Genetic analysis of poliovirus strains isolated from sewage in Poland.

The study describes genetic characterization of poliovirus (PV) strains isolated from sewage samples in Poland. The analyses were performed for the detection of any putative polio revertants and recombinants in three genomic regions by sequencing analysis. Thirty-six strains were analyzed. The analyzed strains were identified by neutralization assay as 7 strains of serotype P1, 10 strains of serotype P2, and 19 strains of serotype P3. Sewage isolates were sequenced in 5'UTR, VP1, and 3D genomic regions. All detected PVs were classified as vaccine strains on the basis of VP1 sequence. Mutational differences in the VP1 sequences of isolated viruses ranged from 0.0% to 0.4%, indicating a limited replication period. The genetic analysis of the 3D region showed that some strains have recombinant genomes. Nine strains were found as dipartite recombinants (seven strains--S3/S2, one strain--S2/S1, one strain--S3/S1), while one strain was found as tripartite recombinant (S3/S2/S1). No recombinants with non-PV enteroviruses were identified. None of wild-type PVs or vaccine-derived polioviruses (VDPVs) were detected. This study showed the absence of wild or VDPV circulation in the country and demonstrated the usefulness of environmental surveillance in addition to acute flaccid paralysis (AFP) surveillance in support of polio eradication initiatives.

Parvovirus B19 infection in five European countries: seroepidemiology, force of infection and maternal risk of infection.

We conducted a seroprevalence survey in Belgium, Finland, England & Wales, Italy and Poland on 13 449 serum samples broadly representative in terms of geography and age. Samples were tested for the presence of immunoglobulin G antibody using an enzyme immunoassay. The age-specific risk of infection was estimated using parametric and non-parametric statistical modelling. The age-specific risk in all five countries was highest in children aged 7-9 years and lower in adults. The average proportion of women of child-bearing age susceptible to parvovirus B19 infection and the risk of a pregnant women acquiring B19 infection during pregnancy was estimated to be 26% and 0.61% in Belgium, 38% and 0.69% in England & Wales, 43.5% and 1.24% in Finland, 39.9% and 0.92% in Italy and 36.8% and 1.58% in Poland, respectively. Our study indicates substantial epidemiological differences in Europe regarding parvovirus B19 infection.

Type specific seroprevalence of HSV-1 and HSV-2 in four geographical regions of Poland.

OBJECTIVE: To examine the type specific seroprevalence of herpes simplex virus (HSV) types 1 and 2 infections, stratified by age and gender, and associated risk factors for HSV-2 seropositivity in Poland. METHODS: 2257 serum samples of individuals from 15-65 years were randomly selected from serum banks in four different geographical regions of Poland, including the Zachodnio-pomorskie, Warminsko-mazurskie, Lubelskie, and Mazowieckie districts. Type specific serum antibodies to HSV-1 and HSV-2 were detected using HerpeSelect IgG ELISA tests. RESULTS: Overall prevalences of type specific HSV-1 and HSV-2 serum antibodies were 90.4% and 9.3%, respectively. Age standardised HSV-2 seroprevalence was higher in women (9.7%) than men (8.8%) (p = 0.06), and increased notably with age from 4% in 15-24 year olds to 12% in those aged 50-65 years. HSV-1 seroprevalence was consistently higher than HSV-2 seroprevalence in each specific age group, ranging from 74.5% in 15-24 year olds to 98.8% in 50-65 year olds. HSV-2 seroprevalence varied significantly by geographical region, with the highest prevalence in the Zachodnio-pomorskie district (12%). Significant multivariate risk factors for HSV-2 seropositivity included older age, female gender, and geographical place of residence. CONCLUSION: This large survey found a notably high seroprevalence of HSV-1, even among young female adolescents 15-19 years of age (80%). HSV-2 seropositivity was under 12% in all age groups surveyed in Poland, tending to be among the lowest overall HSV-2 seropositivity rates reported thus far in Europe.

Flow cytometric enumeration of micronucleated reticulocytes: high transferability among 14 laboratories.

This laboratory previously described a single-laser flow cytometric method, which effectively resolves micronucleated erythrocyte populations in rodent peripheral blood samples. Even so, the rarity and variable size of micronuclei make it difficult to configure instrument settings consistently and define analysis regions rationally to enumerate the cell populations of interest. Murine erythrocytes from animals infected with the malaria parasite Plasmodium berghei contain a high prevalence of erythrocytes with a uniform DNA content. This biological model for micronucleated erythrocytes offers a means by which the micronucleus analysis regions can be rationally defined, and a means for controlling interexperimental variation. The experiments described herein were performed to extend these studies by testing whether malaria-infected erythrocytes could also be used to enhance the transferability of the method, as well as control intra- and interlaboratory variation. For these studies, blood samples from mice infected with malaria, or treated with vehicle or the clastogen methyl methanesulfonate, were fixed and shipped to collaborating laboratories for analysis. After configuring instrumentation parameters and guiding the position of analysis regions with the malaria-infected blood samples, micronucleated reticulocyte frequencies were measured (20,000 reticulocytes per sample). To evaluate both intra- and interlaboratory variation, five replicates were analyzed per day, and these analyses were repeated on up to five separate days. The data of 14 laboratories presented herein indicate that transferability of this flow cytometric technique is high when instrumentation is guided by the biological standard Plasmodium berghei.

Detection of EBV infection in different etiologic groups of patients.

The ability of two diagnostic tests (ELISA and IF) to detect of EBV infection in etiologically different group of patients was compared: cases of chronic lymphoadenopatis, confirmed mononucleosis or suspected EBV infection, tumors like leukemia or lymphoma. The presence of specific IgM and IgG antibodies for different EBV antigens was studied. The results obtained indicated that as many as 17 out of 32 tested serum samples presented different interpretation of EBV infection in both tests used. High number of discordant results was observed in detection of EA-IgG. The highest number of discordant results was observed in group of patients with tumors, while the lowest in group of cases diagnosed or suspected for EBV infection.

Human herpesvirus type 8 seroprevalence among patients in immunosuppression state.

Human herpesvirus 8 (HHV 8) is implicated in the etiology of neoplastic disorders, especially in HIV-infected and immunosuppressed people, but knowledge about seroprevalence of HHV 8 in general or selected populations is still insufficient. In this study the presence of IgG antibodies to HHV 8 in groups of immunosuppressed patients was tested.

[Temperature sensitive mutant of Herpes simplex virus type 1. I. Pathogenicity and immunogenicity]. / Temperaturowrazliwy mutant wirusa Herpes simplex typu 1. I. Patogennosc i immunogennosc.

The aim of the study was to characterize biological features of the sensitive mutant of HSV-1, derived from McIntyre strain by numerous virus passages at lowered replication temperature (28 degrees C). Pathogenicity of obtained ts mutant for inbred mice lines, CFW/Pzh and BALB/cPzh, was determined. Statistically significant decrease in virulence of the mutant for these mouse lines was demonstrated, as compared with the native virus strain, propagated at 37 degrees C. Immunogenic activity of ts mutant of HSV-1 defined by the possibility of mouse protection against infection with high virulent was determined. Mice, which at the time of immunization with ts mutant received Depo-Medrol--an immunosuppressive agent--were also found to be capable of inducing defense mechanisms to infection with the native strain.

[Temperature sensitive mutant of Herpes simplex virus type 1. II. Neurovirulence and latency]. / Temperaturowrazliwy mutant wirusa Herpes simplex typu 1. II. Neurowirulencja, i latencja.

The course of acute infection of mice with ts mutant or the native strain DNA and the antigens of HSV in brain nerve cells were determined. Virus DNA was detected in brains of all mice in both animal groups while the virus antigens--only in cells of mice infected with the native strain. It can be suggested, therefore, that the ability of ts mutant to replicate in central nervous system of the infected mice is lacking or much lower. The detection of virus nucleic acid 3-5 months after virus infection might indicate a possibility of establishing latent infection. However, ts mutant showed a significantly lower possibility of latency induction, as compared with highly virulent strains. It was found that the mutant ability to induce latent infection was markedly increased when mice were treated with both ts mutant and Depo-Medrol as immunosuppressive agent. This finding shows both a possibility of increase of frequency of latent infections in the state of immunosuppression, and of activation of the latent infection (recurrence of acute form of infection).

Immunogenic activity of HSV-1 temperature sensitive mutant's proteins in mono- and polyvalent systems of immunization.

Immunogenic activity of herpes simplex type 1 temperature sensitive mutant's (ts HSV-1 mutant) proteins was tested in two systems: monovalent and polyvalent with other attenuated virus strains (measles and mumps). The guinea pigs were used as animal model. In monovalent system the humoral response in animals infected with ts HSV-1 mutant (1 or 2 doses) was studied and compared to results received for HSV-1 native strain. In polyvalent system the immunological response induced by ts HSV-1 mutant in the presence of RNA virus strains was tested.

Usefulness of hybridization and PCR methods in monitoring of CMV infection in renal transplant recipients.

The purpose of this work was to compare hybridization and PCR methods as diagnostic tests in diagnosing and monitoring CMV infection. The investigation was performed in a group of 24 renal transplant recipients treated with ATG. The results we obtained suggest that quantitative variant of hybridization is more useful in diagnosing the infection than PCR, because it enables to monitor the infection. DNA CMV level of about 60 pg/ml or the increasing level in the subsequent samples should be a sign to start antiviral therapy.

Quantitative detection of CMV DNA by PCR and hybridizaton methods in renal transplant recipients.

The comparison of two quantitative tests: hybridization (Murex Hybrid Capture System) and PCR (COBAS AMPLICOR CMV Monitor) detected CMV DNA was made. Investigation of viral load in serum by PCR gave better correlation with clinical manifestation in renal transplant recipients.

[Evaluation of the usefulness of various PCR method variations and nucleic acid hybridization for CMV infection in immunosuppressed patients]. / Ocena przydatnosci róznych wariantów metody PCR oraz metody hybrydyzacji kwasów nukleinowych w wykrywaniu zakazenia CMV u pacjentów z immunosupresja.

In diagnosis of CMV infection various laboratory methods are used. The methods based on detection of viral nucleic acids have been introduced routinely in many laboratories. The aim of this study was to compare nucleic acid hybridisation method and various variants of PCR methods with respect to their ability to detect CMV DNA. The studied material comprised 60 blood samples from 19 patients including 13 renal transplant recipients and 6 with acute leukaemia. The samples were subjected to hybridisation (Murex Hybrid Capture System CMV DNA) and PCR carried out in 3 variants: with one pair of primers (single PCR), nested PCR and Digene SHARP System with detection of PCR product using a genetic probe in ELISA system. The sensitivity of the variants ranged from 10(0) particles of viral DNA in nested PCR to 10(2) in single PCR. The producer claimed the sensitivity of the hybridisation test to be 3 x 10(5) and it seems to be sufficient for detection of CMV infection. The obtained results show that sensitivity of hybridisation was comparable to that of single PCR and the possibility of obtaining quantitative results makes it superior, on efficacy of antiviral therapy, especially in monitoring CMV infection in immunossuppressed patients and in following the efficacy of antiviral treatment.

Identification of cytomegalovirus (CMV) infection by different laboratory methods in renal transplant recipients undergoing triple-drug immunosupressive treatment.

Early diagnosis of CMV infection is very important mainly in transplant recipients because CMV infection is a frequent complication after transplantation. In this work we compared different laboratory methods: ELISA (IgG, IgM), Western blot,shell vial, antigenemia assay (pp65), the immunofluorescent method with epithelial cells from urine (IF), DNA in leukocytes by PCR and DNA in leukocytes by hybridization (HCS) to estimate the most proper method for diagnosis of CMV in renal transplant recipients. This preliminary study showed that HCS, PCR and Western blot are sensitive methods for detecting CMV infection. Using HCS in quantitative variant we obtained a very good correlation between DNA load and clinical symptoms.

[Papillomaviruses and herpesviruses as selected risk factors in the etiopathogenesis of cervix neck cancer. I. Use of nucleic acid hybridization for diagnosing HPV infections]. / Wirusy papilloma i herpes oraz wybrane czynniki ryzyka w etiopatogenezie raka szyjki macicy. I. Zastosowanie hybrydyzacji kwasów nukleinowych do wykrywania zakazen HPV.

HPV infections are regarded as the main etiological factor responsible for the presence of cytological abnormalities and the primary risk factor for cervical cancer development. Diagnostic materials were collected between 1995 and 1997 in the Gynecologic Cancer Prevention, Cancer Center Institute of Oncology, Memorial Hospital Maria Sklodowska-Curie, Warsaw. The patients were divided into three groups: group C--women suspected for viral infection during clinical or cytological examination and two comparative groups A and B--women invited for routine cytological examination. Cytological smears for nucleic acid hybridisation collected before cytological smear sampling with a dacron swab (groups A and C) or collected after cytological smear sampling with a cervical brush (group B) were used. Cytological and clinical data was also used in the investigation. In 52% of the 236 samples tested by nucleic acid hybridisation HPV DNA was found to be present. DNA from the high/intermediate HPV risk group was also present in 36% of the samples and in 11% of the samples from the low risk HPV group. In 5% of the samples we confirmed the presence of mixed infections from both HPV risk groups. The results obtained from nucleic acid hybridisation with pap smear results were compared. It was observed that HPV infections from the high risk group occurred more frequently in pap 3 and pap 4 test results; HPV infections from the low risk group occurred more frequently in pap 1 and pap 2 results, while mixed infections from both risk groups occurred particularly in pap 4 and pap 5 results. Women in the 35-45 age groups suffered more frequently from infections from the high/intermediate risk group. In the 25-35 and 55-65 age groups HPV infections from the low risk group occurred more frequently. In the comparative groups only 2.6% of the women were infected with HPV.

[Papillomaviruses and herpesviruses as selected risk factors in the etiopathogenesis of cervix neck cancer. II. Use of PCR for isolating DNA of human papillomavirus and Herpes simplex]. / Wirusy papilloma i herpes oraz wybrane czynniki ryzyka w etiopatogenezie raka szyjki macicy. II. Zastosowanie PCR do wykrywania DNA wirusów brodawczaka czlowieka i Herpes simplex.

The aims of the study were to compare polymerase chain reaction PCR with nucleic acid hybridisation HC in the routine diagnosis of HPV infections. Smears collected for PCR were digested for 24 hours using proteinase K. After DNA extraction 174 samples were tested by PCR with human bglobin primers PG04-GH20. The PCR products were separated in 2% agarose gel electrophoresis stained with ethidium bromide. In 80.6% of the samples 256 base pair DNA fragments were observed in the gel in UV light. These samples were tested by PCR with HPV primers MY09-MY11. In 40% of the samples the presence of HPV DNA was confirmed. Next we carried out PCR using a mixture of two pairs of primers bglobin PG04-GH20 and HPV MY09-MY11. DNA for this study was extracted from 24 samples in which the presence of human DNA was not confirmed in the first PCR test and from 7 untested samples. In 21 cases HPV DNA was found to be present in gel electrophoresis. The presence of HPV DNA was confirmed in 44.75% of the samples.