ObjectiveDevelopment of obesity and type 2 diabetes (T2D) are associated with gut microbiota (GM) changes. The gut viral community is predominated by bacteriophages (phages), which are viruses that attack bacteria in a host-specific manner. The antagonistic behaviour of phages has the potential to alter the GM. As a proof-of-concept, we demonstrate the efficacy of faecal virome transplantation (FVT) from lean donors for shifting the phenotype of obese mice into closer resemblance of lean mice.DesignThe FVT consisted of viromes with distinct profiles extracted from the caecal content of mice from different vendors that were fed a low-fat (LF) diet for 14 weeks. Male C57BL/6NTac mice were divided into five groups: LF (as diet control), high-fat (HF) diet, HF+ampicillin (Amp), HF+Amp+FVT and HF+FVT. At weeks 6 and 7 of the study, the HF+FVT and HF+Amp+FVT mice were treated with FVT by oral gavage. The Amp groups were treated with Amp 24 hours prior to first FVT treatment.ResultsSix weeks after first FVT, the HF+FVT mice showed a significant decrease in weight gain compared with the HF group. Further, glucose tolerance was comparable between the LF and HF+FVT mice, while the other HF groups all had impaired glucose tolerance. These observations were supported by significant shifts in GM composition, blood plasma metabolome and expression levels of genes associated with obesity and T2D development.ConclusionsTransfer of caecal viral communities from mice with a lean phenotype into mice with an obese phenotype led to reduced weight gain and normalised blood glucose parameters relative to lean mice. We hypothesise that this effect is mediated via FVT-induced GM changes.

Comparative genomics of 11 lactococcal 936-type phages combined with host range analysis allowed subgrouping of these phage genomes, particularly with respect to their encoded receptor binding proteins. The so-called pellicle or cell wall polysaccharide of Lactococcus lactis, which has been implicated as a host receptor of (certain) 936-type phages, is specified by a large gene cluster, which, among different lactococcal strains, contains highly conserved regions as well as regions of diversity. The regions of diversity within this cluster on the genomes of lactococcal strains MG1363, SK11, IL1403, KF147, CV56, and UC509.9 were used for the development of a multiplex PCR system to identify the pellicle genotype of lactococcal strains used in this study. The resulting comparative analysis revealed an apparent correlation between the pellicle genotype of a given host strain and the host range of tested 936-type phages. Such a correlation would allow prediction of the intrinsic 936-type phage sensitivity of a particular lactococcal strain and substantiates the notion that the lactococcal pellicle polysaccharide represents the receptor for (certain) 936-type phages while also partially explaining the molecular reasons behind the observed narrow host range of such phages. Lactococcal phages are classified into 10 groups based on morphology and DNA hybridization studies (1). Of these, three species are most frequently isolated from dairy environments, namely, the 936, c2, and P335 species (1). The 936 phages are strictly lytic and frequently cause problems for the dairy fermentation industry (2). For this reason, members of this phage species have attracted significant attention in recent years, and the genomes of several phages of this species are now available (3-8). The genome organization of these phages is well conserved and consists of three clusters: the early-, middle-, and late-expressed regions. Comparative genomic analysis of these phages has revealed that while most of the structural genes are highly conserved, there are particular regions of diversity within other regions of their genomes. Among these are the early-expressed genes, which are assumed to encode the replication functions of the phage, as well as the late-expressed region, particularly within the genes encoding the receptor binding protein (RBP) and the tail tape measure protein (TMP) (3,4,8). It has been suggested that the genomes of the 936-type phages can be subgrouped based on these variable regions, which appear to correspond to the subspecies grouping (i.e., Lactococcus lactis subsp. lactis or L. lactis subsp. cremoris) of the host(s) which they infect (3, 9). However, there are a number of exceptions to this in terms of phages that are capable of infecting both subspecies of L. lactis. Phages 645 and P475, for example, are capable of infecting certain members of both subspecies and possess a RBP which is different from those of the two major subgroups of the 936 phages (9). Recently, the RBP structures of phages p2 and bIL170 (or domai...

Pectins are plant cell-wall polysaccharides which can be utilized by commensal bacteria in the gut, exhibiting beneficial properties for the host. Knowledge of the impact of pectins on intestinal bacterial communities is insufficient and limited to a few types of pectins. This study characterized the relationship between the structural properties of pectins and their potential to modulate composition and activity of the gut microbiota in a beneficial way. For this purpose we performed in vitro fermentations of nine structurally diverse pectins from citrus fruits and sugar beet, and a pectic derivative, rhamnogalacturonan I (RGI), using a TIM-2 colon model. The composition of microbiota during TIM-2 fermentations was assessed by 16S rRNA gene amplicon sequencing. Both general and pectin-specific changes were observed in relative abundances of numerous bacterial taxa in a time-dependent way. Bacterial populations associated with human health, such as Faecalibacterium prausnitzii , Coprococcus , Ruminococcus , Dorea , Blautia , Oscillospira , Sutterella , Bifidobacterium , Christensenellaceae , Prevotella copri , and Bacteroides spp. were either increased or decreased depending on the substrate, suggesting that these bacteria can be controlled using structurally different pectins. The main structural features linked to the pectin-mediated shifts in microbiota included degree of esterification, composition of neutral sugars, distribution of homogalacturonan and rhamnogalacturonan fractions, degree of branching, and the presence of amide groups. Cumulative production of the total short chain fatty acids and propionate was largest in fermentations of the high methoxyl pectins. Thus, this study indicates that microbial communities in the gut can be specifically modulated by pectins and identifies the features in pectin molecules linked to microbial alterations. This knowledge can be used to define preferred dietary pectins, targeting beneficial bacteria, and favoring more balanced microbiota communities in the gut.

e Biofilms are major causes of impairment of wound healing and patient morbidity. One of the most common and aggressive wound pathogens is Staphylococcus aureus, displaying a large repertoire of virulence factors and commonly reduced susceptibility to antibiotics, such as the spread of methicillin-resistant S. aureus (MRSA). Bacteriophages are obligate parasites of bacteria. They multiply intracellularly and lyse their bacterial host, releasing their progeny. We isolated a novel phage, DRA88, which has a broad host range among S. aureus bacteria. Morphologically, the phage belongs to the Myoviridae family and comprises a large double-stranded DNA (dsDNA) genome of 141,907 bp. DRA88 was mixed with phage K to produce a high-titer mixture that showed strong lytic activity against a wide range of S. aureus isolates, including representatives of the major international MRSA clones and coagulase-negative Staphylococcus. Its efficacy was assessed both in planktonic cultures and when treating established biofilms produced by three different biofilm-producing S. aureus isolates. A significant reduction of biofilm biomass over 48 h of treatment was recorded in all cases. The phage mixture may form the basis of an effective treatment for infections caused by S. aureus biofilms.

BackgroundThe human gut is densely populated with archaea, eukaryotes, bacteria, and their viruses, such as bacteriophages. Advances in high-throughput sequencing (HTS) as well as bioinformatics have opened new opportunities for characterizing the viral communities harbored in our gut. However, limited attention has been given to the efficiency of protocols dealing with extraction of phages from fecal communities prior to HTS and their impact on the metagenomic dataset.ResultsWe describe two optimized methods for extraction of phages from fecal samples based on tangential-flow filtration (TFF) and polyethylene glycol precipitation (PEG) approaches using an adapted method from a published protocol as control (literature-adapted protocol (LIT)). To quantify phage recovery, samples were spiked with low numbers of c2, ϕ29, and T4 phages (representatives of the Siphoviridae, Podoviridae, and Myoviridae families, respectively) and their concentration (plaque-forming units) followed at every step during the extraction procedure. Compared with LIT, TFF and PEG had higher recovery of all spiked phages, yielding up to 16 times more phage particles (PPs) and up to 68 times more phage DNA per volume, increasing thus the chances of extracting low abundant phages. TFF- and PEG-derived metaviromes showed 10 % increase in relative abundance of Caudovirales and unclassified phages infecting gut-associated bacteria (>92 % for TFF and PEG, 82.4 % for LIT). Our methods obtained lower relative abundance of the Myoviridae family (<16 %) as compared to the reference protocol (22 %). This decline, however, was not considered a true loss of Myoviridae phages but rather a greater level of extraction of Siphoviridae phages (TFF and PEG >32.5 %, LIT 22.6 %), which was achieved with the enhanced conditions of our procedures (e.g., reduced filter clogging). A high degree of phage diversity in samples extracted using TFF and PEG was documented by transmission electron microscopy.ConclusionsTwo procedures (TFF and PEG) for extraction of bacteriophages from fecal samples were optimized using a set of spiked bacteriophages as process control. These protocols are highly efficient tools for extraction and purification of PPs prior to HTS in phage-metavirome studies. Our methods can be easily modified, being thus applicable and adjustable for in principle any solid environmental material in dissolution.Electronic supplementary materialThe online version of this article (doi:10.1186/s40168-015-0131-4) contains supplementary material, which is available to authorized users.