Expression of coagulin A with low cytotoxic activity by Pediococcus pentosaceus ST65ACC isolated from raw milk cheese.

AIMS: We aimed to evaluate some specific conditions for growth of Pediococcus pentosaceus ST65ACC and its bacteriocin expression through ABC transporters; to purify the bacteriocin and determine its sequence; and to evaluate the cytotoxicity potential of the purified bacteriocin(s). METHODS AND RESULTS: The results presented for growth behaviour of P. pentosaceus ST65ACC showed that the bacterial growth was slightly influenced when cultured in MRS broth with different amounts of inoculum: 1, 2, 5 and 10%. The bacteriocin activity increased when 5 and 10% inocula were used. The carbon source (glucose) used in different amounts (1, 2, 3 or 4%) had no significant effect on growth and bacteriocin production. The studied strain P. pentosaceus ST65ACC was able to metabolize xylooligosaccharide (XOS) as the sole carbon source, resulting in the production of an antimicrobial peptide. The genes involved in the ABC transport system and sugar metabolism of P. pentosaceus ST65ACC were expressed at different levels. The bacteriocin produced by P. pentosaceus ST65ACC was partially purified by precipitation with ammonium sulphate (40% saturation), followed by reversed-phase liquid chromatography, resulting in the identification of an active bacteriocin. Tandem mass spectrometry was used to identify the partial sequence KYYGNGVTCGKHSCSVDWGK sharing high similarity to coagulin A. The semi-purified bacteriocin had low cytotoxicity based on estimated values for maximal nontoxic concentration (MNC) and cytotoxicity concentration (CC50 ). CONCLUSIONS: The bacteriocin produced by P. pentosaceus ST65ACC is similar to coagulin, with low cytotoxicity, strong antimicrobial activity and possible additional metabolite routes in the producer cell. In addition to MRS broth, bacteriocin was produced also in medium containing XOS (as the single carbon source). SIGNIFICANCE AND IMPACT OF THE STUDY: To the best of our knowledge, this is the first report of evaluation of the role of ABC transporters in the expression of bacteriocin by P. pentosaceus, cultured in MRS and XOS.

Clinical Case of Parvovirus B19 Infection in Pregnant Woman with Β-Thalassemia in Bulgaria.

A pregnant 30-year-old female in the 34th gestational week was admitted at University "Maichin Dom" Hospital prior to childbirth. The patient is diagnosed with ß-thalassemia. During laboratory screening hemoglobin of 98 g/L was established. Blood smear shows mild microcytic hypochromic anemia: RBC 5.15 x 1012/L, HGB 98 g/L, MCV 65.8 fL, MCH 19.4 pg, MCHC 295 g/L. Serum iron concentration is 12.9 µmol/L and ferritin 17.5 µg/L. For the delivery process cesium was considered. Two days after procedure a rash presented on face, hands and breasts. Although the mother was positive for parvovirus B19 infection, the baby was negative. This was confirmed by se-rological and molecular investigations. We discovered only the mother's B19V IgG antibodies in the newborn. In connection to the main disease, namely ß-thalassemia, acute virus infection could cause aplastic crisis. After consultation with a hematologist, serum hepcidin concentration (an iron homeostasis regulator) was quantified: 19.4 µg/L. ELISA test was used to prove B19V IgM antibodies in the mother. PCR analysis shows the presence of B19V DNA. During infection, inflammatory cytokines increase hepcidin secretion, leading to iron deposition into cells.

[The presumption of mental health: is it really proved?] / Prezumptsiia psikhicheskogo zdorov'ia: mozhno li schitat' ee obosnovannoi?

The article is devoted to one of the most complicated and peculiar problems of modern psychiatry - the presumption of mental health. Its originality is seen in the fact that practically all speaking and writing about it noted the importance of this principle for the protection of rights and legal interests of citizens, failing into the field of psychiatry. However, expressing this opinion, doctors and lawyers offer very few arguments in support of this view. Traditional arguments in favor of the presumption of mental health, scattered and unsystematic, prove to be absolutely insolvent. It was also found out that the similarity of the presumption of mental health with the presumption of innocence is capable to entail negative consequences. The noted similarity strengthens stigmatizing effect of psychiatric diagnosis, making it extremely undesirable for the patient, and the basis of the relationship between the patient and the doctor laid counterproductive antagonistic model as the model of mutual relations of the accused with the prosecutor. The main conclusion of authors is that the supporters of the presumption of mental health have yet to prove its right to exist, to work out its content and try to neutralize some of the negative consequences of its possible implementation in practice. Without compliance with the listed conditions to bring the presumption of mental health to the level of one of the fundamental principles of Russian psychiatry would be premature.

Effect of Complexes of Zinc, Cobalt and Copper With D-Aminosugars on the Replication of Herpes Simplex Virus Type 1 (HSV-1).

Our previous results show that Zn(pic)(2) and Zn(asp)(2) inhibit key steps of the replication of HSV-1. Anti-HSV effect of complexes of Co(II) with aminoacids Lys and Ser was also found. In the present study we describe the effect of complexes of Zn(II), Co(II) and Cu(II) with D-aminosugars on the replication of HSV-1 and on the infectivity of free virions. The experiments were done using primary rabbit kidney cells (r.k.), diploid human embryonal fibroblasts (F) and Vero cells. No differences in the toxicity of metal complexes on diploid cells- r.k. and F, were found. Neither metal complexes, nor ligands-galactosoxime and glucosoxime, influenced the viral replication. During 1-4h prolonged contact only Cu(Gl.NOH)(2) inactivated HSV-1 virions up to 90%. The results show that D-aminosugars are not suitable ligands for Zn(II), Cu(II) and Co(II) in respect of the inhibition of viral replication. However, only Cu(Gl.NOH)(2) was able to inhibit the infectivity of free virions.

Bacteriostatic and biological stimulation effect of Mepitel on experimental burns on the skin of rats.

This study was conducted on 200 white Wistar rats weighing about 200 g each. The animals were divided into two equal groups--an experimental one, with Mepitel (SCA Mölnlycke) dressings, and a control group with cotton gauze dressings. Microbiological and histological examinations and measurements of the wounds made on the 3rd, 7th, 14th, 21st and 28th days after the 3rd-degree heat burn was inflicted using the standard method. The observations included burns with the spontaneous elimination of necroses and healing; wounds after operative total necrectomy of the burnt skin and spontaneous healing; and burns followed by total surgical necrectomy and grafting with allotransplants. The quantitative results reveal statistically reliable bacteriostatic activity above and under the Mepitel dressing. The histological examination reveals considerable biological activity in the tissues under the Mepitel. This manifests itself as an acceleration in the healing process in the wounds caused by burns and after necrectomy of the burnt skin. Mepitel on the allotransplant on the wound, which was left following the necrectomy of the burnt skin provokes the rapid and early rejection of the allograft. Mepitel's biological activity has also been confirmed by the statistically-reliable data relating to the dynamic changes in the length and breadth of the wounds. The authors came to the conclusion that, when applied immediately to a burn or a wound left following the necrectomy of skin burns, Mepitel suppresses the development of the microbiological flora and stimulates the normal healing process. Mepitel is not suitable for dressing allotransplants because of the rapid acceleration of their rejection.

Complexes of zinc with picolinic and aspartic acids inactivate free varicella-zoster virions.

Zn(II) picolinate and aspartate, Zn(pic)(2) and Zn(asp)(2), have been shown to inhibit key steps of the replication of HSV-1. In the present study we describe the effect of Zn(pic)(2) and Zn(asp)(2) on the replication of VZV and on the infectivity of free virions. The experiments are done using BHK-21 cells, a clinical isolate of VZV and Zn-complexes in concentration of 10 muM. When Zn-complexes are present during the whole period of infection, the yield of infectious virus progeny decreases up to 98%. The infectivity of VZV is completely restored after the removal of zinc. The virucidal effect is manifested at the 2nd h of contact, when 90% of the virions are inactivated. The results show that both Zn(pic)(2) and Zn(asp)(2) specifically inactivate free VZV virions with no effect on viral replication.

Effect of Complexes of Cobalt With Aminoacids on the Replication of Herpes Simplex Virus Type 1 (HSV-1).

Cobalt, being essential metal, influences different physiological and enzymatic functions. As cobalt does not accumulate in the body, Co-compounds have relatively low toxicity. The aim of the present study is the effect of complexes of Co(II) with aminoacids - lysine, arginine, histidine and serine on HSV-1 replication. No effect of [O(2)Co(his)(4)].nH(2)O and [O(2)Co(arg)(2)].nH(2)O on HSV-1 infection in vitro was found. Both, [O(2)Co(lys)(2)].nH(2)O and [O(2)Co(ser)(2)].nH(2)O suppress the attachement of HSV-1 particles onto target cells and the viral replication as well. Moreover, the properties of the particular Co-complex (charge, stability, structure) are manifestated by their virucidal effect. Thus, [O(2)Co(ser)(2)].nH(2)O irreversibly inhibits the infectious activity of free HSV-1 virions, while virucidal effect of [O(2)Co(lys)(2)].nH(2)O is completely reversible after the 2h of contact.

Mode of action of Zn-complexes on herpes simplex virus type 1 infection in vitro.

The influence of Zn-complexes with biologically active ligands like aminoacids: picolinic acid as Zn(pic)2 and asparaginic acid as Zn(asp)2 on HSV-1 infection in vitro was investigated. Studies on kinetics of Zn-ions using labeled 65Zn(pic)2 as a marker, show that at the 45th min Zn-ions are exposed onto or into cells. When cells were infected 60min after the treatment with 65Zn(pic)2, 1h later the activity of Zn-ions in the cell fraction decreased while in the medium increased. This result shows that soon after the infection cells expelled part of their Zn. If Zn(pic)2 or Zn(asp)2 are added at the end of the adsorption period, ICP4 and ICP8 are localized in the cytoplasm but not in the nucleus of infected cells, while the synthesis of gH is decreased. A sharply increased number of nucleocapsids with low electron density cores was also found.

Effect of combined acyclovir and ribavirin on experimental herpes simplex virus type 1 keratoconjunctivitis in rabbits.

The combination of acyclovir and ribavirin has been established to be more effective on experimental herpes simplex keratoconjunctivitis in rabbits than the treatment with individual drugs. The better therapeutic effect of the combination is proved by the decreased severity of the ocular infection and the reduction of the virus shedding in tear film, as well as by the diminished duration of the infection.

Antiherpes effect of Melissa officinalis L. extracts.

Studies were carried out on the virucidal and antiviral effects of Melissa officinalis L. extracts (M1, M3, M3 and M4) with respect to Herpes simplex virus type 1 (HSV-1). Virucidal effect was registered within 3 and 6 hours of treatment using M4 administered in MTC. The remaining extracts inactivate the virus at the 12th and 24th hour. No significant values of inhibiting activity of M1, M2, and M3 on the same virus in vitro or in vivo were demonstrated. The presence of caffeic, rosmarinic and ferulic acids was demonstrated by thin-layer chromatography. Their role in the antiviral activity of M. officinalis L. is discussed.

[The synergistic antiviral effect of acyclovir and ribavirin against the herpes simplex type-1 virus and the pseudorabies virus in vitro]. / Sinergichen antivirusen efekt na atsiklovir i ribavirin sreshtu virus kherpes simpleks tip 1 i virusa na psvedobesa in vitro.

The antiviral effect of the combination acyclovir (ACV) and ribavirin (R) on herpes simplex virus type 1 (HSV-1) in diploid cell culture of human embryonal skin and muscle fibroblasts and the virus of pseudorabies in secondary culture of chicken embryonal fibroblasts is studied. The antiviral activity is studied under the checker-board combinations of twofold dilutions of the two substances on the basis of inhibition of the virus cytopathic effect in microplates. The viruses are applied at 100 CCID50, per 0.1 cm3 in each well. Dose isobologrammes are made which point out to the synergistic type of interaction of the substances in the combination in relation to the two viruses. The concentrations with maximal synergistic effect in relation to HSV-1-1.1 for ACV and 41 mu for R respectively, and in relation to the virus of pseudorabies of 3.3 and 9.2 M respectively, are established.

[The effect of antiviral substances on herpes simplex virus type 1 in cell culture]. / Efekt na antivirusni veshtestva spriamo virus kherpes simpleks tip 1 v kletuchni kulturi.

Comparative studies were carried out of the antiviral efficacy of 11 substances on the replication of Herpes simplex virus type 1 (strain DA), including compounds with selective antiherpes action--bromovinyldeoxyuridine, virazole, phosphonoformic acid, acycloguanosine, citarabine, vidarabine, iododeoxyuridine, and some nonspecific viral inhibitors with antiherpes effect as distamycin A, tunicamycin and deoxyglucose. The antiherpes activity of the substances was tested in two cell systems: cell strain of human embryonic skin-muscle fibroblasts and a cell line of green monkey kidneys. The dose-response curves and the 50% effective dose were defined for each compound by using a microtissue plate test system. Differences in the values of the 50% effective doses of iododeoxyuridine, phosphonoformic acid and distamycin A for the two types of cultures were found out. Bromovinyldeoxyuridine exhibited the highest antiherpes activity in both types of cells, and it was comparatively more active than citarabine, acycloguanosine or iododeoxyuridine, followed by vidarabine and virazole. The results obtained emphasized the necessity of specifying the experimental conditions for comparative studies.

Comparative determination of riboflavin in milk by HPLC and lumiflavin methods.

A sensitive method for the determination of riboflavin in milk is described. It involves HPLC at room temperature with fluorescence detection. The technique has good recovery, for milk 98% and for yogurt 93%. The VC for milk is 8.5 and for yogurt 6.5%. The method proposed is more rapid, reproducible and technically simple then the commonly used lumiflavin method. The HPLC technique provides opportunity for a large number of measurements using a small sample volume.

[Amino acid composition of white poultry meat contaminated with psychrophilic microorganisms]. / Aminokiselinen sustav na bialo ptiche meso, kontaminirano s piskhrofilni mikroorganizmi.

Studied was the effect of psychrophilic organisms of the Acinetobacter, Aeromonas, Alcaligenes, and Flavobacter genera on the amino acid composition of white poultry meat stored at 0 degrees to 4 degrees C in the course of 48 hours, and at 18 degrees C in the course of 6 to 12 months. The quantitative determination of individual amino acids was carried out by means of an automatic amino acid analyzer, Hitachi type, after the method of Speckman, Stein, and Moore. No changes were established in the amino acid composition of white poultry meat that was treated with psychrophilic organisms as against the composition of untreated meat. Proteins preserved their biologic value.

[Amino acid composition of white poultry meat contaminated with psychotropic microorganisms of the genus Pseudomonas]. / Aminokiselinen sustav na bialo ptiche meso, kontaminirano s psikhrotrofni mikroorganizmi ot rod Pseudomonas.

Investigated was the amino acid content in white poultry meat contaminated with four species of organisms of the Pseudomonas genus and kept for 48 hours at 0-4 degrees C and for 6 and 12 months at -18 degrees C. The content of the individual amino acids was expressed in g/100 g of protein, giving the figures of the essential amino acids. Results showed that there were no differences in the qualitative and quantitative composition of the amino acids both with regard to the various Pseudomonas species and with regard to the different duration of storage.