Concerning the problem of carcinophilia of <Emphasis Type="Italic">Careproctus</Emphasis> species (Scorpaeniformes: Liparidae) in the North Kurils

Along the east coast of North Kurils, the egg deposits of Careproctus snailfishes (Scorpaeniformes: Liparidae) were revealed in the gill cavity of 9.9% of all golden king crabs, mostly in individuals with carapace of 61–80 mm width (61.6%). The number of Careproctus eggs in the gill cavity of crabs increased with the increase of the carapace width. In fact, 97.0% of the crabs had such egg deposits only at one side of their carapaces. Most of the egg deposits, apparently, belonged to C. cypsellurus and C. furcellus. The number of crabs with egg deposits was related to the number of post-molt individuals. Evidently, egg depositing by Careproctus snailfishes out of the gill cavity of crabs is a rather widespread phenomenon.     

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.     

Cloning of <Emphasis Type="Italic">rec</Emphasis>A gene of <Emphasis Type="Italic">Corynebacterium glutamicum</Emphasis> and phenotypic complementation of <Emphasis Type="Italic">Escherichia coli</Emphasis> recombinant deficient strain

Nucleotide and amino acid sequences of Corynebacterium glutamicum recA genes, from GenBank, were compared in silico. On the basis of the identity found between sequences, two degenerate primers were designed on the two sides of the deduced open reading frame (ORF) of the recA gene. PCR experiments, for amplifying the recA ORF region, were done. pGEM^®-T Easy vector was selected to be used for cloning PCR products. Then recA ORF was placed under the control of Escherichia coli hybrid trc promoter, in pKK388-1 vector. pKK388-1 vector, containing recA ORF, was transformed to E. coli DH5α ΔrecA (recombinant deficient strain), in an attempt to phenotypically complement it. Ultraviolet (u.v.) exposure experiments of the transformed and non-transformed E. coli DH5α ΔrecA cells revealed tolerance of transformed cells up to dose 0.24 J/cm², while non-transformed cells tolerated only up to dose 0.08 J/cm². It is concluded that phenotypic complementation of E. coli DH5α ΔrecA with recA ORF of C. glutamicum, could be achieved and RecA activity could be restored.     

Construction of flavocytochrome <Emphasis Type="Italic">b</Emphasis><Subscript>2</Subscript>-overproducing strains of the thermotolerant methylotrophic yeast <Emphasis Type="Italic">Hansenula polymorpha</Emphasis> (<Emphasis Type="Italic">Pichia angusta</Emphasis>)

L-Lactate cytochrome c oxidoreductase (flavocytochrome b ₂, FC b ₂) from the thermotolerant methylotrophic yeast Hansenula polymorpha (Pichia angusta) is, unlike the enzyme form baker’s yeast, a thermostable enzyme potentially important for bioanalytical technologies for highly selective assays of L-lactate in biological fluids and foods. This paper describes the construction of flavocytochrome b ₂ producers with over-expression of the H. polymorpha CYB2 gene, encoding FC b ₂. The HpCYB2 gene under the control of the strong H. polymorpha alcohol oxidase promoter in a plasmid for multicopy integration was transformed into the recipient strain H. polymorpha C-105 (grc1 catX), impaired in glucose repression and devoid of catalase activity. A method was developed for preliminary screening of the transformants with increased FC b ₂ activity in permeabilized yeast cells. The optimal cultivation conditions providing for the maximal yield of the target enzyme were found. The constructed strain is a promising FC b ₂ producer characterized by a sixfold increased (to 3 μmol min^?1 mg^?1 protein in cell-free extract) activity of the enzyme.     

Cytochromes <Emphasis Type="Italic">c</Emphasis> of the anaerobic methacrylate reducer <Emphasis Type="Italic">Geobacter sulfurreducens</Emphasis> AM-1

Cytochromes c were found in the cells of the bacterium Geobacter sulfurreducens AM-1 grown on acetate and methacrylate. The periplasmic extract of G. sulfurreducens AM-1 contained about 88% of the total content of cytochromes c of intact cells. The analysis of cytochromes c from the native cells of G. sulfurreducens AM-1, from the periplasmic extract and from the cells treated by an alkaline solution showed the presence of nine proteins containing heme c. The molecular masses of cytochromes c from G. sulfurreducens AM-1 were 12.5, 15.5, 25.7, 29.5, 34.7, 41.7, 50.1, 63.1, and 67.6 kDa; localization of each cytochrome c was determined. Three heme-containing proteins (15.5 kDa, 25.7 kDa, and 29.5 kDa with the most intensive staining) were present mainly in the periplasm of the bacterium. The other two (50.1 and 67.6 kDa) were supposedly localized in the cell membrane. Cytochromes c with the molecular masses of 12.5, 15.5, and 67.6 kDa are considered as possible components of the methacrylate redox system of G. sulfurreducens AM-1.     

Interactions between the grass shrimp <Emphasis Type="Italic">Palaemonetes pugio</Emphasis> and the salt marsh grasses <Emphasis Type="Italic">Phragmites australis</Emphasis> and <Emphasis Type="Italic">Spartina alterniflora</Emphasis>

The grass shrimp Palaemonetes pugio, a species common to Spartina alterniflora-dominated marshes, may be sensitive to the invasion of the common reed Phragmites australis in northeastern US salt marshes. We examined two questions: (1) Do grass shrimp have a preference for the native plant over the non-native plant? (2) Are grass shrimp more effective foragers on P. australis? We tested the first hypothesis by comparing the amount of time shrimp spend in physical contact with the plant types over a 1-h period. Shrimp were observed under different arrangements of vegetation to control for differences in conspicuous structural features. Additionally, the amount of time shrimp spent foraging on S. alterniflora and P. australis shoots was compared to determine if shrimp graze more often on S. alterniflora. Shrimp spent significantly more time in contact with S. alterniflora only when plant types were grouped at opposite ends of aquaria, but did not exhibit a foraging preference for this plant type. To address our second question, we investigated the effects of shrimp foraging on stem epifauna, an assemblage of semi-aquatic invertebrates associated with macrophyte shoots. Previous research showed that P. australis supports a lower density of stem-dwelling epifauna relative to S. alterniflora. We hypothesized that the primary grazer of this community, P. pugio, can forage on P. australis stems more effectively due to structural differences between the two plants, causing the lower abundance of epifauna through top-down effects. We exposed individual shoots inhabited by epifauna to shrimp and compared faunal densities on exposed shoots to densities on control shoots after 18 h. The reduction of epifauna by predation was proportional on the two plant types. Therefore, top-down effects can be ruled out as an explanation for the patchy distribution of epifauna observed in P. australis–S. alterniflora marshes.     

Mapping of the Ogura fertility restorer gene <Emphasis Type="Italic">Rfo</Emphasis> and development of <Emphasis Type="Italic">Rfo</Emphasis> allele-specific markers in canola (<Emphasis Type="Italic">Brassica napus</Emphasis> L.)

Ogura cytoplasmic male sterility (CMS) and its corresponding nuclear fertility restorer gene, Rfo, have been introduced from radish to Brassica species by interspecific crosses. Rfo restores male fertility by altering the translational expression of Orf138, a mitochondrial gene, whose expression results in the male sterile phenotype. This system has been extensively investigated and breeding restorer lines for the Ogura CMS has become a major objective for hybrid seed production in many canola breeding programs. In this study, we have sequenced genomic clones of Rfo amplified from a canola restorer line R2000, licensed from INRA, France, and a Dow AgroSciences non-restorer line Nexera 705 using primers designed from the radish Rfo sequence (GenBank accession AJ550021). Sequence alignment revealed three homologous sequences of Rfo. Two of the sequences were present in both R2000 and Nexera 705 but the third one was present only in R2000. These results suggested that the first two sequences could be the homoeologous sequences of Rfo already existing in the canola genome and the third one could be the radish Rfo introduced into canola. Based on the sequence differences between the restorer and non-restorer lines, Rfo allele-specific PCR markers were developed. We also developed a high throughput, Rfo allele-specific Invader^® assay through Third Wave Technologies. Linkage analysis revealed a co-segregation between the allele-specific marker and the phenotypes for fertility restoration. This allele-specific marker has been mapped in the linkage group N19 and proved to be very useful for direct selection of Rfo alleles for fertility restoration during marker-assisted introgression of the Ogura restorer for hybrid development in canola.     

Isolation and characterization of nitrogen-fixing bacteria of the genus <Emphasis Type="Italic">Azospirillum</Emphasis> from the soil of a <Emphasis Type="Italic">Sphagnum</Emphasis> peat bog

he presence of nitrogen-fixing bacteria of the genus Azospirillum in the soils of acidic raised Sphagnum bogs is revealed for the first time. Three Azospirillum strains, B2, B21, and B22, were isolated as a component of methane-oxidizing enrichment cultures, whereas attempts to isolate them directly from peat samples have failed. The results of comparative analysis of the nucleotide sequences of 16S rRNA genes, DNA-DNA hybridization, and the analysis of the sequences of the functional genes encoding nitrogenase and ribulose-1, 5-bisphosphate carboxylase reveal that all the newly obtained strains can be classified as Azospirillum lipoferum. Yet, unlike A. lipoferum, the isolates do not require biotin and utilize sucrose, inositol, and glycerol for growth. The cell morphology of strain B2 differs from that of the type strain and strains B21 and B22. The results obtained indicate the variability of morphological, physiological, and biochemical properties in closely related Azospirillum strains and suggest the existence of metabolic relationships between methanotrophic bacteria and the representatives of the genus Azospirillum under peat bog conditions.     

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.     

A new <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis> deletion mutant <Emphasis Type="Italic">apetala1-20</Emphasis>

A new deletion allele of the APETALA1 (AP1) gene encoding a type II MADS-box protein with the key role in the initiation of flowering and development of perianth organs has been identified in A. thaliana. The deletion of seven amino acids in the conserved region of the K domain in the ap1-20 mutant considerably delayed flowering and led to a less pronounced abnormality in the corolla development compared to the weak ap1-3 and intermediate ap1-6 alleles. At the same time, a considerable stamen reduction has been revealed in ap1-20 as distinct from ap1-3 and ap1-6 alleles. These data indicate that the K domain of AP1 can be crucial for the initiation of flowering and expression regulation of B-class genes controlling stamen development.     

On the multicomponent nature of <Emphasis Type="Italic">Halobacterium salinarum</Emphasis> flagella

Filaments of the flagellum of the halophilic archaeon Halobacterium salinarum consist of five flagellins: A1, A2, B1, B2, and B3, which are encoded by five genes localized in tandem in two flgA and flgB operons. While the role of flagellins A1 and A2 has been determined, the role of the proteins, B operon products, is still unclear. A mutant strain of H. salinarum with deleted A and B flagellin genes (ΔflgAΔflgB) has been obtained for the first time. This strain has been used to create and analyze the strains carrying only individual B1 or B3 flagellin genes. Cells of the ΔflgAΔflgB strain were shown to have short filamentous formations, 7–8 nm thick, which we have named as X-filaments. It has been shown that X-filaments consist of a protein immunologically related to flagellins A and B. Expression of the B1 and B3 genes is suppressed in the absence of A1, A2, and B2. It has been shown that flagellins B1 and B3 cannot be substituted for flagellin B2 upon the formation of a curved hook-like structure, which serves as a connecting element between the flagellar filament and the motor axis. The multicomponent nature of flagella is discussed in the light of their possible involvement in other cell processes besides providing motility.     

Development and relations of <Emphasis Type="Italic">Fusarium culmorum</Emphasis> and <Emphasis Type="Italic">Pseudomonas fluorescens</Emphasis> in soil

The development of Fusarium culmorum and Pseudomonas fluorescens in soil, and the relations between them, were studied using membrane filters containing the fungus, the bacterium, or both microorganisms; the filters were incubated in soil. F. culmorum was identified by indirect immunofluorescence; the GUS-labeled strain was used to visualize P. fluorescens. It was found that F. culmorum introduced in soil can develop as a saprotroph, with the formation of mycelium, macroconidia, and a small amount of chlamydospores. Introduction of glucose and cellulose resulted in increased density of the F. culmorum mycelium and macroconidia. P. fluorescens suppressed the development of the F. culmorum mycelium in soil, but stimulated chlamydospore formation. Decreased mycelial density in the presence of P. fluorescens was more pronounced in soil without additions and less pronounced in the case of introduction of glucose or cellulose. F. culmorum had no effect on P. fluorescens growth in soil.     

<Emphasis Type="Italic">Malassezia</Emphasis> spp. in Acoustic Meatus of Bats (<Emphasis Type="Italic">Molossus molossus</Emphasis>) of the Amazon Region,Brazil

The yeasts of the Malassezia genus are opportunistic microorganisms and can cause human and animal infections. They are commonly isolated from the skin and auricular canal of mammalians, mainly dogs and cats. The present study was aimed to isolate Malassezia spp. from the acoustic meatus of bats (Molossus molossus) in the Montenegro region, “Rondônia”, Brazil. From a total of 30 bats studied Malassezia spp. were isolated in 24 (80%) animals, the breakdown by species being as follows (one Malassezia sp. per bat, N = 24): 15 (62.5%) M. pachydermatis, 5 (20.8%) M. furfur, 3 (12.5%) M. globosa and 1 (4.2%) M. sympodialis. This study establishes a new host and anatomic place for Malassezia spp., as it presents the first report ever of the isolation of this genus of yeasts in the acoustic meatus of bats.     

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.     

Thermotolerance and place memory in adult <Emphasis Type="Italic">Drosophila</Emphasis> are independent of natural variation at the <Emphasis Type="Italic">foraging</Emphasis> locus

Natural polymorphisms at the foraging (for) gene influence several behaviors. However, it is seldom clear how different for alleles could be selected. In one case, Drosophila with the rover allele (for ^r ) have higher locomotor activity in the presence of food than animals with the sitter allele (for ^s ), suggesting a complementary feeding strategy. There are, in addition, differences between for ^r and for ^s Drosophila in some tests of short-term memory (for ^r animals generally perform at higher levels) and thermotolerance (for ^s larvae are more resistant to the effects of high-temperature). We asked whether there could be a direct compensating advantages in adult for ^s flies that could maintain the natural for variants. First, are adult for ^s flies more thermotolerant? Second, do for ^r flies have a higher short-term place memory? Third, as an alternative, might for ^s flies have higher place memory? Our results do not confirm these possibilities. Thus, a thermotolerance advantage of for ^s flies does not compensate for a potential for ^r short-term memory advantage; for ^r flies do not have a universal advantage in short-term memory; and for ^s flies do not have an advantage in place memory that could compensate for for ^r advantages in other learning contexts.     

Photosynthetic activity and components of the electron transport chain in the aerobic bacteriochlorophyll <Emphasis Type="Italic">a</Emphasis>-containing bacterium <Emphasis Type="Italic">Roseinatronobacter thiooxidans</Emphasis>

Bioenergetics of the aerobic bacteriochlorophyll a-containing (BCl a) bacterium (ABC bacterium) Roseinatronobacter thiooxidans is a combination of photosynthesis, oxygen respiration, and oxidation of sulfur compounds under alkaliphilic conditions. The photosynthetic activity of Rna. thiooxidans cells was established by the photoinhibition of cell respiration and reversible photobleaching discoloration of the BCl a of reaction centers (RC), connected by the chain of electron transfer with cytochrome c ₅₅₁ oxidation. The species under study, like many purple bacteria and some of the known ABC bacteria, possesses a light-harvesting pigment-protein (LHI) complex with the average number of 30 molecules of antenna BCl a per one photosynthetic RC. Under microaerobic growth conditions, the cells contained bc ₁ complex and two terminal oxidases: cbb ₃-cytochrome oxidase and the alternative cytochrome oxidase of the a ₃ type. Besides, Rna. thiooxidans was shown to have several different soluble low- and high-potential cytochromes c, probably associated with the ability of utilizing sulfur compounds as additional electron donors.     

Leaf spot disease on <Emphasis Type="Italic">Tilia cordata</Emphasis> caused by the fungus <Emphasis Type="Italic">Cercospora microsora</Emphasis>

Increased incidence of leaf spots on many tree species, up to the presence of peripheral importance only, including linden trees was noticed recently. First massive and continuous occurence of the fungus Cercospora microsora Sacc. [teleomorph Mycosphaerella millegrana (Cook.) Schröet., Mycosphaerella microsora Syd.], causal agent of anthracnose on linden trees (Tilia cordata Mill.) grown in urban plantings in Slovakia was reported. Along with this, certain of the important growth characteristics of this fungus were studied under laboratory conditions. To specify Cercospora biology mycelial growth of C. microsora in pure hyphal cultures was observed in relation to medium and locality. One-way ANOVA has confirmed a statistically significant influence of both factors, culture medium and locality on growth rate values of C. microsora. The effect of these factors has not proved unambiguously in all cases. In the case of one locality (Nitra), the significant influence of used media has not been proved (P > 0.05). PDAg showed generally as the most suitable medium, inducing the most intensive growth in three localities (41.06 mm/week on average). Comparing three localities, the effect of this factor is not so unambiguous. Growth rate values from the localities Bratislava and Pribeta indicate unsuitability of medium A for the fast radial growth. A Tukey test separately conducted for the factors medium and the locality revealed the significant combinations of means (P ≤ 0.05).     

Three new species of <Emphasis Type="Italic">Passiflora</Emphasis> subgenus <Emphasis Type="Italic">Decaloba</Emphasis> (Passifloraceae) from Brazil

Three new species from Brazil are described and illustrated. Passiflora cervii, P. jiboiaensis, and P. transversalis all belong to Passiflora subg. Decaloba.     

Impact of the invasive painted bug <Emphasis Type="Italic">Bagrada hilaris</Emphasis> on physiological traits of its host <Emphasis Type="Italic">Brassica oleracea</Emphasis> var botrytis

Bagrada hilaris is a herbivorous insect native of Asia and Africa, which has invaded southern Europe and North America where it causes major damage to cole crops. Laboratory experiments were conducted to assess how the infestation of this invasive species damages the host Brassica oleracea var botrytis, and to evaluate the interaction between plant emission of volatile organic compounds (VOC) and B. hilaris adults. Plant responses to insect feeding were evaluated through changes in photosynthesis, stomatal conductance, VOC emission, and visual damage on leaves. The impact of B. hilaris was compared with that of Nezara viridula, a polyphagous species distributed worldwide. Plant VOC role in host plant detection was tested with electroantennography bioassays on B. hilaris antenna. Photosynthesis and stomatal conductance were consistently reduced in plants infested with 40 B. hilaris adults for 24 h. The feeding activity of a single B. hilaris caused larger discolored spots on host leaves in comparison with N. viridula. VOC emitted by B. oleracea changed significantly in response to B. hilaris and N. viridula infestation. In particular, production of limonene was strongly reduced by the infestation of the two pentatomids, while an increase in the emission of acetic acid and 2-ethyl-1-hexanol was observed. EAG dose–response tests using the main plant VOC showed B. hilaris antennal responses to benzaldehyde, octanal, nonanal, and acetic acid, which indicates a role of these compounds in host location.