Volume 70, Issue 9

Proteolytic marine bacterium designated as strain MS2-1 was isolated from the deep-sea sediment of Bay of Bengal. Strain MS2-1 was taxonomically identified as Marinobacter aquaeolei on the basis of 16S rRNA gene sequence homology analysis. A new alkaline serine protease gene (1,086 bp) was delineated, cloned into pET-28a-(+) vector and overexpressed in Escherichia coli. The enzyme showed 90% amino acid sequence identity towards subtilisin-like protease from a Pseudoalteromonas sp. AS-11. The three-dimensional homology model predicted the active site residues that may be responsible for the proteolytic activity. Molecular weight of the purified protein was calculated as 39 kDa. The enzyme exhibited the stability within a wide range of pH (7.0-12.0) and temperature (40-70°C). Maximum enzyme activity was observed at pH 8.0 and 50°C. The purified enzyme showed stability in the presence of metal ions, solvents, surfactants and detergents. The loss of activity with PMSF and the 3-fold increase of activity with DTT suggested the thiol-dependent nature of this serine protease enzyme. Results reported in this study also suggested that the new alkaline serine protease produced by the strain MS2-1 can be used as an efficient blood stain remover in detergent industries and as a thrombolytic agent in biomedical applications.

Fluorescent Pseudomonas (FP) is an intrinsic group of plant growth promoting rhizobacteria (PGPR) which also has the potential to control phytopathogens. In the present study sixteen FP isolates were subjected to siderophore production and siderotyping. Four DNA finger printing analysis markers (RAPD, ERIC, BOX A1R, REP and GTG 5 ) were used to assess their genetic diversity through DNA finger printing analysis. Outer membrane protein (OMP) homology of FP was carried out by SDS-PAGE analysis. Chemical characterization showed that the siderophores produced were hydroxamate in nature. Production of siderophores ranged from 21.73 ± 2.18 to 182.03 ± 1.66 μg mL −1 after 72 hr incubation. Maximum production was shown by the isolate M139. Dendrogram obtained from the combined binary data of 10 RAPD primers classified FPs into two distinct clusters. Except for M140, all the isolates from maize rhizosphere were grouped into the same cluster. Among three Rep-PCR analyses, REP primer showed maximum diversity in banding pattern. Combined binary data of three Rep primers also classified FP into two clusters as in RAPD and protein electrophenogram of the OMP. To the best of our knowledge this is the first report on genomic and OMP disparity among hydroxamate producing FPs.

In this study, intra-specific genetic variation and inter-specific genetic relation were investigated among Phlomis oppositiflora, P. kurdica, P. × melitenense (P. kurdica × oppositiflora), P. capitata and P. kurdica × capitata by using RAPD and ISSR markers. The hybridity of P. × melitenense and P. kurdica × capitata samples was also surveyed in terms of morphological and molecular aspects. Except for two, all bands obtained from RAPD (707 bands) and ISSR (651 bands) analyses were polymorphic. The lowest genetic distance values obtained from RAPD and ISSR analyses were 0.0156 (between P. × melitenense and P. kurdica) and 0.0142 (between P. × melitenense and P. kurdica) respectively. The highest genetic distance values obtained from RAPD and ISSR analyses were 0.0866 (between P. kotschyana and P. oppositiflora) and 0.1237 (between P. kotschyana and P. kurdica × capitata) respectively. While P. kurdica indicated the highest genetic diversity (H = 0.1572; I = 0.2646) in RAPD analysis, P. capitata displayed the highest genetic diversity (H = 0.1403; I = 0.2329) in ISSR analysis. AMOVA results showed that 86% and 75% of the total variance resided within groups based on RAPD and ISSR markers, respectively. Based on the RAPD and ISSR results, both P. × melitenense and P. kurdica × capitata samples inherited species specific bands from their parental species, which confirms their hybridity. Although both P. × melitenense and P. kurdica × capitata hybrids showed a morphological mosaic between their parental phenotypes in terms of the majority of the quantitative characters examined, P. × melitenense and P. kurdica × capitata exceeded their parental phenotypes in terms of the three and 11 quantitative characters respectively. MANOVA results from the morphological data showed significant distinction among P. kurdica, P. oppositiflora, P. × melitenense, P. capitata and P. kurdica × capitata (Wilks’ Lambda = 0.003; df = 112; P < 0.01). Average pollen fertilities of P. oppositiflora, P. × melitenense, P. capitata, P. kurdica and P. kurdica × capitata were 93.44%, 68.42%, 93.28 %, 90.12% and 92.77% respectively.

The community formed by the dominance of Carex elata ssp. in Belarus was studied. The stands are classified into the associations Caricetum elatae and Caricetum omskianae (Phragmito-Magnocaricetea). Both syntaxons are characterized by species composition, diagnostic species, synmorphology, habitat. Documented by a synoptic table, the syntaxonomy, structure, ecology, distribution and variability of each syntaxon are discussed.

Greenhouse pot experiments were conducted to investigate the effect of salinity stress on survival percentage and physiological parameters in three varieties of Pistacia vera L.; including ‘Sarakhs’, ‘Badami’ and ‘Akbari’. To impose salinity stress, the plants were watered with NaCl solution (250 mM). The physiological characters were investigated at 3 rd , 6 th and 9 th days of salinity induction. Based on our results, Badami showed a greater survival percentage among the other varieties. The highest level of proline, soluble sugar, anthocyanin and glycine-betaine contents and the lowest amount of H 2 O 2 and MDA were also detected in Badami. Large quantities of H 2 O 2 , MDA and phenolic compounds were detected in Sarakhs. The highest level of superoxide dismutase (SOD) and catalase (CAT) activity was detected in Badami, while the highest activity of polyphenol oxidase (PPO) was seen in Sarakhs. All physiological characters increased during 3 rd to 9 th days of salinity induction, except CAT and SOD. In conclusion, Badami was more tolerant to salinity stress in comparison with other varieties due to it’s high capacity for osmotic adjustment and antioxidant protection. The physiological parameters including proline and H 2 O 2 contents, besides the activity of CAT can be used as biochemical markers for identification of tolerant varieties of Pistacia vera.

The Starch Branching Enzyme I (SBEI) gene plays an important role in amylopectin synthesis. Here, we isolated and characterized the full-length cDNA and DNA sequences of SBEI gene from diploid Triticeae species, Triticum monococcum, T. urartu, Aegilopsspeltoides, and Ae. tauschii. Then we predicted its protein structure, analyzed its evolutionary relationship with other species, and explored its expression patterns using real-time quantitative PCR. The SBEI cDNA includes a 2,490-bp open reading frame (ORF) encoding 829 amino acids. The genomic DNA of SBEI is 5,526-bp in length, containes fourteen exons and thirteen introns, and shares a similar structure with its homologous genes from other cereal plants. Sequence similarity ranging from 70.50% to 98.02% in exons and from 15.50% to 83.63% in introns was detected. Results of phylogenetic tree based on the deduced amino acid sequences from T. monococcum and other plants indicated that T. monococcum SBEI is more closely related to T. boeoticum and T. urartu. Expression analysis revealed that T. monococcum SBEI and AGPase genes were highly expressed in the seeds at middle developmental stage. This is the first report on characterization of the SBEI gene in T. monococcum. These results could be used to explore the roles of this enzyme in amylopectin synthesis.

The effect of specific microclimatic conditions and temporary flooding on terrestrial and aquatic invertebrate community composition as well as on parietal fauna was investigated in a cave located on the Krakow-Wieluń Upland. Studies of the fauna started after partial water retreat from the cave. Microclimate conditions and water chemistry parameters were also investigated. Due to the cave geometry and its size, temperatures recorded during summer were relatively high, whereas almost all its parts were frozen in winter. Temporary cave flooding promoted reproducing populations of Asellus aquaticus, Pseudocandona sarsi and copepods (Diacyclops bicuspidatus, Megacyclops viridis). Completely depigmented (including eyes), slightly, as well as fully pigmented individuals of A. aquaticus were found. Some dipterans started to reproduce in the flooded cave but drying stopped their development. Some taxa typical for terrestrial habitats (Collembola and Acari) and for parietal fauna (spider Meta menardi, moth Triphosa dubitata, dipterans from families Helomyzidae and Mycethophilidae) were not observed. Moreover, cave freezing significantly diminished the number of Culicidae, Oniscus sp., gastropods and Nesticus cellulanus wintering in the cave. The recolonization process was possible due to the cave location at the shallow subsurface and multiple entrances. The diversity of terrestrial invertebrates in the cave is very low as the effect of periodical flooding and freezing.

We present the first data from Poland concerning parasitism of Hydrachnidia on Chironomidae. Larvae of Hygrobates sp., H. setosus, Unionicola aculeata and Piona stjoerdalensis were found attached to adult Chironomidae (Cricotopus triannulatus, Tanytarsus sp., Paratanytarsus sp., Critocopus sp. and Orthocladius sp.). Intensity of infestation ranged between 1 to 2 larvae. All of the parasites were attached to abdomens of the hosts. The determination of parasitism of U. aculeata on Paratanytarsus and Tanytarsus genera extends the list of the hosts of this water mite species to the mentioned genera of Chironomidae. In case of H. setosus, the parasitism of this water mite on currently known hosts from the family Chironomidae has been confirmed.

Within the investigated river basins the deforestation ranged from 0-45.5%, dependent on the amount of windstorm damage. Our water temperature readings revealed that the canopy elimination above the streams in the areas damaged by the windstorm caused increase in daily and annual water temperature and also wider daily water temperature range, than those in the undisturbed reference stream, which caused the decline of cold stenotherm species abundance. The stream basins deforestation was collinear with FPOM and UFPOM concentrations, water temperature gradient and nitrate concentrations. Statistical analysis showed that size of stream, conductivity, catchment area and discharge were negatively correlated with the stonefly mesotherm ratio and nitrate contents. Higher average concentrations of nutrients (like NO − 3 ) in samples from some streams are a consequence of their mobilization from disturbed forest soils due to the windstorm. Most streams located in the deforested area were found by us in a higher trophic status, especially due the particulate organic matter (POM) content and biofilms amount, what caused increase in some functional feeding groups occurrence (collectorgatherers, scrapers and mainly predators). Stream degradation positively correlated with the percentage of deforested area, ratio of eurytherm taxa, transported organic matter, dissolved organic matter, and biofilms occurrence. The conductivity values measured in water samples were collinear with alkalinity and pH values and increased with watersheds areal extent. We established environmental variables influenced by the windstorm event along the catchment degradation gradient using measurable characteristics of stonefly assemblages and metrics. The proportion of the Leuctridae family and the Nemoura genus expressed as the LN index (1) shows a negative correlation with part undisturbed forested areas and channel stability, and positive correlation with stream erosion (expressed as the TAM quantity) in river basins. Proportion of shredders among the investigated streams was significantly higher in deforested streams, and shows lower stability of stonefly community in these habitats, where significantly fluctuate channel stability, water temperature and discharge of these streams.

Body size dimorphism between genders is a commonly observed phenomenon in insects, usually manifested in larger female body size. Sexual Size Dimorphism (SSD) varies from species to species, the degree and direction influenced by certain evolutionary pressures. Intraspecific variation in SSD may also occur between populations. The Hypera postica (Gyllenhal, 1813) is a well-known alfalfa plant pest that shows a degree of morphological divergence in its populations. The female alfalfa weevils are very fecund and have a larger body size compared to males. To improve our knowledge on magnitude and direction of SSD in alfalfa weevil, we studied 200 specimens of H. postica from four Iranian populations (Karaj1, Karaj2, Tuyserkan and Jovein). 10 morphological variables from three external anatomic parts (pronotum, elytra and rostrum) and 45 ratio characters were statistically analyzed in order to determine the amount of SSD in Iranian populations. In addition we investigated for morphological divergence pattern in mentioned populations. The results of this study show that a low degree of morphological divergence occurs in Iranian populations. Measured variables indicate that the SSD pattern of H. postica is compatible with the Rensch’s rule, and is related to high fecundity of females and a lack of strong sexual selection. Also it is mentioned that the larger rostrum in females may correspond to its unique role in egg laying.

A 12-month zooplankton survey (September 2008 to August 2009) in the hypoxic, stratified and seasonally eutrophic Amvrakikos Gulf (western Greece) revealed the presence of larvae of the Cephalochordate Branchiostoma lanceolatum. The larvae were found at almost all sampling occasions, except September and February, and presented their peak of abundance in April (426.8 ind. m −3 ). The oxygen depletion in the water column affected their vertical and horizontal distribution. Thus, their abundance was greater in the subsurface layers (10-20 m) which corresponds to the well oxygenated thermocline layer. An east to west increase of their abundance in the deeper layers was associated with the severity of oxygen depletion. The lower oxygen limit for their presence seems to have been 2 mg L −1 which corresponds to hypoxia. A size-specific depth distribution was noticed, with larger larvae residing in deeper strata. A diel vertical migration of the larvae in the summer was recorded, with a night ascent in shallower depths and a day descent in deeper strata. There was an apparent coexistence in space and time between the larvae and the dinoflagellate Ceratium sp., though this could not indicate a trophic interrelation between them due to the large size of the latter. The results suggest that, during their planktonic stage, the larvae utilize the water stratification within the Amvrakikos Gulf by inhabiting the depths within or close to the thermocline, where they can satisfy their oxygen and energy demands while being protected from predation.

Clarias gariepinus is an economically important fish belonging to Siluriformes. In order to understand the characterization of catfishes, the complete mitochondrial genome sequence of the African catfish C. gariepinus was determined using 17 pairs of primers. The complete mitogenome is 16,508 bp in length and contains 37 mitochondrial genes (13 protein-coding, two rRNA, and 22 tRNA genes) and a control region as in most other fishes. Moreover, a comparison of C. gariepinus with Clarias batrachus, Clarias fuscus and Clarias sp. NM-2010 indicated that the four mitogenomics were similar in length, total nucleotide compositions and codon usage patterns, but different in 13 protein-coding genes, control region and other aspects. The complete mitochondrial genome sequence of C. gariepinus would contribute to better understanding of population genetics, conservation, biogeography and evolution of this species.

Vitellogenin (VTG) is the major precursor of yolk proteins and could transport many components into oocytes, and hence it was thought to be a potential marker of reproduction of oviparious animal. Andrias davidianus is a critically endangered amphibian species, of which the wild population decreased dramatically and the artificial breeding has been successfully established. In this study, a full-length VTG cDNA was cloned and characterized from A. davidianus (designated as AdVTG) using 5’ and 3’ rapid amplification cDNA ends and RT-PCR. The AdVTG cDNA is 5,418 base pair in length and it encodes 1,805 amino acid long protein with typical quadripartite domains and several conserved cleavage sites. The tissue expression pattern indicated that AdVTG is mainly expressed in liver in one-year old individual. The threedimensional model of AdVTG constructed by homology modelling revealed the presence of conserved N-terminal β-sheet and lipid-binding cavity. Moreover, the phylogenetic analysis of VTG in metazoa was carried out, which revealed that Anuraspecific gene duplication occurred after the divergence of Caudata. All these results would improve the understanding of the physiological roles of AdVTG in A. davidianus and contribute to elucidating the evolution of VTG in vertebrates.

Using RT-PCR we examined expression of dopamine and adrenergic receptors in undifferentiated and spontaneously differentiating mouse embryonic stem (ES) cells. We also examined expression of dopamine receptor subtypes in mouse ovulated oocytes and preimplantation embryos. Comparing the expression of catecholamine receptors in undifferentiated mouse ES cells and in blastocysts (from which ES cells are derived), we found that transcripts of all five dopamine receptors were expressed in both cell types. In contrast, we detected eight adrenergic receptor subtypes in undifferentiated mouse ES cells, but only three subtypes were found in mouse blastocysts. In three adrenergic receptors (α1D, α2B, β1), we found higher expression in the spontaneously differentiating ES cells than in undifferentiated ES cells, and the α1B adrenoceptor was not even detectable in the undifferentiated cells. These results indicate that genes encoding all types of catecholamine receptors are transcribed in mouse ES cells, and some of them are differentially expressed during ES cell differentiation. We found several profiles of dopamine receptor mRNA expression during the preimplantation period. The DR3 transcript was present in all examined stages (oocytes, 4-cell embryos, 8- to 16-cell embryos, blastocysts). DR1 and DR4 transcripts were not found in oocytes, but we detected them in preimplantation embryos. The DR2 receptor transcript was found in all examined stages except for the 4-cell embryos, and the DR5 receptor transcript was found in all examined stages except for the 8- to 16-cell embryos. The expression profiles of dopamine receptor transcripts suggest different roles of some receptor subtypes in particular preimplantation developmental stages.