efficient vectors for genetic transformation in insects. In combination with piggyBac-based transgenesis, our work allows the introduction of sophisticated multicomponent genetic tools in Tribolium.     

Genome-wide prediction of genetic interactions in a metazoan

Genetic interactions provide information about genes and processes with overlapping functions in biological systems. For Saccharomyces cerevisiae, computational integration of multiple types of functional genomic data is used to generate genome-wide predictions of genetic interactions. However, this methodology cannot be applied to the vastly more complex genome of metazoans, and only recently has the first metazoan genome-wide prediction of genetic interactions been reported. The prediction for Caenorhabditis elegans was generated by computationally integrating functional genomic data from S. cerevisiae, C. elegans and Drosophila melanogaster. This achievement is an important step toward system-level understanding of biological systems and human diseases.     

USING POPULATION GENOMICS TO DETECT SELECTION IN NATURAL POPULATIONS: KEY CONCEPTS AND METHODOLOGICAL CONSIDERATIONS

Natural selection shapes patterns of genetic variation among individuals, populations, and species, and it does so differentially across genomes. The field of population genomics provides a comprehensive genome-scale view of the action of selection, even beyond traditional model organisms. However, even with nearly complete genomic sequence information, our ability to detect the signature of selection on specific genomic regions depends on choosing experimental and analytical tools appropriate to the biological situation. For example, processes that occur at different timescales, such as sorting of standing genetic variation, mutation-selection balance, or fixed interspecific divergence, have different consequences for genomic patterns of variation. Inappropriate experimental or analytical approaches may fail to detect even strong selection or falsely identify a signature of selection. Here we outline the conceptual framework of population genomics, relate genomic patterns of variation to evolutionary processes, and identify major biological factors to be considered in studies of selection. As data-gathering technology continues to advance, our ability to understand selection in natural populations will be limited more by conceptual and analytical weaknesses than by the amount of molecular data. Our aim is to bring critical biological considerations to the fore in population genomics research and to spur the development and application of analytical tools appropriate to diverse biological systems.     

Postgenomics of Tribolium: Targeting the endocrine regulation of diuresis

Coleopteran insects comprise a highly successful taxon, representing more than 25% of eukaryotic species, many of which are economically important. One of these, the red flour beetle (Tribolium castaneum H.), is a major pest of stored grain and cereal products and is also an excellent genetic model for the Coleoptera. The Tribolium genome sequencing project was recently completed, the first for an agricultural insect pest species. This genome sequence and annotation, in combination with the tractability of Tribolium for genetic dissection and functional genomic analysis, makes it one of the most useful subjects for studies of insect development, genetics and physiology. Tribolium has a number of interesting physiological adaptations, including desiccation tolerance associated with specialized cryptonephridial organs for active rectal absorption of atmospheric water. The study of endocrine regulation of diuresis in this species has led to the identification of a number of osmoregulatory peptides, including the novel arginine‐vasopressin‐like peptide.     

Genetic analysis of the homeotic gene complex (HOM-C) in the beetle Tribolium castaneum

Our laboratories have undertaken both genetic and molecular studies of the homeotic gene complex (HOM-C) of the beetle Tribolium castaneum, and this paper discusses results from our genetic analyses. We describe here the adult phenotypes and complementation behavior of over 50 new mutations. Many of these homeotic phenotypes resemble those of Drosophila melanogaster, but few precisely parallel the segmental transformations seen in this fly. Analysis of putative loss-of-function mutations affecting the head and thorax suggests that the maxillopedia and Cephalothorax genes most closely resemble proboscipedia and Sex combs reduced of Drosophila. In the abdomen, putative loss-of-function alleles of Abdominal affect a domain corresponding to those of the combined abdominal-A and Abdominal-B genes of Drosophila. In contrast to the situation in flies, Abdominal loss-of-function variants in Tribolium cause anteriorward transformations in A3-A5a, but posteriorward transformations in A5p-A7. The implications of the differences in developmental strategies evolved in Tribolium vs Drosophila are discussed.