A new locality of the Haemaphysalis concinna tick (Koch, 1844) in Poland and its role as a potential vector of infectious diseases

The Haemaphysalis concinna tick is a rare species in Poland. To date, it was found only once a few decades ago. During tick collection for epidemiological studies, a stable population of this arachnid was found in a military training area near Nowa Deba. This report is particularly important, given the role of Haemaphysalis concinna in the spread of dangerous vector-borne diseases.

Prevalence of Coxiella burnetii in environmental samples collected from cattle farms in Eastern and Central Poland (2011-2012).

Coxiella burnetii is the etiologic agent of Q fever. It may occur as two different morphological forms, a large cell variant (LCV) and a small cell variant (SCV). The SCV is characterized by unique resistance to physical and chemical factors and may survive in the environment for many months. The objective of this study was to examine environmental samples for the presence of C. burnetii using real-time PCR in areas where Q fever was previously reported and in randomly selected animal farms where Q fever was not reported. The samples were collected in the following provinces in Poland: Lublin, Subcarpathian and Masovian. Monitoring was performed with real-time PCR and serological methods. Of the 727 environmental samples, 33 (4.54%) contained the multi-copy insertion sequence IS1111, which is specific for C. burnetii. Subsequently, the presence of C. burnetii antibodies was determined using serological tests in selected herds in which positive genetic results were obtained. Serological analyses of 169 serum samples using CFT and ELISA were performed on Polish black-and-white Holstein-Friesian cows and one cow imported from Denmark. Using the CFT method, 11 samples were positive for phase I antibodies and six were positive for phase II antibodies. Moreover, in two cases, the presence of antibodies specific for both phase I and phase II antigens of C. burnetii was detected. However, of the 169 examined serum samples, 20 were positive by ELISA test, of which six were also positive by CFT. Additionally, multi spacer typing (MST) of isolated C. burnetii strains was performed. The MST results identified two new genotypes in Poland, ST3 and ST6. The results indicate that continued research regarding spread of this pathogen within a country is necessary.

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.

Q fever--selected issues.

Q fever is an infectious disease of humans and animals caused by Gram-negative coccobacillus Coxiella burnetii, belonging to the Legionellales order, Coxiellaceae family. The presented study compares selected features of the bacteria genome, including chromosome and plasmids QpH1, QpRS, QpDG and QpDV. The pathomechanism of infection--starting from internalization of the bacteria to its release from infected cell are thoroughly described. The drugs of choice for the treatment of acute Q fever are tetracyclines, macrolides and quinolones. Some other antimicrobials are also active against C. burnetii, namely, telitromycines and tigecyclines (glicylcycline). Q-VAX vaccine induces strong and long-term immunity in humans. Coxevac vaccine for goat and sheep can reduce the number of infections and abortions, as well as decrease the environmental transmission of the pathogen. Using the microarrays technique, about 50 proteins has been identified which could be used in the future for the production of vaccine against Q fever. The routine method of C. burnetii culture is proliferation within cell lines; however, an artificial culture medium has recently been developed. The growth of bacteria in a reduced oxygen (2.5%) atmosphere was obtained after just 6 days. In serology, using the IF method as positive titers, the IgM antibody level >1:64 and IgG antibody level >1:256 (against II phase antigens) has been considered. In molecular diagnostics of C. burnetii infection, the most frequently used method is PCR and its modifications; namely, nested PCR and real time PCR which detect target sequences, such as htpAB and IS1111, chromosome genes (com1), genes specific for different types of plasmids and transposase genes. Although Q fever was diagnosed in Poland in 1956, the data about the occurrence of the disease are incomplete. Comprehensive studies on the current status of Q fever in Poland, with special focus on pathogen reservoirs and vectors, the sources of infection and molecular characteristics of bacteria should be conducted.

[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.

[Yersinia pestis pathogenesis and diagnostics]. / Chorobotwórczosc i diagnostyka Yersinia pestis.

Plague is an acute bacterial infection caused by Gram negative organism Yersinia pestis. This bacteria is subdivided into three classical biotypes: Orientalis, Medievalis and Antiqua. Plague is transmitted via flea vectors from rodents to humans and by respiratory droplets from animals to humans or humans to humans. This agent is on the top of the CDC list of "Critical Biological Agents"--category A. It appears to be a good candidate agent for a bioterrorist attack. Type III secretion (TTS) is a mechanism by which Y. pestis communicates with eukaryotic cells by injecting bacterial proteins across cellular membranes into the cytosol of these cells. These bacterial proteins take control of the host cells by hijacking their intracellular machinery. A laboratory diagnostics of plague is based on: staining techniques, culture on media, immunochromatography, hemagglutination, immunofluorescence, ELISA, phage tests and genetical techniques including: PCR, multiplex and nested PCR, real time PCR, VNTR, PFGE, ISBF and Microarray.

[Francisella tularensis--feature of pathogen, pathogenesis, diagnostics]. / Francisella tularensis--cechy zarazka, patogeneza, diagnostyka.

Francisella tularensis belongs to the Francisellaceae family. There are four known subspecies of Francisella tularensis: tularensis, holarctica, mediasiatica and novicida. Fully virulent strains possess a capsule, which protects F. tularensis from bactericidal action of serum. The main virulen factors of F. tularensis are 23-kDa cytoplasmatic protein and LPS. F tularensis mechanism of pathogenecity is very unique. F. lularensis affect macrophages using a cytochalasin B intensive pathway. Bacteria live within macrophage in a phagosome. Acidification of the phagosome and acquisition of iron is essential for growth of F. tularensis. An acid pH promotes the release of iron from host-cell transferrin. An acid phosphatase function protein, AcpA, has been identified in F. tularensis. AcpA is capable of inhibiting the respiratory burst. A laboratory diagnostics of tularemia is based on classical microbiology and molecular biology techniques: PCR, nested-PCR, PCR-ELISA, Real-Time - PCR, ALFP, ERIC-PCR, PFGE, LR-REP-PCR and microarray techniques.