Genetic diversity of Listeria monocytogenes strains from a high-prevalence dairy farm

Listeria monocytogenes is a significant food-borne human and veterinary pathogen. Contaminated silage commonly leads to disease in livestock, but the pervasive nature of the bacterium can make it difficult to identify the source of infection. An investigation of bovine listeriosis that occurred on a Pacific Northwest dairy farm ("farm A") revealed that the clinical strain was closely related to fecal strains from asymptomatic cows, and that farm environment was heavily contaminated with a diversity of L. monocytogenes strains. In addition, the farm A clinical strain was closely related to clinical and environmental strains obtained 1 year prior from a second Northwest dairy farm ("farm B"). To investigate the source(s) of contamination on farm A, environmental samples were collected from farm A at two time points. Pulsed-field gel electrophoresis characterization of 538 isolates obtained from that farm identified 57 different AscI pulsovars. Fecal isolates obtained from individual cows were the most genetically diverse, with up to 94% of fecal samples containing more than one pulsovar. The maximum numbers of pulsovars and serotypes isolated from a fecal sample of one cow were 6 and 4, respectively. Serotype 1/2a was isolated most frequently at both time points. Microarray genotyping of bovine listeriosis, fecal, and silage strains from both farms identified four probes that differentiated listeriosis strains from environmental strains; however, no probe was common to both bovine listeriosis strains.     

A MINIATURIZED ENRICHMENT SEROLOGY METHOD FOR RAPID DETECTION of SALMONELLA FROM POULTRY MEAT and REARING FARMS ENVIRONMENT

A miniaturization of the enrichment serology method for the detection of Salmonella was improved in order to make the technique more reliable, cheaper, and faster. the miniaturized method ("Micromethod") was compared to the Sperber and Deibel's method ("Macromethod") and with a classical isolation method; 1062 samples including 700 rearing farms environment samples, 247 poultry meat samples, and 115 nonfat dry milk samples were analyzed. Specificity of both enrichment serology methods was about 92–99.4%. Sensitivity of Micromethod was better than that of the Macromethod for the environmental samples (86.8 and 74.1%, respectively) and the poultry meat samples (87.5 and 77.5%, respectively) but was the same for the nonfat dry milk samples (82.5%). the costs of both methods were respectively 0.43 US $ for the Macromethod and 0.20 US $ for the Micromethod. This "Micromethod" could be proposed for the screening of Salmonella positive batches in the food industries.     

Cellular receptors for lymphotoxin: correlation of binding and cytotoxicity in sensitive and resistant target cells.

The binding of radioactively labeled lymphotoxin (LT) to both lymphotoxin-sensitive and -resistant cell clones was examined. The sensitive clone had a low- capacity, high-affinity ("specific") binding component, the curve of which closely followed the cytotoxicity curve of the lymphocyte mediator. The capacity of this binding component was calculated to be about 600 molecules of LT/cell. In addition, there was a low-affinity, high-capacity ("nonspecific") binding component. In striking contrast, the high-affinity, low-capacity ("specific") component was absent or greatly diminished from the resistant clone, whereas the low-affinity, high-capacity ("nonspecific") component was present at a similar level as in the sensitive cells.These binding characteristics closely resemble those observed by us and other investigators working with a variety of steroid hormones in steroid-sensitive and- resistant cell lines.     

High-performance liquid chromatography analysis of naturally occurring D-amino acids in sake

We measured all of the D- and L-amino acids in 141 bottles of sakes using HPLC. We used two precolumn derivatization methods of amino acid enantiomer detection with o-phthalaldehyde and N-acetyl-L-cysteine, as well as (+)-1-(9-fluorenyl)ethyl chloroformate/1-aminoadamantane and one postcolumn derivatization method with o-phthalaldehyde and N-acetyl-L-cysteine. We found that the sakes contained the D-amino acids forms of Ala, Asn, Asp, Arg, Glu, Gln, His, Ile, Leu, Lys, Ser, Tyr, Val, Phe, and Pro. We were not able to detect D-Met, D-Thr D-Trp in any of the sakes analyzed. The most abundant D-Ala, D-Asp, and D-Glu ranged from 66.9 to 524.3 μM corresponding to relative 34.4, 12.0, and 14.6% D-enantiomer. The basic parameters that generally determine the taste of sake such as the sake meter value (SMV; "Nihonshudo"), acidity ("Sando"), amino acid value ("Aminosando"), alcohol content by volume, and rice species of raw material show no significant relationship to the D-amino acid content of sake. The brewing water ("Shikomimizu") and brewing process had effects on the D-amino acid content of the sakes: the D-amino acid contents of the sakes brewed with deep-sea water "Kaiyoushinosousui", "Kimoto yeast starter", "Yamahaimoto", and the long aging process "Choukijukusei" are high compared with those of other sakes analyzed. Additionally, the D-amino acid content of sakes that were brewed with the adenine auxotroph of sake yeast ("Sekishoku seishu kobo", Saccharomyces cerevisiae) without pasteurization ("Hiire") increased after storage at 25 °C for three months.     

A study of water autodiffusion in model biological membranes, oriented lipid bilayers

The autodiffusion of water in a multibilayer structure formed by dipalmitoyl phosphatidylcholine and oriented on glass plates was studied by the method of NMR with magnetic field pulse gradient. It was shown that water molecules occur in several states differing in the degree of interaction with lipid molecules. A spectrum of the coefficients of water autodiffusion in a direction transversal to bilayers was found. The use of samples with different distances between the plates and an analysis of the dependence of the mode of diffuse decay of spin echo on diffusion time and the orientation of the sample, as well as measurements at temperatures above and below the gel-liquid crystal phase transition in cholesterol-containing samples enabled one to discriminate the diffuse decay component responsible for the transbilayer movement of water. The coefficient of bilayer permeability was estimated using the Tanner model. It was shown that the formation of mechanical defects ("cracks") in plane oriented bilayers is the most probable reason for the presence of the water component with the relatively high coefficient of diffusion.     

Resource-based models of competitive interactions. I. Intraspecific competition in Ratibida columnifera ( Asteraceae)

We constructed and tested a series of regression models of intraspecific competition in Ratibida columnifera (Asteraceae), based on the growth and water use of individual plants. Models were constructed from a set of plants ("model") grown without competition under three watering regimes. Each model was then tested on another set of plants ("test") grown, singly or in pairs, under two watering regimes, one of them different than those of the "model" plants. Both sets of plants were grown simultaneously. Models that used only estimates of plant dry mass (ISON [interval by interval size only], and SON [size only]) were outperformed (i.e., the difference between predicted values of final dry mass and their true values were larger on average) by models that incorporated direct measures of water uptake and usage (ISAW [interval by interval size and water], WON [water only], SAW [size and water]). Harvest biomass predictions given by these three last models deviated from values of true biomass by an average of only 6.1%.     

Activation of ryanodine receptors by flash photolysis of caged Ca2+.

Flash photolysis of DM-nitrophen generates an extremely large [Ca2+] transient ("Ca2+ spike") at the start of each Ca2+ "step." The Ca2+ spike greatly increases the speed of activation of the ryanodine receptor channel ("supercharging") and could be responsible for apparent channel adaptation.     

Field-scale transplantation experiment to investigate structures of soil bacterial communities at pioneering sites

Studies on the effect of environmental conditions on plants and microorganisms are a central issue in ecology, and they require an adequate experimental setup. A strategy often applied in geobotanical studies is based on the reciprocal transplantation of plant species at different sites. We adopted a similar approach as a field-based tool to investigate the relationships of soil bacterial communities with the environment. Soil samples from two different (calcareous and siliceous) unvegetated glacier forefields were reciprocally transplanted and incubated for 15 months between 2009 and 2010. Controls containing local soils were included. The sites were characterized over time in terms of geographical (bedrock, exposition, sunlight, temperature, and precipitation) and physicochemical (texture, water content, soluble and nutrients) features. The incubating local ("home") and transplanted ("away") soils were monitored for changes in extractable nutrients and in the bacterial community structure, defined through terminal restriction fragment length polymorphism (T-RFLP) of the 16S rRNA gene. Concentrations of soluble ions in most samples were more significantly affected by seasons than by the transplantation. For example, NO(3)(-) showed a seasonal pattern, increasing from 1 to 3 μg NO(3)(-) (g soil dry weight)(-1) after the melting of snow but decreasing to <1 μg NO(3)(-) (g soil dry weight)(-1) in autumn. Seasons, and in particular strong precipitation events occurring in the summer of 2010 (200 to 300 mm of rain monthly), were also related to changes of bacterial community structures. Our results show the suitability of this approach to compare responses of bacterial communities to different environmental conditions directly in the field.     

Costs of parturition and rearing in female sika deer (Cervus nippon)

The costs of parturition and lactation of female sika deer on Kinkazan Island (9.6 km(2) in size), northern Japan, which live at a high density (about 50 deer/km(2)), were evaluated by comparison of body weights of 481 females measured during a 15 year study (1993-2008). Weight data were chosen from only females that did not give birth in the preceding year. The mean body weight of females that did not give birth ("yelds") was significantly lower (P < 0.001) than that of females who gave birth ("milks"); yelds' body weight was 93.1% and 83.5% that of milks in the preceding and parturition years, respectively. The yelds increased in body weight by the following March by 8.2% (P < 0.001), whereas milks did not. Among the milks, those whose fawns survived until the following May ("rearing milks") lost body weight by 14.9% (P < 0.001). Milks who lost fawns within a week after birth ("early fawn-less milks") did not lose body weight (P = 0.583), while those whose fawns died after the first autumn but died before May ("late fawn-less milks") lost body weight by 19.9% (P < 0.001). These results indicate that sika deer females do not enter estrus unless they are heavy enough, and that both parturition and rearing are costly for sika deer mothers living in high-density conditions.     

Dissociable influences of auditory object vs. spatial attention on visual system oscillatory activity

Given that both auditory and visual systems have anatomically separate object identification ("what") and spatial ("where") pathways, it is of interest whether attention-driven cross-sensory modulations occur separately within these feature domains. Here, we investigated how auditory "what" vs. "where" attention tasks modulate activity in visual pathways using cortically constrained source estimates of magnetoencephalograpic (MEG) oscillatory activity. In the absence of visual stimuli or tasks, subjects were presented with a sequence of auditory-stimulus pairs and instructed to selectively attend to phonetic ("what") vs. spatial ("where") aspects of these sounds, or to listen passively. To investigate sustained modulatory effects, oscillatory power was estimated from time periods between sound-pair presentations. In comparison to attention to sound locations, phonetic auditory attention was associated with stronger alpha (7-13 Hz) power in several visual areas (primary visual cortex; lingual, fusiform, and inferior temporal gyri, lateral occipital cortex), as well as in higher-order visual/multisensory areas including lateral/medial parietal and retrosplenial cortices. Region-of-interest (ROI) analyses of dynamic changes, from which the sustained effects had been removed, suggested further power increases during Attend Phoneme vs. Location centered at the alpha range 400-600 ms after the onset of second sound of each stimulus pair. These results suggest distinct modulations of visual system oscillatory activity during auditory attention to sound object identity ("what") vs. sound location ("where"). The alpha modulations could be interpreted to reflect enhanced crossmodal inhibition of feature-specific visual pathways and adjacent audiovisual association areas during "what" vs. "where" auditory attention.     

Structure of peptostreptococcal protein L and identification of a repeated immunoglobulin light chain-binding domain.

The gene for protein L, an immunoglobulin (Ig) light chain-binding protein expressed by some strains of the anaerobic bacterial species Peptostreptococcus magnus, was cloned and sequenced. The gene translates into a protein of 719 amino acid residues. Following a signal sequence of 18 amino acids and a NH2-terminal region ("A") of 79 residues, the molecule contains five homologous "B" repeats of 72-76 amino acids each. Further, toward the COOH terminus, two additional repeats ("C") were found. These are not related to the "B" repeats, but are highly homologous to each other. After the C repeats (52 amino acids each), a hydrophilic, proline-rich putative cell wall-spanning region ("W") was found, followed at the COOH-terminal end by a hydrophobic membrane anchor ("M"). Fragments of the gene were expressed, and the corresponding peptides were analyzed for Ig-binding activity. The B repeats were found to be responsible for the interaction with Ig light chains. An Escherichia coli high level expression system was adapted for the production of large amounts of two Ig-binding protein L fragments comprising one and four B repeats, respectively.     

Hydrogen ion dependent reversible fibrillarization and hydration of chromatin in divalent cation-depleted isolated rat liver nuclei

Rat liver cell nuclei isolated in the presence of Ca++ and Mg++ ions were exposed to citrate buffer at different pH levels. Their morphology was studied with the electron microscope and the water content measured. At a low pH the chromatin remained compact, but at neutral pH an intense fibrillarization of the chromatin was observed; and, seemed to be reversible, because of deoxyribonucleoproteins (DNP) filaments which dispersed at a pH of 7 were repacked to their previous site at a pH of 3. The ribonucleo-proteins (RNP)-containing structures did not show the same reversible fibrillarization. In addition to the morphological changes, a large increase in the water content ("water-holding capacity") of the chromatin structures was observed.     

The yields of the processes of DNA radiation-chemical destruction in the chromatin

The methods of gel electrophoresis and spectrophotometry were used to study DNA destruction in the chromatin (a nucleosome set of 1 to 8 units) irradiated with 60Co-gamma quanta in water solutions at doses ranging from 1 to 40 kGy, depending on the chromatin concentration, the nature of gas that saturated the solution (Ar, N2O), and the protein/DNA ratio in the chromatin samples. In conditions of a complete involvement of water radiolysis products, OH and eaq, in the reaction with the chromatin (the chromatin concentrations in the solution exceeding 0.04 weight % when calculated per DNA) determined were G ("DNA-protein") cross-links = 6.10(-5), the G (chromophoric group degradation) = 0.4, and G (double-strand breaks) less than 10(-5), as well as G of the process of free DNA release into the solution. The latter depend upon the protein/DNA ratio in the sample and increased with increasing protein content of the chromatin with the protein/DNA ratio being greater than 1.1/1.     

Chronogenetic control or the continuity of morphological organization during postnatal ontogenesis of man

Chronological peculiarities of genetic regulation of growth processes in morphological structures of postcranial skeleton ("body") and cranium ("head") were studied at different phases of peripubertal stage of boys ontogenesis. The component partitioning of the phenotype variance of individual scores of the structures considered revealed relative autonomy of chronogenetic systems controlling morphogenesis in the "body" and the "head". Succession of morphological organization is ensured by the stable level of correlation between morphological structures analysed. Raise in correlation is registered at the "critical" phase (13-14 years) which is connected with the "switching on" genetic programme of growth processes control in the adolescent period of human postnatal ontogenesis.     

Different Tridacnins in different Tridacnid clams: a comparative study.

1. Haemolymph samples from different Tridacnid clams were tested for haemagglutinin activity, using normal and enzyme-treated red cells from different sources. 2. Precipitin reactions with these haemolymph fluids were performed in agar gel, using several defined glycosubstances of different origin. 3. In several experiments it could be demonstrated that in all Tridacnid haemolymph samples anti-galactosyl ("anti-galactan") specific lectins do occur, however, they definitely differ from each other.     

Proximal femur articulation in Pliocene hominids

The supposed "nonhuman anthropoid"-type femur head articular surface described for the Pliocene hominid specimen A.L.288-1 ("Lucy") by Stern and Susman in 1983 is present in significant numbers of modern human femora. This nonmetric skeletal trait was also found to be sex-related in modern human samples examined.     

Morphological aspects of lymphocyte-macrophage interactions in human tumoral effusions "in vivo" and "in vitro"

"In vivo" and "in vitro" morphological analysis of associations of cells ("rosettes") involved in immune response in human tumoral effusions revealed the existence of cell interactions either by simple membrane apposition between the cell projections or by gap-like junctions between two adjacent cells; endocytotic phenomena were also observed. The giant fibroblastic cells seen "in vitro" ("myofibronoblasts") reacting positively to anti-human macrophage Mabs, might be the cells presenting antigen to lymphocytes.     

Lipid changes during natural aging of soybean seeds

When soybean seeds [ Glycine max (L.) Merr. cv. Wayne] were stored at approximately 4°C and low humidity for 44 months ("natural aging") there was a marked decline in vigor and viability which was associated with a decrease in the proportion of the polyunsaturated fatty acids. Other lipid parameters showed little change. Seeds subjected to high humidity at 40°C for several days ("accelerated aging") experience a comparable decline in vigor and viability, but without the change in fatty acid polyunsaturation. These results indicate that accelerated aging may cause loss of vigor in a manner quite different from natural aging. The accelerated aging treatment is therefore of limited usefulness in investigations of the mechanism of natural seed aging.     

Direct measurement of actin polymerization rate constants by electron microscopy of actin filaments nucleated by isolated microvillus cores

We used actin filament bundles isolated from intestinal brush-border microvilli to nucleate the polymerization of pure muscle actin monomers into filaments. Growth rates were determined by electron microscopy by measuring the change in the length of the filaments as a function of time. The linear dependence of the growth rates on the actin monomer concentration provided the rate constants for monomer association and dissociation at the two ends of the growing filament. The rapidly growing ("barbed") end has higher association and dissociation rate constants than the slowly growing ("pointed") end. The values of these rate constants differ in 20 mM KCl compared with 75 mM KCl, 5 mM MgSO4. 2 microM cytochalasin B blocks growth entirely at the barbed end, apparently by reducing both association and dissociation rate constants to near zero, but inhibits growth at the pointed end to only a small extent.     

Selection for nonbuoyant morphological mutants of Caulobacter crescentus.

Morphological mutants of Caulobacter crescentus were isolated by selecting for cells that did not possess normal, buoyancy-conferring stalks. The most prevalent phenotype enriched by the selection was structurally deficient stalks (designated "Cds"), observable as crumpled or flattened stalks. The second phenotype ("Abs") was observed as spontaneous shedding of normal-appearing stalks. Third, one mutant ("Ecs") was isolated that sheds not only stalks, but also miniature stalked and nonstalked cells.