Calcifying Bacteria Flexibility in Induction of CaCO3 Mineralization.

Microbially induced CaCO3 precipitation (MICP) is considered as an alternative green technology for cement self-healing and a basis for the development of new biomaterials. However, some issues about the role of bacteria in the induction of biogenic CaCO3 crystal nucleation, growth and aggregation are still debatable. Our aims were to screen for ureolytic calcifying microorganisms and analyze their MICP abilities during their growth in urea-supplemented and urea-deficient media. Nine candidates showed a high level of urease specific activity, and a sharp increase in the urea-containing medium pH resulted in efficient CaCO3 biomineralization. In the urea-deficient medium, all ureolytic bacteria also induced CaCO3 precipitation although at lower pH values. Five strains (B. licheniformis DSMZ 8782, B. cereus 4b, S. epidermidis 4a, M. luteus BS52, M. luteus 6) were found to completely repair micro-cracks in the cement samples. Detailed studies of the most promising strain B. licheniformis DSMZ 8782 revealed a slower rate of the polymorph transformation in the urea-deficient medium than in urea-containing one. We suppose that a ureolytic microorganism retains its ability to induce CaCO3 biomineralization regardless the origin of carbonate ions in a cell environment by switching between mechanisms of urea-degradation and metabolism of calcium organic salts.

Understanding complex dynamics of behavioral, neurochemical and transcriptomic changes induced by prolonged chronic unpredictable stress in zebrafish.

Stress-related neuropsychiatric disorders are widespread, debilitating and often treatment-resistant illnesses that represent an urgent unmet biomedical problem. Animal models of these disorders are widely used to study stress pathogenesis. A more recent and historically less utilized model organism, the zebrafish (Danio rerio), is a valuable tool in stress neuroscience research. Utilizing the 5-week chronic unpredictable stress (CUS) model, here we examined brain transcriptomic profiles and complex dynamic behavioral stress responses, as well as neurochemical alterations in adult zebrafish and their correction by chronic antidepressant, fluoxetine, treatment. Overall, CUS induced complex neurochemical and behavioral alterations in zebrafish, including stable anxiety-like behaviors and serotonin metabolism deficits. Chronic fluoxetine (0.1 mg/L for 11 days) rescued most of the observed behavioral and neurochemical responses. Finally, whole-genome brain transcriptomic analyses revealed altered expression of various CNS genes (partially rescued by chronic fluoxetine), including inflammation-, ubiquitin- and arrestin-related genes. Collectively, this supports zebrafish as a valuable translational tool to study stress-related pathogenesis, whose anxiety and serotonergic deficits parallel rodent and clinical studies, and genomic analyses implicate neuroinflammation, structural neuronal remodeling and arrestin/ubiquitin pathways in both stress pathogenesis and its potential therapy.

A Comparative Characterization of the Mitochondrial Genomes of Paramoeba aparasomata and Neoparamoeba pemaquidensis (Amoebozoa, Paramoebidae).

Marine amebae of the genus Paramoeba (Amoebozoa, Dactylopodida) normally contain a eukaryotic endosymbiont known as Perkinsela-like organism (PLO). This is one of the characters to distinguish the genera Neoparamoeba and Paramoeba from other Dactylopodida. It is known that the PLO may be lost, but PLO-free strains of paramoebians were never available for molecular studies. Recently, we have described the first species of the genus Paramoeba which has no parasome-Paramoeba aparasomata. In this study, we present a mitochondrial genome of this species, compare it with that of Neoparamoeba pemaquidensis, and analyze the evolutionary dynamics of gene sequences and gene order rearrangements between these species. The mitochondrial genome of P. aparasomata is 46,254 bp long and contains a set of 31 protein-coding genes, 19 tRNAs, two rRNA genes, and 7 open reading frames. Our results suggest that these two mitochondrial genomes within the genus Paramoeba have rather similar organization and gene order, base composition, codon usage, the composition and structure of noncoding, and overlapping regions.

The Increase of the Magnitude of Spontaneous Viral Blips in Some Participants of Phase II Clinical Trial of Therapeutic Optimized HIV DNA Vaccine Candidate.

We developed a candidate DNA vaccine called "DNA-4"consisting of 4 plasmid DNAs encoding Nef, Gag, Pol(rt), and gp140 HIV-1 proteins. The vaccine was found to be safe and immunogenic in a phase I clinical trial. Here we present the results of a phase II clinical trial of "DNA-4". This was a multicenter, double-blind, placebo-controlled clinical trial of safety, and dose selection of "DNA-4" in HIV-1 infected people receiving antiretroviral therapy (ART). Fifty-four patients were randomized into 3 groups (17 patients-group DNA-4 0.25 mg, 17 patients-group DNA-4 0.5 mg, 20 patients-the placebo group). All patients were immunized 4 times on days 0, 7, 11, and 15 followed by a 24-week follow-up period. "DNA-4" was found to be safe and well-tolerated at doses of 0.25 mg and 0.5 mg. We found that the amplitudes of the spontaneous viral load increases in three patients immunized with the candidate DNA vaccine were much higher than that in placebo group-2800, 180,000 and 709 copies/mL, suggesting a possible influence of therapeutic DNA vaccination on viral reservoirs in some patients on ART. We hypothesize that this influence was associated with the reactivation of proviral genomes.

The complete mitochondrial genome of Paravannella minima (Amoebozoa, Discosea, Vannellida).

We present a complete sequence and describe the organization of the mitochondrial genome of the amoeba Paravannella minima (Amoebooza, Discosea, Vannellida). This tiny species represents a branch at the base of Vannellida tree, to the moment being its earliest-branching lineage. The circular mitochondrial DNA of this species has 53,464 bp in length and contains 30 protein-coding genes, 2 ribosomal RNAs, 23 transfer RNAs, and 15 open reading frames. This genome is significantly longer and contains more protein-coding genes than any yet sequenced mitochondrial genome of vannellid amoebae. Unlike the previously sequenced mitochondrial genomes of Vannellida, which should be translated using the "Table 4" (the mold, protozoan, and coelenterate mitochondrial code), that of P. minima can be properly translated using the universal genetic code.

Scytalidium candidum 3C is a new name for the Geotrichum candidum Link 3C strain.

In the 1970s, the strain Geotrichum candidum Link 3C was isolated from rotting rope and since then has been extensively studied as a source of cellulose and xylan-degrading enzymes. The original identification of the strain was based only on morphological characters of the fungal mycelium in culture. Recent comparison of the internal transcribed spacer (ITS) fragments derived from the draft genome published in 2015 did not show its similarity to G. candidum species. Given the value of the strain 3C in lignocellulosic biomass degradation, we performed morphological and molecular studies to find the appropriate taxonomic placement for this fungal strain within the Ascomycota phylum. ITS, 18S rDNA, 28S rDNA sequences, and RPB2 encoding genes were used to construct phylogenetic trees with Maximum likelihood and Bayesian inference methods. Based on sequence comparison and multiple gene sequencing, we conclude that the fungal strain designated as Geotrichum candidum Link 3C should be placed into the genus Scytalidium (Pezizomycotina, Leotiomycetes) and is redescribed herein as Scytalidium candidum 3C comb. nov.

Characterization of the Transmitted Virus in an Ongoing HIV-1 Epidemic Driven by Injecting Drug Use.

Understanding features of the HIV-1 transmission process has the potential to inform biological interventions for prevention. We have examined the transmitted virus in a cohort of people who inject drugs and who are at risk of HIV-1 infection through blood contamination when injecting in a group. This study focused on seven newly infected participants in St. Petersburg, Russia, who were in acute or early infection. We used end-point dilution polymerase chain reaction to amplify single viral genomes to assess the complexity of the transmitted virus. We also used deep sequencing to further assess the complexity of the virus. We interpret the results as indicating that a single viral variant was transmitted in each case, consistent with a model where the exposure to virus during transmission was limited. We also looked at phenotypic properties of the viral Env protein in isolates from acute and chronic infection. Although differences were noted, there was no consistent pattern that distinguished the transmitted variants. Similarly, despite the reduced genetic heterogeneity of the more recent subtype A HIV-1 epidemic in St. Petersburg, we did not see reduced variance in the neutralization properties compared to isolates from the more mature subtype C HIV-1 epidemic. Finally, in looking at members of injecting groups related to the acute HIV-1 infection/early subjects, we found examples of sequence linkage consistent with ongoing and rapid spread of HIV-1 in these groups. These studies emphasize the dynamic nature of this epidemic and reinforce the idea that improved prevention methods are needed.

Mitochondrial Genome of Vannella croatica (Amoebozoa, Discosea, Vannellida).

Mitochondrial genome sequence of Vannella croatica (Amoebozoa, Discosea, Vannellida) was obtained using pulse-field gel electrophoretic isolation of the circular mitochondrial DNA, followed by the next-generation sequencing. The mitochondrial DNA of this species has the length of 28,933 bp and contains 12 protein-coding genes, two ribosomal RNAs, and 16 transfer RNAs. Vannella croatica mitochondrial genome is relatively short compared to other known amoebozoan mitochondrial genomes but is rather gene-rich and contains significant number of open reading frames.

The complete mitochondrial genome of Vannella simplex (Amoebozoa, Discosea, Vannellida).

Vannella simplex (Amoebozoa, Discosea, Vannellida) is one of the commonest freshwater free-living lobose amoebae, known from many locations worldwide. In the present study, we describe the complete mitochondrial genome of this species. The circular mitochondrial DNA of V. simplex has 34,145öbp in length and contains 27 protein-coding genes, 2 ribosomal RNAs, 16 transfer RNAs and 4 open reading frames. Mitochondiral genome of V. simplex is one of the most gene compact due to overlapping genes and reduced intergenic space. It has much in common with its closest relative, mitochondrial genome of V. croatica GenBank number MF508648. In the same time, both of them show considerable differences in length and in gene order from the next close relative - that of Neoparamoeba pemaquidensis KX611830 (deposited as Paramoeba) and even more - from other sequenced amoebozoan mitochondrial genomes. The present study confirms the opinion that the level of synteny between the mitochondrial genomes across the entire Amoebozoa clade is low. More or less considerable similarity yet was found only between members of the same clade of the genera or family level, but hardly - among more distant lineages.

Disruption of Transcriptional Coactivator Sub1 Leads to Genome-Wide Re-distribution of Clustered Mutations Induced by APOBEC in Active Yeast Genes.

Mutations in genomes of species are frequently distributed non-randomly, resulting in mutation clusters, including recently discovered kataegis in tumors. DNA editing deaminases play the prominent role in the etiology of these mutations. To gain insight into the enigmatic mechanisms of localized hypermutagenesis that lead to cluster formation, we analyzed the mutational single nucleotide variations (SNV) data obtained by whole-genome sequencing of drug-resistant mutants induced in yeast diploids by AID/APOBEC deaminase and base analog 6-HAP. Deaminase from sea lamprey, PmCDA1, induced robust clusters, while 6-HAP induced a few weak ones. We found that PmCDA1, AID, and APOBEC1 deaminases preferentially mutate the beginning of the actively transcribed genes. Inactivation of transcription initiation factor Sub1 strongly reduced deaminase-induced can1 mutation frequency, but, surprisingly, did not decrease the total SNV load in genomes. However, the SNVs in the genomes of the sub1 clones were re-distributed, and the effect of mutation clustering in the regions of transcription initiation was even more pronounced. At the same time, the mutation density in the protein-coding regions was reduced, resulting in the decrease of phenotypically detected mutants. We propose that the induction of clustered mutations by deaminases involves: a) the exposure of ssDNA strands during transcription and loss of protection of ssDNA due to the depletion of ssDNA-binding proteins, such as Sub1, and b) attainment of conditions favorable for APOBEC action in subpopulation of cells, leading to enzymatic deamination within the currently expressed genes. This model is applicable to both the initial and the later stages of oncogenic transformation and explains variations in the distribution of mutations and kataegis events in different tumor cells.

Two Independent HIV Epidemics in Saint Petersburg, Russia Revealed by Molecular Epidemiology.

The HIV epidemic in Russia, one of the world's fastest growing, has been concentrated mostly among people who inject drugs (PWID). We sought to explore the epidemiology of the epidemic in St. Petersburg by sampling from the highest risk groups of PWID and men who have sex with men (MSM) and use viral sequencing data to better understand the nature of the city's epidemic. Serological testing confirmed an HIV prevalence among PWID in excess of 40%. All but 1 of 110 PWID whose blood samples were tested for genetic diversity were infected by subtype A virus, specifically by the AFSU strain. The remaining person was infected with a CRF-06cpx recombinant. Analysis of pairwise genetic distance among all PWID studied revealed an average of 3.1% sequence divergence, suggesting clonal introduction of the AFSU strain and/or constraints on sequence divergence. The HIV prevalence was less than 10% among MSM. All 17 sequences from HIV-infected MSM were found to be a clade B virus with a much higher average sequence diversity of 15.7%. These findings suggest two independent epidemics with little overlap between the two highest at-risk populations, which will require different HIV prevention approaches.

A substantial transmission bottleneck among newly and recently HIV-1-infected injection drug users in St Petersburg, Russia.

There are limited data on the genetic complexity of human immunodeficiency virus type 1 (HIV-1) after transmission among a cohort of injection drug users (IDUs). We used single-genome amplification of HIV-1 env to determine the genotypic characteristics of virus among IDUs with acute infection in St Petersburg, Russia. Our results indicate that a single variant was transmitted in a majority of cases (9 of 13 participants), which is analogous to what is observed in sexual transmission. These data are most consistent with a genetic bottleneck during transmission by injection drug use that is due to a small inoculum, which most often results in the transmission of a low-complexity viral population.

The feasibility of HIV vaccine efficacy trials among Russian injection drug users.

IDU exposure remains a primary driver of the Russian HIV epidemic, and recent incidence data provide little evidence that this epidemic is slowing. While there are multiple important challenges that need to be further explored before starting vaccine trials, most importantly access to evidence-based drug treatment services for trial participants, the current context of high HIV incidence and low genetic diversity of HIV strains, suggests the need for intensified prevention strategies and supports the feasibility of mounting efficacy trials of HIV vaccines among IDUs in the Russian Federation.

Immunogenicity of candidate DNA vaccine based on subtype A of human immunodeficiency virus type 1 predominant in Russia.

Human immunodeficiency virus (HIV)-1 subtype A strains circulating among the majority of HIVinfected individuals in the former Soviet Union (FSU) countries demonstrate low genetic diversity. The consensus sequence of the FSU region-specific isolate has been used for the candidate DNA vaccine development. We constructed recombinant plasmids with four viral genes: env (gp140), gag, pol (reverse transcriptase), and nef. We immunized BALB/c mice intramuscularly using equimolar mixture of four recombinant plasmids, and observed significant cytotoxic T lymphocyte response and specific CD8(+) cell production against cells presenting HIV-1 peptides. Overall, the Th1 pathway of immune response clearly dominated. Immunological properties of this candidate DNA vaccine against HIV-1 suggest the possibility of its further study in clinical trials.

The rate of epidemiological and virological changes during the transition from nascent to concentrated HIV epidemic stage in the former Soviet Union countries.

The rate of processes accompanying the transition of the HIV-1 epidemic from nascent stage to concentrated one in the Former Soviet Union (FSU) during intravenous drug user (IDU)-associated HIV infection outbreaks in 1994-1999 has not been analyzed. To define the rates, we studied susceptible populations and circulating viruses before, during, and after the outbreaks. Our findings included the following: (1) the pattern of high HIV-1 genetic diversity characteristic of the nascent epidemic changed to a concentrated one within 1 year in St. Petersburg and in Moscow; (2) different FSU regions were at different stages of the HIV-1 epidemic in 1994-1996; (3) the change of serotypic patterns characteristic of different stages of the HIV/AIDS epidemic for the non-IDU risk group occurred within 1 year in Moscow, suggesting an extremely high rate of IDU-associated epidemic pattern distributions in regions and susceptible populations in the FSU.