Partial Opsin Sequences Suggest UV-Sensitive Vision is Widespread in Caudata

Ultraviolet (UV) vision exists in several animal groups. Intuitively, one would expect this trait to be favoured in species living in bright environments, where UV light is the most present. However, UV sensitivity, as deduced from sequences of UV photoreceptors and/or ocular media transmittance, is also present in nocturnal species, raising questions about the selective pressure maintaining this perceptual ability. Amphibians are among the most nocturnal vertebrates but their visual ecology remains poorly understood relative to other groups. Perhaps because many of these species breed in environments that filter out a large part of UV radiation, physiological and behavioural studies of UV sensitivity in this group are scarce. We investigated the extent of UV vision in Caudata, the order of amphibians with the most nocturnal habits. We could recover sequences of the UV sensitive SWS1 opsin in 40 out of 58 species, belonging to 6 families. In all of these species, the evidence suggests the presence of functional SWS1 opsins under purifying selection, potentially allowing UV vision. Interestingly, most species whose opsin genes failed to amplify exhibited particular ecological features that could drive the loss of UV vision. This likely wide distribution of functional UV photoreceptors in Caudata sheds a new light on the visual ecology of amphibians and questions the function of UV vision in nocturnal animal species.     

Development of Vaccine Prototype Against Zika Virus Disease of Peptide-Loaded PLGA Nanoparticles and Evaluation of Cytotoxicity

Zika virus has recently evolved from an obscure mosquito-borne pathogen to an international public health concern. People with Zika virus disease can have indications including mild fever, skin rash, conjunctivitis, muscle pain, malaise or headache. Effective vaccines are needed for controlling and preventing the disease. In the current study, we aim to design the substructure for vaccine against Zika virus by forming antigenic peptide epitope of the disease. Zika peptide loaded poly (lactic-co-glycolic acid) (PLGA) nanoparticles have been fabricated in the present work as a potential artificial vaccine. UV and FT-IR Spectrophotometers and ZetaSizer were used for studying the nanoparticles, and Scanning Electron Microscope was used for morphological examination. The nanoparticles (NPs) yield, encapsulation efficiency, the peptide loading capacity were determined and in vitro release of the peptide was studied. Cytotoxic effects of the various concentrations of Zika peptide, blank PLGA nanoparticles and Zika peptide loaded PLGA nanoparticles on ECV304 human epithelial cells were determined via MTT assay. The present paper could be considered as one of the important steps in the use of nanoparticles for constructing the new generation of vaccination systems.

     

Antioxidant and antimicrobial actions of the clubmoss <Emphasis Type="Italic">Lycopodium clavatum</Emphasis> L.

Purpose of the present study was to evaluate antioxidant, antibacterial, antifungal, and antiviral activities of the petroleum ether, chloroform, ethyl acetate and methanol extracts as well as the alkaloid fraction of Lycopodium clavatum L. (LC) from Lycopodiaceae growing in Turkey. Antioxidant activity of the LC extracts was evaluated by 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical-scavenging method at 0.2 mg/ml using microplate-reader assay. Antiviral assessment of LC extracts was evaluated towards the DNA virus Herpes simplex (HSV) and the RNA virus Parainfluenza (PI-3) using Madin-Darby Bovine Kidney (MDBK) and Vero cell lines. Antibacterial and antifungal activities of the extracts were tested against standard and isolated strains of the following bacteria; Escherichia coli, Pseudomonas aeruginosa, Proteus mirabilis, Acinobacter baumannii, Klebsiella pneumoniae, Staphylococcus aureus, Bacillus subtilis as well as the fungi; Candida albicans and C. parapsilosis. All of the extracts possessed noteworthy activity against ATCC strain of S. aureus (4 μg/ml), while the LC extracts showed reasonable antifungal effect. On the other hand, we found that only the chloroform extract was active against HSV (16–8 μg/ml), while petroleum ether and alkaloid extracts inhibited potently PI-3 (16–4 μg/ml and 32–4 μg/ml, respectively). However, all of the extracts had insignificant antiradical effect on DPPH. In addition, we also analyzed the content of the alkaloid fraction of the plant by capillary gas chromatography-mass spectrometry (GC-MS) and identified lycopodine as the major alkaloid.     

Autophagy and mTOR pathways in mouse embryonic stem cell,lung cancer and somatic fibroblast cell lines

Embryonic developmental stages and regulations have always been one of the most intriguing aspects of science. Since the cancer stem cell discovery, striking for cancer development and recurrence, embryonic stem cells and control mechanisms, as well as cancer cells and cancer stem cell control mechanisms become important research materials. It is necessary to reveal the similarities and differences between somatic and cancer cells which are formed of embryonic stem cells divisions and determinations. For this purpose, mouse embryonic stem cells (mESCs), mouse skin fibroblast cells (MSFs) and mouse lung squamous cancer cells (SqLCCs) were grown in vitro and the differences between these three cell lines signalling regulations of mechanistic target of rapamycin (mTOR) and autophagic pathways were demonstrated by immunofluorescence and real-time polymerase chain reaction. Expressional differences were clearly shown between embryonic, cancer and somatic cells that mESCs displayed higher expressional level of Atg10, Hdac1 and Cln3 which are related with autophagic regulation and Hsp4, Prkca, Rhoa and ribosomal S6 genes related with mTOR activity. LC3 and mTOR protein levels were lower in mESCs than MSFs. Thus, the mechanisms of embryonic stem cell regulation results in the formation of somatic tissues whereas that these cells may be the causative agents of cancer in any deterioration.     

Antioxidative metabolism in down syndrome

Down syndrome is the most common cause of mental retardation, affecting 1 in 700–800 liveborn infants. Although numerous biochemical abnormalities accompanying the syndrome have not yet been completely clarified, the antioxidant defense system enzymes have shown to be altered due to increased gene dosage on chromosome 21 and overproduction of superoxide dismutase (SOD-1 or Cu/Zn SOD). The purpose of this study was to investigate the activities of SOD-1 and glutathione peroxidase (GSH-Px) enzymes and the levels of their cofactors zinc (Zn), copper (Cu) and selenium (Se) in plasma of 20 Down syndrome patients. In comparison with age and sex-matched controls (n=15), plasma GSH-Px, SOD, and Cu levels were significantly decreased in the patient group, but Zn and Se concentrations remained unchanged. This study was presented as a poster in 29th Annual Meeting of European Society of Human Genetics held in Genoa in May, 1997.     

Bioluminescence Imaging of Clavibacter michiganensis subsp. michiganensis Infection of Tomato Seeds and Plants

Clavibacter michiganensis subsp. michiganensis is a Gram-positive bacterium that causes wilting and cankers, leading to severe economic losses in commercial tomato production worldwide. The disease is transmitted from infected seeds to seedlings and mechanically from plant to plant during seedling production, grafting, pruning, and harvesting. Because of the lack of tools for genetic manipulation, very little is known regarding the mechanisms of seed and seedling infection and movement of C. michiganensis subsp. michiganensis in grafted plants, two focal points for application of bacterial canker control measures in tomato. To facilitate studies on the C. michiganensis subsp. michiganensis movement in tomato seed and grafted plants, we isolated a bioluminescent C. michiganensis subsp. michiganensis strain using the modified Tn1409 containing a promoterless lux reporter. A total of 19 bioluminescent C. michiganensis subsp. michiganensis mutants were obtained. All mutants tested induced a hypersensitive response in Mirabilis jalapa and caused wilting of tomato plants. Real-time colonization studies of germinating seeds using a virulent, stable, constitutively bioluminescent strain, BL-Cmm17, showed that C. michiganensis subsp. michiganensis aggregated on hypocotyls and cotyledons at an early stage of germination. In grafted seedlings in which either the rootstock or scion was exposed to BL-Cmm17 via a contaminated grafting knife, bacteria were translocated in both directions from the graft union at higher inoculum doses. These results emphasize the use of bioluminescent C. michiganensis subsp. michiganensis to help better elucidate the C. michiganensis subsp. michiganensis-tomato plant interactions. Further, we demonstrated the broader applicability of this tool by successful transformation of C. michiganensis subsp. nebraskensis with Tn1409::lux. Thus, our approach would be highly useful to understand the pathogenesis of diseases caused by other subspecies of the agriculturally important C. michiganensis.Clavibacter michiganensis subsp. michiganensis is a Gram-positive, aerobic bacterium that belongs to a group of plant-pathogenic actinomycetes (37). Infections by C. michiganensis subsp. michiganensis cause bacterial canker and wilt in tomato, which is considered one of the most destructive and economically significant diseases of this crop. Severe epidemics can cause up to 80% yield loss, mainly due to wilting and death of plants and lesions on fruit. Bacterial canker was first discovered in Michigan greenhouses in 1909 and has now been reported to occur in most tomato production areas around the world (11, 40).Plant wounds facilitate but are not required for infection by C. michiganensis subsp. michiganensis, which invades the xylem vessels and causes vascular disease with high titers (10⁹ bacteria/g of plant tissue) (2, 29), impairing water transport and leading to plant wilting, canker stem lesions, and death (17, 23). Alternatively, asymptomatic infections can be induced by C. michiganensis subsp. michiganensis during late stages of plant development, resulting in the production of contaminated seeds, a major source of outbreaks of C. michiganensis subsp. michiganensis infections in tomato production (13, 34). Traditional bacterial-disease management measures, such as applications of antibiotics and copper bactericides, have not been successful against this disease, and canker-resistant tomato cultivars are not available. As a result, C. michiganensis subsp. michiganensis has been included under international quarantine regulation (10, 11). Consequently, seed testing and maintaining pathogen-free seeds and transplants is currently the most appropriate approach to minimize the spread of disease (23). However, even a low C. michiganensis subsp. michiganensis transmission rate (0.01%) from seed to seedling can cause a disease epidemic under favorable conditions (5). Due to overcrowding of seedlings during transplant production, the pathogen can easily spread through splashing of irrigation water and leaf contact. Despite its apparent significance in C. michiganensis subsp. michiganensis epidemiology, the mechanism of seed-to-seedling transmission of C. michiganensis subsp. michiganensis is not well understood.Another critical point for disease spread is the grafting process, which is now a common practice for the majority of plants used in production greenhouses. Desirable tomato cultivars (scions) are grafted onto rootstocks that provide greater vigor, longevity, or, in some cases, disease resistance (26). Grafting requires cutting both rootstock and scion, providing a quick way for C. michiganensis subsp. michiganensis to spread from plant to plant. However, grafting is a relatively recent innovation in tomato production, and little is known about how grafting affects the dynamics of C. michiganensis subsp. michiganensis infection. Developing adequate control measures for C. michiganensis subsp. michiganensis is complicated by the complexity of genetic manipulation of Gram-positive bacteria, which impairs analysis and characterization of pathogenesis mechanisms (23). Consequently, there is a need to develop molecular techniques that would allow a better understanding of C. michiganensis subsp. michiganensis infections.One method of interest is using engineered bioluminescent bacteria to monitor plant-pathogen interactions in real time. By exploiting natural light-emitting reactions that are encoded by the luxCDABE genes, bioluminescent bacteria have been used to assess gene expression and to monitor the internalization and distribution of bacteria in hosts (3, 6, 7, 8, 9, 12, 15, 24, 31, 35, 36). In particular, bioluminescent phytopathogenic Xanthomonas campestris pathovars and Pseudomonas spp. have been used to track bacterial movement and distribution in host plants (7, 8, 15, 31, 36), as well as to assess host susceptibility quantitatively (15). Likewise, the lux genes have also been transferred to beneficial bacteria, such as Rhizobium leguminosarum and Pseudomonas spp. to visualize colonization patterns in rhizospheres (3, 9).The genes that carry the function of light emission are luxAB, which express luciferase enzymes that catalyze the bioluminescent reaction, while luxCDE encode the enzymes required for biosynthesis of a fatty aldehyde substrate necessary for the reaction (28, 39). Bioluminescence involves an intracellular oxidation of the reduced form of flavin mononucleotide and the fatty aldehyde by luciferase in the presence of molecular oxygen; therefore, bacterial bioluminescence also requires oxygen, a source of energy (38). Cells that express the lux operon spontaneously emit photons that can be captured by a sensitive charge-coupled-device (CCD) camera, enabling imaging and visualization of bacterial cells (22). Luciferase activity depends on the metabolic integrity of the cell, while the number of photons emitted correlates with the biomass of living bacteria (12, 31). Furthermore, since the half-life of luciferase binding to its substrate is several seconds (28), captured light events reflect processes in real time and are not artifacts of accumulated signals. Consequently, live imaging of bioluminescence provides a sensitive means of visualizing bacterial colonization and invasion of hosts and allows real-time representation and examination of pathogen-plant interactions (24, 36).Very little information is available about the mechanisms of C. michiganensis subsp. michiganensis pathogenesis and its colonization of seeds and subsequent transmission to seedlings. This is largely attributable to a lack of tools and difficulties in genetically manipulating this Gram-positive bacterium (30). However, recent development of an insertion sequence element IS1409 (Tn1409)-based efficient transposon mutagenesis system for C. michiganensis subsp. michiganensis has increased our knowledge of the pathogenesis of tomato canker (16, 25). To better understand the dynamics of seed-to-seedling transmission of C. michiganensis subsp. michiganensis, as well as movement of C. michiganensis subsp. michiganensis in grafted plants, we constructed a bioluminescent C. michiganensis subsp. michiganensis strain using the Tn1409 transposon mutagenesis system. Our results demonstrated the utility of using a bioluminescent C. michiganensis subsp. michiganensis strain as a novel approach to elucidate the interaction of plants with this economically important pathogen.     

Structural investigation of the glandular trichomes of <Emphasis Type="Italic">Salvia argentea</Emphasis>

The morphology, anatomy and distribution of glandular trichomes on the aerial organs of Salvia argentea L. has been investigated. Two morphologically distinct types of glandular trichomes were determined. Capitate glandular trichomes forming a base 1–7 celled, a stalk 1–5 celled or no stalk and a head uni- or bicellular had various types. In capitate trichomes, the neck cell that has an important role especially for xeroformic plants, acting to prevent the backflow of secreted substance through the apoplast has been distinctively observed in the investigated species. The capitate trichomes were present abundantly on all aerial organs of S. argentea. Peltate glandular trichomes had a large secretory head forming 1–5, 8 central and 8–10, 12, 14 peripheral cells. Peltate trichomes are present on all aerial organs, except petiole, being the most abundant on calyx and corolla. Results were shown by tables and photographs.     

Sleep stage and obstructive apneaic epoch classification using single-lead ECG

Background  

Polysomnography (PSG) is used to define physiological sleep and different physiological sleep stages, to assess sleep quality and diagnose many types of sleep disorders such as obstructive sleep apnea. However, PSG requires not only the connection of various sensors and electrodes to the subject but also spending the night in a bed that is different from the subject's own bed. This study is designed to investigate the feasibility of automatic classification of sleep stages and obstructive apneaic epochs using only the features derived from a single-lead electrocardiography (ECG) signal.     

Ultrastructural Changes in the Kidney of Rats with Acute Exposure to Cadmium and Effects of Exogenous Metallothionein

Ultrastructural changes in the kidneys of rats after acute cadmium exposure and the effects of exogenous metallothionein (MT) were studied by transmission electron microscopy. Thirty-six adult Wistar rats were divided into three groups. Cadmium chloride (CdCl₂) (3.5 mg/kg/day) was injected subcutaneously in the first group. In the second group, 30 μmol/kg MT was administered in addition to CdCl₂. Control rats received 0.5 ml subcutaneous saline solution. Four rats from each group were killed on days 1, 3, 5, and 7 after administration of the compounds. Kidney tissues were taken and fixed in 2.5% glutaraldehyde solution for electron microscopic observations. Tissue damage in kidney increased as time passed since the administration of CdCl₂ in the first group. Degeneration in the proximal and distal tubules was observed. Increased apoptosis was seen in the proximal tubules epithelium, especially on day 7. Peritubular capillaries became dilated, there was degeneration of the endothelial cells, and the amount of intertubular collagen fibers was increased. On day 1, irregular microvilli in the proximal tubules, deepening of the basal striations, and myelin figures; on day 3, multiple vesicular mitochondria and regions of edema around tubules; on days 5 and 7, increased apoptotic cell in the proximal tubules and widened rough endoplasmic reticulum of the endothelial cells of glomerular capillaries were observed. We observed that the structural alterations that increased depending on the day of Cd administration decreased after exogenous MT administration, the dilation of the peritubular capillaries persisted, and there were degenerated proximal tubules. It was established that cadmium chloride was toxic for kidney cortex and caused structural damage. Exogenous MT partly prevents CdCl₂-induced damage.