The first complete <Emphasis Type="Italic">Mag</Emphasis> family retrotransposons discovered in <Emphasis Type="Italic">Drosophila</Emphasis>

A retrotransposon of the Mag family was found in the Drosophila simulans genome for the first time. We also identified novel transposable elements representing the Mag family in seven Drosophila species. The high similarity between the 3’ and 5’ long terminal repeats in the found copies of transposable elements indicates that their retrotransposition has occurred relatively recently. Thus, the Mag family of retrotransposons is quite common for the genus Drosophila.     

Long terminal repeat retrotransposons of <Emphasis Type="Italic">Mus musculus</Emphasis>

Background

Long terminal repeat (LTR) retrotransposons make up a large fraction of the typical mammalian genome. They comprise about 8% of the human genome and approximately 10% of the mouse genome. On account of their abundance, LTR retrotransposons are believed to hold major significance for genome structure and function. Recent advances in genome sequencing of a variety of model organisms has provided an unprecedented opportunity to evaluate better the diversity of LTR retrotransposons resident in eukaryotic genomes.

Results

Using a new data-mining program, LTR_STRUC, in conjunction with conventional techniques, we have mined the GenBank mouse (Mus musculus) database and the more complete Ensembl mouse dataset for LTR retrotransposons. We report here that the M. musculus genome contains at least 21 separate families of LTR retrotransposons; 13 of these families are described here for the first time.

Conclusions

All families of mouse LTR retrotransposons are members of the gypsy-like superfamily of retroviral-like elements. Several different families of unrelated non-autonomous elements were identified, suggesting that the evolution of non-autonomy may be a common event. High sequence similarity between several LTR retrotransposons identified in this study and those found in distantly-related species suggests that horizontal transfer has been a significant factor in the evolution of mouse LTR retrotransposons.

     

Homology-dependent inactivation of LTR retrotransposons in <Emphasis Type="Italic">Aspergillus fumigatus</Emphasis> and <Emphasis Type="Italic">A. nidulans</Emphasis> genomes

The repeat-induced point mutation mechanism (RIP) is the most intriguing among the known mechanisms of homology-dependent gene inactivation (silencing) because of its ability to produce irreversible mutations in repetitive DNA sequences. Discovered for the first time in Neurospora crassa, RIP is characterized by C:G to T:A transitions in duplicated sequences. The mechanisms and range of occurrence of RIP are still poorly understood. Mobile elements, including retrotransposons, are a common target for the processes that lead to homology-dependent silencing because of their ability to propagate themselves. Comparative analysis of LTR retrotransposons was performed throughout the genomes of two ascomycetes, Aspergillus fumigatus and A. nidulans. “De-RIP” retroelements were reconstructed on the basis of several copies. CpG, CpA, and TpG sites, which are potential targets for mutagenesis, were found at a much lower frequency in mobile elements than in structural genes. The dinucleotide targets of the two species are affected by RIP at different frequencies: mutagenesis occurs at both CpG and CpA sites in A. fumigatus and is confined to CpG dinucleotides in A. nidulans. This work provides a theoretical background for planning the experimental investigation of RIP inactivation in aspergilli.     

Polymorphism of <Emphasis Type="Italic">yellow</Emphasis> locus in <Emphasis Type="Italic">Drosophila melanogaster</Emphasis> mutants from natural populations

We have studied the molecular characteristics of the yellow locus (y; 1–0.0), which determines the body color of phenotypically wild-type and mutant alleles isolated in different years from geographically distant populations of Drosophila melanogaster. According to the Southern blot, data restriction maps of the yellow locus of all examined strains differ from one another, as well as from Oregon stock. FISH analysis shows that, in the neighborhood of the yellow locus in the X chromosome, neither P nor hobo elements are found in y^1–775 stock, while only hobo is found in these region in y^1–859 and y^1–866 stocks, only the P element is found in y⁺sn⁸⁴⁹ stock, and both elements are found in y^1–719 stock. Thus, all yellow mutants studied are of independent origin. Locus yellow located on the end of X chromosome (region 1A5–8 on the cytologic map) carries significantly more transposon than retrotransposon induced mutations compared to the white locus (region 3C2). It is possible that, at the ends of Drosophila melanogaster chromosomes, transposons are more active than retrotransposons.     

Retrotransposons in <Emphasis Type="Italic">Betula nana</Emphasis>, and interspecific relationships in the Betuloideae,based on inter-retrotransposon amplified polymorphism (IRAP) markers

The Betulaceae family comprises two subfamilies, Betuloideae and Corylaceae. The subfamily Betuloideae contains two genera, Alnus Mill. and Betula L. Twenty putative long terminal repeat (LTR) retrotransposons were mined from 171 scaffolds containing 5,208,995 bp of dwarf birch (Betula nana) genome sequences. Five retrotransposons were finally selected after filtering the retrotransposon canonical features and nucleotide similarities between left and right LTR sequences. Of the five retroelements, three elements were found to be Ty1/Copia retrotransposons; identity of the other two elements could not be ascertained due to sequence undetermined ‘N’ bases in the sequence database. Inter-retrotranposon amplified polymorphism (IRAP) analysis, based on the LTR sequences of the mined LTR-retrotransposons, produced 179 discernible IRAP bands among the Alnus and Betula genera. Sequence analysis revealed no size homoplasy among the homologous IRAP bands. Phylogenetic and principle coordinate analysis, based on the band sharing among the taxa, showed the species in two different genera were clearly separated. The subgenera in each genus of Alnus and Betula were also distinguishable from the IRAP profiles. In the genus Betula, the species in subgenus Betula showed mixed clustering between species. This is incongruent with the phylogeographical distribution of the species.     

Emerging Antifungal Drug Resistance in <Emphasis Type="Italic">Aspergillus fumigatus</Emphasis> and Among Other Species of <Emphasis Type="Italic">Aspergillus</Emphasis>

Purpose of Review

The purpose of this review is to give an overview of recent findings on antifungal resistance in Aspergillus fumigatus (the major causative agent of aspergillosis) and sibling Aspergillus species, which can be hidden agents of aspergillosis.

Recent Findings

Azole resistance by Cyp51A mutation in A. fumigatus is a growing problem worldwide. The resistance can occur in patients or in the environment. The former occurs by drug selection in the host, inducing mutations in Cyp51A. The latter is characterized by a tandem repeat in the promoter region of cyp51A gene and mutation(s) in Cyp51A. Environmental resistant strains are prevailing rapidly and globally. Moreover, efflux pump and biofilm formation are closely related with antifungal resistance of A. fumigatus. Finally, sibling species of Aspergillus are described with regard to antifungal resistance.

Summary

Environmental azole-resistant strains have newly emerged and been dispersed globally, and continuous survey and countermeasures are urgently needed against these strains. Although the contributions of Cyp51A and efflux pumps to antifungal resistance are becoming clear, other resistance mechanisms remain unclear. Further investigations including genome comparisons will help to clarify the novel resistant mechanisms and to develop countermeasures or novel antifungal drugs against resistant strains of A. fumigatus and other Aspergillus species that have low susceptibility to antifungal therapeutics.

     

Molecular evolution of mobile elements of the <Emphasis Type="Italic">gypsy</Emphasis> group: A homolog of the <Emphasis Type="Italic">gag</Emphasis> gene in <Emphasis Type="Italic">Drosophila</Emphasis>

Retrotransposons of the gypsy group of Drosophila melanogaster that are structurally similar to retroviruses of vertebrates occupy an important place among retroelements of eukaryotes. The infectious abilities of some retrotransposons of this group (gypsy, ZAM, and Idefix) have been demonstrated experimentally, and therefore they are true retroviruses. It is supposed that retrotransposons can evolve acquiring new components, the sources of which remain to be elucidated. In this work, the CG4680gene (Gag related protein, Grp) homologous to gag of retrotransposons of the gypsy group has been identified in the genome of D. melanogaster and characterized. The Grp gene product has a highly conserved structure in different species of the Drosophilidae family and is under of purifying selection, which suggests its important genomic function in Drosophila. In view of the earlier data, it can be concluded that homologous genes of all components of gypsy retrotransposons are present in the Drosophila genome. These genes can be both precursors and products of domestication of retrovirus genes.     

Regularory elements of the <Emphasis Type="Italic">copia</Emphasis> retrotransposon determine different levels of expression in different organs of males and females of <Emphasis Type="Italic">Drosophila melanogaster</Emphasis>

Using a model system in which the expression of the reporter gene lacZ is under the control of five deleted variants of the copia retrotransposon regulatory region, which includes the 5′-long terminal repeat (LTR) and the 5′-untranslated region (5′-UTR), their contribution to the control of retrotransposon activity in different organs of males and females of Drosophila melanogaster was analyzed. The whole regulatory region provides expression of the reporter gene at the embryonic stage, and in larvae and adult flies only in generative organs. The 5′-end of LTR harbors a positive regulator that determines expression of the retrotransposon in organs of all types. The 3′-end of LTR harbors a negative regulator, which is sex- and time-specific: it represses copia expression in generative organs of males at all stages of development, and only at the imaginal stage in somatic tissues, without any effect on the expression of the retrotransposon in females. 5′-UTR contains a negative regulator of copia expression: it decreases the expression in embryos and generative organs and blocks it in somatic tissues. It may be suggested that a complex set of regulatory elements was formed in the course of the evolution of the retrotransposon, which made it possible to maintain a certain level of its expression in different types of cells and tissues and at different stages of development and, thus, to limit the harm caused to the host and provide the possibility for the retrotransposon to exist in the host genome over many generations.     

Study of the Interaction between HP1 Family Proteins and Untranslated Regulatory Regions of the <Emphasis Type="Italic">Gypsy</Emphasis> Retrotransposons in <Emphasis Type="Italic">Drosophila melanogaster</Emphasis>

In vitro interaction between recombinant protein paralogs (HP1a, HP1b, and HP1c) of the HP1 family and 5′-untranslated regulatory regions of the gypsy retrotransposon group in Drosophila melanogaster—gypsy, Springer, Tirant, ZAM, Rover, and 17.6—was studied. Using competitive DNA, the conditions that enable specific binding with a matrix were identified. It was found that HP1 family proteins efficiently bind to the 5′-untranslated regulatory region of retrotransposons with tandem repeats. It was found that repeats are absolutely necessary for HP1a to bind to the 5′-untranslated region of the Tirant and ZAM mobile genetic elements. The absence of repeats (ZAM) or the presence of fewer than two repeats (Tirant) makes such interaction impossible. Thus, the presence of tandem repeats in the 5′-untranslated region of the gypsy retrotransposons is an important tool for regulating their transposition by heterochromatin proteins.     

LTR retrotransposons from the <Emphasis Type="Italic">Citrus x clementina</Emphasis> genome: characterization and application

Long terminal repeat retrotransposons (LTR-RTs) are a large portion of most plant genomes, and can be used as a powerful molecular marker system. The first citrus reference genome (Citrus x clementina) has been publicly available since 2011; however, previous studies in citrus have not utilized the whole genome for LTR-RT marker development. In this study, 3959 full-length LTR-RTs were identified in the C. x clementina genome using structure-based (LTR_FINDER) and homology-based (RepeatMasker) methods. LTR-RTs were first classified by protein domain into Gypsy and Copia superfamilies, and then clustered into 1074 families based on LTR sequence similarity. Three hundred fifty Copia families were grouped into four lineages: Retrofit, Tork, Sire, and Oryco. One hundred seventy-eight Gypsy families were sorted into six lineages: Athila, Tat, Renia, CRM, Galadriel, and Del. Most LTR-RTs (3218 or 81.3%) were anchored to the nine Clementine mandarin linkage groups, accounting for 9.74% of chromosomes currently assembled. Accessions of 25 Rutaceae species were genotyped using 17 inter-retrotransposon amplified polymorphism (IRAP) markers developed from conserved LTR regions. Sequence-specific amplified polymorphism (SSAP) makers were used to distinguish ‘Valencia’ and ‘Pineapple’ sweet oranges (C. x sinensis), and 24 sweet orange clones. LTR-RT markers developed from the Clementine genome can be transferred within the Rutaceae family demonstrating that they are an excellent tool for citrus and Rutaceae genetic analysis.     

New sterigmatocystin-producing species of <Emphasis Type="Italic">Aspergillus</Emphasis> section <Emphasis Type="Italic">Versicolores</Emphasis> from indoor air in Croatia

Multilocus DNA sequence-based identification methods raised the number of known species assigned to the Aspergillus section Versicolores. Currently, there are 16 species accepted in the section, including A. amoenus, A. austroafricanus, A. creber, A. cvjetkovicii, A. fructus, A. griseoaurantiacus, A. hongkongensis, A. jensenii, A. protuberus, A. puulaauensis, A. subversicolor, A. sydowii, A. tabacinus, A. tennesseensis, A. venenatus, and A. versicolor. Based on morphological identifications, most of these species were identified as either A. sydowii or A. versicolor, with the latter reported to have a world-wide distribution, growing in many habitats. Aspergillus versicolor has been implicated in health hazards including sick building syndrome, human and animal mycoses, and contamination of food and feed were assigned primarily to this species. A. versicolor is still commonly isolated from indoor surveys, even though species such as A. jensenii and A. creber seem more common. From indoor air samples collected at a grain mill in Croatia, we isolated an undescribed species assigned to the Aspergillus section Versicolores. A polyphasic approach, including sequence-based methods, morphological and physiological studies, was used for species characterization and in this paper is described as Aspergillus pepii. Additionally, sterigmatocystin producing abilities have been confirmed. Based on a combined phylogenetic tree, morphological features and sterigmatocystin producing abilities, A. pepii is closely related to A. versicolor. Further studies should explore the frequency of the species in indoor environments and its medical, industrial, and environmental significance.     

The role of putrescine against the long terminal repeat (LTR) retrotransposon polymorphisms induced by salinity stress in <Emphasis Type="Italic">Triticum aestivum</Emphasis>

This study aimed to research the impact of putrescine against the long terminal repeat (LTR) retrotransposon polymorphisms (Nikita-E2647, Sukkula, Stowaway, WLTR2105 and 5′LTR) induced by salinity stress in Triticum aestivum using inter-retrotransposon amplified polymorphism (IRAP) assay. The results showed that the LTR retrotransposon polymorphisms can be induced by all treated sodium chloride (NaCl) doses (0, 50, 100, 200 and 300 mM NaCl). On the other hand, the LTR retrotransposons polymorphisms were decreased effectively by treatment with putrescine (0, 0.01, 0.1 and 1 mM) together with NaCl. These results suggest that putrescine could effectively inhibit salt-induced LTR retrotransposon polymorphisms, and putrescine positively contributed to salt stress tolerance.     

<Emphasis Type="Italic">SnRK2</Emphasis> Homologs in <Emphasis Type="Italic">Gossypium</Emphasis> and <Emphasis Type="Italic">GhSnRK2.6</Emphasis> Improved Salt Tolerance in Transgenic Upland Cotton and <Emphasis Type="Italic">Arabidopsis</Emphasis>

SnRK2s are a large family of plant-specific protein kinases, which play important roles in multiple abiotic stress responses in various plant species. But the family in Gossypium has not been well studied. Here, we identified 13, 10, and 13 members of the SnRK2 family from Gossypium raimondii, Gossypium arboreum, and Gossypium hirsutum, respectively, and analyzed the locations of SnRK2 homologs in chromosomes based on genome data of cotton species. Phylogenetic tree analysis of SnRK2 proteins showed that these families were classified into three groups. All SnRK2 genes were comprised of nine exons and eight introns, and the exon distributions and the intron phase of homolog genes among different cotton species were analogous. Moreover, GhSnRK2.6 was overexpressed in Arabidopsis and upland cotton, respectively. Under salt treatment, overexpressed Arabidopsis could maintain higher biomass accumulation than wild-type plants, and GhSnRK2.6 overexpression in cotton exhibited higher germination rate than the control. So, the gene GhSnRK2.6 could be utilized in cotton breeding for salt tolerance.     

A Review of Onychomycosis Due to <Emphasis Type="Italic">Aspergillus</Emphasis> Species

Aspergillus spp. are emerging causative agents of non-dermatophyte mould onychomycosis (NDMO). New Aspergillus spp. have recently been described to cause nail infections. The following criteria are required to diagnose onychomycosis due to Aspergillus spp.: (1) positive direct microscopy and (2) repeated culture or molecular detection of Aspergillus spp., provided no dermatophyte was isolated. A review of 42 epidemiological studies showed that onychomycosis due to Aspergillus spp. varies between < 1 and 35% of all cases of onychomycosis in the general population and higher among diabetic populations accounting for up to 71% and the elderly; it is very uncommon among children and adolescence. Aspergillus spp. constitutes 7.7–100% of the proportion of NDMO. The toenails are involved 25 times more frequently than fingernails. A. flavus, A. terreus and A. niger are the most common aetiologic species; other rare and emerging species described include A. tubingensis, A. sydowii, A. alliaceus, A. candidus, A. versicolor, A. unguis, A. persii, A. sclerotiorum, A. uvarum, A. melleus, A. tamarii and A. nomius. The clinical presentation of onychomycosis due to Aspergillus spp. is non-specific but commonly distal–lateral pattern of onychomycosis. A negative culture with a positive KOH may point to a NDM including Aspergillus spp., as the causative agent of onychomycosis. Treatment consists of systemic therapy with terbinafine or itraconazole.     

Genome-wide identification and evolution of <Emphasis Type="Italic">TC1</Emphasis>/<Emphasis Type="Italic">Mariner</Emphasis> in the silkworm (<Emphasis Type="Italic">Bombyx mori</Emphasis>) genome

TC1/Mariner transposons belong to class II transposable elements (TEs) that use DNA-mediated “cut and paste” mechanism to transpose, and they have been identified in almost all organisms. Although silkworm (Bombyx mori) has a large amount of TC1/Mariner elements, the genome wide information of this superfamily in the silkworm is unknown. In this study, we have identified 2670 TC1/Mariner (Bmmar) elements in the silkworm genome. All the TEs were classified into 22 families by means of fgclust, a tool of repetitive sequence classification, seven of which was first reported in this study. Phylogenetic and structure analyses based on the catalytic domain (DDxD/E) of transposase sequences indicated that all members of TC1/Mariner were grouped into five subgroups: Mariner, Tc1, maT, DD40D and DD41D/E. Of these five subgroups, maT rather than Mariner possessed most members of TC1/Mariner (51.23%) in the silkworm genome. In particular, phylogenetic analysis and structure analysis revealed that Bmmar15 (DD40D) formed a new basal subgroup of TC1/Mariner element in insects, which was referred to as bmori. Furthermore, we concluded that DD40D appeared to intermediate between mariner and Tc1. Finally, we estimated the insertion time for each copy of TC1/Mariner in the silkworm and found that most of members were dramatically amplified during a period from 0 to 1 mya. Moreover, the detailed functional data analysis showed that Bmmar1, Bmmar6 and Bmmar9 had EST evidence and intact transposases. These implied that TC1/Mariner might have potential transpositional activity. In conclusion, this study provides some new insights into the landscape, origin and evolution of TC1/Mariner in the insect genomes.     

Regulatory activity of heterologous gene-activator <Emphasis Type="Italic">xlnR</Emphasis> of <Emphasis Type="Italic">Aspergillus niger</Emphasis> in <Emphasis Type="Italic">Penicillium canescens</Emphasis>

The gene encoding the xlnR xylanolytic activator of the heterologous fungus Aspergillus niger was incorporated into the Penicillium canescens genome. Integration of the xlnR gene resulted in the increase in a number of activities, i.e. endoxylanase, β-xylosidase, α-L-arabinofuranosidase, α-galactosidase, and feruloyl esterase, compared to the host P. canescens PCA 10 strain, while β-galactosidase, β-glucosidase, endoglucanase, and CMCase activities remained constant. Two different expression constructs were developed. The first consisted of the nucleotide sequence containing the mature P. canescens phytase gene under control of the axhA promoter region gene encoding A. niger (1,4)-β-D-arabinoxylan-arabinofuranohydrolase. The second construct combined the P. canescens phytase gene and the bgaS promoter region encoding homologous β-galactosidase. Both expression cassettes were transformed into P. canescens host strain containing xlnR. Phytase synthesis was observed only for strains with the bgaS promoter on arabinose-containing culture media. In conclusion, the bgaS and axhA promoters were regulated by different inducers and activators in the P. canescens strain containing a structural tandem of the axhA promoter and the gene of the xlnR xylanolytic activator.     

Chromosomal differentiation of the sibling species <Emphasis Type="Italic">Pichia membranifaciens</Emphasis> and <Emphasis Type="Italic">Pichia manshurica</Emphasis>

The molecular karyotyping analysis of 21 strains within the taxonomic complex Pichia membranifaciens allowed the sibling species P. membranifaciens and P. manshurica, as well as P. deserticola and P. punctispora, to be differentiated. Heterogeneity of the species P. membranifaciens at the variety level is discussed.     

Investigation of stored wheat mycoflora,reporting the<Emphasis Type="Italic">Fusarium</Emphasis> cf.<Emphasis Type="Italic">langsethiae</Emphasis> in three provinces of Iran during 2007

Wheat is the most important cereal produced in Iran. A mycological survey was carried out for the first time, on the stored wheat samples in Tehran, East Azarbayejan and Mazandaran provinces in 2007. Exogenous and endogenous fungi, were isolated by the method of flotation with Malachite green agar (MGA 0.25) and Freeze blotter techniques respectively. In this study, 46 species belonging to 23 different genera were isolated.Cladosporium spp. (57.1–89.2%) andAlternaria spp. (82.4–100%) species were the predominant fungal species identified as endogenous mycoflora. The predominant exogenous fungi werePenicillium spp. (78.4–92.8%) andAspergillus spp. (71.4–85.7%) species.Fusarium proliferatum was the most prevalent species ofFusarium isolates.Aspergillus niger (39.4%) andAspergillus flavus (36.7%) were the predominantAspergillus species identified as exogenous mycoflora.Aspergillus flavus (26.6%) was the predominantAspergillus species identified as endogenous mycoflora. Flotation method with MGA 0.25 recommended for isolating of hyaline fungi from wheat cereals. In this study one isolate fromFusarium species was isolated on the basis of morphology and ribosomal internal transcribed spacer classified asFusarium langsethiae but on the basis of partial translation elongation factor-1alpha gene grouped withFusarium sporotrichioides. To our knowledge, this is the first report aboutF. cf.langsethiae in Iran and Asia.