The metastable associations of bacteriophages and Erwinia amylovora.

Bacteriophages are often considered as possible agents of biological control of unwanted bacterial populations in medicine, agriculture and food industry. Although the virulent phages can efficiently kill the infected host cells but at the population level phage attack not always leads to the host population collapse but may result in establishment of a more or less stable co-existence. The mechanism of the long-term stabilization of the mixed phage-host cultures is poorly understood. Here we describe bacteriophages VyarbaL and Hena2, the members of the Molineuxvirinae and the Ounavirinae subfamilies, respectively, that are able to form the pseudolysogenic associations (PA) with their host Erwinia amylovora 1/79Sm on solid media. These PAs were stable through multiple passages. The phenomenon of the PA formation between a bacterial culture and bacteriophages decreases the effectiveness of bacteriophage-mediated biological control agents based on lytic bacteriophages.

The Burden of Survivors: How Can Phage Infection Impact Non-Infected Bacteria?

The contemporary understanding of complex interactions in natural microbial communities and the numerous mechanisms of bacterial communication challenge the classical concept of bacteria as unicellular organisms. Microbial populations, especially those in densely populated habitats, appear to behave cooperatively, coordinating their reactions in response to different stimuli and behaving as a quasi-tissue. The reaction of such systems to viral infection is likely to go beyond each cell or species tackling the phage attack independently. Bacteriophage infection of a fraction of the microbial community may also exert an influence on the physiological state and/or phenotypic features of those cells that have not yet had direct contact with the virus or are even intrinsically unable to become infected by the particular virus. These effects may be mediated by sensing the chemical signals released by lysing or by infected cells as well as by more indirect mechanisms.

Genetic analysis of the cold-sensitive growth phenotype of Burkholderia pseudomallei/thailandensis bacteriophage AMP1.

Bacteriophages related to phage Bp_AMP1 are the most widely spread group of phages infecting Burkholderia pseudomallei-the causative agent of melioidosis. These viruses are also infective against the nonpathogenic host Burkholderia thailandensis, allowing experimental work with them without any special safety precautions. The indirect data as well as the results of the mathematical modelling suggest that the AMP1-like viruses may act as natural biocontrol agents influencing the population levels of B. pseudomallei in soil and water habitats in endemic regions. The cold sensitivity of the lytic growth (CSg) of these phages was suggested to be an important feature modulating the effect of viral infection on host populations in nature. We performed genetic analysis to determine the molecular background of the CSg phenotype of the AMP1 phage. The results indicate that CSg is not due to the lack of any function or product missing at low temperature (25 °C) but results in growth inhibition by a phage-encoded temperature-sensitive genetic switch. We identified phage ORF3 and ORF14 to be involved in the genetic determination of this mechanism.

Inhibition of Class A ß-Lactamase (TEM-1) by Narrow Fractions of Humic Substances.

Antimicrobial resistance is a global threat. The use of biologically active natural products alone or in combination with the clinically proven antimicrobial agents might be a useful strategy to fight the resistance. The scientific hypotheses of this study were twofold: (1) the natural humic substances rich in dicarboxyl, phenolic, heteroaryl, and other fragments might possess inhibitory activity against ß-lactamases, and (2) this inhibitory activity might be linked to the molecular composition of the humic ensemble. To test these hypotheses, we used humic substances (HS) from different sources (coal, peat, and soil) and of different fractional compositions (humic acids, hymatomelanic acids, and narrow fractions from solid-phase extraction) for inhibiting serine ß-lactamase TEM-1. Fourier transform ion cyclotron resonance mass spectrometry (FTICR MS) was used to characterize the molecular composition of all humic materials used in this study. The kinetic assay with chromogenic substrate CENTA was used for assessment of inhibitory activity. The inhibition data have shown that among all humic materials tested, a distinct activity was observed within apolar fractions of hymatomelanic acid isolated from lignite. The decrease in the hydrolysis rate in the presence of most active fractions was 42% (with sulbactam-87%). Of particular importance is that these very fractions caused a synergistic effect (2-fold) for the combinations with sulbactam. Linking the observed inhibition effects to molecular composition revealed the preferential contribution of low-oxidized aromatic and acyclic components such as flavonoid-, lignin, and terpenoid-like molecules. The binding of single low-molecular-weight components to the cryptic allosteric site along with supramolecular interactions of humic aggregates with the protein surface could be considered as a major contributor to the observed inhibition. We believe that fine fractionation of hydrophobic humic materials along with molecular modeling studies on the interaction between humic molecules and ß-lactamases might contribute to the development of novel ß-lactamase inhibitors of humic nature.

Equine Intestinal O-Seroconverting Temperate Coliphage Hf4s: Genomic and Biological Characterization.

Tailed bacteriophages constitute the bulk of the intestinal viromes of vertebrate animals. However, the relationships between lytic and lysogenic lifestyles of phages in these ecosystems are not always clear and may vary between the species or even between the individuals. The human intestinal (fecal) viromes are dominated mostly by temperate phages, while in horse feces virulent phages are more prevalent. To our knowledge, all the previously reported isolates of horse fecal coliphages are virulent. Temperate coliphage Hf4s was isolated from horse feces, from the indigenous equine Escherichia coli 4s strain. It is a podovirus related to the Lederbergvirus genus (including the well-characterized Salmonella bacteriophage P22). Hf4s recognizes the host O antigen as its primary receptor and possesses a functional O antigen seroconversion cluster that renders the lysogens protected from superinfection by the same bacteriophage and also abolishes the adsorption of some indigenous equine virulent coliphages, such as DT57C, while other phages, such as G7C or phiKT, retain the ability to infect E. coli 4s (Hf4s) lysogens. IMPORTANCE The relationships between virulent and temperate bacteriophages and their impact on high-density symbiotic microbial ecosystems of animals are not always clear and may vary between species or even between individuals. The horse intestinal virome is dominated by virulent phages, and Hf4s is the first temperate equine intestinal coliphage characterized. It recognizes the host O antigen as its primary receptor and possesses a functional O antigen seroconversion cluster that renders the lysogens protected from superinfection by some indigenous equine virulent coliphages, such as DT57C, while other phages, such as G7C or phiKT, retain the ability to infect E. coli 4s (Hf4s) lysogens. These findings raise questions on the significance of bacteriophage-bacteriophage interactions within the ecology of microbial viruses in mammal intestinal ecosystems.

Genome Sequences of a Green-Colored Chlorobium phaeovibrioides Strain Containing Two Plasmids and a Closely Related Plasmid-Free Brown-Colored Strain.

Here, we report the draft genome sequences of the green sulfur bacterium Chlorobium phaeovibrioides strains GrTcv12 and PhvTcv-s14, isolated from the chemocline zone from meromictic Lake Trekhtzvetnoe, separated from the White Sea, in Russia. This is the first report showing the presence of plasmids containing antiphage systems in the Chlorobium sp. genome.

Sulfobacillus thermotolerans: new insights into resistance and metabolic capacities of acidophilic chemolithotrophs.

The first complete genome of the biotechnologically important species Sulfobacillus thermotolerans has been sequenced. Its 3 317 203-bp chromosome contains an 83 269-bp plasmid-like region, which carries heavy metal resistance determinants and the rusticyanin gene. Plasmid-mediated metal resistance is unusual for acidophilic chemolithotrophs. Moreover, most of their plasmids are cryptic and do not contribute to the phenotype of the host cells. A polyphosphate-based mechanism of metal resistance, which has been previously unknown in the genus Sulfobacillus or other Gram-positive chemolithotrophs, potentially operates in two Sulfobacillus species. The methylcitrate cycle typical for pathogens and identified in the genus Sulfobacillus for the first time can fulfill the energy and/or protective function in S. thermotolerans Kr1 and two other Sulfobacillus species, which have incomplete glyoxylate cycles. It is notable that the TCA cycle, disrupted in all Sulfobacillus isolates under optimal growth conditions, proved to be complete in the cells enduring temperature stress. An efficient antioxidant defense system gives S. thermotolerans another competitive advantage in the microbial communities inhabiting acidic metal-rich environments. The genomic comparisons revealed 80 unique genes in the strain Kr1, including those involved in lactose/galactose catabolism. The results provide new insights into metabolism and resistance mechanisms in the Sulfobacillus genus and other acidophiles.

Sharp water column stratification with an extremely dense microbial population in a small meromictic lake, Trekhtzvetnoe.

Located on the shore of Kandalaksha Bay (the White Sea, Russia) and previously separated from it, Trekhtzvetnoe Lake (average depth 3.5 m) is one of the shallowest meromictic lakes known. Despite its shallowness, it features completely developed water column stratification with high-density microbial chemocline community (bacterial plate) and high rates of major biogeochemical processes. A sharp halocline stabilizes the stratification. Chlorobium phaeovibrioides dominated the bacterial plate, which reached a density of 2 × 108 cell ml-1 and almost completely intercepts H2 S diffusion from the anoxic monimolimnion. The resulting anoxygenic photosynthesis rate reached 240 µmol C l-1 day-1 , exceeding the oxygenic photosynthesis rate in the mixolimnion. The rates of other processes are also high, reaching 4.5 µmol CH4 l-1 day-1 for methane oxidation and 35 µmol S l-1 day-1 for sulfate reduction. Metagenomic analysis demonstrated that the Chl. phaeovibrioides population in the bacterial plate layer had nearly clonal homogeneity, although some fraction of these cells harbour a plasmid. The Chlorobium population was associated with bacteriophages that share homology with CRISPR spacers in the host. These features make the ecosystem of the Trekhtzvetnoe Lake a valuable model for studying regulation and evolution processes in natural high-density microbial systems.

Genomic sequencing and biological characteristics of a novel Escherichia coli bacteriophage 9g, a putative representative of a new Siphoviridae genus.

Bacteriophage 9 g was isolated from horse feces using Escherichia coli C600 as a host strain. Phage 9 g has a slightly elongated capsid 62 × 76 nm in diameter and a non-contractile tail about 185 nm long. The complete genome sequence of this bacteriophage consists of 56,703 bp encoding 70 predicted open reading frames. The closest relative of phage 9 g is phage PhiJL001 infecting marine alpha-proteobacterium associated with Ircinia strobilina sponge, sharing with phage 9 g 51% of amino acid identity in the main capsid protein sequence. The DNA of 9 g is resistant to most restriction endonucleases tested, indicating the presence of hypermodified bases. The gene cluster encoding a biosynthesis pathway similar to biosynthesis of the unusual nucleoside queuosine was detected in the phage 9 g genome. The genomic map organization is somewhat similar to the typical temperate phage gene layout but no integrase gene was detected. Phage 9 g efficiently forms stable associations with its host that continues to produce the phage over multiple passages, but the phage can be easily eliminated via viricide treatment indicating that no true lysogens are formed. Since the sequence, genomic organization and biological properties of bacteriophage 9 g are clearly distinct from other known Enterobacteriaceae phages, we propose to consider it as the representative of a novel genus of the Siphoviridae family.