An experience in the clinical use of specific immunoglobulin from horse blood serum for prophylaxis of Ebola haemorrhagic fever.

The aim of this work was to estimate the efficacy and safety of single intramuscular introduction of specific heterologous immunoglobulin as prophylactic drug against Ebola hemorrhagic fever. Materials and methods. The specific heterologous immunoglobulin was introduced as a special prophylactic drug to 28 patients in epidemic situations, after skin hurt with infectious materials or contact with infectious blood. Clinico-laboratory observation was performed in 24 subjects after single intramuscular introduction of heterologous immunoglobulin Ebola. The samples of blood serum were investigated for immunoglobulin Ebola and antibodies to horse gamma-globulin on the 30th and 60th days after prophylaxis. Results. None of the subjects of the study contracted Ebola fever. There were no anaphylactic reactions after special prophylaxis with specific heterologous immunoglobulin. Among the subjects with normal allergic state 31% responded with local reactions; 13%, with a general reaction (mild case of the serum disease). Almost no reaction was observed in patients with unfavorable allergic state subjected to desensitizing therapy; in the absence of desensitizing therapy, 50% of patients with unfavorable allergic state exhibited local reactions; 17%, mild cases of the serum disease; 33%, moderate cases of the serum disease. In summary, if the tactics of immunoglobulin application was right, the quantity of local allergic reactions was 28%; of wide spread reactions, 6%. Weak serum disease was observed in 11% of the subjects. The prognostic period of resistance to Ebola fever was less than 30 days. Conclusion. The prophylactic use of specific immunoglobulin from horse blood serum against hemorrhagic Ebola fever is effective and relatively safe in patients subjected to desensitizing therapy.

[Sensitivity and specificity of the ELISA Kit for the detection of antidobies to Junin virus].

The goal of this work was to describe methodological approaches to determination of sensitivity and specificity of the enzyme-linked immunosorbent assay kit (ELISA Kit) for detection of the specific anti-Junin virus (JV) antibody. Comparison of ELISA to plaque reduction neutralization test (PRNT) showed direct relationship between antibody titers in the samples of serum of immunized animals, determined by either PRNT or ELISA methods. The obtained results provided an opportunity to form the panels of positive and negative serum samples to determine the sensitivity and specificity of the ELISA Kit. Sensitivity of the ELISA Kit was at least 98% when studying the samples of serum of immunized guinea pigs and rabbits (determined as positive in PRNT). The sensitivity of the ELISA Kit was at least 68% when studying the samples determined by PNRT as uncertain positive. The specificity was 98%. The specificity of the ELISA Kit was 98%.

[Evaluation of safety of prophylactic use of immunoglobulins against viral hemorrhagic fevers from horse blood sera].

AIM: Evaluate safety of prophylaxis of viral hemorrhagic fevers by specific heterologous immunoglobulins. MATERIALS AND METHODS: Clinical-laboratory examination of 24 individuals after intramuscular administration of heterologous Ebola immunoglobulin was carried out. Anaphylactogenicity of the immunoglobulins was studied by WD 42-28-8-89 in guinea pigs compared with commercial preparations. RESULTS: Immediate type reactions were not observed. In individuals with normal anamnesis the number of local reactions was 31%, general in the form of lung serum disease - 13%. In individuals with unfavorable anamnesis against the background of desensitization therapy there were almost no reactions; without it local reactions were present in 50%, mild severity serum lung disease - in 17%, medium - in 33%. Immunoglobulins against especially dangerous viral agents by anaphylactogenic properties did not differ from commercial heterologous preparations. CONCLUSION: Application of specific immunoglobulins from horse blood sera (the main means of protection from dangerous and especially dangerous exotic viral infections) with compliance by desensitization principles is relatively safe. Safe level of sensitization properties is characterized by anaphylaxis index up to 3.7 for guinea pigs.

[Influence of cultivation temperature and Yersinia pestis fra-operon carriage on morphological features of Yersinia pseudotuberculosis].

AIM: Studies of influence of Yersinia pestis fra-operon carriage on morphological properties of Y. pseudotuberculosis recipient strain cells and colonies at different temperature and cultivation time. MATERIALS AND METHODS: Cultures of Y. pseudotuberculosis isogenic variants were grown on solid nutrient medium at 4 - 6, 9 - 11, 26 - 28 and 36 - 38 degrees C. Indirect hemagglutination was used to determine F1 antigen production. Cytorefractometry was used to determine live cell percentage in colonies. Size and dividing cells percentage was evaluated by using phase-contrast microscope "Lyumam-I2". RESULTS: Time period between appearance of micro-colonies and achievement of maximum colony size increased with cultivation temperature decrease. Size of fra+ and fra- variant colony was not significantly different for every temperature regiment used and was significantly lower at the temperature of 36 - 38 degrees C. CONCLUSION: Temperature has a significant influence on population growth, colony size and form.

[Stabilization of peroxidase conjugates used in enzyme immunoassay systems to detect Ebola and Marburg virus antigens].

The time course of changes in the activity of solutions of horseradish peroxidase conjugates with immunoglobulins against Ebola and Marburg fevers was studied in the presence of different components. The series of the conjugates of ELISA kits for the detection of Ebola and Marburg virus antigens, which were prepared on the basis of the designed stabilizing solution, preserved at less than 90% of its baseline activity during 10 months at a storage temperature of 2 to 8 degrees C.

[Use of antibiotics in the treatment of ulceronecrotic tonsillitis due to vaccination by tabletted smallpox vaccine].

Clinical trials of tabletted pox vaccine revealed development of tonsillitis as a postvaccinal reaction in some volunteers: ulceronecrotic lesions in the tonsils, lymphadenitis, hyperthermia and asthenia. The main cause of the local inflammatory reactions was activation of the host opportunistic microflora including hemolytic streptococci and Staphylococcus aureus. For the treatment of the infectious complications systemic antimicrobials, such as benzylpenicillin, amoxicillin, ampicillin, cefazolin and fluoroquinolones (ciprofloxacin) in combination with the symptomatic therapy were used. The treatment course of 9 days provided complete elimination of the postvaccinal reactions, the specific antibody generation being not affected.

[Genotyping of the Bacillus anthracis vaccine strains using the multiloci VNTR-analysis].

The genotyping variety of 5 known anthracis vaccine strains using 18 variable loci of the chromosomal localization taken from a microbe culture collection of 48 Research Institute of Ministry of Defense was revealed in the research. The stability of the VNTR-loci was shown to be inherited from the B. anthracis strains with common origin and an opportunity of their gene-identification application. The gene profile of each analyzed vaccine strain using every 18 polymorphic loci was determined and the amplification products were sequenced. The variation of electrophoretic mobility of the amplifiers was found to be caused by the presence of the replication elements with various numbers of copies in their structure.

[Use of guinea pigs to evaluate the efficacy of a heterological immunoglobulin against Bolivian hemorrhagic fever].

The use of guinea pigs as a laboratory model was proven to be appropriate in investigating the protective properties of a heterological immunoglobulin against Bolivian hemorrhagic fever at the preclinical stage of the study. A highly pathogenic Machupo virus strain that caused guinea pigs' death with respect with an agent's dose was cultivated. Injection of 1.0 ml of the immunoglobulin provided a 100% protective effect for the guinea pigs infected with the highly pathogenic Machupo virus strain in a dose of 10 LD50.

[Study of efficacy of modern fluoroquinolones against agent of glanders in experiments in vivo].

AIM: Efficacy of modern fluoroquinolones for urgent prophylaxis and treatment of experimental glanders was studied. In experiments on laboratory animals in vivo, it was shown that sparfloxacin, gemifloxacin, levofloxacin and moxifloxacin were highly effective for urgent prophylaxis and treatment of glanders. MATERIALS AND METHODS: Golden hamsters of both sexes, with weight 80 - 100 g, were inoculated with 100 LD50 of 48-hour agar culture of Burkholderia mallei (strain ts-5). Commercial preparations of 2 - 4th generations of fluoroquinolones: sparfloxacin (Sparflo, India), gemifloxacin (Faktiv, Russia), moxifloxacin (Avelox, Germany), pefloxacin (Abactal, Slovenia), levofloxacin (Eleflox, India), lomefloxacin (Lomeflox, India), ofloxacin (Russia). Urgent prophylaxis started 3 hours after inoculation with duration of 10 days, whereas treatment started 24 hours after inoculation with duration of 15 days. Daily dose of pefloxacin, ciprofloxacin, of loxacin was divided on 2 parts, which were administered with 12-hour interval; other drugswere administered once a day. RESULTS: All studied drugs, excluding lomefloxacin, were highly effective for urgent prophylaxis and treatment of experimental glanders and provided 80 - 100% protection. CONCLUSION: Third-fourth generations of fluoroquinolones: sparfloxacin, levofloxacin, moxifloxacin, gemifloxacin were highly effective against agent of glanders in in vivo experiments. They are promising drugs for the development of schemes for urgent prophylaxis and treatment of glanders in humans.

[The time course of changes in cell immunological parameters during administration of live dry plague vaccine].

The study of the time course of changes in cell immunological parameters by a magnetic separation technique in human beings during the administration of plague vaccine in relation to the immunological load revealed the higher blood levels of all T lymphocyte subpopulations on day 14 after vaccination. These changes are most typical of a primary vaccinated cohort. The increased frequency of plague vaccine administration and multiple immunizations with live plague, anthrax, and tularemia vaccines produce the time-course of changes in T lymphocyte populations (subpopulations) in response to the regular administration of plague vaccine. A high immunological load in man also promotes a significant reduction in the level of B lymphocytes.

[Design of equine serum-based Marburg virus immunoglobulin].

Immunoglobulin (Ig) against Marburg fever (MF) has been obtained from the equine serum. In terms of physicochemical and immunobiological properties, the obtained preparation corresponds to the quality of heterologous commercial immunoglobulins. The application of Marburg virus (MV) Ig with a titer of no less than 1:2048 by the emergency prevention scheme 1-2 hours after intraperitoneal inoculation of guinea pigs with MV in a dose of 20-50 LD50 protected 88-100% of the animals from death. MV Ig is recommended for emergency prevention of human MF.

[Temperate Legionella bacteriophage: discovery and characteristics].

For the first time, temperate Legionella bacteriophage was isolated from organs of guinea pig infected with Philadelphia 1 strain of Legionella pneumophila. Negative colonies of bactriophage from 1.5 to 2.5 mm in diameter were detected. Central part of them was transparent and surrounded by peripheral zone of partial lysis. Electron microscopy showed that corpuscles of the phage consist from multifaceted elongated head of stretched hexagonal form and short tail. The bacteriophage lyzed bacteria, which cause Legionnaires' disease, and also had certain lytic activity against causative agent of tularemia.

[Fluoroquinolones: antimicrobial activity and chemotherapeutic efficacy with respect to various pathogens in highly dangerous diseases].

Comparative in vitro and in vivo efficacy of new fluoroquinolones with respect to pathogens of tularemia, glanders, melioidosis and anthrax was estimated. It was shown that the strains of the tularemia, glanders, melioidosis and anthrax pathogens were in vitro highly susceptible to the new agents. The experiments on laboratory animals demonstrated that pefloxacin and sparfloxacin had extremely broad spectra and were of special value in emergency prophylaxis of tularemia, glanders, melioidosis and anthrax.

[Hemorrhagic (Marburg, Ebola, Lassa, and Bolivian) fevers: epidemiology, clinical pictures, and treatment].

The evaluation of the biological and epidemiological properties of Ebola, Marburg, Lassa, and Machupo viruses suggests that they are of social importance for health care authorities. The studies have created prerequisites to the development of reliable biosafety means against these pathogens. Particular emphasis is laid on the methods for infection diagnosis and on the studies to design specific protective agents--immunoglobulins and inactivated vaccines.

A molecular and cytogenetic analysis of lambda 20p7 fragment DNA from the proximal beta-heterochromatin of Drosophila melanogaster.

A DNA fragment from the Drosophila melanogaster genome, cloned in lambda 20p7, was derived independently from clones lambda 20 and lambda L [Baiborodin et al., Genetika 29 (1993) 403-416; Sharakhov et al., Genetika 29 (1993) 392-402]. In situ hybridization of lambda 20p7 DNA to the chromosomes of D. melanogaster demonstrated preferential hybridization of the fragment to the chromocenter of polytene chromosomes and to pericentric heterochromatin of chromosomes II, IV and X at the metaphase plate. Copy number per haploid genome for lambda 20p7 was estimated as approximately 200. Based on Southern blotting, the major portion of this moderate repeat was localized in the region of a 5.5-kb HindIII digest. In situ hybridization to polytene chromosomes from strain fs(2)B trophocytes revealed that repeats homologous to lambda 20p7 are located in the proximal heterochromatin which undergoes structural reorganization during tissue differentiation. The nucleotide sequence of two segments of the clone lambda 20p7, Dm0.9 and Dm270, was determined. Sequence analysis of the 300-bp Dm0.9 clone revealed that it contains 21-bp and 30-bp d(GT/CA) sequences, a 12-bp AT box, recognition sites for nuclear factors NFI and SpI, and a set of inverted repeats. Clone Dm270 contains an open reading frame (ORF). The deduced amino acid (aa) sequence shares homology with the gag-like gene from type-I (R1) ribosomal DNA insertion and may code for a polypeptide of 10 kDa. The Dm270 sequence was found to contain two direct repeats showing homology to the human CENP-B box.

DNA from Drosophila melanogaster beta-heterochromatin binds specifically to nuclear lamins in vitro and the nuclear envelope in situ.

A DNA fragment designated lambda 20p1.4 binds in vitro to polymerized Drosophila melanogaster lamin. In situ hybridization of lambda 20p1.4 to isolated polytene chromosomes revealed localization at the chromocenter and to the 49 CD region on the right arm of chromosome 2. About 120 copies of sequences homologous to lambda 20p1.4 were detected per haploid genome. Nucleotide (nt) sequence analysis demonstrates that lambda 20p1.4 is an A + T-rich, 1327-bp fragment containing four repeated units between nt 595 and 919. Results suggest that lamin interacts with a region of lambda 20p1.4 between nt 300 and 1000. Confocal immunofluorescence co-localization demonstrates that in situ, the major locus of lambda 20p1.4 hybridization, the chromocenter, is found juxtaposed to the nuclear envelope (lamina). This is the first demonstration that a DNA sequence that binds specifically to nuclear lamins in vitro, is located at or near the nuclear envelope in situ and, presumably, in vivo.

[Beta-heterochromatin in Drosophila: molecular organization and function. Molecular biological analysis of a MAR/SAR DNA sequence in centromere heterochromatin of Drosophila melanogaster]. / Beta-geterokhromatin Drosophila: molekuliarnaia organizatsiia i funktsionirovanie. Molekuliarno-biologicheskii analiz MAR/SAR posledovatel'nosti DNK iz tsentromernogo geterokhromatina Drosophila melanogaster.

A molecular biological characteristic of a DNA fragment delta 20p1.4, which is a moderately repetitive sequence of the Drosophila melanogaster genome, is presented. The fragment is present in about 120 copies per haploid genome. The main pool of the delta 20p1.4 homologous DNA can be isolated, along with the nuclear matrix DNA, and consists of Hind III-EcoR I monomers 1.4-1.6 kb in length. The monomers may occur in the genome as both single copies and tandem clusters forming chromosomes fragments up to 6-10 kb in length. The region of the delta 20p1.4 fragment between nucleotides 350 and 905 polymerizes with purified lamin from D. melanogaster. The sites of the fragment that had physical contact with lamin in vitro were determined using an Exo III protection. It was demonstrated that ATATTT, A, and T boxes located in four nonperfect tandem repeats were involved in the contact, both DNA strains reacting with lamin.

[Drosophila beta-heterochromatin: molecular organization and function. Characteristics of the DNA sequences from proximal beta-heterochromatin, associated with the nuclear envelope of Drosophila melanogaster]. / beta-Geterochromatin Drosophila: molekuliarnaia organizatsiia i funktsionirovanie. Kharakteristika posledovatel'nosti DNK iz proksimal'nogo beta-geterokhromatina, assotsiirovannogo s iadernoi obolochkoi Drosophila melanogaster.

To study the nucleotide sequence from the pericentric heterochromatin associated with the nuclear envelope, a residual DNA was extracted from the DNAse-treated nuclear lamins of Drosophila melanogaster tissue culture cell line Kc. The isolated DNA was cloned in lambda vector. The DNA library obtained was screened for the clones homologous to the pericentric heterochromatin. The experiments on in situ hybridization to the polytene chromosome of the nurse cell nuclei of the strain fs(2) B assigned the reiterated sequence, homologous to the lamin DNA clone, to the nuclear envelope associated regions of the proximal beta-heterochromatin which is known to undergo structural reorganization during cell differentiation. The nucleotide analysis of 300 bp from this sequence has established the presence of 21 bp and 300 bp d(GT/CA), 12 bp AT-box, the regions of recognition of the nuclear factor and the inverted repeats.

[Drosophila beta-heterochromatin: molecular organization and function. Cloning and molecular biological analysis of the lambda 20 DNA fragment from Drosophila melanogaster beta-heterochromatin]. / beta-Geterokhromatin Drosophila: molekuliarnaia organizatsiia i funktsionirovanie. Klonirovanie i molekuliarno-biologicheskii analiz lamda 20 fragmenta DNK iz beta-geterokhromatina Drosophila melanogaster.

To isolate the DNA sequences specific for the pericentric heterochromatin of Drosophila we used two CREST-autoimmune sera which bind in the Western-blot analysis the nuclear antigens of 30 kDa, 43 kDa and 45 kDa molecular weight. Cloning of the DNA fragments associated with these CREST-specific proteins of Drosophila resulted in obtaining 8 clones. One of them, lambda 20, hybridized mainly to the chromomcenter of polytene chromosomes. The further analysis indicated that the lambda 20 DNA might belong to the proximal beta-heterochromatin of the polytene chromosomes of D. melanogaster.

[Dynamics of chromatin position within the interphase nucleus]. / Dinamika polozheniia khromatina v ob''eme interfaznogo iadra.

In this article we attempt to analyze the relationship between the processes, which occur in the nucleus, and the dynamics of chromatin, as well as to classify the changes in the position of chromatin in the cell nucleus during the lifetime of the cell. The proposed concept integrates possible types of chromatin movement within the nucleus.