O-chromosome lethal frequencies in Serbian and Montenegrin Drosophila subobscura populations

Lethal chromosomal frequencies were obtained from three Drosophila subobscura samples from the Mt. Avala (Serbia) population in September 2003 (0.218), June 2004 (0.204) and September 2004 (0.250). These values and those from other Balkan populations studied previously (Petnica, Kamariste, Zanjic and Djerdap) were used to analyze the possible effect of population, year, month and altitude above sea level on lethal chromosomal frequencies. According to ANOVAS no effect were observed. Furthermore, the lethal frequencies of the Balkan populations did not vary according to latitude. This is probably due to the relative proximity and high gene flow between these populations. From a joint study of all the Palearctic D. subobscura populations so far analyzed, it can be deduced that the Balkan populations are located in the central area of the species distribution. Finally, it seems that lethal chromosomal frequencies are a consequence of the genetic structure of the populations.     

Genetic constraints for thermal coadaptation in <Emphasis Type="Italic">Drosophila subobscura</Emphasis>

Background  

Behaviour has been traditionally viewed as a driver of subsequent evolution because behavioural adjustments expose organisms to novel environments, which may result in a correlated evolution on other traits. In Drosophila subobscura, thermal preference and heat tolerance are linked to chromosomal inversion polymorphisms that show parallel latitudinal clines worldwide, such that "cold-climate" ("warm-climate") chromosome arrangements collectively favour a coherent response to colder (warmer) settings as flies carrying them prefer colder (warmer) conditions and have lower (higher) knock out temperatures. Yet, it is not clear whether a genetic correlation between thermal preference and heat tolerance can partially underlie such response.     

Climatic adaptation of chromosomal inversions in <Emphasis Type="Italic">Drosophila subobscura</Emphasis>

Drosophila subobscura is a species with a rich chromosomal polymorphism which is adaptive to different climatic conditions. Five samples of the Font Groga population (Barcelona, Spain) were sampled in autumn during 5 consecutive years (2011–2015) to obtain their inversion chromosomal polymorphism, and climatic data of several meteorological variables were also collected. The aim was to analyze the adaptive potential of inversions with regard to climatic variables, being the most relevant: mean temperature (T_mean), maximum temperature (T_max), minimum temperature (T_min), humidity (Hm) and rainfall (Rf). As expected, no significant variation in inversion frequencies were detected over this short period of time. However, from a climatic point of view it was possible to differentiate ‘warm’ and ‘dry’ from ‘cold’ and ‘humid’ samples. The joint study of maximum (T_max) and minimum (T_min) temperatures was a key element to understand the effect on adaptation of many inversions. It was also observed that temperature had to be considered in conjunction with humidity and rainfall. All these factors would condition the biota of D. subobscura habitat, and chromosomal inversions could provide an adaptive response to it.     

Quantitative genetics of speciation: additive and non-additive genetic differentiation between <Emphasis Type="Italic">Drosophila madeirensis</Emphasis> and <Emphasis Type="Italic">Drosophila subobscura</Emphasis>

The role of dominance and epistasis in population divergence has been an issue of much debate ever since the neoDarwinian synthesis. One of the best ways to dissect the several genetic components affecting the genetic architecture of populations is line cross analysis. Here we present a study comparing generation means of several life history-traits in two closely related Drosophila species: Drosophila subobscura, D. madeirensis as well as their F ₁ and F ₂ hybrids. This study aims to determine the relative contributions of additive and non-additive genetic parameters to the differentiation of life-history traits between these two species. The results indicate that both negative dominance and epistatic effects are very important in the differentiation of most traits. We end with considerations about the relevance of these findings for the understanding of the role of non-additive effects in speciation.     

The endosymbiont <Emphasis Type="Italic">Wolbachia</Emphasis> in Eurasian populations of <Emphasis Type="Italic">Drosophila melanogaster</Emphasis>

The endosymbiotic α-proteobacteria Wolbachia is widely spread among arthropods and Filariidae nematodes. This bacterium is transmitted vertically via a transovarian route. Wolbachia is a cause of several reproductive abnormalities in the host species. We analyzed the isofemale lines created using flies collected from Drosophila melanogaster natural populations for infection with the endosymbiont Wolbachia. Wolbachia were genotyped according to five variable markers: the presence of insertion sequence IS5 in two loci, the copy number of two minisatellite repeats, and an inversion. Overall, 665 isofemale lines isolated from the populations of D. melanogaster from Ukraine, Belarus, Moldova, Caucasus, Central Asia, Ural, Udmurtia, Altai, West and East Siberia, and Far East in 1974 through 2005 were used in the work. The samples from Ukrainian, Altaian, and Middle Asian populations were largest. The infection rate of D. melanogaster populations from Middle Asia, Altaian, and Eastern Europe (Ukraine, Moldavia, and Belarus) with Wolbachia amounted to 64, 56, and 39%, respectively. The D. melanogaster population from the Caucasus displayed heterogeneity in the genotypes of this cytoplasmic infection. The Wolbachia genotype wMel, detected in all the populations studied, was the most abundant. The genotype wMelCS2 was always present in the populations from Middle Asia and Altai and was among the rare variants in the D. melanogaster populations from the Eastern Europe. Single instances of the Wolbachia genotype wMelCS occurred in a few flies from the Central Asian and Altai populations, but was not found this genotype in the other regions.     

The <Emphasis Type="Italic">Adh</Emphasis> locus and adaptation of <Emphasis Type="Italic">cn</Emphasis> and <Emphasis Type="Italic">vg</Emphasis> mutants in experimental populations of <Emphasis Type="Italic">Drosophila melanogaster MEIG</Emphasis>

A complex study on the adaptation of cn and vn mutants and the allozymes of alcoholdehydrogenase (ADH) was carried out in initially pure lines, and their panmixia populations during exchange of the mutant genotype with that of wild-type flies (C-S) and D) through saturating crossings. The relative adaptation of the genotypes was estimated by their effect on reproductive efficiency in the experimentally obtained population. Fecundity, lifespan, and the resistance of the studied genotypes to hyperthermia were investigated individually. It was shown that the high level of adaptation of the cn mutants and the low level of adaptation of the vg mutants was correlated with the presence of different ADH allozymes. In the studied population, the F-allozyme of ADH accompanied the vg mutation, while the S-allozyme of the enzyme was detected in cn mutants. Saturating crossings of C-S(Adh ^S)×vg(Adh ^F) and D(Adh ^F) × cn(Adh ^S), along with the parallel determination of the allele composition of the Adh locus, demonstrated that the complete substitution of the F-allozyme of ADH in the vg mutants by the S-allozyme in D flies, as well as the substitution of the S-allozyme of ADH in the cn mutants by the F-allozyme in D flies was realized only after the 15th–20th backcrosses. These results favor the coadaptation of cn and vg marker genes with alleles of the Adh locus and indicate the important role of the latter in the adaptation of genotypes. In the studied population, selection acted primarily against the vg mutants, which were inferior to the cn mutants, and heterozygote genotypes in indices of the main adaptation components.     

The polytene dot chromosome of <Emphasis Type="Italic">Drosophila</Emphasis>: <Emphasis Type="Italic">D. melanogaster</Emphasis> and <Emphasis Type="Italic">D. subobscura</Emphasis>

The chromosome arms are assumed to be homologous within the genus Drosophila. Homology at the level of the polytene chromosome banding pattern between non-sibling species is, however, almost impossible to establish as different processes such as inversion, transposition and unequal crossing over, have disturbed it. Even though the band sequences cannot be followed, we may ask whether there is a correlation in the total number of bands between species. The polytene dot chromosome is an excellent starting point for such an approach. Here we present the detailed cytology of polytene chromosome 4 of D. melanogasterand the polytene dot chromosome of D. subobscura using electron microscopy. The results show that the number of bands is about the same, around 30, in both species. We predict that by using thin sections and electron microscopy for the longer polytene chromosome arms, both species will turn out to have approximately equal band numbers.     

<Emphasis Type="Italic">Ty</Emphasis>1<Emphasis Type="Italic">/copia</Emphasis>- and <Emphasis Type="Italic">Ty</Emphasis>3<Emphasis Type="Italic">/gypsy</Emphasis>-like DNA sequences in <Emphasis Type="Italic">Helianthus </Emphasis>species

Two repeated DNA sequences isolated from a partial genomic DNA library of Helianthus annuus, p HaS13 and p HaS211, were shown to represent portions of the int gene of a Ty3 /gypsy retroelement and of the RNase-Hgene of a Ty1 /copia retroelement, respectively. Southern blotting patterns obtained by hybridizing the two probes to BglII- or DraI-digested genomic DNA from different Helianthus species showed p HaS13 and p HaS211 were parts of dispersed repeats at least 8 and 7 kb in length, respectively, that were conserved in all species studied. Comparable hybridization patterns were obtained in all species with p HaS13. By contrast, the patterns obtained by hybridizing p HaS211 clearly differentiated annual species from perennials. The frequencies of p HaS13- and p HaS211-related sequences in different species were 4.3x10(4)-1.3x10(5) copies and 9.9x10(2)-8.1x10(3) copies per picogram of DNA, respectively. The frequency of p HaS13-related sequences varied widely within annual species, while no significant difference was observed among perennial species. Conversely, the frequency variation of p HaS211-related sequences was as large within annual species as within perennials. Sequences of both families were found to be dispersed along the length of all chromosomes in all species studied. However, Ty3 /gypsy-like sequences were localized preferentially at the centromeric regions, whereas Ty1/ copia-like sequences were less represented or absent around the centromeres and plentiful at the chromosome ends. These findings suggest that the two sequence families played a role in Helianthusgenome evolution and species divergence, evolved independently in the same genomic backgrounds and in annual or perennial species, and acquired different possible functions in the host genomes.     

Prevalence of full-size <Emphasis Type="Italic">P</Emphasis> and <Emphasis Type="Italic">KP</Emphasis> elements in North American populations of <Emphasis Type="Italic">Drosophila melanogaster</Emphasis>

The P transposable element invaded the Drosophila melanogaster genome in the middle of the twentieth century, probably from D. willistoni in the Caribbean or southeastern North America. P elements then spread rapidly and became ubiquitous worldwide in wild populations of D. melanogaster by 1980. To study the dynamics and long-term fate of transposable genetic elements, we examined the molecular profile of genomic P elements and the phenotype in the P-M system of the current North American natural populations collected in 2001-2003. We found that full-size P and KP elements were the two major size classes of P elements present in the genomes of all populations ("FP + KP predominance") and that the P-related phenotypes had largely not changed since the 1980s. Both FP + KP predominance and phenotypic stability were also seen in other populations from other continents. As North American populations did not show many KP elements in earlier samples, we hypothesize that KP elements have spread and multiplied in the last 20 years in North America. We suggest that this may be due to a transpositional advantage of KP elements, rather than to a role in P-element regulation.     

Patterns of <Emphasis Type="Italic"> Hermes </Emphasis>transposition in <Emphasis Type="Italic"> Drosophila melanogaster</Emphasis>

Transposable elements are being developed as tools for genomics and for the manipulation of insect genotypes for the purposes of biological control. An understanding of their transposition behavior will facilitate the use of these elements. The behavior of an autonomous Hermes transposable element from Musca domestica in the soma and germ-line of Drosophila melanogaster was investigated using the method of transposon display. In the germ-line, Hermes transposed at a rate of approximately 0.03 jumps per element per generation. Within the soma Hermes exhibited markedly non-random patterns of integration. Certain regions of the genome were distinctly preferred over others as integration targets, while other regions were underrepresented among the integration sites used. One particular site accounted for 4.4% of the transpositions recovered in this experiment, all of which were located within a 2.5-kb region of the actin5C promoter. This region was also present within the Hermes element itself, suggesting that this clustering is an example of transposable element "homing". Clusters of integration sites were also observed near the original donor sites; these represent examples of local hopping. The information content (sequence specificity) of the 8-bp target site was low, and the consensus target site resembles that determined from plasmid-based integration assays.     

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.     

Genetics and geometry of canalization and developmental stability in <Emphasis Type="Italic">Drosophila subobscura</Emphasis>

Background  

Many properties of organisms show great robustness against genetic and environmental perturbations. The terms canalization and developmental stability were originally proposed to describe the ability of an organism to resist perturbations and to produce a predictable target phenotype regardless of random developmental noise. However, the extent to which canalization and developmental stability are controlled by the same set of genes and share underlying regulatory mechanisms is largely unresolved.     

<Emphasis Type="Italic">Agrobacterium</Emphasis><Emphasis Type="Italic">tumefaciens</Emphasis>-mediated transformation of callus cells of <Emphasis Type="Italic">Crataegus</Emphasis><Emphasis Type="Italic">aronia</Emphasis>

A genetic transformation system has been developed for callus cells of Crataegus aronia using Agrobacterium tumefaciens. Callus culture was established from internodal stem segments incubated on Murashige and Skoog (MS) medium supplemented with 5 mg l⁻¹ Indole-3-butyric acid (IBA) and 0.5 mg l⁻¹ 6-benzyladenine (BA). In order to optimize the callus culture system with respect to callus growth and coloration, different types and concentrations of plant growth regulators were tested. Results indicated that the best average fresh weight of red colored callus was obtained on MS medium supplemented with 2 mg l⁻¹ 2,4-dichlorophenoxyacetic acid (2,4-D) and 1.5 mg l⁻¹ kinetin (Kin) (callus maintenance medium). Callus cells were co-cultivated with Agrobacterium harboring the binary plasmid pCAMBIA1302 carrying the mgfp5 and hygromycin phosphotransferase (hptII) genes conferring green fluorescent protein (GFP) activity and hygromycin resistance, respectively. Putative transgenic calli were obtained 4 weeks after incubation of the co-cultivated explants onto maintenance medium supplemented with 50 mg l⁻¹ hygromycin. Molecular analysis confirmed the integration of the transgenes in transformed callus. To our knowledge, this is the first time to report an Agrobacterium-mediated transformation system in Crataegus aronia.     

<Emphasis Type="Italic">Galleria</Emphasis><Emphasis Type="Italic">mellonella</Emphasis> are Resistant to <Emphasis Type="Italic">Pneumocystis</Emphasis><Emphasis Type="Italic">murina</Emphasis> Infection

Studying Pneumocystis has proven to be a challenge from the perspective of propagating a significant amount of the pathogen in a facile manner. The study of several fungal pathogens has been aided by the use of invertebrate model hosts. Our efforts to infect the invertebrate larvae Galleria mellonella with Pneumocystis proved futile since P. murina neither caused disease nor was able to proliferate within G. mellonella. It did, however, show that the pathogen could be rapidly cleared from the host.     

Revision of the <Emphasis Type="Italic">Serrulati</Emphasis> species of <Emphasis Type="Italic">Penstemon</Emphasis> section <Emphasis Type="Italic">Saccanthera</Emphasis> subsection <Emphasis Type="Italic">Serrulati</Emphasis>

A revision of Penstemon sect. Saccanthera subsect. Serrulati includes a new species (P. salmonensis), a new variety (P. triphyllus var. infernalis), and the elevation of a subspecies to species (P. curtiflorus), bringing the total number of species to eight, which are keyed and described, complete with nomenclature and type citations.     

The <Emphasis Type="Italic">Caenorhabditis</Emphasis><Emphasis Type="Italic">briggsae</Emphasis> genome contains active <Emphasis Type="Italic">CbmaT1</Emphasis> and <Emphasis Type="Italic">Tcb1</Emphasis> transposons

The maT clade of transposons is a group of transposable elements intermediate in sequence and predicted protein structure to mariner and Tc transposons, with a distribution thus far limited to a few invertebrate species. We present evidence, based on searches of publicly available databases, that the nematode Caenorhabditis briggsae has several maT-like transposons, which we have designated as CbmaT elements, dispersed throughout its genome. We also describe two additional transposon sequences that probably share their evolutionary history with the CbmaT transposons. One resembles a fold back variant of a CbmaT element, with long (380-bp) inverted terminal repeats (ITRs) that show a high degree (71%) of identity to CbmaT1. The other, which shares only the 26-bp ITR sequences with one of the CbmaT variants, is present in eight nearly identical copies, but does not have a transposase gene and may therefore be cross mobilised by a CbmaT transposase. Using PCR-based mobility assays, we show that CbmaT1 transposons are capable of excising from the C. briggsae genome. CbmaT1 excised approximately 500 times less frequently than Tcb1 in the reference strain AF16, but both CbmaT1 and Tcb1 excised at extremely high frequencies in the HK105 strain. The HK105 strain also exhibited a high frequency of spontaneous induction of unc-22 mutants, suggesting that it may be a mutator strain of C. briggsae.     

New combinations and typifications in <Emphasis Type="Italic">Bistorta</Emphasis>, <Emphasis Type="Italic">Persicaria</Emphasis>, <Emphasis Type="Italic">Polygonum,</Emphasis> and <Emphasis Type="Italic">Rumex</Emphasis> (Polygonaceae)

New combinations are proposed in anticipation of the Polygonaceae treatment in the forthcoming volume of Intermountain Flora: Polygonum kelloggii var. esotericum, P. kelloggii var. watsonii , Rumex densiflorus var. pycnanthus , R. salicifolius var. utahensis, and R. occidentalis var. tomentellus. Typifications are proposed to facilitate ongoing studies in Polygonaceae and to maintain current usage.     

<Emphasis Type="Italic">Drosophila melanogaster</Emphasis> as a polymicrobial infection model for <Emphasis Type="Italic">Pseudomonas aeruginosa</Emphasis> and <Emphasis Type="Italic">Staphylococcus aureus</Emphasis>

Non-mammalian infection models have been developed over the last two decades, which is a historic milestone to understand the molecular basis of bacterial pathogenesis. They also provide small-scale research platforms for identification of virulence factors, screening for antibacterial hits, and evaluation of antibacterial efficacy. The fruit fly, Drosophila melanogaster is one of the model hosts for a variety of bacterial pathogens, in that the innate immunity pathways and tissue physiology are highly similar to those in mammals. We here present a relatively simple protocol to assess the key aspects of the polymicrobial interaction in vivo between the human opportunistic pathogens, Pseudomonas aeruginosa and Staphylococcus aureus, which is based on the systemic infection by needle pricking at the dorsal thorax of the flies. After infection, fly survival and bacteremia over time for both P. aeruginosa and S. aureus within the infected flies can be monitored as a measure of polymicrobial virulence potential. The infection takes ~24 h including bacterial cultivation. Fly survival and bacteremia are assessed using the infected flies that are monitored up to ~60 h post-infection. These methods can be used to identify presumable as well as unexpected phenotypes during polymicrobial interaction between P. aeruginosa and S. aureus mutants, regarding bacterial pathogenesis and host immunity.     

Male accessory gland secretory protein polymorphism in natural populations of <Emphasis Type="Italic">Drosophila nasuta nasuta</Emphasis> and <Emphasis Type="Italic">Drosophila sulfurigaster neonasuta</Emphasis>

Male accessory gland secretory protein polymorphism was analysed in natural populations of Drosophila nasuta nasuta and D. sulfurigaster neonasuta for the first time, using SDS-PAGE to score polymorphism of these proteins in 2788 individuals of D. n. nasuta and 2232 individuals of D. s. neonasuta from 12 different populations from southern India. A total of 25 and 18 variant protein phenotypes were identified in D. n. nasuta and D. s. neonasuta, respectively. Protein fractions of group III were more polymorphic than those from groups I and II. The results show that accessory gland secretory proteins show high levels of polymorphism, irrespective of species or habitat. Moreover, we have used the variation in the accessory gland proteins to assess the extent of divergence between the species and to infer their population structure. The study suggests that though both D. n. nasuta and D. s. neonasuta belong to the same subgroup, they differ in population structure, as far as accessory gland protein polymorphism is concerned.