Phosphoenolpyruvate Carboxylase from the Crassulacean PlantBryophyllum fedtschenkoiHametet Perrier: ACTIVITY CHANGES AND KINETIC BEHAVIOUR IN CRUDE EXTRACTS

The total activity (capacity) of phosphoenolpyruvate carboxylasein crude extracts of leaves of Bryophyllum fedtschenkoi, measuredon a fresh weight basis, varies considerably with leaf positionand with photoperiodic treatment. No diurnal variation in capacitywas detected under long – day or short – day conditions.The enzyme in freshly prepared crude extracts is about 50 timesmore sensitive to inhibition by malate at pH 7 than that inaged extracts, or in the fully purified state. Desensitizationin extracts proceeds rapidly unless protective measures aretaken, and appears to be irreversible. A pH-dependence studyshows that the effect of desensitization on the kinetic parametersof malate inhibition is identical to that of increasing thepH by 2.0 units over a wide range, but the maximum velocityat pH 7.8 is virtually unaffected. The significance of the resultsfor both the experimental determination of cyclic changes inenzyme capacity, and the theories concerning the mechanism ofa circadian rhythm of phosphoenolpyruvate carboxylase activityin vivo are discussed.     

The effect of pH on host plant 'preference' for strains of Rhizobium trifolii using fluorescent ELIS A for strain identification

The effect of pH on host plant ‘preference’ for strains of R. trifolii was studied using the fluorescent ELISA technique. Four white clover cultivars were compared growing at pH 5, 6 and 7 inoculated with 1:1 mixtures of two strains of R. trifolii, CP3B and R4. The growth of these two bacterial strains was also studied at the same pH levels in pure culture. At pH 5, in pure culture, CP3B grew very well but R4 failed to reach the log phase. CP3B also produced the majority of nodules at this pH (86%). At pH 7, in pure culture, R4 grew better than CP3B and also produced 66% of the nodules in the nodulation experiment. However, there was good evidence of host cultivar ‘preference’ with cv. Milkanova having no nodules inhabited by CP3B at pH 7 but cv. S100 having 32%. The results are discussed from the point of view of the establishment of white clover in acid soils and the usefulness of the fluorescent ELISA technique for Rhizobium strain identification is also emphasised.     

Characterization of 59 canine single nucleotide polymorphisms in the Italian wolf (Canis lupus) population

We characterized 59 canine single nucleotide polymorphisms (SNPs) in the endangered Italian wolf (Canis lupus) population, which were discovered by resequencing sequence‐tagged‐site (STS) DNA sequences that are known to contain SNPs in domestic dogs. Dog SNPs were usually found also in wolves. Additional SNPs unique in dogs or wolves were discovered, which is important for detecting hybrids between dogs and wolves. We developed new primer sets and analysed 15 SNPs by Pyrosequencing. The characterized SNPs will provide an important addition to the genetic markers that are currently available for studying wild populations of canids.     

IDENTIFICATION OF BRITISH BAT SPECIES BY MULTIVARIATE ANALYSIS OF ECHOLOCATION CALL PARAMETERS

ABSTRACT

Previous studies have found variability and individual distinctiveness in the echolocation calls of bats. We consider two implications of individually distinct echolocation calls: 1) whether bats may be able to use such variation to recognise familiar conspecifics, and 2) whether investigators could use such variation to identify known individuals or to census populations. We compared the discriminability of the echolocation calls of big brown bats (Eptesicus fuscus) recorded in three situations: (a) while held in the hand, (b) while perched on a platform, and (c) while flying in an anechoic chamber. Using variables describing each sonar call, we employed discriminant function analysis (DFA) to assign calls to recording situation or to bat. Discrimination of calls by recording situation was largely unsuccessful, although flying calls could be distinguished from platform calls. Assignment of calls to individual bat across recording situations yielded 72% success, and, within a given recording situation, yielded 87% success. Stepwise DFA reduced the number of variables needed to discriminate between individuals with only a slight decrease in correct classification. These results suggest that bats (or researchers) may be able to use the information contained in the echolocation calls for individual recognition. Individual distinctiveness raises the possibility of censusing bats by sound. We used cluster analysis in an attempt to determine whether, given a sample of calls from an unknown number of bats, a reasonable estimate of the number of bats could be obtained. Results were unsatisfactory, suggesting that cluster analysis probably will not permit acoustic censusing of bats in the field.     

Identification of senescent cells in multipotent mesenchymal stromal cell cultures: Current methods and future directions

Regardless of their tissue of origin, multipotent mesenchymal stromal cells (MSCs) are commonly expanded in vitro for several population doublings to achieve a sufficient number of cells for therapy. Prolonged MSC expansion has been shown to result in phenotypical, morphological and gene expression changes in MSCs, which ultimately lead to the state of senescence. The presence of senescent cells in therapeutic MSC batches is undesirable because it reduces their viability, differentiation potential and trophic capabilities. Additionally, senescent cells acquire senescence-activated secretory phenotype, which may not only induce apoptosis in the neighboring host cells following MSC transplantation, but also trigger local inflammatory reactions. This review outlines the current and promising new methodologies for the identification of senescent cells in MSC cultures, with a particular emphasis on non-destructive and label-free methodologies. Technologies allowing identification of individual senescent cells, based on new surface markers, offer potential advantage for targeted senescent cell removal using new-generation senolytic agents, and subsequent production of therapeutic MSC batches fully devoid of senescent cells. Methods or a combination of methods that are non-destructive and label-free, for example, involving cell size and spectroscopic measurements, could be the best way forward because they do not modify the cells of interest, thus maximizing the final output of therapeutic-grade MSC cultures. The further incorporation of machine learning methods has also recently shown promise in facilitating, automating and enhancing the analysis of these measured data.     

Characterization of tetranucleotide microsatellites for Rio Grande cutthroat trout and rainbow trout,and their cross‐amplification in other cutthroat trout subspecies

We describe the isolation and characterization of 12 tetranucleotide microsatellites for Rio Grande cutthroat trout (Oncorhynchus clarkii virginalis) and rainbow trout (Oncorhynchus mykiss), and subsequently investigate their performance in Colorado River cutthroat trout (Oncorhynchus clarkii pleuriticus), greenback cutthroat trout (Oncorhynchus clarkii stomias) and Yellowstone cutthroat trout (Oncorhynchus clarki bouvieri). All 12 loci are polymorphic in all subspecies of O. clarkii examined.