Prevalence of Transmitted Drug-Resistance Mutations and Polymorphisms in HIV-1 Reverse Transcriptase, Protease, and gp41 Sequences Among Recent Seroconverters in Southern Poland.

BACKGROUND Monitoring of drug resistance-related mutations among patients with recent HIV-1 infection offers an opportunity to describe current patterns of transmitted drug resistance (TDR) mutations. MATERIAL AND METHODS Of 298 individuals newly diagnosed from March 2008 to February 2014 in southern Poland, 47 were deemed to have recent HIV-1 infection by the limiting antigen avidity immunoassay. Proviral DNA was amplified and sequenced in the reverse transcriptase, protease, and gp41 coding regions. Mutations were interpreted according to the Stanford Database algorithm and/or the International Antiviral Society USA guidelines. TDR mutations were defined according to the WHO surveillance list. RESULTS Among 47 patients with recent HIV-1 infection only 1 (2%) had evidence of TDR mutation. No major resistance mutations were found, but the frequency of strains with ≥1 accessory resistance-associated mutations was high, at 98%. Accessory mutations were present in 11% of reverse transcriptase, 96% of protease, and 27% of gp41 sequences. Mean number of accessory resistance mutations in the reverse transcriptase and protease sequences was higher in viruses with no compensatory mutations in the gp41 HR2 domain than in strains with such mutations (p=0.031). CONCLUSIONS Despite the low prevalence of strains with TDR mutations, the frequency of accessory mutations was considerable, which may reflect the history of drug pressure among transmitters or natural viral genetic diversity, and may be relevant for future clinical outcomes. The accumulation of the accessory resistance mutations within the pol gene may restrict the occurrence of compensatory mutations related to enfuvirtide resistance or vice versa.

Beauveria bassiana Water Extracts' Effect on the Growth of Wheat.

For a long time, entomopathogenic fungi were considered alternative biological control factors. Recently, these organisms were shown to fulfill additional roles supporting plants' development, improving their resistance to disease and survival under stress conditions. Considering the documented interactions of B. bassiana with a wide range of plants, we aimed to evaluate the impact of aqueous extracts of the fungus on the growth of an agriculturally significant plant-wheat. The usage of fungal extracts instead of fungi could be beneficial especially in unfavorable, environmentally speaking, regions. Selected dilutions of the crude extract obtained under different pH and temperature conditions were used to establish the optimal method of extraction. Plant growth parameters such as length, total fresh weight, and chlorophyll composition were evaluated. Additionally, the antibacterial activity of extracts was tested to exclude negative impacts on the beneficial soil microorganisms. The best results were obtained after applying extracts prepared at 25 °C and used at 10% concentration. Enhancement of the tested wheat's growth seems to be related to the composition of the extracts, which we documented as a rich source of macro- and microelements. Our preliminary results are the first confirming the potential of fungal water extracts as factors promoting plant growth. Further detailed investigation needs to be carried out to confirm the effects in real environment conditions. Additionally, the consistency of the plant growth stimulation across different entomopathogenic fungi and agriculturally used plant species should be tested.

Molecular epidemiology of recent HIV-1 infections in southern Poland.

The genetic diversity of human immunodeficiency virus type 1 (HIV-1) offers an opportunity to track the development of the epidemic across different populations. Viral pol gene fragments from 55 individuals of Polish origin with recent HIV-1 infection identified in 2008-2010 in four Polish cities were analyzed. Viral sequences were compared with sequences from 100 individuals (reference group) infected before 2004. Viral spread among groups with different HIV transmission categories was compared using a phylogenetic approach. The majority of sequences from individuals with recent infection were subtype B (93%) within which four transmission clusters (18% of samples) were detected. Samples from men infected through sex between men and from persons infected through injecting drugs were broadly separated (P < 0.0001), while samples from individuals infected by heterosexual contacts were dispersed uniformly within phylogenetic tree (P = 0.244) inferred from viral sequences derived from individuals infected recently and the reference group. The percentage of samples from persons infected by heterosexual contacts which clustered with samples from men infected through sex between men was not significantly higher for those with recent infection (47%), compared to the reference group (36%). In conclusion, men infected by sex between men and individuals infected through injecting drugs appear to form separate HIV transmission networks in Poland. The recent spread of HIV-1 among persons infected with subtype B by heterosexual contacts appears to be linked to both these groups.

Retrospective analysis of the HIV-1 reverse transcriptase inhibitors' resistance in Silesia, Poland.

BACKGROUND: This study was a retrospective analysis of drug resistance mutations among HIV-1 strains prevalent in Silesia, Poland, from the origin of the epidemic to 2004. The investigations included both type and frequency of the reverse transcriptase inhibitors' resistance mutations and estimation of the drugs' resistance levels. MATERIAL/METHODS: Proviral DNA, obtained from peripheral blood mononuclear cells of the 101 HIV-1-infected patients, was amplified and sequenced in the pol gene fragment covering the first 256 codons of the reverse transcriptase (RT). Reverse transcriptase inhibitors resistance mutations were determined and interpreted with the HIVdb: Genotypic Resistance Interpretation Algorithm available from the Stanford University HIV Drug Resistance Database. In the examined population, 35 subjects (34.7%) received no antiretroviral treatment by the time of specimen collection. RESULTS: The overall frequency of the RT inhibitors resistance mutations in the studied population was 15.8%. Substitutions related to the reverse transcriptase inhibitors resistance were identified in 10 pol gene sequences (9.9%), all of them were present in the HIV-1 sequences obtained from persons receiving antiretroviral therapy. CONCLUSIONS: Lack of drug-resistant viruses among treatment-naïve Silesian patients HIV-1-infected before the year 2004 may indicate that there was no transmission of the drug-resistant viruses in the studied population to that time.

Transmission patterns of HIV-1 non-R5 strains in Poland.

HIV-1 env sequencing enables predictions of viral coreceptor tropism and phylogenetic investigations of transmission events. The aim of the study was to estimate the contribution of non-R5 strains to the viral spread in Poland. Partial proviral env sequences were retrieved from baseline blood samples of patients with newly diagnosed HIV-1 infection between 2008-2014, including 46 patients with recent HIV-1 infection (RHI), and 246 individuals with long-term infection (LTHI). These sequences were subjected to the genotypic coreceptor tropism predictions and phylogenetic analyses to identify transmission clusters. Overall, 27 clusters with 57 sequences (19.5%) were detected, including 15 sequences (26.3%) from patients with RHI. The proportion of non-R5 strains among all study participants was 23.3% (68/292), and was comparable between patients with RHI and LTHI (11/46, 23.9% vs 57/246, 23.2%; p = 1.000). All 11 patients with non-R5 strains and RHI were men having sex with men (MSM). Among these patients, 4 had viral sequences grouped within phylogenetic cluster with another sequence of non-R5 strain obtained from patient with LTHI, indicating potential acquisition of non-R5 HIV-1 for at least 4/46 (8.7%) patients with RHI. We were unable to confirm the contribution of patients with RHI to the forward transmission of non-R5 strains, but a relatively high proportion of non-R5 strains among them deserves attention due to the limited susceptibility to CCR5 antagonists.

[Effects of CCR5-delta32, CCR2-64I and SDF-1-3'A polymorphic alleles on human immunodeficiency virus 1 (HIV-1) infection in the Polish population]. / Rola polimorficznych alleli CCR5-delta32, CCR2-64I i SDF-1-3'A w zakazeniu ludzkim wirusem uposledzenia odpornosci typu 1 (HIV-1) w populacji polskiej.

Among many factors that can influence vulnerability to infection and disease progression are genetic host factors together with the phenotype/genotype of the transmitting virus and the route of infection. Each of these factors alone or in combination could determine susceptibility to infection and subsequent rate of progression towards AIDS. Between host genetic factors identified and analyzed for their role in HIV-1 transmission and disease progression are polymorphisms in the genes encoding chemokine receptors and CCR5, CCR2 and SDF-1 a natural ligand for CXCR4 receptor. It has been shown that the distribution of this genetic polymorphisms and their role in the course of disease varies between different racial, ethnic and risk groups. The aim of present study was to examine the frequencies of polymorphic alleles CCR5-delta32, CCR2-64I and SDF-1-3'A and their role in human immunodeficiency virus (HIV-1) transmission in Polish population. The allelic and genotype distribution was studied in 103 HIV-1 infected patients (group HIV+) and 59 seronegative participants (group HIV-). Genotyping was done by the use of polymerase chain reaction with sequence-specific primers and restriction fragment length polymorphism. We found higher prevalence of CCR5-delta32 mutant allele among seronegative participants (13.6%) compared with HIV-infected patients (9.7%), although this did not attain statistical significance (p = 0.29). The CCR2-64I allelic frequency was almost identical in the HIV- and HIV+ groups (12.7% vs. 12.6%; respectively; p = 0.98). In contrast, the SDF-1-3'A allelic frequency was slightly lower among seronegative participants (15.3%) compared with HIV-infected patients (16.5%), and observed difference was not statistically significant (p = 0.77). Furthermore, we found that the genotype or allelic frequencies among HIV-1 infected patients were independent on the participant's sex, age at HIV-1 infection and the transmission route. Our results showed no significant differences in the prevalence of examined alleles and genotypes between HIV-1 infected patients and seronegative participants, which indicates that in the examined population they are not influencing host susceptibility to the HIV-1 infection.

[Coding sequence analysis of CYP1B1 exon III in the Polish population of patients with primary congenital and juvenile glaucoma]. / Analiza sekwencji kodujacej eksonu III genu CYP1B1 w polskiej populacji pacjentów chorych na jaskre wrodzona i mlodziencza.

Several mutations of CYP1B1 gene, a member of cytochrome P450 gene family, have been associated with occurrence of primary congenital glaucoma. The aim of presented work was to investigate CYP1B1 gene exon III mutations in 72 unrelated Polish patients with primary congenital glaucoma (C), juvenile glaucoma (J) and juvenile glaucoma suspects (P). In one patient we have detected homozygous duplication of 10bp at nucleotide 8046 resulting in synthesis of truncated protein, first found among the polish population. Furthermore, sequence analysis revealed three distinct single nucleotide polymorphisms at nucleotides 8131 (C-->G), 8184 (C-->T) and 8195 (A-->G,) in examined populations. The haplotypes in homozygous state (C/C/A, C/C/G, and G/T/A) were present in 52.6% cases. Since it is suggested that the presence of these haplotypes may be linked to mutations in exon II of the CYP1B1 gene, we plan to analyze that exon in the future.

Prevalence of tick-borne encephalitis virus in a highly urbanized and low risk area in Southern Poland.

The knowledge of the exact distribution of tick-borne encephalitis virus (TBEV) endemic foci is crucial to plan and implement the optimal prevention of tick-borne encephalitis (TBE), including a vaccination program. In Poland, however, there is still no data on the local distribution of TBEV in many areas of the country. Silesian agglomeration area (Southern Poland) is a highly urbanized and industrialized region of the country, where TBE cases are only sporadically recorded. In this study, a total of 4350 adult Ixodes ricinus were collected between September 2010 and June 2012 at twelve locations. The screening using real-time PCR was carried out on 854 tick pools of five specimens, and the positive pools were verified by pyrosequencing. TBEV was identified in 13 pools (1.52%) at 4 sites, of which 9 pools were verified by pyrosequencing. An overall pool prevalence was estimated at 0.31% ranging from 0.19% to 1.11% for positive locations [95% CI 0.16-0.52], which is comparable with regions with high number of TBE cases reported annually.

Transmission of drug-resistant HIV-1 variants among individuals with recent infection in southern Poland.

Transmitted drug resistance (TDR) is an important public health issue, because it may affect the outcome of antiretroviral treatment. The prevalence of human immunodeficiency virus type 1 (HIV-1) with TDR mutations defined according to the list of the World Health Organization was investigated among 53 therapy-naïve persons with confirmed recent HIV-1 infection diagnosed in Poland, in the years 2008-2010. Proviral DNA was amplified, sequenced, and screened for the TDR mutations in the pol gene fragments coding for the whole protease and the initial 256 residues of the reverse transcriptase. The frequency of sequences with at least one TDR mutation was 11.3%. In four (7.5%) sequences at least one resistance mutation related to reverse transcriptase inhibitors was identified, and in further two (3.8%) sequences one mutation related to protease inhibitors' resistance was present. The moderate rate of TDR highlights the need for a continuous surveillance and resistance testing among treatment-naïve individuals to optimize treatment effects within a country.

HIV type 1 genetic diversity in Silesia, Poland: a retrospective analysis.

To characterize the genetic diversity of HIV-1 strains circulating among patients with different transmission risk behaviors in Silesia, Poland, from the origin of the epidemic to the year 2004, we have sequenced and analyzed the p24 coding region of the gag gene and part of the pol gene covering the first 256 codons for the reverse transcriptase (RT). The proviral DNA was obtained from the 101 HIV-1-infected patients, 80 of whom (79.2%) were intravenous drug users (IDUs) and 21 of whom (20.8%) reported sexual transmission risk practices (STs) with 11 (10.9%) being heterosexuals and 10 (9.9%) being homosexual men, which corresponds to the population's epidemiological data. All of the investigated viral sequences were classified as HIV-1 subtype B with low genetic heterogeneity. There was an association between HIV-1 genetic diversity and the risk of virus transmission in the investigated population. The mean nucleotide distances were significantly lower among sequences derived from IDUs than among sequences obtained from STs. Additionally, strains present among IDUs, as opposed to viruses circulating among STs, were genetically more distinct from HIV-1 subtype B strains found in other populations worldwide. Our findings that HIV-1 strains circulating among IDUs were closely related to each other, but were distinct from viruses prevalent in other geographic regions, allow further tracing of the spread of these strains.