Phylogenomic insights into the diversity and evolution of Palearctic vipers.

Despite decades of molecular research, phylogenetic relationships in Palearctic vipers (genus Vipera) still essentially rely on a few loci, such as mitochondrial barcoding genes. Here we examined the diversity and evolution of Vipera with ddRAD-seq data from 33 representative species and subspecies. Phylogenomic analyses of âˆ¼ 1.1 Mb recovered nine major clades corresponding to known species/species complexes which are generally consistent with the mitochondrial phylogeny, albeit with a few deep discrepancies that highlight past hybridization events. The most spectacular case is the Italian-endemic V. walser, which is grouped with the alpine genetic diversity of V. berus in the nuclear tree despite carrying a divergent mitogenome related to the Caucasian V. kaznakovi complex. Clustering analyses of SNPs suggest potential admixture between diverged Iberian taxa (V. aspis zinnikeri and V. seoanei), and confirm that the Anatolian V. pontica corresponds to occasional hybrids between V. (ammodytes) meridionalis and V. kaznakovi. Finally, all analyzed lineages of the V. berus complex (including V. walser and V. barani) form vast areas of admixture and may be delimited as subspecies. Our study sets grounds for future taxonomic and phylogeographic surveys on Palearctic vipers, a group of prime interest for toxinological, ecological, biogeographic and conservation research.

Investigation of the effect of different environmental conditions on biofilm structure of Salmonella enterica serotype Virchow via FTIR spectroscopy.

This study aims to describe the content of polymeric matrix components under different incubation temperatures and pH levels. Optimal biofilm production of 15 S. Virchow isolates occurred following the incubation in LB-NaCl for 72 h, at pH 6.6 and 20 °C. The expression of csgA, csgD, adrA and bcsA genes at 20 °C, 25 °C and 30 °C in S. Virchow DMC18 was analyzed, and it was discovered that the maximum production of cellulose and curli fimbriae occurred at 20 °C. The physical characteristics of pellicle structure of S. Virchow DMC18 was determined as rigid at 20 °C, while becoming fragile at higher temperatures. FTIR analyses confirmed the obtained molecular findings. The intensities of the 16 different peaks originating from carbohydrate, protein, and nucleic acid in the spectra of biofilm samples significantly diminished (p < 0.05) with the increasing temperature. The highest intensities of lipids and carbohydrates were observed at 20 °C indicating the changes in cell surface properties.

Examination of cervical swabs of patients with endometriosis using Fourier transform infrared spectroscopy.

PURPOSE: There is no established non-invasive method to diagnose patients with endometriosis. As a nondestructive type of radiation, infrared light might be used for discrimination by causing vibration of the covalent bonds of the molecules when absorbed by the tissues. The aim of the study was to test whether cervical swab can be used to diagnose women with endometriosis using Fourier transform infrared spectroscopy (FTIR). METHODS: In this prospective case-control study, women between 18-45 years old and undergoing laparoscopy due to various reasons were recruited (n = 20). According to the findings during laparoscopy, patients were stratified as stage I-II or stage III-IV endometriosis groups. Women lacking any visible lesions of endometriosis were recruited as controls. A cervical swab was taken from all patients just before the surgical procedure and pulled into a tube containing saline solution. FTIR spectra were obtained and the fingerprint region (1750-850 cm-1) was used for analyses. RESULTS: Finally, three samples in stage I-II, five samples in stage III-IV and five samples in the control group were analyzed. Hierarchical cluster analysis and principal component analysis were performed as the chemometric method. A total of ten observable peaks were detected in the absorbance spectra of samples. The peaks at 1450 and 1405 cm-1 originating from lipids and proteins significantly increased in the stage III-IV endometriosis group when compared with controls. In addition, nucleic acid/carbohydrate ratio was significantly lower in the stage I-II group indicating that the alteration of the carbohydrate level might be important. CONCLUSIONS: Examination of cervical swab with FTIR spectroscopy might be a proper candidate for a non-invasive diagnostic approach of endometriosis.

Variations in age structure and growth in congeners Lacerta viridis and Lacerta media.

Determining the age of any species allows it to be analyzed from the ontogenetic, demographic, and ecological perspectives. In the present study, we tested the hypothesis that the age structure of congener species (Lacerta media and Lacerta viridis) with the same ecological niche may vary in different areas. In this context, we applied skeletochronology method to reveal various demographic parameters, such as age structure, longevity, age at sexual maturity, growth rate, survival rate, adult life expectancy, and the relationship between age and body size in the green lizard, L. viridis, and the medium lizard, L. media. In L. media and L. viridis, the maximum lifespan was 10 and 8 years, respectively. The mean age and body size of females were significantly greater than those of males in L. media. However, in the examined L. viridis population, no appreciable variation in mean age or body size was found to exist between the sexes. It was estimated that the green lizards reach maturity at the age of 2 or 3 years. However, the L. media reached sexual maturity approximately 1 year later than the congener. The body size markedly increased with age in males for both studied populations. However, in females, body size positively increased with age only in L. media. The approach of skeletochronology that we utilized in this study to assess age structure makes it simple to gather a variety of time-dependent ecological data for such ectothermic species.

A preliminary investigation into the venom proteome of Macrovipera lebetina obtusa (Dwigubsky, 1832) from Southeastern Anatolia by MALDI-TOF mass spectrometry and comparison of venom protein profiles with Macrovipera lebetina lebetina (Linnaeus, 1758) from Cyprus by 2D-PAGE.

We compared venoms of two subspecies of blunt-nosed viper Macrovipera lebetina (Linnaeus, 1758) from Southeastern Anatolia and Cyprus by two-dimensional gel electrophoresis (2D-PAGE). Additionally, peptide mass fingerprinting analysis was carried out using matrix-assisted laser desorption ionization/time of flight (MALDI-TOF) mass spectrometry in order to achieve preliminary protein identification from M. lebetina obtusa venom from Turkey. As a result of 2D-PAGE, statistical tests revealed some significant differences that can be considered as subspecies-specific biomarker candidates between two subspecies. Using bioinformatic analyses, proteins belonging to 11 families were identified from the venom of M. l. obtusa: phospholipase A(2), metalloproteinase, serin proteinase, disintegrin, cysteine-rich secretory protein, C-type lectin, vascular endothelial growth factor, nerve growth factor, hyaluronidase, L: -amino acid oxidase, and trypsin inhibitor. Venom of M. lebetina was studied by 2D-PAGE for the first time in the literature, and also this is the first work aiming to determine regional variations of snake venoms by this method in Turkey and Cyprus. Our preliminary results show that snake venom research deserves more attention in Turkey as well as in the toxinology field in general.

Modern venomics-Current insights, novel methods, and future perspectives in biological and applied animal venom research.

Venoms have evolved >100 times in all major animal groups, and their components, known as toxins, have been fine-tuned over millions of years into highly effective biochemical weapons. There are many outstanding questions on the evolution of toxin arsenals, such as how venom genes originate, how venom contributes to the fitness of venomous species, and which modifications at the genomic, transcriptomic, and protein level drive their evolution. These questions have received particularly little attention outside of snakes, cone snails, spiders, and scorpions. Venom compounds have further become a source of inspiration for translational research using their diverse bioactivities for various applications. We highlight here recent advances and new strategies in modern venomics and discuss how recent technological innovations and multi-omic methods dramatically improve research on venomous animals. The study of genomes and their modifications through CRISPR and knockdown technologies will increase our understanding of how toxins evolve and which functions they have in the different ontogenetic stages during the development of venomous animals. Mass spectrometry imaging combined with spatial transcriptomics, in situ hybridization techniques, and modern computer tomography gives us further insights into the spatial distribution of toxins in the venom system and the function of the venom apparatus. All these evolutionary and biological insights contribute to more efficiently identify venom compounds, which can then be synthesized or produced in adapted expression systems to test their bioactivity. Finally, we critically discuss recent agrochemical, pharmaceutical, therapeutic, and diagnostic (so-called translational) aspects of venoms from which humans benefit.

Peptidomic characterization and bioactivity of Protoiurus kraepelini (Scorpiones: Iuridae) venom.

Protoiurus kraepelini is a scorpion species found in parts of Turkey and Greece. In this study, the peptide profile of its venom was determined for the first time. The electrophoretic profile of the crude venom showed a protein distribution from 2 to 130 kDa. MALDI-TOF MS analysis of the venom peptide fraction yielded 27 peptides between 1059 and 4623 Da in mass. Several ion channelblocking and antimicrobial peptides were identified by peptide mass fingerprinting analysis. Cytotoxic and antimicrobial effects of the venom were also demonstrated on Jurkat cells and Escherichia coli, respectively. As the first peptidomic characterization study on P. kraepelini venom, this report lays the foundation for detailed future studies that may lead to the discovery of novel bioactive peptides.

Application of Fourier transform infrared spectroscopy to biomolecular profiling of cultured fibroblast cells from Gaucher disease patients: A preliminary investigation.

BACKGROUND: Gaucher disease (GD) is defined as an autosomal recessive disorder resulting from the deficiency of glucocerebrosidase (E.C. 3.2.1.45). Glucocerebrosidase is responsible for the degradation of glucosylceramide into ceramide and glucose. The deficiency of this enzyme results in the accumulation of undegraded glucosylceramide, almost exclusively in macrophages. With Fourier transform infrared (FTIR) spectroscopy, the complete molecular diversity of the samples can be studied comparatively and the amount of the particular materials can be determined. Also, the secondary structure ratios of proteins can be determined by analysing the amide peaks. OBJECTIVES: The primary aim of this study is to introduce FTIR-ATR spectroscopy technique to GD research for the first time in the literature and to assess its potential as a new molecular method. MATERIAL AND METHODS: Primary fibroblast cell cultures obtained from biopsy samples were used, since this material is widely used for the diagnosis of GD. Intact cells were placed onto a FTIR-ATR crystal and dried by purging nitrogen gas. Spectra were recorded in the mid-infrared region between 4500-850 cm-1 wavenumbers. Each peak in the spectra was assigned to as organic biomolecules according to their chemical bond information. A quantitative analysis was performed using peak areas and we also used a hierarchical cluster analysis as a multivariate spectral analysis. RESULTS: We obtained FTIR spectra of fibroblast samples and assigned the biomolecule origins of the peaks. We observed individual heterogeneity in FTIR spectra of GD fibroblast samples, confirming the well-known phenotypic heterogeneity in GD at the molecular level. Significant alterations in protein, lipid and carbohydrate levels related to the enzyme replacement therapy were also observed, which is also supported by cluster analysis. CONCLUSIONS: Our results showed that the application of FTIR spectroscopy to GD research deserves more attention and detailed studies with an increased sample size in order to evaluate its potential in the diagnosis and follow-up of GD patients.

Combined venom profiling and cytotoxicity screening of the Radde's mountain viper (Montivipera raddei) and Mount Bulgar Viper (Montivipera bulgardaghica) with potent cytotoxicity against human A549 lung carcinoma cells.

Here we report the first characterization of the endemic Mount Bulgar Viper (Montivipera bulgardaghica) and Radde's mountain viper (Montivipera raddei) venom by a combined approach using intact mass profiling and bottom-up proteomics. The cytotoxicity screening of crude venom as well as isolated serine proteases revealed a high activity against A549 human lung carcinoma cells. By means of intact mass profiling of native and reduced venom we observed basic and acidic phospholipases type A2. Moreover, the analysis revealed snake venom metalloproteases, cysteine-rich secretory proteins, disintegrins, snake venom serine proteases, C-type lectins, a vascular endothelial growth factor and an L-amino acid oxidase.

Prohemostatic and antithrombin activities of Ankaferd hemostat are linked to fibrinogen gamma chain and prothrombin by functional proteomic analyses.

Ankaferd blood stopper (ABS) is a novel topical hemostatic agent of plant origin registered for the management of external hemorrhages, in Turkey. The ABS-induced formation of the protein network with vital erythroid aggregation covers the whole physiological hemostatic process. The aim of this study is to assess prohemostatic and antithrombin effects of ABS on the basis of functional proteomic analyses performed in ABS-treated plasma and serum samples based on the previous hypotheses about ABS action. For this purpose, serum and plasma proteins were separated by 2-dimensional (2D) gel electrophoresis, and proteins were identified using reference plasma gel on Swiss-2DPAGE database. Our results indicated that fibrinogen gamma chain and prothrombin levels just initially decreased first and thereafter enhanced following the ABS exposure. Dual effects of ABS on those critical hemostatic molecules seem to be associated with prohemostatic and antithrombin activities of the hemostatic agent.