Characterization of CoCas9 nuclease from Capnocytophaga ochracea.

Cas9 nucleases are widely used for genome editing and engineering. Cas9 enzymes encoded by CRISPR-Cas defence systems of various prokaryotic organisms possess different properties such as target site preferences, size, and DNA cleavage efficiency. Here, we biochemically characterized CoCas9 from Capnocytophaga ochracea, a bacterium that inhabits the oral cavity of humans and contributes to plaque formation on teeth. CoCas9 recognizes a novel 5'-NRRWC-3' PAM and efficiently cleaves DNA in vitro. Functional characterization of CoCas9 opens ways for genetic engineering of C. ochracea using its endogenous CRISPR-Cas system. The novel PAM requirement makes CoCas9 potentially useful in genome editing applications.

[State of corneal nerve fibers in systemic amyloidosis]. / Sostoyanie nervnykh volokon rogovitsy pri sistemnom amiloidoze.

The term systemic amyloidosis unites a group of diseases with a single pathogenetic mechanism involving diffuse deposition of a pathological fibrillar protein (amyloid) in the intercellular space of various organs. Among the systemic forms of amyloidosis, light chain amyloidosis (AL-amyloidosis) occurs most often in clinical practice, while transthyretin amyloidosis (TTR-amyloidosis) is its most common hereditary form. Laser corneal confocal microscopy (CCM) allows for in vivo and non-invasive assessment of the state of corneal nerve fibers (CNF). PURPOSE: To assess the state of CNF in systemic amyloidosis by confocal microscopy data obtained in vivo. MATERIAL AND METHODS: The main study group included 16 patients (6 men and 10 women, mean age 60.5±11.6 years) with morphologically confirmed primary AL-amyloidosis, and 14 patients (5 men and 9 women, mean age 59.4±11.3 years) with genetically and morphologically confirmed hereditary TTR-amyloidosis. The control group included 23 healthy volunteers of the same age range without any neurological pathologies. The state of CNF was assessed by in vivo CCM data recorded on the HRT III system and its consequently processing using authors' self-developed program Liner 1.2. The criteria for neuropathy intensity was the degree of CNF tortuosity characterized by coefficients of anisotropy (KΔL) and symmetry (Ksym) of CNF orientation. RESULTS: According to the NIS scale, the manifestations of neuropathy in the subgroup of patients with TTR-amyloidosis were significantly more pronounced compared to AL-amyloidosis patients. The severity of clinical manifestations of neuropathy did not depend on the duration of TTR-amyloidosis and AL-amyloidosis (Spearman R rs=0.21, p=0.58 and rs= -0.49, p=0.055, respectively). Changes in the quantitative indicators (a decrease in the anisotropy coefficient and an increase in the symmetry coefficient of the fibers orientation) confirm increased tortuosity of CNF in systemic amyloidosis. CONCLUSION: The clinical picture of systemic amyloidosis is characterized by polymorphism of neurological manifestations that include various symptoms of damage to the peripheral somatic and autonomic nervous system. In vivo CCM can be used to reveal qualitative and quantitative changes in CNF in patients with systemic amyloidosis. However, statistical unreliability of the identified quantitative changes allows considering the state of CNF in amyloidosis only as a component of the disease monitoring algorithm, but not as a biomarker of the disease.

[The prospects of using bacteria of the genus Rhodococcus and microbial surfactants for the degradation of oil pollutants].

The possibility of accelerating oil degradation by an enrichment culture of oil-oxidizing microorganisms in the presence of bacteria of the genus Rhodococcus and microbial surfactants was studied. It was shown that the degree of consumption of crude oil (2% v/v) after 192 h of enrichment culture growth reached 84%. Inoculation of the active hydrocarbon-oxidizing strain Rhodococcus erythropolis EK-1 and exogenous surfactants produced by Pseudomonas sp. PS-27 increased this degree to 90% and 93-94%, respectively. On the grounds of these results, efficient methods of purification of the environment from oil pollutants can be developed.

[The surface-active compounds of a Pseudomonas sp. S-27 culture]. / Poverkhnostno-aktivnye soedineniia kul'tury Pseudomonas sp. S-27.

Pseudomonas species S-27 was grown on various substrates. It was established that the Pseudomonas species S-27 strain can produce biosurfactants of ramnolipid nature decreasing the surface and interfacial tension to 29.2 and 0.05 mN/m. respectively, as well as a biopolymer stabilizing the emulsions with hydrocarbons and oils. The biosurfactant and bioemulsifier synthesis is shown to depend of the substrate nature.

[The influence of the Soviet antimalarial preparation dabequin on the hemostatic system and on the anticoagulant effect of heparin]. / Issledovanie vliianiia otechestvennogo protivomaliariinoi preparaty dabekhina na sistemu gemostaza i antikoaguliantnyi éffekt geparina.

In experiments on rabbits and experiments in vitro dabechin was shown to decrease the blood coagulation status and to change the anticoagulant effect of heparin. The latter depends on the route and duration of administration of the antimalarial drug. Injections of heparin in the period of the maximal blood content of dabechin decrease the specific action of the anticoagulant. A prolonged oral administration of dabechin enhances the anticoagulant effect of heparin. During in vitro experiments the ability of dabechin to neutralize heparin with a simultaneous decrease of its specific action was shown.

[The influence of ciprofloxacin and pefloxacin on blood coagulation and the anticoagulant effect of heparin]. / Vliianie tsiprofloksatsina i pefloksatsina na svertyvaemost' krovi i antikoaguliantnyi éffekt geparina.

At a single intravenous administration to rabbits ciprofloxacin (10 and 20 mg/kg) and pefloxacin (40 mg/kg) increased the blood coagulability. Administered in a dose of 20 mg/kg pefloxacin exerted no effect on the parameter. Ciprofloxacin (20 mg/kg) and pefloxacin (40 mg/kg) decreased the anticoagulant effect of heparin (100 U/kg).

[Interaction of quinidine, lidocaine, tricaine and pyromecaine with heparin]. / Issledovanie vzaimodeistviia khinidina, lidokaina, trikaina i piromekaina s geparinom.

In the experiments on rabbits it was shown that quinidine administered intravenously in a dose of 5 mg/kg exerts no effect on the blood coagulability but decreases the degree and duration of the anticoagulant effect of heparin. Lidocaine, trimecaine and pyromecaine at the same dose alter neither the blood coagulability nor the anticoagulant effect of heparin.

[Comparative evaluation of the anticoagulant activity of the sodium and calcium salts of heparin, isolated from swine intestinal mucosa]. / Sravnitel'naia otsenka antikoaguliantnoi aktivnosti natrievoi i kal'tsievoi solei heparina, vydelennogo iz slizistoi obolochki kishechnika svinei.

During in vitro experiments it was shown that at intravenous administration to rabbits and dogs in a dose of 100 IU/kg the anticoagulant effect of mucosal heparin sodium was greater than that of heparin calcium. At subcutaneous administration to rabbits in a dose of 750 IU/kg calcium heparin exhibited a stable and even anticoagulant effect for 12 hours. Mucosal sodium heparin administered subcutaneously caused marked peaks of hypocoagulation and rebound effect. At subcutaneous injection sodium heparin action duration was 10 hours.

[Interaction of the calcium salt of heparin with specific antagonists and ethacizine]. / Izuchenie vzaimodeistviia kal'tsievoi soli geparina so spetsificheskimi antagonistami i étatsizinom.

Protamine sulfate was shown to bind calcium heparinate isolated from the pig intestinal mucosa during in vitro experiments in 0.6:1 ration and during experiments on rabbits and dogs in 1.5:1 ratio. 2,5-ionen neutralizes the anticoagulant effect of calcium heparinate during in vitro and animal experiments in 0.6:1 ratio. Neutralization of calcium heparinate requires by approximately 30% more specific antagonists than neutralization of the same amount of sodium heparinate. Ethacizine was found in experiments on rabbits to decrease the anticoagulant effect of calcium heparinate to the same degree as does sodium heparinate.

[Influence of isoptin on the anticoagulant effect of the sodium and calcium salts of heparin]. / Vliianie izoptina na antikoaguliantnyi éffekt natrievoi i kal'tsievoi solei geparina.

It was found that in buffer medium isoptin at concentrations of 0.025-0.0015 mg/ml failed to interact with heparin calcium. In experiments on rabbits isoptin (0.05-0.1 mg/kg) was shown to exert no effect on the time of whole blood coagulation, the time of recalcification and thrombin time and to produce no changes in the anticoagulant effect of heparin calcium but to increase the effect of heparin sodium.

[Influence of bigumal on the blood coagulation system and on the anticoagulant effect of heparin]. / Vliianie bigumalia na sistemu svertyvaniia krovi i antikoaguliantnyi éffekt geparina.

It was found that in the blood plasma bigumal even at high dose (15 mg/kg) exerted no effect on the parameters of partial thromboplastin and thrombin time but increased the blood plasma tolerance to heparin. Both in buffer medium and blood plasma bigumal effectively decreased the specific effect of heparin. Oral acute (5 mg/kg) and long-term (3 mg/kg daily for 5 days) administration of bigumal to rabbits decreased the time of whole blood coagulation, partial thromboplastin time and reduced the sensitivity of animals to heparin.