Colonization of Raphanus sativus by human pathogenic microorganisms.

Contamination of vegetables with human pathogenic microorganisms (HPMOs) is considered one of the most important problems in the food industry, as current nutritional guidelines include increased consumption of raw or minimally processed organic vegetables due to healthy lifestyle promotion. Vegetables are known to be potential vehicles for HPMOs and sources of disease outbreaks. In this study, we tested the susceptibility of radish (Raphanus sativus) to colonization by different HPMOs, including Escherichia coli PCM 2561, Salmonella enterica subsp. enterica PCM 2565, Listeria monocytogenes PCM 2191 and Bacillus cereus PCM 1948. We hypothesized that host plant roots containing bactericidal compounds are less prone to HPMO colonization than shoots and leaves. We also determined the effect of selected pathogens on radish growth to check host plant-microbe interactions. We found that one-week-old radish is susceptible to colonization by selected HPMOs, as the presence of the tested HPMOs was demonstrated in all organs of R. sativus. The differences were noticed 2 weeks after inoculation because B. cereus was most abundant in roots (log10 CFU - 2.54), S. enterica was observed exclusively in stems (log10 CFU - 3.15), and L. monocytogenes and E. coli were most abundant in leaves (log10 CFU - 4.80 and 3.23, respectively). The results suggest that E. coli and L. monocytogenes show a higher ability to colonize and move across the plant than B. cereus and S. enterica. Based on fluorescence in situ hybridization (FISH) and confocal laser scanning microscopy (CLSM) approach HPMOs were detected in extracellular matrix and in some individual cells of all analyzed organs. The presence of pathogens adversely affected the growth parameters of one-week-old R. sativus, especially leaf and stem fresh weight (decreased by 47-66 and 17-57%, respectively). In two-week-old plants, no reduction in plant biomass development was noted. This observation may result from plant adaptation to biotic stress caused by the presence of HPMOs, but confirmation of this assumption is needed. Among the investigated HPMOs, L. monocytogenes turned out to be the pathogen that most intensively colonized the aboveground part of R. sativus and at the same time negatively affected the largest number of radish growth parameters.

AMF species do matter: Rhizophagus irregularis and Funneliformis mosseae affect healthy and PVY-infected Solanum tuberosum L. in a different way.

Arbuscular mycorrhizal fungi (AMF) were documented to positively influence plant growth and yield, which is extremely important for the production of many crops including potato. However, the nature of the interaction between arbuscular mycorrhiza and plant virus that share the same host is not well characterized. In this study, we examined the effect of different AMF, Rhizophagus irregularis and Funneliformis mosseae, on healthy and potato virus Y (PVY)-infected Solanum tuberosum L. The analyses conducted included the measurement of potato growth parameters, oxidative stress indicators, and photosynthetic capacity. Additionally, we evaluated both the development of AMF in plant roots and the virus level in mycorrhizal plants. We found that two AMF species colonized plant roots to varying degrees (ca. 38% for R. irregularis vs. 20% for F. mosseae). Rhizophagus irregularis had a more positive effect on potato growth parameters, causing a significant increase in the total fresh and dry weight of tubers, along with virus-challenged plants. Furthermore, this species lowered hydrogen peroxide levels in PVY-infected leaves and positively modulated the levels of nonenzymatic antioxidants, i.e., ascorbate and glutathione in leaves and roots. Finally, both fungal species contributed to reduced lipid peroxidation and alleviation of virus-induced oxidative damage in plant organs. We also confirmed an indirect interaction between AMF and PVY inhabiting the same host. The two AMF species seemed to have different abilities to colonize the roots of virus-infected hosts, as R. irregularis showed a stronger drop in mycorrhizal development in the presence of PVY. At the same time, arbuscular mycorrhiza exerted an effect on virus multiplication, causing increased PVY accumulation in plant leaves and a decreased concentration of virus in roots. In conclusion, the effect of AMF-plant interactions may differ depending on the genotypes of both symbiotic partners. Additionally, indirect AMF-PVY interactions occur in host plants, diminishing the establishment of arbuscular mycorrhiza while changing the distribution of viral particles in plants.

Microbial degradation of polyhydroxybutyrate with embedded polyhexamethylene guanidine derivatives.

The aim of this study was to isolate biofilm-forming bacteria that are capable of degrading polyhydroxybutyrate (PHB) with polyhexamethylene guanidine (PHMG) derivatives. The three types of derivatives incorporated in PHB and their concentration affected the biodegradability of the tested films in both water and compost. The PHMG derivative granular polyethylene wax at the highest concentration significantly inhibited BOD in both environments. At the same time, in water, PHB with PHMG stearate at 1% concentration was also found to inhibit biodegradation but to a lesser extent than PHMG polyethylene wax granulate. Analyzing the values of biofilm abundance and their hydrolytic activity in water, low concentrations of PHMG derivatives (0.2 and 0.6%) slightly inhibited biofilm abundance on the surface of the tested composites. Only granular polyethylene wax PHMG (at 1% concentration) significantly reduced biofilm formation and hydrolase activity in the compost to the greatest extent. Bacteria from biofilm were isolated and identified. Based on the 16S rRNA gene sequence, the strains belong to Bacillus toyonensis HW1 and Variovorax boronicumulans HK3. Introduction of the tested isolates to the environment can enhance composites degradation. However, this requires further research.

Biofilm formation during biodegradation of polylactide, poly (3,4 hydroxybutyrate) and poly(ε-caprolactone) in activated sludge.

Biodegradable materials, namely pure polylactide (PLA), poly (3,4-hydroxybutyrate) (PHB), poly(ε-caprolactone) (PCL) were investigated to assess their degradability by activated sludge. The study aimed at the isolation of biofilm-forming bacteria and the determination of their hydrolytic activity toward the PLA, PHB, and PCL with embedded PHMG derivatives. The biological oxygen demand and physical properties (tensile strength, water vapor permeability, surface structure) of materials indicated that PCL was the best biodegradable film. Aeromonas and Rhodococcus isolated from the polymers' surface during the process of decomposition showed the ability to form biofilms. The introduction of PHMG derivatives into PLA, PCL, and PHB films did not affect biofilm formation and hydrolase activity for most of the isolates. PHMG derivatives at the concentration of 1% disturbed the degradation process.

Microbial Diversity in Deep-Subsurface Hot Brines of Northwest Poland: from Community Structure to Isolate Characteristics.

Deep-subsurface hot brines in northwest Poland, extracted through boreholes reaching 1.6 and 2.6 km below the ground surface, were microbiologically investigated using culture-independent and culture-dependent methods. The high-throughput sequencing of 16S rRNA gene amplicons showed a very low diversity of bacterial communities, which were dominated by phyla Proteobacteria and Firmicutes Bacterial genera potentially involved in sulfur oxidation and nitrate reduction (Halothiobacillus and Methylobacterium) prevailed in both waters over the sulfate reducers ("Candidatus Desulforudis" and Desulfotomaculum). Only one archaeal taxon, affiliated with the order Thermoplasmatales, was detected in analyzed samples. Bacterial isolates obtained from these deep hot brines were closely related to Bacillus paralicheniformis based on the 16S rRNA sequence similarity. However, genomic and physiological analyses made for one of the isolates, Bacillus paralicheniformis strain TS6, revealed the existence of more diverse metabolic pathways than those of its moderate-temperature counterpart. These specific traits may be associated with the ecological adaptations to the extreme habitat, which suggest that some lineages of B. paralicheniformis are halothermophilic.IMPORTANCE Deep-subsurface aquifers, buried thousands of meters down the Earth's crust, belong to the most underexplored microbial habitats. Although a few studies revealed the existence of microbial life at the depths, the knowledge about the microbial life in the deep hydrosphere is still scarce due to the limited access to such environments. Studying the subsurface microbiome provides unique information on microbial diversity, community structure, and geomicrobiological processes occurring under extreme conditions of the deep subsurface. Our study shows that low-diversity microbial assemblages in subsurface hot brines were dominated by the bacteria involved in biogeochemical cycles of sulfur and nitrogen. Based on genomic and physiological analyses, we found that the Bacillus paralicheniformis isolate obtained from the brine under study differed from the mesophilic species in the presence of specific adaptations to harsh environmental conditions. We indicate that some lineages of B. paralicheniformis are halothermophilic, which was not previously reported.

The Contribution of Endomycorrhiza to the Performance of Potato Virus Y-Infected Solanaceous Plants: Disease Alleviation or Exacerbation?

Solanaceae, comprising meaningful crops (as potato, tomato, pepper, eggplant, and tobacco), can benefit from a symbiosis with arbuscular mycorrhizal fungi (AMF), which improve plant fitness and support plant defense against pathogens. Currently, those crops are likely the most impacted by Potato virus Y (PVY). Unfortunately, the effects of AM symbiosis on the severity of disease induced by PVY in solanaceous crops remain uncertain, partly because the interplay between AMF and PVY is poorly characterized. To shed some light on this issue, available studies on interactions in tripartite association between the host plant, its fungal colonizer, and viral pathogen were analyzed and discussed. Although the best-documented PVY transmission pathway is aphid-dependent, PVY infections are also observed in the absence of insect vector. We hypothesize the existence of an additional pathway for virus transmission involving AMF, in which the common mycorrhizal network (CMN) may act as a potential bridge. Therefore, we reviewed (1) the significance of AM colonization for the course of disease, (2) the potential of AMF networks to act as vectors for PVY, and (3) the consequences for crop breeding and production of AM biofertilizers.

Rapid Microbial Community Changes During Initial Stages of Pine Litter Decomposition.

Plant litter decomposition is a process enabling biogeochemical cycles closing in ecosystems, and decomposition in forests constitutes the largest part of this process taking place in terrestrial biomes. Microbial communities during litter decomposition were studied mainly with low-throughput techniques not allowing detailed insight, particularly into coniferous litter, as it is more difficult to obtain high quality DNA required for analyses. Motivated by these problems, we analyzed archaeal, bacterial, and eukaryotic communities at three decomposition stages: fresh, 3- and 8-month-old litter by 16/18S rDNA pyrosequencing, aiming at detailed insight into early stages of pine litter decomposition. Archaea were absent from our libraries. Bacterial and eukaryotic diversity was greatest in 8-month-old litter and the same applied to bacterial and fungal rDNA content. Community structure was different at various stages of decomposition, and phyllospheric organisms (bacteria: Acetobacteraceae and Pseudomonadaceae members, fungi: Lophodermium, Phoma) were replaced by communities with metabolic capabilities adapted to the particular stage of decomposition. Sphingomonadaceae and Xanthomonadaceae and fungal genera Sistotrema, Ceuthospora, and Athelia were characteristic for 3-month-old samples, while 8-month-old ones were characterized by Bradyrhizobiaceae and nematodes (Plectus). We suggest that bacterial and eukaryotic decomposer communities change at different stages of pine litter decomposition in a way similar to that in broadleaf litter. Interactions between bacteria and eukaryotes appear to be one of the key drivers of microbial community structure.

Arbuscular Mycorrhiza Changes the Impact of Potato Virus Y on Growth and Stress Tolerance of Solanum tuberosum L. in vitro.

Under the field conditions crop plants interact with diverse microorganisms. These include beneficial (symbiotic) and phytopathogenic microorganisms, which jointly affect growth and productivity of the plants. In last decades, production of potato (Solanum tuberosum L.) suffers from increased incidence of potato virus Y (PVY), which is one of most important potato pests. Arbuscular mycorrhizal fungi (AMF) are common symbionts of potato, however the impact of mycorrhizal symbiosis on the progression of PVY-induced disease is scarcely known. Therefore, in the present study we investigated the effect of joint PVY infection and mycorrhizal colonization by Rhizophagus irregularis on growth traits of the host potato plant (cv. Pirol). The tested PVY isolate belonged to N-Wilga strain group, which is considered to be predominant in Europe and many other parts of the world. The viral particles were concentrated in the leaves, but decreased the root growth. Furthermore, the infection with PVY evoked prolonged oxidative stress reflected by increased level of endogenous H2O2. AMF alleviated oxidative stress in PVY-infected host plants by a substantial decrease in the level of shoot- and root-derived H2O2, but still caused asymptomatic growth depression. It was assumed that mycorrhizal symbiosis of potato might mask infection by PVY in field observations.

Comamonadaceae OTU as a Remnant of an Ancient Microbial Community in Sulfidic Waters.

Intraterrestrial waters harbor microbial communities being extensively studied to understand microbial processes underlying subsurface ecosystem functioning. This paper provides the results of an investigation on the microbiomes of unique, subsurface sulfidic waters associated with Upper Jurassic, Cretaceous, and Miocene sediments. We used high-throughput 16S rDNA amplicon sequencing to reveal the structure of bacterial and archaeal communities in water samples differing in sulfide content (20-960 mg/dm3), salinity (1.3-3.2%), and depth of extraction (60-660 m below ground level). Composition of the bacterial communities strongly varied across the samples; however, the bacteria participating in the sulfur cycle were common in all sulfidic waters. The shallowest borehole water (60 m bgl) was dominated by sulfur-oxidizing Epsilonproteobacteria (Sulfurimonas, Sulfurovum). In the waters collected from greater depths (148-300 m bgl), the prevalence of Betaproteobacteria (Comamonadaceae) and sulfate/sulfur-reducing Deltaproteobacteria (Desulfopila, Desulfomicrobium, MSBL7) was observed. Sulfate reducers (members of Clostridia: Candidatus Desulforudis) were the most abundant bacteria in the deepest borehole water (660 m bgl). Out of 850 bacterial OTUs, only one, affiliated with the Comamonadaceae family, was found abundant (> 1% of total bacterial sequences) in all samples. Contribution of Archaea to the whole microbial communities was lower than 0.5%. Archaeal communities did not differ across the samples and they consisted of Halobacteriaceae. Out of 372 archaeal OTUs, five, belonging to the four genera Natronomonas, Halorubrum, Halobellus, and Halorhabdus, were the most numerous.

Salino-alkaline lime of anthropogenic origin a reservoir of diverse microbial communities.

This paper presents study on the microbiome of a unique extreme environment - saline and alkaline lime, a by-product of soda ash and table salt production in Janikowo, central Poland. High-throughput 16S rDNA amplicon sequencing was used to reveal the structure of bacterial and archaeal communities in the lime samples, taken from repository ponds differing in salinity (2.3-25.5% NaCl). Surprisingly abundant and diverse bacterial communities were discovered in this extreme environment. The most important geochemical drivers of the observed microbial diversity were salinity, calcium ions, nutrients, and water content. The bacterial and archaeal communities in saline, alkaline lime were similar to those found in natural haloalkaline environments. Although the archaeal contribution to the whole microbial community was lower than 4%, the four archaeal genera Natronomonas, Halorubrum, Halobellus, and Halapricum constituted the core microbiome of saline, alkaline lime - a set of OTUs (> 0.1% of total archaeal relative abundance) present in all samples under study. The high proportion of novel, unclassified archaeal and bacterial sequences (not identified at 97% similarity level) in the 16S rRNA gene libraries indicated that potentially new genera, especially within the class of Thermoplasmata inhabit this unique environment.

Changes in bacterial and archaeal communities during the concentration of brine at the graduation towers in Ciechocinek spa (Poland).

This study evaluates the changes in bacterial and archaeal community structure during the gradual evaporation of water from the brine (extracted from subsurface Jurassic deposits) in the system of graduation towers located in Ciechocinek spa, Poland. The communities were assessed with 16S rRNA gene sequencing (MiSeq, Illumina) and microscopic methods. The microbial cell density determined by direct cell count was at the order of magnitude of 107 cells/mL. It was found that increasing salt concentration was positively correlated with both the cell counts, and species-level diversity of bacterial and archaeal communities. The archaeal community was mostly constituted by members of the phylum Euryarchaeota, class Halobacteria and was dominated by Halorubrum-related sequences. The bacterial community was more diverse, with representatives of the phyla Proteobacteria and Bacteroidetes as the most abundant. The proportion of Proteobacteria decreased with increasing salt concentration, while the proportion of Bacteroidetes increased significantly in the more concentrated samples. Representatives of the genera Idiomarina, Psychroflexus, Roseovarius, and Marinobacter appeared to be tolerant to changes of salinity. During the brine concentration, the relative abundances of Sphingobium and Sphingomonas were significantly decreased and the raised contributions of genera Fabibacter and Fodinibius were observed. The high proportion of novel (not identified at 97% similarity level) bacterial reads (up to 42%) in the 16S rRNA gene sequences indicated that potentially new bacterial taxa inhabit this unique environment.

Molecular phylogeny of Miltogramminae (Diptera: Sarcophagidae): Implications for classification, systematics and evolution of larval feeding strategies.

Miltogramminae is one of the phylogenetically most poorly studied taxa of the species-rich family Sarcophagidae (Diptera). Most species are kleptoparasites in nests of solitary aculeate wasps and bees, although parasitoids and saprophagous species are also known, and the ancestral miltogrammine life habit remains unsettled. Here, we present for the first time a comprehensive phylogenetic tree consisting of 58 representatives of Miltogramminae, reconstructed using sequence data from three mitochondrial (COI, cytB, ND4) and one nuclear (Ef-1α) genes. Our phylogenetic hypothesis suggests that: (1) Miltogramminae are sister to Paramacronychiinae, (2) Miltogramminae can be divided into the "lower miltogrammines" containing two clades of mainly saprophages and a clade of "higher miltogrammines" with mainly kleptoparasitic species, (3) only three genera turn out to be non-monophyletic: Miltogramma, Senotainia and Pterella and (4) the genus Sarcotachina, which traditionally has been considered as belonging to the Paramacronychiinae, is placed in one of the clades of "lower miltogrammines". Ancestral state reconstruction of larval feeding strategy and five larval characters reveals that the ancestor of Miltogramminae was likely a saprophage retaining plesiomorphic oral ridges and a cephaloskeleton with sclerotized dorsal bridge. Synapomorphies like large pseudocephalic sensory organs and well-developed cuticular sculpture suggest that the ancestral first instar larva actively searched for a buried food supply.

Microbial communities associated with the anthropogenic, highly alkaline environment of a saline soda lime, Poland.

Soda lime is a by-product of the Solvay soda process for the production of sodium carbonate from limestone and sodium chloride. Due to a high salt concentration and alkaline pH, the lime is considered as a potential habitat of haloalkaliphilic and haloalkalitolerant microbial communities. This artificial and unique environment is nutrient-poor and devoid of vegetation, due in part to semi-arid, saline and alkaline conditions. Samples taken from the surface layer of the lime and from the depth of 2 m (both having pH ~11 and ECe up to 423 dS m-1) were investigated using culture-based (culturing on alkaline medium) and culture-independent microbiological approaches (microscopic analyses and pyrosequencing). A surprisingly diverse bacterial community was discovered in this highly saline, alkaline and nutrient-poor environment, with the bacterial phyla Proteobacteria (representing 52.8% of the total bacterial community) and Firmicutes (16.6%) showing dominance. Compared to the surface layer, higher bacterial abundance and diversity values were detected in the deep zone, where more stable environmental conditions may occur. The surface layer was dominated by members of the genera Phenylobacterium, Chelativorans and Skermanella, while in the interior layer the genus Fictibacillus was dominant. The culturable aerobic, haloalkaliphilic bacteria strains isolated in this study belonged mostly to the genus Bacillus and were closely related to the species Bacillus pseudofirmus, B. cereus, B. plakortidis, B. thuringensis and B. pumilus.

Biofilm formation on the surface of polylactide during its biodegradation in different environments.

The research was aimed at determining the abundance and viability of biofilm formed on the surface of polylactide (PLA) during its biodegradation in different environments. It was also aimed at isolating biofilm forming bacteria, determining their hydrolytic activity and taxonomic status. The first step was to evaluate PLA biodegradability in lake water, compost and soil, using OxiTop Control. The next step was to assess the ability of isolated bacteria to form biofilm in the investigated environments and to evaluate the biofilm structure. The results indicate that PLA is sensitive to biodegradation in any environment, particularly in compost. During this process biofilm of high viability was observed on the surface of PLA. Based on the 16S rRNA gene sequence, the biofilm-forming bacteria were classified as the following species: Acidovorax sp. LW9, Chryseobacterium sp. LW2, Aeromonas veronii LW8, Arthrobacter aurescens LG2, Arthrobacter sp. LG12, A. aurescens LG9, Elizabethkingia meningoseptica LK3, A. aurescens LK9, A. aurescens and LK7. The results show that different bacterial species formed biofilm of different abundance and hydrolytic activitiy levels.

16S rDNA pyrosequencing analysis of bacterial community in heavy metals polluted soils.

Soil contamination with heavy metals is a widespread problem, especially prominent on grounds lying in the vicinity of mines, smelters, and other industrial facilities. Many such areas are located in Southern Poland; they are polluted mainly with Pb, Zn, Cd, or Cu, and locally also with Cr. As for now, little is known about most bacterial species thriving in such soils and even less about a core bacterial community--a set of taxa common to polluted soils. Therefore, we wanted to answer the question if such a set could be found in samples differing physicochemically and phytosociologically. To answer the question, we analyzed bacterial communities in three soil samples contaminated with Pb and Zn and two contaminated with Cr and lower levels of Pb and Zn. The communities were assessed with 16S rRNA gene fragments pyrosequencing. It was found that the samples differed significantly and Zn decreased both diversity and species richness at species and family levels, while plant species richness did not correlate with bacterial diversity. In spite of the differences between the samples, they shared many operational taxonomic units (OTUs) and it was possible to delineate the core microbiome of our sample set. The core set of OTUs comprised members of such taxa as Sphingomonas, Candidatus Solibacter, or Flexibacter showing that particular genera might be shared among sites ~40 km distant.