Antiviral activity of novel oseltamivir derivatives against some influenza virus strains.

The aim of this study was to investigate the in vitro cytotoxicity of oseltamivir derivatives and determine their activity against A/H1N1/PR/8/34 and A/H3N2/HongKong/8/68 - strains of influenza virus. Antiviral activity of these compounds was determined by using two methods. MTT staining was used to assess the viability of MDCK cells infected with influenza viruses and treated with various concentrations of drugs. In parallel, the effect of drugs on viral replication was assessed using the hemagglutination test. The most toxic compounds were: OS-64, OS-35, OS-29, OS-27 and OS-25, whereas OS-11, OS-20 and OS-23 were the least toxic ones. Statistically significant antiviral effect at a higher virus dose was shown by compounds: OS-11, OS-20, OS-27, OS-35, and OS-64. H3N2 virus was sensitive to 10-times lower concentrations of OS-11 and OS-35 than H1N1. At a lower infection dose, the antiviral activity was observed for OS-11, OS 27, OS-35 and OS-20. OS-64 turned out to be effective only at a high concentration. OS-23 showed no antiviral effect.

Diversity of influenza-like illness etiology in Polish Armed Forces in influenza epidemic season.

The aim of this study was to conduct an epidemiological and laboratory surveillance of Influenza-Like Illnesses (ILI) in Polish Armed Forces, civilian military personnel and their families in 2011/2012 epidemic season, under the United States Department of Defense-Global Emerging Infections Surveillance and Response System (DoD-GEIS). ILI incidence data were analyzed in relation to age, gender, patient category as well as pathogen patterns. Multiple viral, bacterial and viral-bacterial co-infections were identified. Nose and throat swabs of active duty soldiers in the homeland country and in the NATO peacekeeping forces KFOR (Kosovo Force), as well as members of their families were tested for the presence of viral and bacterial pathogens. From October 2011 to May 2012, 416 specimens from ILI symptoms patients were collected and analyzed for the presence of viral and bacterial pathogens. Among viruses, coronavirus was the most commonly detected. In the case of bacterial infections, the most common pathogen was Staphylococcus aureus.

Surveillance of hantaviruses in Poland: a study of animal reservoirs and human hantavirus disease in Subcarpathia.

The first cluster of hemorrhagic fever with renal syndrome (HFRS) in Poland was identified in 2007 in the Subcarpathian region. The natural environment of this area is a key habitat for hantavirus vectors. The animal reservoir of existing human HFRS clusters was studied to assess the occurrence of viruses (including Tula virus, Puumala virus, and Dobrava-Belgrade virus) among rodents. We examined 70 suspected human cases with symptoms corresponding to the clinical picture of HFRS. Serological analysis (indirect immunofluorescence assay and immunoblot) confirmed the presence of anti-hantavirus antibodies in 18 patients, which were surveyed with regard to developed symptoms and presumed rodent contact. Seroepidemiological analysis of newly confirmed human cases was performed, putative areas of human exposure were studied, and 194 rodents were subsequently captured from identified areas. Internal organs (lungs, heart, spleen, bladder, and kidneys) were collected from 64 Apodemus flavicollis, 55 Apodemus agrarius, 40 Myodes glareolus, 21 Mus musculus, and 14 Microtus arvalis and tested for the presence of hantavirus RNA by reverse transcription and subsequent real-time PCR. Positive samples were also tested by indirect immunofluorescence. Animal reservoir surveillance enabled the first detection of Puumala virus and Dobrava-Belgrade virus among animals in Poland. Furthermore, some places where rodents were captured correlated with areas of residence of laboratory-confirmed human cases and likely detected virus species. Moreover, three species of hantaviruses coexisting in a relatively small area were identified.

Acute respiratory distress syndrome (ARDS) in the course of influenza A/H1N1v infection--genetic aspects.

Influenza is a contagious respiratory disease caused by viruses belonging to the family Ortomyxoviridae. Among the influenza viruses type A, B and C, the A type virus shows the most pathogenic potential. Its surface receptor glycoproteins, hemagglutinin (HA) and neuraminidase (NA), are characterized by high antigenic variation, thus a host organism cannot develop permanent resistance. The case is described of a male patient with severe acute respiratory distress syndrome in the course of influenza A/N1H1v infection, confirmed by virological molecular analysis. During diagnostic procedures based on the MSSCP genotyping it was observed that the WHO recommended RT-PCR kits and/or procedure of sample collection from patients for molecular investigation could lead to false positive A/H1N1 pandemic strain detection because of the co-amplification during the RT-PCR fragments of the human genome.

Severe influenza outbreak in Western Ukraine in 2009--a molecular-epidemiological study.

INTRODUCTION: In the autumn of 2009 the authors participated in a humanitarian operation in Western Ukraine by undertaking an epidemiological investigation of an influenza-like-illness (ILI) in the L'viv Oblast region. Mobile biological survey teams took samples from civilian patients with severe acute respiratory distress syndrome, rapid transportation of the samples, and their molecular analysis in Poland to provide accurate results. OBJECTIVE: The aim of the study was the molecular and epidemiological analysis of the biological samples collected. MATERIAL AND METHODS: Real-time reverse transcriptase polymerase chain reaction (real-time RT-PCR), multiplex PCR techniques, traditional Sanger Sequencing and classical viral culture methods were used. RESULTS: Among the 124 influenza-like illness cases, ~50% (58) were positive for influenza A virus in WHO-CDC molecular assay, including subtyping. The specimens were further analyzed to confirm results and determine the genetic sequence. Phylogenetically, the nucleotide similarity of both the Ukraine specimens and reference A/California/7/2009 (pH1N1) was 99.2-99.3%. Oseltamivir resistance was not registered. HA1 region characterization showed an overall protein identity of 98.5-99.4%. CONCLUSIONS: An unexpected high contribution of influenza A was confirmed among ILI patients, as well as a very limited number of other detected viruses, indicate that the 2009 epidemic in western Ukraine was strongly related to novel influenza A/H1N1. The importance of swift sharing of information and reference laboratories networking in surveillance, as well as serving governments and international agencies in pursuing adequate actions, should be stressed.

Research on substances with activity against orthopoxviruses.

Although smallpox was eradicated over 30 years ago, the disease remains a major threat. High mortality, high infectivity and low resistance of the contemporary population make the smallpox virus very attractive to terrorists. The possible presence of illegal stocks of the virus or risk of deliberate genetic modifications cause serious concerns among experts. Hence, it is reasonable to seek effective drugs that could be used in case of smallpox outbreak. This paper reviews studies on compounds with proven in vitro or in vivo antipoxviruses potential, which show various mechanisms of action. Nucleoside analogues, such as cidofovir, can inhibit virus replication. Cidofovir derivatives are developed to improve the bioavailability of the drug. Among the nucleoside analogues under current investigation are: ANO (adenozine N1-oxide) and its derivatives, N-methanocarbothymidine [(N)-MCT], or derivatitives of aciklovir, peninclovir and brivudin. Recently, ST-246 - which effectively inhibits infection by limiting release of progeny virions - has become an object of attention. It has been also been demonstrated that compounds such as: nigericin, aptamers and peptides may have antiviral potential. An interesting strategy to fight infections was presented in experiments aimed at defining the role of individual genes (E3L, K3L or C6L) in the pathogenesis, and looking for their potential blockers. Additionally, among substances considered to be effective in the treatment of smallpox cases, there are factors that can block viral inhibitors of the human complement system, epidermal growth factor inhibitors or immunomodulators. Further studies on compounds with activity against poxviruses are necessary in order to broaden the pool of available means that could be used in the case of a new outbreak of smallpox.

Research on prevention and treatment of hemorrhagic fevers.

Viral hemorrhagic fevers are severe zoonotic diseases caused by RNA-viruses classified into 4 families: Arenaviridae, Bunyaviridae, Filoviridae, and Flaviviridae. They are present on all continents except Antarctica, their person-to-person spread is easy, and there is a high risk of them being used as weapon by bioterrorists. So far, efforts to develop effective drugs against these viruses have failed, and typical therapy usually relies on symptomatic treatment. Search for substances that could effectively inhibit this type of infections is now a priority. The presented paper gives an overview of different approaches used in combating the viral hemorrhagic fevers. Researchers look for safe antiviral agents with appropriate properties among natural sources, such as various types of herbs plants, algae, or essential oils obtained from trees, as well as investigate the use of various synthetic substances. The aim is to broaden the pool of available antiviral drugs that could replace hitherto applied medicines such as ribavirin, which is not always sufficiently effective and may have side-effects. The scientists focus not only on combating the diseases, but also on their prevention. For this purpose, recombinant vaccines or various types of immunomodulators may serve as a useful tools. Results of the latest studies are promising and encourage further work which may eventually lead to the solution of the urgent problem of hemorrhagic fevers.

[Peptides--a new strategy for combating viral infections]. / Peptydy--nowe mozliwosci zwalczania zakazen wirusowych.

The efficiency of peptides against many species of bacteria, fungi and parasites has been widely described. Recent studies on peptides have also demonstrated their antiviral activity. Some peptides exhibit direct virucidal activity, others disturb attachment of virus particles to the cell membrane surface or interfere with intracellular replication of virus. Due to limited effectiveness of commonly used drugs and emerging resistance of viruses, antiviral peptides may have the potential to be developed as putative therapeutic agents.

[Real time PCR hybridization for the rapid and specific identification of Francisella tularensis]. / Identyfikacja Francisella tularensis technika real-time PCR z wykorzystaniem sond hybrydyzujacych zaprojektowanych dla fragmentów sekwencji genów fopA i tul4.

Tularemia is highly infectious and fatal zoonotic disease caused by Gram negative bacteria Francisella tularensis. The necessity to undergo medical treatment in early phase of illness in humans and possibility of making use of bacterial aerosol by terrorists in an attack create an urgent need to implement a rapid and effective method which enables to identify the agent. In our study two primers FopA F/R and hybridization probes FopA S1/S2 designed from fopA gene sequence, were tested for their potential applicability to identify F. tularensis. In this research 50 strains of F. tularensis were used and the test gave positive results. Reaction specificity was confirmed by using of non-Francisella tularensis bacterial species. The results obtained in the real-time PCR reaction with primers Tul4 F/R and hybridization probes Tul4 S1/S2, designed from tul4 gene, were comparable to the results from previous experiment with fopA - primers set. Investigation of fopA and tul4 primers and hybridization probes properties revealed characteristic Tm (melting temperature) value of the products--61 degrees C and 60 degrees C, respectively. Detection sensitivity was remarkably higher when fopA primers set was used 1 fg/microl, and for tul4 primers set, minimal detectable concentration is 10 fg/microl.