The human gastric lumen is one of the most hostile environments of the human body suspected to be sterile until the discovery of Helicobacter pylori (H.p.). State of the art next generation sequencing technologies multiply the knowledge on H.p. functional genomics as well as on the colonization of supposed sterile human environments like the gastric habitat. Here we studied in a prospective, multicenter, clinical trial the 16S rRNA gene amplicon based bacterial microbiome in a total of 30 homogenized and frozen gastric biopsy samples from eight geographic locations. The evaluation of the samples for H.p. infection status was done by histopathology and a specific PCR assay. CagA status was determined by a CagA-specific PCR assay. Patients were grouped accordingly as H.p.-negative, H.p.-positive but CagA-negative and H.p.-positive and CagA-positive (n = 10, respectively). Here we show that H.p. infection of the gastric habitat dominates the gastric microbiota in most patients and is associated with a significant decrease of the microbial alpha diversity from H.p. negative to H.p. positive with CagA as a considerable factor. The genera Actinomyces, Granulicatella, Veillonella, Fusobacterium, Neisseria, Helicobacter, Streptococcus, and Prevotella are significantly different between the H.p.-positive and H.p.-negative sample groups. Differences in microbiota found between CagA-positive and CagA-negative patients were not statistically significant and need to be re-evaluated in larger sample cohorts. In conclusion, H.p. infection dominates the gastric microbiome in a multicentre cohort of patients with varying diagnoses.

Viruses shape a diversity of ecosystems by modulating their microbial, eukaryotic, or plant host metabolism. The complexity of virus-host interaction networks is progressively fathomed by novel metagenomic approaches. By using a novel metagenomic method, we explored the virome in mammalian cell cultures and clinical samples to identify an extensive pool of mobile genetic elements in all of these ecosystems. Despite aseptic treatment, cell cultures harbored extensive and diverse phage populations with a high abundance of as yet unknown and uncharacterized viruses (viral dark matter). Unknown phages also predominated in the oropharynx and urine of healthy individuals and patients infected with cytomegalovirus despite demonstration of active cytomegalovirus replication. The novelty of viral sequences correlated primarily with the individual evaluated, whereas relative abundance of encoded protein functions was associated with the ecologic niches probed. Together, these observations demonstrate the extensive presence of viral dark matter in human and artificial ecosystems.-Thannesberger, J., Hellinger, H.-J., Klymiuk, I., Kastner, M.-T., Rieder, F. J. J., Schneider, M., Fister, S., Lion, T., Kosulin, K., Laengle, J., Bergmann, M., Rattei, T., Steininger, C. Viruses comprise an extensive pool of mobile genetic elements in eukaryote cell cultures and human clinical samples.

Mosquitoes are the most important vectors for arthropod-borne viral diseases. Mixed viral infections of mosquitoes allow genetic recombination or reassortment of diverse viruses, turning mosquitoes into potential virologic mixing bowls. In this study, we field-collected mosquitoes of different species (Aedes aegypti and Culex pipiens complex), from different geographic locations and environments (central Europe and the Caribbean) for highly sensitive next-generation sequencing-based virome characterization. We found a rich virus community associated with a great diversity of host species. Among those, we detected a large diversity of novel virus sequences that we could predominately assign to circular Rep-encoding single-stranded (CRESS) DNA viruses, including the full-length genome of a yet undescribed Gemykrogvirus species. Moreover, we report for the first time the detection of a potentially zoonotic CRESS-DNA virus (Cyclovirus VN) in mosquito vectors. This study expands the knowledge on virus diversity in medically important mosquito vectors, especially for CRESS-DNA viruses that have previously been shown to easily recombine and jump the species barrier.

Anatomically inhomogeneous infection with different, multiple H. pylori strains is common. Prospective clinical study design, collection of samples from multiple sites and microbiologic methods that allow the detection of coinfections are mandatory for collection of reliable data on antimicrobial resistance patterns in representative patient populations. (ClinicalTrials.gov identifier: NCT02925091).

Background noise in metagenomic studies is often of high importance and its removal requires extensive post-analytic, bioinformatics filtering. This is relevant as significant signals may be lost due to a low signal-to-noise ratio. The presence of plasmid residues, that are frequently present in reagents as contaminants, has not been investigated so far, but may pose a substantial bias. Here we show that plasmid sequences from different sources are omnipresent in molecular biology reagents. Using a metagenomic approach, we identified the presence of the (pol) of equine infectious anemia virus in human samples and traced it back to the expression plasmid used for generation of a commercial reverse transcriptase. We found fragments of multiple other expression plasmids in human samples as well as commercial polymerase preparations. Plasmid contamination sources included production chain of molecular biology reagents as well as contamination of reagents from environment or human handling of samples and reagents. Retrospective analyses of published metagenomic studies revealed an inaccurate signal-to-noise differentiation. Hence, the plasmid sequences that seem to be omnipresent in molecular biology reagents may misguide conclusions derived from genomic/metagenomics datasets and thus also clinical interpretations. Critical appraisal of metagenomic data sets for the possibility of plasmid background noise is required to identify reliable and significant signals.