Purification and characterization of two cold-adapted extracellular tannin acyl hydrolases from an Antarctic strain <Emphasis Type="Italic">Verticillium</Emphasis> sp. P9

Two extracellular tannin acyl hydrolases (TAH I and TAH II) produced by an Antarctic filamentous fungus Verticillium sp. P9 were purified to homogeneity (7.9- and 10.5-fold with a yield of 1.6 and 0.9%, respectively) and characterized. TAH I and TAH II are multimeric (each consisting of approximately 40 and 46 kDa sub-units) glycoproteins containing 11 and 26% carbohydrates, respectively, and their molecular mass is approximately 155 kDa. TAH I and TAH II are optimally active at pH of 5.5 and 25 and 20°C, respectively. Both the enzymes were activated by Mg²⁺and Br⁻ ions and 0.5–2.0 M urea and inhibited by other metal ions (Zn²⁺, Cu²⁺, K⁺, Cd²⁺, Ag⁺, Fe³⁺, Mn²⁺, Co²⁺, Hg²⁺, Pb²⁺ and Sn²⁺), anions, Tween 20, Tween 60, Tween 80, Triton X-100, sodium dodecyl sulphate, β-mercaptoethanol, α-glutathione and 4-chloromercuribenzoate. Both tannases more efficiently hydrolyzed tannic acid than methyl gallate. E _a of these reactions and temperature dependence (at 0–30°C) of k _cat, k _cat/K _m, ΔG*, ΔH* and ΔS* for both the enzymes and substrates were determined. The k _cat and k _cat/K _m values (for both the substrates) were considerably higher for the combined preparation of TAH I and TAH II.     

The influence of glutamic and aminoacetic acids on the excitability of the liverwort Conocephalum conicum

Intracellular microelectrode measurements revealed that a resting potential (RP), an action potential (AP) and a calcium component of AP (named voltage transient, VT) can be influenced by glutamic acid (Glu) and aminoacetic acid (glycine, Gly) in the liverwort Conocephalum conicum. In the continuous presence of 5mM Glu or 5mM Gly, the RP hyperpolarized constantly and the plants became desensitized to the excitatory amino acids (Glu or Gly). Under such circumstances, the amplitudes of APs evoked by stimuli other than Glu or Gly grew, as did their calcium components (VTs). The sudden application of 1-15 mM Glu or Gly to a thallus not yet desensitized resulted in an excitation, i.e. a single AP or AP series. Aspartate (Asp) could not substitute for Glu in any way. Simultaneous action of both amino acids acted synergically to trigger APs. The same phenomenon was observed when glycine solution was enriched with N-methyl-D-aspartic acid (NMDA). Gly-induced APs were totally hindered by 1mM D-amino-5-phosphonopentanoic acid (AP5)--an inhibitor of ionotropic glutamate receptors of the NMDA kind. Glu-induced APs could be totally suppressed by 1mM AP5 as well as by 1mM 6,7-dinitroquinoxaline-2,3-dione (DNQX)--an inhibitor of AMPA/KA receptors. DNQX also completely blocked the calcium component of Glu-evoked APs. After DNQX treatment, the only response to Glu was a membrane potential hyperpolarization (like the Glu response in a desensitized plant). It was concluded that the Glu-induced depolarization and hyperpolarization are separate phenomena. The stimulatory effects of both Glu and Gly on liverwort excitability may be the consequences of an activation of a variety of ionotropic Glu receptor subtypes.     

Multidrug-resistant Salmonella enterica serovar paratyphi A harbors IncHI1 plasmids similar to those found in serovar typhi

Salmonella enterica serovars Typhi and Paratyphi A cause systemic infections in humans which are referred to as enteric fever. Multidrug-resistant (MDR) serovar Typhi isolates emerged in the 1980s, and in recent years MDR serovar Paratyphi A infections have become established as a significant problem across Asia. MDR in serovar Typhi is almost invariably associated with IncHI1 plasmids, but the genetic basis of MDR in serovar Paratyphi A has remained predominantly undefined. The DNA sequence of an IncHI1 plasmid, pAKU_1, encoding MDR in a serovar Paratyphi A strain has been determined. Significantly, this plasmid shares a common IncHI1-associated DNA backbone with the serovar Typhi plasmid pHCM1 and an S. enterica serovar Typhimurium plasmid pR27. Plasmids pAKU_1 and pHCM1 share 14 antibiotic resistance genes encoded within similar mobile elements, which appear to form a 24-kb composite transposon that has transferred as a single unit into different positions into their IncHI1 backbones. Thus, these plasmids have acquired similar antibiotic resistance genes independently via the horizontal transfer of mobile DNA elements. Furthermore, two IncHI1 plasmids from a Vietnamese isolate of serovar Typhi were found to contain features of the backbone sequence of pAKU_1 rather than pHCM1, with the composite transposon inserted in the same location as in the pAKU_1 sequence. Our data show that these serovar Typhi and Paratyphi A IncHI1 plasmids share highly conserved core DNA and have acquired similar mobile elements encoding antibiotic resistance genes in past decades.     

Genetic variability of body weight in two goose strains under long-term selection

Body weight is one of the most important traits in any genetic improvement program in geese for at least 2 reasons. First, measurements of the trait are very easy. Second, body weight is correlated with a number of other meat performance traits. However, the genetic background of body weight shows considerable complexity. Three genetic models (with direct, maternal genetic and permanent maternal environmental effects) were employed in this study. Records of 3076 individuals of maternal strain W11 and 2656 individuals of paternal strain W33 over 6 consecutive generations, kept in the pedigree farm of Ko?uda Wielka, were analysed. Body weight (in kilograms) was measured in weeks 8 (BW8) and 11 (BW11). The inbreeding levels in both populations were relatively low (0.14% and 0.02% for W11 and W33, respectively), therefore these effects were not included in the linear models to estimate genetic parameters. Three fixed effects (hatch period, sex and year) were included in each linear model. Two criteria (AIC, BIC) were used to check the goodness of fit of the models. The computations were performed by WOMBAT software. In general, the genetic parameter estimates varied across the traits, models and strains studied. Direct additive heritability estimates ranged from 0.0001 (for BW11 of W33) to 0.55 (for BW11 of W33). Maternal and total heritabilities were also variable. Estimates of ratios of direct-maternal effect covariance in phenotypic variance were both positive and negative, but they were negligible, whereas ratios of the permanent environmental maternal variance to phenotypic variance were close to zero. Both of the applied criteria of model adequacy indicate that the model with maternal genetic and environmental effects should be considered as optimal. Genetic trends were close to zero. It seems that they were influenced by long-term selection. Similar tendencies have been observed for phenotypic trends, as well.     

Centric fusion polymorphism in captive animals of family Bovidae

Chromosomes of 228 captive specimens of the family Bovidae have been investigated. The examined animals were classified into the subfamilies Aepycerotinae, Reduncinae, Antilopinae, Alcelaphinae, Hippotraginae and Bovinae. Polymorphism for one fusion was identified in the species: Aepyceros melampus, 2n = 59–60; Redunca fulvorufula, 2n = 56–57; Kobus e. ellipsiprymnus, 2n = 50–52; Kobus e. defassa, 2n = 52–54 and Syncerus c. nanus, 2n = 54–55. This is the first study to reveal fusion 7;29 in Kobus e. defassa and simultaneously the respective polymorphism. Variation in the diploid number of chromosomes is also known in species: Oryx g. dammah and Oryx g. leucoryx but in this study only fusion 1;25 was identified in both karyotyped species. Our study showed that 13% of investigated individuals were polymorphic for the centric fusion and demonstrated the important role of cytogenetic screening in captive animals at zoological gardens.     

Hydroxyl group as a substituent with varying electronic properties: Effect of strength of H-bonding on charge density changes in Ph–OH…F<Superscript>−</Superscript> complexes

Due to gradual and controlled changes of interatomic distances between heavy atoms in OH…F⁻ of C₆H₅OH…F⁻ systems it was possible to study the electronic structure evolution. Computation at B3LYP/6-311+G(d,p) level of theory was performed for this purpose. Changes in charges at atoms and characteristics at bond critical points (BCPs) of the H-bond region and also in distant parts of the systems were investigated by means of natural bond orbitals (NBO) and atoms in molecules (AIM) analyses. It is shown that at the border line between partially covalent and non-covalent H-bonding (Espinosa et al. in J Chem Phys 117:5529, 2002; Grabowski et al. in J Phys Chem B 110:6444, 2006) with the H…F interatomic distance ∼1.8 Ǻ the hydrogen atom has the most positive charge. In addition, the change in the atomic charge values in the interacting region affects the phenyl ring properties. The decrease of the sum of atomic charges as well as of the aromaticity was noticed when the OH….F distance is shortened.     

Identification of the chromosome complement and the spontaneous 1R/1V translocations in allotetraploid <Emphasis Type="Italic">Secale cereale</Emphasis> × <Emphasis Type="Italic">Dasypyrum villosum</Emphasis> hybrids through cytogenetic approaches

Genome modifications that occur at the initial interspecific hybridization event are dynamic and can be consolidated during the process of stabilization in successive generations of allopolyploids. This study identifies the number and chromosomal location of ribosomal DNA (rDNA) sites between Secale cereale, Dasypyrum villosum, and their allotetraploid S. cereale × D. villosum hybrids. For the first time, we show the advantages of FISH to reveal chromosome rearrangements in the tetraploid Secale × Dasypyrum hybrids. Based on the specific hybridization patterns of ribosomal 5S, 35S DNA and rye species-specific pSc200 DNA probes, a set of genotypes with numerous Secale/Dasypyrum translocations of 1R/1V chromosomes were identified in successive generations of allotetraploid S. cereale × D. villosum hybrids. In addition we analyse rye chromosome pairs using FISH with chromosome-specific DNA sequences on S. cereale × D. villosum hybrids.     

Dysmyelinating and demyelinating Charcot–Marie–Tooth disease associated with two myelin protein zero gene mutations

Mutations in the myelin protein zero (MPZ) gene are the third most frequent cause of hereditary motor and sensory neuropathies (HMSN), also called Charcot–Marie–Tooth disorders (CMT). Only in case of recurrent mutations occurring in the MPZ gene is it possible to draw phenotype–genotype correlations essential for establishing the prognosis and outcomes of CMT1. We have surveyed a cohort of 67 Polish patients from CMT families with demyelinating neuropathy for mutations in the MPZ gene. In this study, we report two CMT families in which the Ile135Thr and Pro132Leu mutations have been identified for the MPZ gene. These MPZ gene mutations had not been identified hitherto in the Polish population. The Pro132Leu mutation segregates with a severe early-onset dysmyelinating–hypomyelinating neuropathy, whereas the Ile135Thr substitution is associated with the classical phenotype of CMT1. To the best of our knowledge, we present here, for the first time, morphological data obtained in two sural nerve biopsies pointing to a hypomyelination–dysmyelination process in a family harboring the Pro132Leu mutation in the MPZ gene.     

Detection of Giardia intestinalis assemblages A, B and D in domestic cats from Warsaw, Poland

Giardia intestinalis is a complex species divided into 7 assemblages (A - G). Two of them (A and B) are infective for both humans and animals. In cats four assemblages can occur: A, B, D, and F Assemblages A and B infect either cats, dogs and humans, assemblage D infects cats and dogs and assemblage F only cats. The purpose of this study was to determine the prevalence and genotypes of G. intestinalis in cats from Warsaw. From November 2006 to March 2007 a hundred sixty samples of stool were collected and examined by light microscopy. G. intestinalis cysts were detected in 3.75% of samples. DNA extracted from positive samples was used as template for PCR-RFLP using Giardia specific primers and the amplicons were sequenced. A comparison of the obtained DNA sequences with the Giardia sequences in the GeneBank database revealed assemblage A in 1.25% of the investigated cats, assemblage B in 1.25% and D in 1.25%.     

I – V characteristics and electrical conductivity of synthetic melanin

I – V characteristics for synthetic melanin were investigated at – 10 and 25°C. It was found that the Ohm law is valid for low voltages whereas the Child law holds for higher ones. Furthermore, the steady state electrical conductivity was estimated. The change of activation energy from 0.76 eV for low temperatures to 1.58 eV was noticeable near 0°C.