High diversity of thermophilic cyanobacteria in Rupite hot spring identified by microscopy,cultivation, single-cell PCR and amplicon sequencing

Extremophiles - Genotypic and morphological diversity of cyanobacteria in the Rupite hot spring (Bulgaria) was investigated by means of optical microscopy, cultivation, single-cell PCR, and 16S...     

Vibrational and electronic circular dichroism study of the interactions of cationic porphyrins with (dG-dC)10 and (dA-dT)10

The interactions of two different porphyrins, without axial ligands-5,10,15,20-tetrakis(1-methylpyridinium-4-yl)porphyrin-Cu(II) tetrachloride (Cu(II)TMPyP) and with bulky meso substituents-5,10,15,20-tetrakis(N,N,N-trimethylanilinium-4-yl)porphyrin tetrachloride (TMAP), with (dG-dC)10 and (dA-dT)10 were studied by combination of vibrational circular dichroism (VCD) and electronic circular dichroism (ECD) spectroscopy at different [oligonucleotide]/[porphyrin] ratios, where [oligonucleotide] and [porphyrin] are the concentrations of oligonucleotide per base-pair and porphyrin, respectively. The combination of VCD and ECD spectroscopy enables us to identify the types of interactions, and to specify the sites of interactions: The intercalative binding mode of Cu(II)TMPyP with (dG-dC)(10), which has been well described, was characterized by a new VCD "marker" and it was shown that the interaction of Cu(II)TMPyP with (dA-dT)10 via external binding to the phosphate backbone and major groove binding caused transition from the B to the non-B conformer. TMAP interacted with the major groove of (dG-dC)10, was semi-intercalated into (dA-dT)10, and caused significant variation in the structure of both oligonucleotides at the higher concentration of porphyrin. The spectroscopic techniques used in this study revealed that porphyrin binding with AT sequences caused substantial variation of the DNA structure. It was shown that VCD spectroscopy is an effective tool for the conformational studies of nucleic acid-porphyrin complexes in solution.     

Cytogenetic evidence for diversity of two nuclei within a single diplomonad cell of <Emphasis Type="Italic">Giardia</Emphasis>

Giardia intestinalis is an ancient protist that causes the most commonly reported human diarrheal disease of parasitic origin worldwide. An intriguing feature of the Giardia cell is the presence of two morphologically similar nuclei, generally considered equivalent, in spite of the fact that their karyotypes are unknown. We found that within a single cell, the two nuclei differ both in the number and the size of chromosomes and that representatives of two major genetic groups of G. intestinalis possess different karyotypes. Odd chromosome numbers indicate aneuploidy of Giardia nuclei, and their stable occurrence is suggestive of a long-term asexuality. A semi-open type of Giardia mitosis excludes a chromosome interfusion between the nuclei. Differences in karyotype and DNA content, and cell cycle-dependent asynchrony are indicative of diversity of the two Giardia nuclei.     

Phenotypic and genotypic analysis of<Emphasis Type="Italic">Borrelia</Emphasis> spp. isolated from<Emphasis Type="Italic">Ixodes ricinus</Emphasis> ticks by using electrophoretic chips and real-time polymerase chain reaction

The genotype of Borrelia burgdorferi sensu lato was detected in 371 out of 1244 ticks. Borrelia determination was based on partial sequencing of the 16S rRNA gene and real-time polymerase chain reactions for identification and quantitation of ospA and recA genes. Different Borrelia spp. were identified; B. garinii in 40% ticks followed by B. afzelii (36.3%), B. burgdorferi sensu stricto (12.9%), B. valaisiana (3.5%), B. lusitaniae (0.8%), B. bissettii (0.5%) and B. miyamotoi-like (0.5%). Cultivation of 30 borrelia strains in BSK-H medium, among them B. valaisiana, B. bissettii-like and B. miyamotoi-like strains was unique in Czechia. Calibrated microfluidic-based quantification showed differences in the concentration of the nucleic acids and molar mass of the outer surface proteins of different Borrelia spp. with standard sensitivity and specificity and was helpful for their identification. The outer surface protein OspA was absent in B. miyamotoi-like and the OspB protein in B. valaisiana, B. lusitaniae and in three subtypes of B. garinii.     

Antimicrobial susceptibility of<Emphasis Type="Italic">Enterococcus</Emphasis> species isolated from slovak bryndza cheese

Three hundred and ten enterococcal isolates (178 Enterococcus faecium, 68 E. durans, 49 E. faecalis, 8 E. italicus, 3 E. gallinarum, 3 E. casseliflavus, and 1 E. hirae) from Slovak Bryndza cheese were evaluated for susceptibility to nine antimicrobial agents (vancomycin, teicoplanin, ampicillin, streptomycin, gentamicin, erythromycin, rifampicin, nitrofurantoin, and ciprofloxacin). All enterococcal isolates from Bryndza cheese were susceptible to ampicillin, streptomycin, gentamicin, vancomycin, and teicoplanin as determined by the disk diffusion method. Vancomycin resistance genes vanA and vanB were not detected. Resistance rates of enterococcal isolates to rifampicin, erythromycin, ciprofloxacin, and nitrofurantoin were 24, 26, 2, and 1 %, respectively. Thirty-six % of E. faecium isolates and 22 % of the E. faecalis isolates were resistant to erythromycin. Resistance to rifampicin was similar in E. faecium (31 %) and E. faecalis (29 %). Both E. faecium and E. faecalis strains showed the same resistance to ciprofloxacin (2 %). E. durans isolates showed low levels of resistance to rifampicin, erythromycin, ciprofloxacin, and nitrofurantoin (1-4 %). Forty-eight (30 %) of the E. faecium isolates, two (3 %) of the E. durans isolates, and six (12 %) of the E. faecalis isolates exhibited multidrug resistance. The highest frequency of resistant enterococci was observed in Bryndza produced in winter season.     

Radiochromatographic assay of metabolites of the oostatic peptide labeled in different positions of the peptide chain

Reversed-phase high-performance liquid radio-chromatography (radio-HPLC) was set up to detect the time course of labeled degradation product formation of the pentapeptide H-Tyr-Asp-Pro-Ala-Pro-OH (5P), which has oostatic effects in different insect species. The detection limit of the system was in the range of 80-150 Bq. To follow formation of the degradation products, three amino acid residues in 5P were independently tritiated: Tyr1, Pro3 and Pro5. Each of the three tritiated peptides was analyzed after incubation with fresh hemolymph or ovaries of Neobellieria bullata. In the incubation mixture, free terminal amino acids and shortened sequences of 5P were identified. A metabolite of tyrosine represented the only exception; it was finally identified as water using degradation of [3H]Tyr by tyrosinase. Metabolic degradation of [3H]Tyr-5P was found to be considerably quicker than that of H-[3H]Tyr-Asp-Pro-Ala-OH (4P). The degradation of 5P was considerably slower in ovaries in comparison to hemolymph.     

The platinum-olefin binding energy in series of (PH<Subscript>3</Subscript>)<Subscript>2</Subscript>Pt(olefin) complexes - a theoretical study

Theoretical investigation of Pt(0)-olefin organometallic complexes containing tertiary phosphine ligands was focused on the strength of platinum-olefin electronic interaction. DFT theoretical study of electronic effects in a substantial number of ethylene derivatives was evaluated in terms of the Pt-olefin binding energy using MP2 correlation theory. Organometallics bearing coordinated olefins with general formula (R¹R²C = CR³R⁴)Pt(PH₃)₂ [R = various substituents] had been selected, including olefins containing both electron-donor substituents as well as electron-withdrawing groups. The stability of the corresponding complexes increases with a strengthening electron-withdrawal ability of the olefin substituents. Figure Representation of (CH₂ = CHR)Pt(PPh₃)₂ and the stability chart     

CD8+ T lymphocytes protect SCID mice against Encephalitozoon cuniculi infection

Microsporidia are obligate intracellular parasites that cause opportunistic infections in immunocompromised patients. The role of two main T cell subsets in anti-microsporidial immunity has been studied using an Encephalitozoon cuniculi-severe combined immunodeficient (SCID) mouse model. Whereas SCID mice reconstituted with CD4+ T lymphocyte-depleted naive BALB/c splenocytes resolved the infection, adoptive transfer of CD8+ T cell-depleted splenocytes failed to protect the animals against a lethal E. cuniculi infection. Splenocytes from E. cuniculi-immune mice specifically killed syngeneic infected macrophages in a short-term 51Cr-release assay. These results suggest the crucial role of cytotoxic T lymphocytes in the protection against E. cuniculi infection.     

Reaction mechanism and stereochemistry of gamma-hexachlorocyclohexane dehydrochlorinase LinA

gamma-Hexachlorocyclohexane dehydrochlorinase (LinA) catalyzes the initial steps in the biotransformation of the important insecticide gamma-hexachlorocyclohexane (gamma-HCH) by the soil bacterium Sphingomonas paucimobilis UT26. Stereochemical analysis of the reaction products formed during conversion of gamma-HCH by LinA was investigated by GC-MS, NMR, CD, and molecular modeling. The NMR spectra of 1,3,4,5,6-pentachlorocyclohexene (PCCH) produced from gamma-HCH using either enzymatic dehydrochlorination or alkaline dehydrochlorination were compared and found to be identical. Both enantiomers present in the racemate of synthetic gamma-PCCH were converted by LinA, each at a different rate. 1,2,4-trichlorobenzene (1,2,4-TCB) was detected as the only product of the biotransformation of biosynthetic gamma-PCCH. 1,2,4-TCB and 1,2,3-TCB were identified as the dehydrochlorination products of racemic gamma-PCCH. delta-PCCH was detected as the only product of dehydrochlorination of delta-HCH. LinA requires the presence of a 1,2-biaxial HCl pair on a substrate molecule. LinA enantiotopologically differentiates two 1,2-biaxial HCl pairs present on gamma-HCH and gives rise to a single PCCH enantiomer 1,3(R),4(S),5(S),6(R)-PCCH. Furthermore, LinA enantiomerically differentiates 1,3(S),4(R),5(R),6(S)-PCCH and 1,3(R),4(S),5(S),6(R)-PCCH. The proposed mechanism of enzymatic biotransformation of gamma-HCH to 1,2,4-TCB by LinA consists of two 1,2-anti conformationally dependent dehydrochlorinations followed by 1,4-anti dehydrochlorination.