Long-run monitoring of bacteria,yeasts and other micromycetes in the air of an industrial conurbation

Three different methods were used for the monitoring of airborne microorganisms: (1). Cultivation of microbes trapped in a single-stage biological impactor directly on a solid agar nutrient medium (meat-pepton agar, Sabouraud's agar, blood agar) in Petri dishes. The repeated yearly course of concentrations of cultivable organisms, or colony-forming units (CFU), was obtained by long-run measurements. (2) Aeresol was trapped by impact on membrane filters, and the microorganisms were cultivated by placing the filters on the agar media as above. (3) Direct microorganism counting in a fluorescence microscope; air was sampled in a four-stage impactor where the aerosol was trapped on microscope slides, and the microorganisms were subsequently stained with fluorescent dyes (fluorescein diacctate, 4;6-diamidino-2-phenylindole and, particular, ethidium bromide).

The highest microorganism counts were obtained by using the fluorescence method, the direct cultivation method gave counts an order of magnitude lower, and the method of cultivation on filters gave values approximately 10 times lower than the conventional cultivation.

High variations in the airborne CFU concentrations over the year were observed in Prague. Over the winter season the variations in the amounts of airborne bacteria and other micromycetes as well as the amounts themselves were lower than in the remaining seasons. In the spring and in the summer, the concentrations of yeasts and other micromycetes were highest, whereas in the autumn the concentrations of the microorganisms decreased. Among the bacteria cultivated form the airborne aerosol, the genera Micrococcus, Bacillus, Neisseria and Corynebacterium predominated. The prevailing genera of micromycetes were Penicillium, Aspergillus and Cladosporium.

The concentrations of microorganisms in free air were also affected by the local weather conditions, temperature in particular, the overall air pollution by aerosols was of minor importance in this respect.     

Mosaic structure of the small cryptic plasmid pKST23 from <Emphasis Type="Italic">Escherichia coli</Emphasis>

Small plasmid pKST23 was isolated from sheep ruminal Escherichia coli population. Plasmid sequence was determined to be 2,779 bp in length and was found to have an overall 42 % of GC pairs. However, its sequence can be divided into two regions based on genetic composition and the GC content. It was found that the high GC region spanning approximately from nucleotide 1,300 to 2,750 was identical to a group of small Escherichia coli plasmids and encoded a putative replication protein identical to plasmid pKL1 Rep protein. The part with lower GC pairs seemed to be more specific as it showed no similarity to the GenBank database. Computational analysis revealed four open reading frames, two of which showed considerable homology to replication proteins. PCR primers targeting parts of the two different regions of plasmid pKST23 were used to assess the occurrence of related plasmids within ruminal E. coli population.     

In vitro characteristics of 1-phenyl-3-methyl-4-acylpyrazol-5-ones iron chelators

Iron chelators represent a group of structurally different compounds sharing the ability of iron binding. The group has been evolving in recent years mainly due to novel experimental indications associated with variable requirements for iron chelators. A group of synthetic 1-phenyl-3-methyl-4-acyl-pyrazol-5-ones has been known for many years but data on their potential biological activity are rather limited.In this study, we analysed a series of these compounds for their iron-chelating properties as well as for their effects on iron based Fenton chemistry. For the former ferrozine spectrophotometric method and for the latter HPLC method with salicylic acid were used.All of the tested compounds were very efficient ferric chelators but their ferrous-chelating effects differed according to the acyl substitution. Notwithstanding various ferrous chelation activities, the individual Fe²⁺-affinities were not significantly different through pathophysiologically relevant pH conditions and some of the tested substances were more potent ferrous chelators at pH 4.5 than clinically used standard deferoxamine. Of particular interest is H₂QpyQ /2,6-bis[4(1-phenyl-3-methylpyrazol-5-one)carbonyl]pyridine/ which iron-chelating affinity increased when pH was decreasing. In spite of ferrous chelation differences, most of the tested acylpyrazolones were similarly active powerful inhibitors of Fenton chemistry as deferoxamine.Conclusively, acylpyrazolones are efficient iron chelators and H₂QpyQ may represent a prototype of novel specific chelators designated particularly for chelation at acidic conditions.     

The growth of dermatophytes at 4° C and 37° C; The relation of this character to others

Summary The ability of the generaEpidermophyton, Microsporon andTrichopyton to grow on some media at 4° C and 37° C was studied. It has been shown that specific differences exist among these fungi in the capability or rapidity of the growth at extreme temperatures.There is high positive correlation among perfect state production, isolation from the soil and growth at 4° C (group of characters A) and between pathogenicity and growth at 37° C (group of characters B). Between the groups A and B of characters exists a slighter negative correlation. Some prognosis about the five characters by certain species of dermatophytes may be given.     

Modern analogues from the Southern Urals provide insights into biodiversity change in the early Holocene forests of Central Europe

Aim The diversity changes that occurred in Central European forests during the early Holocene can be better understood using ecological knowledge of modern analogues of these forests, which occur in far Eastern Europe. Here we compare the diversity of vascular plants, bryophytes and snails among different forest types of the Southern Urals to provide insights into the palaeoecology of the vanished Central European forests. Location Southern Ural Mountains, Bashkortostan, Russia. Methods We recorded all species of vascular plants, ground-dwelling bryophytes and land snails, and measured biotic and abiotic variables, in 100-m² plots located in four forest types dominated by different trees: Pinus sylvestris–Larix sibirica, Betula pendula–Populus tremula, Quercus robur and Acer platanoides–Tilia cordata–Ulmus glabra. These types correspond to the chronosequence of forest communities that occurred in Central Europe in the early Holocene. Results The species richness of herb-layer plants was most affected by the canopy-transmitted light. The former three forest types had open canopy and were rich in species, whereas the Acer–Tilia–Ulmus forests were darker and poor in species. The species richness of ground-dwelling bryophytes decreased from coniferous to broad-leaved deciduous forests. In contrast, the highest species richness of snails was in the Acer–Tilia–Ulmus forests owing to the higher calcium content in the litter of these trees. Main conclusions Based on the modern analogue of the Southern Ural forests, we propose the hypothesis that the late-glacial open-canopy pine and larch forests of Central European lowlands were rich in light-demanding species of vascular plants, many of which were also typical of tall-grass steppes or mesic grasslands. They also contained several species of ground-dwelling bryophytes. The spread of birch, aspen and oak in the early Holocene reduced the local species richness of ground-dwelling bryophytes but not of vascular plants. The subsequent spread of elm, lime, maple and ash caused canopy closure, a retreat of the light-demanding herbs and a decline in the local species richness of vascular plants. Besides the increased shading by these tree species, their litter enriched soils in calcium, and enhanced decomposition and nutrient cycling. This supported an increase in the species richness of land snails.     

Direct determination of total soil carbohydrate content

A direct procedure for the determination of total soil carbohydrate content and a classic determination after acid hydrolysis, both employing the phenol-sulphuric acid method, are compared. The direct procedure enables simultaneous determination of mono-, oligo- and polysaccharides without prior hydrolysis. This procedure is reproducible and takes only a short period of time. The correlation between the proposed method and the classic one with hydrolysis was high (r²=0.9843; n=11; p=0.05).     

Carbohydrate levels in rhizomes ofPhragmites australis at an oligotrophic and a eutrophic site: A preliminary study

Seasonal dynamics of total nonstructural carbohydrates (TNC) and of carbohydrate species (starch, sucrose, glucose, fructose) were followed in rhizomes ofPhragmites australis at two sites of the T?eboň Biosphere Reserve (Czech Republic): Branná sand pit and Ro?mberk fishpond-western shore, which were classified (according to plant species composition and phosphorus availability) as oligotrophic and hyper-eutrophic, respectively.Phragmites stands at these sites were expanding and retreating, respectively. Rhizomes were sampled within terrestrial parts of the reed stands (at a water depth of about 10 cm). The levels of total non-structural carbohydrates were determined mainly by levels of starch and sucrose, while glucose and fructose were present at comparatively low levels. The most conspicuous differences between the sites were associated with autumnal and March levels of carbohydrates. In March, i.e. at the beginning of vegetative development, TNC and starch levels were lower at the hyper-eutrophic, as compared with the oligotrophic, site. Starch and TNC levels fell from August to September at the hyper-eutrophic, but not at the oligotrophic, site. At the low water depth investigated, the differences between stands in carbohydrate levels do not seem to be large enough to account solely for their different vigour. It is suggested that the effect of water depth needs to be evaluated in more detail.     

Competitive canalization of PIN-dependent auxin flow from axillary buds controls pea bud outgrowth

Shoot branching is one of the major determinants of plant architecture. Polar auxin transport in stems is necessary for the control of bud outgrowth by a dominant apex. Here, we show that following decapitation in pea (Pisum sativum L.), the axillary buds establish directional auxin export by subcellular polarization of PIN auxin transporters. Apical auxin application on the decapitated stem prevents this PIN polarization and canalization of laterally applied auxin. These results support a model in which the apical and lateral auxin sources compete for primary channels of auxin transport in the stem to control the outgrowth of axillary buds.     

Excretome of the chitinolytic bacterium <Emphasis Type="Italic">Clostridium paraputrificum</Emphasis> J4

A strictly anaerobic mesophilic chitinolytic bacterial strain identified as Clostridium paraputrificum J4 was isolated from human feces. In response to various types of growth substrates, the bacterium produced an array of chitinolytic enzymes representing significant components of the J4 strain secretome. The excreted active proteins were characterized by estimating the enzymatic activities of endochitinase, exochitinase, and N-acetylglucosaminidase induced by cultivation in medium M-10 with colloidal chitin. The enzyme activities produced by J4 strain cultivated in medium M-10 with glucose were significantly lower. The spectrum of extracellularly excreted proteins was separated by SDS-PAGE. The chitinase variability was confirmed on zymograms of renatured SDS-PAGE. The enzymes were visualized under ultraviolet light by using 4-methylumbelliferyl derivatives of N-acetyl-β-d-glucosaminide, N,N′-diacetyl-β-d-chitobiose, or N,N′,N˝-triacetyl-β-d-chitotriose for β-N-acetylglucosaminidase, chitobiosidase, or endochitinase activities, respectively. Protein components of the secretome were separated by 2D-PAGE analysis. The distinct protein bands were excised, isolated, and subsequently characterized by using MALDI-TOF/TOF tandem mass spectrometry. The final identification was performed according to sequence homology by database searching.     

Synthetic substrates in the histochemical demonstration of intestinal disaccharidases

Summary The splitting of 6-Br-2-naphthyl-, -naphthyl-, and 4-Cl-5-Br-3-indolyl-glycosides which proved useful for the assessment of cytological localization of intestinal enzymes in previous studies was investigated using isolated human and rat intestinal disaccharidases as a source of enzyme activities.Previous findings based on histochemical studies were confirmed and extended. 6-Br-2naphthyl-D-glucoside is cleaved by glucoamylase and sucrase-isomaltase. The participatio of trehalase in splitting of this substrate is very low and can be neglected. The mentioned -glucosidases are responsible for the brush border staining of enterocytes with this substrate when unfixed cold microtome sections are used. Even when a differential heat inactivation of sucrase-isomaltase and of glucoamylase occurs during paraffin embedding (so that the staining in paraffin sections is due mostly to glucoamylase) the use of natural substrates is desirable for a more precise assessment of sucrase-isomaltase activity (but without the possibility of a correct localization).4-Cl-5-Br-3-indolyl--D-fucoside is the substrate of choice for the demonstration of lactase. Even when this substrate is split also by hetero--galactosidase and by acid (lysosomal) -galactosidase these activities do not disturb the histochemical demonstration of lactase. If however some doubts arise, the inhibition with p-Cl-mercuribenzoate (2 · 10^–4 M) is to be emloyed (lactase activity is not inhibited). Due to a low K_m and a high V_max of indolyl-fucoside and due to its extreme stability in solution (which enables to use the substrate solution repeatidly) this substrate is suitable in routine practice even though it is expensive. -naphthyl- and 4-Cl-5-Br-3-indolyl--D-glucosides are split by lactase and -glucosidase. Due to the fact that the mutual delineation of these activities is not easy and that K_m an V_max for lactase are not so favourable as in the case of fucoside these substrates are not recommended for the assessment of lactase.6-Br-2-naphthyl--D-glucoside is the substrate of choice for the histochemical studies concerned with hetero--galactosidase and 4-Cl-5-Br-3-indolyl--D-galactoside for acid -galactosidase.