Genomic characterization of the Taylorella genus

The Taylorella genus comprises two species: Taylorella equigenitalis, which causes contagious equine metritis, and Taylorella asinigenitalis, a closely-related species mainly found in donkeys. We herein report on the first genome sequence of T. asinigenitalis, analyzing and comparing it with the recently-sequenced T. equigenitalis genome. The T. asinigenitalis genome contains a single circular chromosome of 1,638,559 bp with a 38.3% GC content and 1,534 coding sequences (CDS). While 212 CDSs were T. asinigenitalis-specific, 1,322 had orthologs in T. equigenitalis. Two hundred and thirty-four T. equigenitalis CDSs had no orthologs in T. asinigenitalis. Analysis of the basic nutrition metabolism of both Taylorella species showed that malate, glutamate and alpha-ketoglutarate may be their main carbon and energy sources. For both species, we identified four different secretion systems and several proteins potentially involved in binding and colonization of host cells, suggesting a strong potential for interaction with their host. T. equigenitalis seems better-equipped than T. asinigenitalis in terms of virulence since we identified numerous proteins potentially involved in pathogenicity, including hemagluttinin-related proteins, a type IV secretion system, TonB-dependent lactoferrin and transferrin receptors, and YadA and Hep_Hag domains containing proteins. This is the first molecular characterization of Taylorella genus members, and the first molecular identification of factors potentially involved in T. asinigenitalis and T. equigenitalis pathogenicity and host colonization. This study facilitates a genetic understanding of growth phenotypes, animal host preference and pathogenic capacity, paving the way for future functional investigations into this largely unknown genus.     

Survival of Haemophilus equigenitalis in different antibiotic-containing semen extenders.

The ability of Haemophilus equigenitalis, the causal agent of contagious equine metritis 1977, to survive in various antibiotic-containing semen extenders was studied at different environmental temperatures. Gentamicin sulphate was found to be markedly superior to ampicillin or a combination of sodium benzyl penicillin and polymyxin B sulphate, Semen treated with the former antibiotic was either sterile at cultural examination or else yielded appreciably fewer colonies of H. equigenitalis than the untreated semen control. Ampicillin had no observable effect on the survival of this organism. Gentamicin was most effective when semen-extender mixtures were held at room temperature rather than at 37 or 4 degrees C. No detrimental effects on sperm motility were observed following the use of the different antibiotic-containing semen extenders in the presence or absence of H. equigenitalis.     

Diagnosis and treatment of four stallions, carriers of the contagious metritis organism--case report

Contagious Equine Metritis (CEM), a venereal disease of horses caused by the bacterium Taylorella equigenitalis, was first diagnosed in 1977 and subsequently spread to many nations [Proc 24th AM Assoc Equine Pract (1979) 287]. The disease was confirmed in the United States in 1978 [Proc Am Assoc Equine Pract (1983) 295]. Specific regulatory procedures for this disease have been established in the United States and 37 other countries. From 1999 through 2001, four of 120 imported European stallions tested positive for CEM at a quarantine facility in Darlington, MD, USA. Two stallions were identified by positive bacterial cultures for T. equigenitalis on arrival. The other two positive stallions were negative on initial bacterial cultures, but were identified as CEM carriers when test mares (that they had mated) were culture-positive for T. equigenitalis. Since T. equigenitalis, is a fastidious slow-growing coccobacillus, additional sets of samples taken over a interval might be required to ensure positive stallions are detected before mating test mares. Likewise, additional sets of samples taken over a long interval after treatment of a stallion for CEM might be required to ensure that positive stallions treated for CEM are detected before mating test mares. Aggressive systemic antibiotic therapy accompanied by routine topical therapy might be required to treat some CEM-positive stallions.     

Epidemiologic aspects of Taylorella equigenitalis

Contagious equine metritis (CEM) is a sexually transmissible disease in mares. Although the disease is commonly diagnosed by culturing the causative bacterium Taylorella equigenitalis (T. equigenitalis) . false negative results do occur. A recently developed Polymerase Chain Reaction (PCR) assay, however, appeared to be much more sensitive, with initial results indicating an unexpected high incidence of the agent in selected horses. In this study, samples from 107 randomly selected mares with no clinical signs of CEM submitted for conventional culture were all negative for T. equigenitalis . but in the PCR-assay 54 (49%) were positive for Taylorella -DNA. Positives in the PCR-assay were found in all breeds tested, even in horses imported from the isolated population in Iceland. These findings suggest that T. equigenitalis was present long before it was first isolated in 1977, The high incidence of Taylorella in horse populations without apparent clinical signs of CEM, the occurrence of incidental clinical case and the known variability between strains, all indicate that Taylorella is endemic in the horse population. In order to explore whether the organism is present in species other than the horse, we also used the PCR-assay on clinically health donkeys (n = 14), zebras (n = 15), Przewalski horses (n = 2) and cows (n = 21). All the animals showed negative results except one of the Przewalski horses, and one cow that was repeatedly found to give positive reaction. We also found that the fertility of 7 stallions with cultures positive for Taylorella (6 used in an AI-program and 1 by natural breeding) was not affected, as shown by the normal range of foaling rates in mares inseminated or bred by these stallions. The overall results may be interpreted to mean that Taylorella is of limited significance in horse breeding.     

Structure of the O-polysaccharide of the lipopolysaccharide produced by Taylorella asinigenitalis type strain (ATCC 700933).

Taylorella asinigenitalis sp. nov is a nonpathogenic gram-negative bacterium recently isolated from the genital tract of male donkeys. The bacterium is phenotypically indistinguishable from Taylorella equigenitalis, a pathogen that is the cause of contagious equine metritis, a highly communicable venereal disease of horses. The structural analysis of the lipopolysaccharide produced by T. asinigenitalis sp. nov (ATCC 700933) demonstrated that its O-polysaccharide (O-PS) component is a linear unbranched polymer of repeating disaccharide units composed of 1,3-linked pyranosyl residues of 2,4-diacetamido-2,4-dideoxy-beta-D-quinovose (bacillosamine) and 2-acetamidino-2-deoxy-beta-D-glucuronic acid, and has the structure [-->3)-beta-D-QuipNAc4NAc-(1-->3)-beta-D-GlcpNAmA-(1-->]n. The chemical structure and serological characteristics of the T. asinigenitalis O-PS are distinct from those of the O-PS of the T. equigenitalis type strain, thus providing a cell-surface target macromolecule that can be used to distinguish pathogenic from nonpathogenic Taylorella sp. clinical isolates.     

The role of international transport of equine semen on disease transmission.

Despite the numerous benefits of having the capability to transport semen internationally, there are serious potential ramifications if that semen is contaminated with a communicable disease. BACTERIA: Many commensal bacteria colonize the exterior of the stallion penis and are not regarded as pathogenic. They may be cultured from an ejaculate. Alterations of the normal bacterial flora on the exterior genitalia may cause the growth of opportunistic bacteria such as Klebsiella pneumonia, Pseudomonas aeruginosa, Streptococcus zooepidemicus, which, if inseminated, may cause infertility in susceptible mares. Contagious equine metritis (CEM), a highly transmissible, true venereal disease of horses, is caused by the gram-negative coccobacillis, Taylorella equigenitalis. Even with the use of rigorous testing protocols, the current techniques used may not ensure accuracy of results. VIRUSES: Equine coital exanthema (equine herpes virus type 3; EHV-3) is a highly contagious virus that causes painful lesions on the stallion's penis and mare's vulva. Although it is primarily transmitted through coitus, infected fomites have also been implicated in its spread. Therefore, it is possible that the virus can potentially be transmitted to the ejaculate through penile contact with an artificial vagina or sleeve. Equine arteritis virus appears to be becoming more prevalent in recent years. The most common method of transmission is through respiratory disease, but the organism can also be shed in the semen of asymptomatic stallions. Equine infectious anemia virus has also been found to be present in the semen of an infected stallion, although no evidence exists at this time that there is venereal transmission of this disease. PROTOZOA: Dourine, caused by Trympanosoma equiperidum, is a venereal disease found only in Africa, South and Central America and the Middle East. Serological testing using complement fixation is recommended for diagnosis. Piroplasmosis, a disease caused by Babesia equi or by a less severe strain, Babesia caballi, has received a great deal of attention in recent years due to the increased transfer of horses between countries. It is considered to be enzootic in many areas of the southern US, and is found throughout the world. The protozoal agent is most often spread by ticks, but mechanical transmission has also been documented; therefore, there is concern for venereal transmission if blood from an infected horse contaminates the semen.     

Shapes of curves of pH-dependence of reactions

A simple case is considered in which the rate of a two-step reaction depends on pH because the intermediate formed in the first step has to gain (or lose) a proton before it can react in the second step, and in which the rate-determining step therefore changes with pH. The curves of reaction rate against pH are shown to be symmetrical, and the sharpest peak possible has a width at half its height of 1.53pH units, i.e. of 2log(3+2 radical2). Any particular curve for this situation proves to be identical with a curve that could be generated for the pH-dependence of a single-step reaction in which the rate is proportional to the concentration of a particular ionic form of a reactant. Curves for the latter situation, however, can have forms impossible for the former case in which the rate-determining step changes, but only if the protonations that activate and deactivate the reactant are co-operative. The peak can then become even sharper, and its width at half its height can fall to 1.14pH units, i.e. to 2log(2+ radical3).     

Comparison of models of insulin release

A model for insulin secretion with a storage and a labile compartment, as well as a provisionary factor, is combined with a signal model in which the signal can be the difference between an excitation and an inhibition, or the difference in concentrations inside and outside some cell components. The model, using a single set of values for the parameters, accounts in a semiquantitative manner for all of the regularly appearing features of the insulin secretion from thein vitro perfused pancreas to a wide range of patterns of glucose and tolbutamide stimulation. Among the features which can be accounted for are: early and late secretion of insulin as a function of glucose in terms of a single parameter; the apparent depletion and recovery during a pulsed pattern of stimulation by tolbutamide; the hypersecretion following a short period of rest during a prolonged stimulation by glucose; the negative spike which occurs when the concentration of glucose, which has been maintained for a period of time, is suddenly reduced to a lower level; and the appropriate responses to slow and fast ramp functions of glucose concentration.     

Findings of the importance of dietetic fiber in the regulation of cholesterolemia

We studied the variations of some lipid indexes in cholesterolemia, lipidemia and triglycerides in relation to a diet rich in bulk. Seven adult male subjects in good health were tested for cholesterol, lipid and triglycerides before and after a two week period of a dialy diet of 40 g of whole wheat bran containing about 10g of fiber. At the some time the lipid, cholesterol and triglycerides limits were measured in 3 groups of rabbits. The first groups was subjected to a normal diet, the second group was subjected to a diet of bleached flour and vitamins, and the third group had a diet similar to the second group but with the addition of 40 g whole wheat bran. The results of our investigation demonstrated a statistically significant decrease of cholesterol lipid and triglycerides levels the special diets at the some time we did not see in increase in the third group of rabbit. Our study seems to indicate that dietetic fibers exercise a vital role in regulating the concentration of plasma lipids even if the subject continues to consume a diet rich in fats. From this we see to importance of the dietetic fibers as a protective factor in the prevention of ateriosclerotic.     

The effect of a complex of meditation exercises on the psychophysiological state of young men

Using a simple motor response, a complex sensorimotor response, and a tapping test, the effect of a complex of meditation exercises on the neuromuscular coordination in healthy male students who were preliminarily trained in methods of self-regulation based on meditation techniques was studied. As a result of a 6-min session of self-regulation, a considerable decrease (by 30% or more) in the time of a simple motor response and a complex sensorimotor response (due to a decrease in the latent time at constant motor time) and a simultaneous decrease in the number of errors, as well as a considerable increase in the result of the tapping test, were recorded. A short-term effect expressed in an increase in the rate and accuracy of sensorimotor responses can be a result of a change in the functional state of the nervous system under the impact of transcendental meditation techniques.     

Pathogenicity of two species of Fusarium on some cultivars of bean in greenhouse

Twenty isolates of Fusarium oxysporum and F. solani were isolated from the infected roots of bean in different farms of east Azarbaijan and Tehran Provinces and their pathogenicity determined. Most isolates of the fungi were identified as F. oxysporun. They caused root rot, yellowing and wilting of bean in the field. In this test, the roots of 6 cultivars of bean seedlings soaked in suspension of the 7 isolates of the fungi (a1, Gogan, a2, Bilverdi, a3, Savojbolagh-Hashtgerd, a4, field of Agr. Coll. a5, Khomein, a6, Ramjin of F. oxysporum and a7 of F. solani of Varamin, Iran) for 5 minute (106 spores/ml.) then transplanted into the sterilized soil in 4 pots (as replication). For control (a8) the roots soaked in distilled water. The results showed that percentage average of necrotic roots and crowns of isolates al, a2, a3, a5, a6, a7 was %20.31 in group a, a4 was %43.52 in group b and a8 was %2.77 in group c after 3 weeks. The isolate a4 (from the field of Agricultural College, Karaj) was more infectious than the other because it caused wilting, yellowing the leaves and decreased the growth very soon, followed by a5 with %25.32 rate was more pathogenic. Bean cultivar Goli-Red was more tolerant with %10.02 than the others of 16.29 (Naz Red) to 25.15 percent of necrotic the roots & stems.     

Analysis of the molecular interaction of the farnesyl moiety of transducin through the use of a photoreactive farnesyl analogue

Farnesylation of the gamma-subunit of the retinal G-protein, transducin (Talpha/Tbetagamma), is indispensable for light-initiated signaling in photoreceptor cells. However, the farnesyl-mediated molecular interactions important for signaling are not well understood. To explore this issue, we created a functional Tbetagamma analogue in which the farnesyl group was replaced with a (3-azidophenoxy)geranyl (POG) group, a novel farnesyl analogue with a distal photoreactive azido group. In the presence of lipid membranes and/or Talpha-GDP, UV irradiation of POG-modified Tbetagamma (POG-Tbetagamma) invariably yielded a cross-linked product Tgamma-Tbeta, reflecting a constitutive interaction of the Tgamma C-terminal lipid with Tbeta. In addition to the Tgamma-Tbeta adduct, a Tgamma-Talpha cross-link was detected in the aqueous fraction. Reconstitution of POG-Tbetagamma with Talpha and light-activated rhodopsin (Rh) in photoreceptor membranes resulted in cross-linking of Tgamma with a glycerophospholipid, indicating molecular interaction of the farnesyl group with cellular membranes. The Tgamma-phospholipid cross-link was observed only in the presence of both Talpha-GDP and Rh, and was abolished by the addition of GTPgammaS or by replacing Rh with opsin. These findings suggest a transient farnesyl-membrane interaction occurs only in a signaling state formed in a transducin-Rh ternary complex. On the other hand, UV irradiation of POG-Tbetagamma in a soluble complex with phosducin, a negative regulator of G-protein, yielded a Tgamma-phosducin adduct in addition to the Tgamma-Tbeta cross-link. These results illustrate that, rather than being a static membrane anchor, the farnesyl moiety plays an active role in the dynamics of protein-protein and protein-membrane interactions at defined steps in the signal transduction process.     

Spatial division of labour of Schoenoplectus americanus

If connected ramets are growing in heterogeneous environments, Division of Labour (DoL) among ramets potentially will result in more efficient sharing of resources and an overall benefit to the plants. As a result of DoL, connected ramets growing in a heterogeneous environment might achieve more biomass than ramets growing in a homogeneous environment. DoL has been demonstrated to occur in a few clonal plant species, although most studies simply focussed on biomass allocation, not on actual resource capturing such as water and nutrient consumption. The model system for our study is one in which two connected ramet groups of Schoenoplectus americanus were placed into contrasting environments. In one treatment, the connected ramets grew in heterogeneous environments and in the other treatment, the connected ramets grew in the same (i.e. homogeneous) environment. We manipulated two variables (light and salinity) in the experiment. We hypothesized that ramets growing in a shaded and fresh water condition in a heterogeneous environment would use more water than ramets growing in a similar condition but in a homogeneous environment. We further hypothesized that ramets growing in a light and saline condition in a heterogeneous environment would assimilate less water than ramets growing in a similar condition but in a homogeneous environment. These hypotheses are based on the assumption that ramets in a heterogeneous environment would translocate water from ramets growing in a shaded and fresh water condition to ramets growing in a light and saline water condition. We also hypothesized that ramets growing in heterogeneous environments achieve larger biomass than ramets in homogeneous environments. Ramets grown in light and saline conditions in heterogeneous environments allocated more biomass to aboveground parts, had taller shoots, larger Specific Green (leaf) Area and consumed less water than ramets grown in similar conditions but in a homogeneous environment. Results confirm the hypothesis that connected ramets in heterogeneous environments are specialised to capture locally abundant resources and share these with connected ramets growing in other habitats. The result of DoL is that the entire connected ramet system benefits and achieves higher biomass.     

Estimation of age and rate of increase of rare variants.

The problem considered is that of estimating the age or rate of increase of a variant on the basis of the present number of replicates observed in a population. In place of previous diffusion equation analyses of age probability distributions, the likelihood for the age is studied on the basis of a discrete branching process model. It is shown that variations inherent in the process of gene evolution in natural populations make it impossible to provide a reliable point estimate of the age of a specified variant, although the likelihood analysis provides a confidence interval which may place useful bounds on the period in which a variant originated. The observed distribution of numbers of several variants may also provide useful information. The problems of estimation are discussed with reference to rare variants arising in American Indian populations.     

Isolation of domains of the plasma membrane of hepatocytes

Several recent studies have demonstrated the ability of techniques based on immunoadsorption to selectively isolate specialized subregions of membranes, termed domains, which are derived from a larger more complex parent membrane like the plasma membrane. The immunoadsorbent is directed against a specific antigen that resides exclusively or predominantly in the membrane domain to be isolated. Thus, a monospecific antibody to the domain-specific antigen is required. In the present study we developed a method employing a modified immunoblotting strategy which could utilize polyspecific antibodies to isolate membrane vesicles derived from a specific membrane domain of the hepatocyte plasma membrane. We also used specific cell surface labeling of the hepatocyte plasma membrane by lactoperoxidase-catalyzed iodination at 4 degrees C and preparation of different sized vesicles by sonication to facilitate isolation of the specific domain. For this study, polyspecific antisera were raised in goats against a membrane fraction, denoted N2u, which is enriched in bile canalicular proteins. This antiserum recognizes, among other antigens, a 110,000 Mr polypeptide previously shown to be localized in the bile canaliculus (J. Cook et al. (1983) J. Cell. Biol. 97, 1823-1833). A monospecific antiserum was raised in rabbits against the rat hepatocyte asialoglycoprotein receptor, a sinusoidal domain-specific set of glycoproteins whose major form has a Mr of 43,000. These antisera were each coupled indirectly to different pieces of nitrocellulose by the immunoblotting protocol and were used to isolate membrane vesicles from a crude extract of liver plasma membrane prepared by sonication. The ratio of iodinated asialoglycoprotein receptor to the 110,000 Mr polypeptide in vesicles isolated by the affinity nitrocellulose immunoadsorbent method indicate a 10- to 15-fold enrichment of sinusoidal-derived vesicles relative to bile canalicular-derived membrane vesicles. These results show that the affinity nitrocellulose immunoadsorbent method can be used to isolate domain-specific vesicles. Further, the affinity immunoadsorbent method described here for the isolation of domains of the plasma membrane is an integrative one allowing isolation of vesicles present in relatively small concentration in crude cell extracts and it requires minimal ultracentrifugation time.     

Importance of preservation of biophysical organization of isolated mitochondria for revealing physiological regulation of their functions

A method has been elaborated that preserves the mitochondrial-reticular network in lymphocytes in composition to the physiological one. Physiologicalby the immobilization of a blood smear on glass and its subsequent incubation in a medium closeresponses of respiration to excitation in the ition of early responses of ions. The recogn organism are well pronounced on these preparat mitochondria to pathogenic agents in the organism is a timely problem of basic and medicinal e- investigations since they play a leading role in the development of pathological states.     

Description of rearrangement of distraction reclaim in the extension of the lower extremities and elimination of their deformity in patients with various etiology of shortening

The data of X-ray and computed tomographic studies of a long tube distraction reclaim were analyzed in 80 patients who had limb shortening and deformity of varying etiology. The analysis has shown that during distraction there is a positive correlation between the increase in the connective-tissue interlayer area of a distraction reclaim and the overall increase in the area of the distraction reclaim (their ratio is not greater than 30% under the optimal elongation conditions). During fixation there is a decrease in the ratio of the area of a growth zone to the total area of a reclaim and in its total density with a parallel increase in the density of cortical laminae of a newly formed bone. Complete organotypic rearrangement comes to the end in the range of 1-3 years depending on the magnitude of pathomorphological and biomechanical changes and limb function.     

Mechanical control of the time-course of contraction of the frog heart

Changes in load during most phases of an isotonic contraction of the frog and turtle heart increased or decreased the duration of the twitch. It was abbreviated by a maintained increase or by a brief decrease in load. The relaxing effect of these procedures developed with a delay lasting more than a second under some conditions and will be called lengthening deactivation. The reverse procedures, a maintained diminution or a brief increase in load, increased the duration of the twitch. This effect will be called shortening activation. Although the termination of relaxation may be delayed or advanced by the mechanical interventions mentioned, the normal time- course of isotonic relaxation was always resumed later, regardless of the starting level of the load, making it possible to measure accurately changes in the duration of the twitch. The responses to changes in load produce positive feedback during the isotonic contraction and explain, at least in part, the difference in the time- course of isotonic and isometric contraction. The effects of changes in load were much smaller and briefer in the atrium than the ventricle.     

Effect of conditioning polarization of soma of giant neurons of mollusks on the mechanism of action-potential generation

To ascertain the properties of an excitable membrane of the soma of giant neurons of mollusks, experiments were carried out to study the effect of conditioning shift of the membrane potential on the mechanism of action-potential generation. The effect of conditioning was assessed from changes in the action-potential curve and its first derivative, as well as from the curve of transmembrane currents under voltage clamp conditions. It was found that a change in membrane potential evokes at least two reactions which have opposite effects on the mechanism of generation of action potentials. These reactions evidently have different time characteristics. One of these does not differ notably from the reaction recorded for other excitable structures, and is manifested in the activation (with hyperpolarization) or inactivation (with depolarization) of the mechanism generating action potentials. The other reaction contributes either to an increase (with depolarization) or a decrease (with hyperpolarization) in the efficiency of this mechanism. Conditioning polarization also has a marked effect on the system responsible for repolarization of the membrane during generation of action potentials. This effect is manifested in a change in the reaction of this system to tetraethylammonium ions. The specific membrane systems sustaining excitability and reacting to changes in the strength of the membrane's electrical field were found to be very inert. After a shift in the potential to a given stable level a rearrangement, lasting sometimes tens of seconds, takes place in the membrane.A. A. Bogomolets Institute of Physiology, Academy of Sciences of the Ukrainian SSR, Kiev. Translated from Neirofiziologiya, Vol. 2, No. 1, pp. 91–99, January–February, 1970.