Epigenetics and transgenerational inheritance.

Epigenetic modifications can alter the function of genes. The epigenetics changes are caused by environmental effects, which lead to chemical modifications of the DNA or the chromatin. The mechanisms involve the influence of small interfering siRNAs on gene silencing. Epigenetic changes normally last only during the life-time of an individual and are erased in embryos and eggs for a naive progeny. The genomes are reprogrammed and the chemical modifications removed to restart the next generation. However, there are mechanisms that allow the genome to escape from such a clearing effect so that modifications can be transmitted to one or more subsequent generations. In the germline of animal cells small RNAs, including piRNAs, have evolved which guarantee a higher degree of fidelity for transmission of genetic information, guarding especially against the detrimental effect caused by transposon activity. piRNA is essential for transposon silencing for survival of a species and protection of subsequent generations. Inactivation of piRNA results in abundant transposon activity and sperm infertility. The effect in humans has been described but is less distinct. Some stress-induced epigenetic changes are transitory in mice and can be reversed by a change of environment or lifestyle.

Phage Therapy in Germany-Update 2023.

Bacteriophage therapy holds promise in addressing the antibiotic-resistance crisis, globally and in Germany. Here, we provide an overview of the current situation (2023) of applied phage therapy and supporting research in Germany. The authors, an interdisciplinary group working on patient-focused bacteriophage research, addressed phage production, phage banks, susceptibility testing, clinical application, ongoing translational research, the regulatory situation, and the network structure in Germany. They identified critical shortcomings including the lack of clinical trials, a paucity of appropriate regulation and a shortage of phages for clinical use. Phage therapy is currently being applied to a limited number of patients as individual treatment trials. There is presently only one site in Germany for large-scale good-manufacturing-practice (GMP) phage production, and one clinic carrying out permission-free production of medicinal products. Several phage banks exist, but due to varying institutional policies, exchange among them is limited. The number of phage research projects has remarkably increased in recent years, some of which are part of structured networks. There is a demand for the expansion of production capacities with defined quality standards, a structured registry of all treated patients and clear therapeutic guidelines. Furthermore, the medical field is still poorly informed about phage therapy. The current status of non-approval, however, may also be regarded as advantageous, as insufficiently restricted use of phage therapy without adequate scientific evidence for effectiveness and safety must be prevented. In close coordination with the regulatory authorities, it seems sensible to first allow some centers to treat patients following the Belgian model. There is an urgent need for targeted networking and funding, particularly of translational research, to help advance the clinical application of phages.

The Roles of the Virome in Cancer.

Viral infections as well as changes in the composition of the intestinal microbiota and virome have been linked to cancer. Moreover, the success of cancer immunotherapy with checkpoint inhibitors has been correlated with the intestinal microbial composition of patients. The transfer of feces-which contain mainly bacteria and their viruses (phages)-from immunotherapy responders to non-responders, known as fecal microbiota transplantation (FMT), has been shown to be able to convert some non-responders to responders. Since phages may also increase the response to immunotherapy, for example by inducing T cells cross-reacting with cancer antigens, modulating phage populations may provide a new avenue to improve immunotherapy responsiveness. In this review, we summarize the current knowledge on the human virome and its links to cancer, and discuss the potential utility of bacteriophages in increasing the responder rate for cancer immunotherapy.

Viroids and the Origin of Life.

Viroids are non-coding circular RNA molecules with rod-like or branched structures. They are often ribozymes, characterized by catalytic RNA. They can perform many basic functions of life and may have played a role in evolution since the beginning of life on Earth. They can cleave, join, replicate, and undergo Darwinian evolution. Furthermore, ribozymes are the essential elements for protein synthesis of cellular organisms as parts of ribosomes. Thus, they must have preceded DNA and proteins during evolution. Here, we discuss the current evidence for viroids or viroid-like RNAs as a likely origin of life on Earth. As such, they may also be considered as models for life on other planets or moons in the solar system as well as on exoplanets.

Within-Host and Between-Host Evolution in SARS-CoV-2-New Variant's Source.

Some of the newly emerging corona viral variants show high numbers of mutations. This is unexpected for a virus with a low mutation rate due to an inherent proof-reading system. Could such a variant arise under very special conditions occurring in a host where the virus replicates and mutates in a rather unlimited fashion, such as in immune compromised patients? The virus was shown to replicate in an immunosuppressed cancer patient for more than 105 days and might be a source of new variants. These patients are asymptomatic and the virus may therefore escape detection and attention and be high-risk. Similarly, HIV-infected individuals may be immunocompromised and support coronavirus replication with increased mutation rates. The patients may promote "within-host evolution". Some of the viruses present in such a highly mutagenic swarm or quasispecies within one patient may become founders and cause a pandemic by further "between-host evolution". B.1.1.7 with 23 mutations may be such a case. Immunosuppressed patients can be identified and treated by the synthetic antibody cocktails as passive immunization and kept under control. Immunosuppressed patients can be easily identified and supervised by healthcare workers-once they become aware of the risk-to avoid new variants with pandemic potential.

Phages Needed against Resistant Bacteria.

Phages have been known for more than 100 years. They have been applied to numerous infectious diseases and have proved to be effective in many cases. However, they have been neglected due to the era of antibiotics. With the increase of antibiotic-resistant microorganisms, we need additional therapies. Whether or not phages can fulfill this expectation needs to be verified and tested according to the state-of-the-art of international regulations. These regulations fail, however, with respect to GMP production of phages. Phages are biologicals, not chemical compounds, which cannot be produced under GMP regulations. This needs to be urgently changed to allow progress to determine how phages can enter routine clinical settings.

Air Microbiome and Pollution: Composition and Potential Effects on Human Health, Including SARS Coronavirus Infection.

Polluted air poses a significant threat to human health. Exposure to particulate matter (PM) and harmful gases contributes to cardiovascular and respiratory diseases, including allergies and obstructive lung disease. Air pollution may also be linked to cancer and reduced life expectancy. Uptake of PM has been shown to cause pathological changes in the intestinal microbiota in mice and humans. Less is known about the effects of pollution-associated microbiota on human health. Several recent studies described the microbiomes of urban and rural air samples, of the stratosphere and sand particles, which can be transported over long distances, as well as the air of indoor environments. Here, we summarize the current knowledge on airborne bacterial, viral, and fungal communities and discuss their potential consequences on human health. The current data suggest that bacterial pathogens are typically too sparse and short-lived in air to pose a significant risk for infecting healthy people. However, airborne fungal spores may exacerbate allergies and asthma. Little information is available on viruses including phages, and future studies are likely to detect known and novel viruses with a yet unknown impact on human health. Furthermore, varying experimental protocols have been employed in the recent microbiome and virome studies. Therefore, standardized methodologies will be required to allow for better comparisons between studies. Air pollution has been linked to more severe outcomes of SARS (severe acute respiratory syndrome) coronavirus (SARS-CoV) infections. This may have contributed to severe SARS-CoV-2 outbreaks, especially those in China, Northern Italy, Iran, and New York City.

Viruses and Evolution - Viruses First? A Personal Perspective.

The discovery of exoplanets within putative habitable zones revolutionized astrobiology in recent years. It stimulated interest in the question about the origin of life and its evolution. Here, we discuss what the roles of viruses might have been at the beginning of life and during evolution. Viruses are the most abundant biological entities on Earth. They are present everywhere, in our surrounding, the oceans, the soil and in every living being. Retroviruses contributed to about half of our genomic sequences and to the evolution of the mammalian placenta. Contemporary viruses reflect evolution ranging from the RNA world to the DNA-protein world. How far back can we trace their contribution? Earliest replicating and evolving entities are the ribozymes or viroids fulfilling several criteria of life. RNA can perform many aspects of life and influences our gene expression until today. The simplest structures with non-protein-coding information may represent models of life built on structural, not genetic information. Viruses today are obligatory parasites depending on host cells. Examples of how an independent lifestyle might have been lost include mitochondria, chloroplasts, Rickettsia and others, which used to be autonomous bacteria and became intracellular parasites or endosymbionts, thereby losing most of their genes. Even in vitro the loss of genes can be recapitulated all the way from coding to non-coding RNA. Furthermore, the giant viruses may indicate that there is no sharp border between living and non-living entities but an evolutionary continuum. Here, it is discussed how viruses can lose and gain genes, and that they are essential drivers of evolution. This discussion may stimulate the thinking about viruses as early possible forms of life. Apart from our view "viruses first", there are others such as "proteins first" and "metabolism first."

What viruses tell us about evolution and immunity: beyond Darwin?

We describe mechanisms of genetic innovation mediated by viruses and related elements that, during evolution, caused major genetic changes beyond what was anticipated by Charles Darwin. Viruses and related elements introduced genetic information and have shaped the genomes and immune systems of all cellular life forms. None of these mechanisms contradict Darwin's theory of evolution but extend it by means of sequence information that has recently become available. Not only do small increments of genetic information contribute to evolution, but also do major events such as infection by viruses or bacteria, which can supply new genetic information to a host by horizontal gene transfer. Thereby, viruses and virus-like elements act as major drivers of evolution.

Evolution of Immune Systems From Viruses and Transposable Elements.

Virus-derived sequences and transposable elements constitute a substantial portion of many cellular genomes. Recent insights reveal the intimate evolutionary relationship between these sequences and various cellular immune pathways. At the most basic level, superinfection exclusion may be considered a prototypical virus-mediated immune system that has been described in both prokaryotes and eukaryotes. More complex immune mechanisms fully or partially derived from mobile genetic elements include CRISPR-Cas of prokaryotes and the RAG1/2 system of vertebrates, which provide immunological memory of foreign genetic elements and generate antibody and T cell receptor diversity, respectively. In this review, we summarize the current knowledge on the contribution of mobile genetic elements to the evolution of cellular immune pathways. A picture is emerging in which the various cellular immune systems originate from and are spread by viruses and transposable elements. Immune systems likely evolved from simple superinfection exclusion to highly complex defense strategies.

A Wake-Up Call: We Need Phage Therapy Now.

The rise of multidrug-resistant bacteria has resulted in an increased interest in phage therapy, which historically preceded antibiotic treatment against bacterial infections. To date, there have been no reports of serious adverse events caused by phages. They have been successfully used to cure human diseases in Eastern Europe for many decades. More recently, clinical trials and case reports for a variety of indications have shown promising results. However, major hurdles to the introduction of phage therapy in the Western world are the regulatory and legal frameworks. Present regulations may take a decade or longer to be fulfilled. It is of urgent need to speed up the availability of phage therapy.

RNase H As Gene Modifier, Driver of Evolution and Antiviral Defense.

Retroviral infections are 'mini-symbiotic' events supplying recipient cells with sequences for viral replication, including the reverse transcriptase (RT) and ribonuclease H (RNase H). These proteins and other viral or cellular sequences can provide novel cellular functions including immune defense mechanisms. Their high error rate renders RT-RNases H drivers of evolutionary innovation. Integrated retroviruses and the related transposable elements (TEs) have existed for at least 150 million years, constitute up to 80% of eukaryotic genomes and are also present in prokaryotes. Endogenous retroviruses regulate host genes, have provided novel genes including the syncytins that mediate maternal-fetal immune tolerance and can be experimentally rendered infectious again. The RT and the RNase H are among the most ancient and abundant protein folds. RNases H may have evolved from ribozymes, related to viroids, early in the RNA world, forming ribosomes, RNA replicases and polymerases. Basic RNA-binding peptides enhance ribozyme catalysis. RT and ribozymes or RNases H are present today in bacterial group II introns, the precedents of TEs. Thousands of unique RTs and RNases H are present in eukaryotes, bacteria, and viruses. These enzymes mediate viral and cellular replication and antiviral defense in eukaryotes and prokaryotes, splicing, R-loop resolvation, DNA repair. RNase H-like activities are also required for the activity of small regulatory RNAs. The retroviral replication components share striking similarities with the RNA-induced silencing complex (RISC), the prokaryotic CRISPR-Cas machinery, eukaryotic V(D)J recombination and interferon systems. Viruses supply antiviral defense tools to cellular organisms. TEs are the evolutionary origin of siRNA and miRNA genes that, through RISC, counteract detrimental activities of TEs and chromosomal instability. Moreover, piRNAs, implicated in transgenerational inheritance, suppress TEs in germ cells. Thus, virtually all known immune defense mechanisms against viruses, phages, TEs, and extracellular pathogens require RNase H-like enzymes. Analogous to the prokaryotic CRISPR-Cas anti-phage defense possibly originating from TEs termed casposons, endogenized retroviruses ERVs and amplified TEs can be regarded as related forms of inheritable immunity in eukaryotes. This survey suggests that RNase H-like activities of retroviruses, TEs, and phages, have built up innate and adaptive immune systems throughout all domains of life.

Stable core virome despite variable microbiome after fecal transfer.

We recently described the 4.5-year time course of the enteric bacterial microbiota and virome of a patient cured from recurrent Clostridium difficile infection (rCDI) by fecal microbiota transplantation (FMT). Here, we extended the virome analyses and found the patient's phage population to exhibit highly donor-similar characteristics following FMT, which remained stable for the whole period tested (up to 7 months). Moreover, the detected viral populations of donor and patient exhibited comparable diversity and richness. These findings were unexpected since enteric viromes are normally highly variable, assumed to influence the bacterial host community and change with environmental conditions. In contrast to the virome, the bacterial microbiota varied indeed for more than 7 months with ongoing dysbiosis before it reached donor similarity. Our findings that are based on sequence information and protein domain analysis seem to suggest that stable phage properties correlate with successful FMT better than the changing bacterial communities. We speculate that we here preferentially detected a stable core virome, which dominated over a variable flexible virome that may have been too heterogeneous for experimental detection or was underrepresented in the databases. It will be interesting to analyze whether the enteric virome allows predictions for the clinical outcome of FMT for rCDI and other diseases such as inflammatory bowel disease or obesity.

The intron-enriched HERV-K(HML-10) family suppresses apoptosis, an indicator of malignant transformation.

BACKGROUND: Human endogenous retroviruses (HERVs) constitute 8% of the human genome and contribute substantially to the transcriptome. HERVs have been shown to generate RNAs that modulate host gene expression. However, experimental evidence for an impact of these regulatory transcripts on the cellular phenotype has been lacking. RESULTS: We characterized the previously little described HERV-K(HML-10) endogenous retrovirus family on a genome-wide scale. HML-10 invaded the ancestral genome of Old World monkeys about 35 Million years ago and is enriched within introns of human genes when compared to other HERV families. We show that long terminal repeats (LTRs) of HML-10 exhibit variable promoter activity in human cancer cell lines. One identified HML-10 LTR-primed RNA was in opposite orientation to the pro-apoptotic Death-associated protein 3 (DAP3). In HeLa cells, experimental inactivation of HML-10 LTR-primed transcripts induced DAP3 expression levels, which led to apoptosis. CONCLUSIONS: Its enrichment within introns suggests that HML-10 may have been evolutionary co-opted for gene regulation more than other HERV families. We demonstrated such a regulatory activity for an HML-10 RNA that suppressed DAP3-mediated apoptosis in HeLa cells. Since HML-10 RNA appears to be upregulated in various tumor cell lines and primary tumor samples, it may contribute to evasion of apoptosis in malignant cells. However, the overall weak expression of HML-10 transcripts described here raises the question whether our result described for HeLa represent a rare event in cancer. A possible function in other cells or tissues requires further investigation.

Abolishing HIV-1 infectivity using a polypurine tract-specific G-quadruplex-forming oligonucleotide.

BACKGROUND: HIV is primarily transmitted by sexual intercourse and predominantly infects people in Third World countries. Here an important medical need is self-protection for women, particularly in societies where condoms are not widely accepted. Therefore, availability of antiviral microbicides may significantly reduce sexual HIV transmission in such environments. METHODS: Here, we investigated structural characteristics and the antiviral activity of the polypurine tract (PPT)-specific ODN A, a 54-mer oligodeoxynucleotide (ODN) that has been previously shown to trigger the destruction of viral RNA genomes by prematurely activating the retroviral RNase H. The stability of ODN A and mutants thereof was tested at various storage conditions. Furthermore, antiviral effects of ODN A were analyzed in various tissue culture HIV-1 infection models. Finally, circular dichroism spectroscopy was employed to gain insight into the structure of ODN A. RESULTS: We show here that ODN A is a powerful tool to abolish HIV-1 particle infectivity, as required for a candidate compound in vaginal microbicide applications. We demonstrate that ODN A is not only capable to prematurely activate the retroviral RNase H, but also prevents HIV-1 from entering host cells. ODN A also exhibited extraordinary stability lasting several weeks. Notably, ODN A is biologically active under various storage conditions, as well as in the presence of carboxymethylcellulose CMC (K-Y Jelly), a potential carrier for application as a vaginal microbicide. ODN A's remarkable thermostability is apparently due to its specific, guanosine-rich sequence. Interestingly, these residues can form G-quadruplexes and may lead to G-based DNA hyperstructures. Importantly, the pronounced antiviral activity of ODN A is maintained in the presence of human semen or semen-derived enhancer of virus infection (SEVI; i.e. amyloid fibrils), both known to enhance HIV infectivity and reduce the efficacy of some antiviral microbicides. CONCLUSIONS: Since ODN A efficiently inactivates HIV-1 and also displays high stability and resistance against semen, it combines unique and promising features for its further development as a vaginal microbicide against HIV.

Long-term microbiota and virome in a Zürich patient after fecal transplantation against Clostridium difficile infection.

Fecal microbiota transplantation (FMT) is an emerging therapeutic option for Clostridium difficile infections that are refractory to conventional treatment. FMT introduces fecal microbes into the patient's intestine that prevent the recurrence of C. difficile, leading to rapid expansion of bacteria characteristic of healthy microbiota. However, the long-term effects of FMT remain largely unknown. The C. difficile patient described in this paper revealed protracted microbiota adaptation processes from 6 to 42 months post-FMT. Ultimately, bacterial communities were donor similar, suggesting sustainable stool engraftment. Since little is known about the consequences of transmitted viruses during C. difficile infection, we also interrogated virome changes. Our approach allowed identification of about 10 phage types per sample that represented larger viral communities, and phages were found to be equally abundant in the cured patient and donor. The healthy microbiota appears to be characterized by low phage abundance. Although viruses were likely transferred, the patient established a virome distinct from the donor. Surprisingly, the patient had sequences of algal giant viruses (chloroviruses) that have not previously been reported for the human gut. Chloroviruses have not been associated with intestinal disease, but their presence in the oropharynx may influence cognitive abilities. The findings suggest that the virome is an important indicator of health or disease. A better understanding of the role of viruses in the gut ecosystem may uncover novel microbiota-modulating therapeutic strategies.