Pseudomonas mosselii improves cold tolerance of Asian rice (Oryza sativa L.) in a genotype-dependent manner by increasing proline in japonica and reduced glutathione in indica varieties.

Cold stress is an important factor limiting rice production and distribution. Identifying factors that contribute to cold tolerance in rice is of primary importance. While some plant specific genetic factors involved in cold tolerance have been identified, the role of the rice microbiome remains unexplored. In this study, we evaluated the influence of plant growth promoting bacteria (PGPB) with the ability of phosphate solubilization on rice cold tolerance and survival. To reach this goal, inoculated and uninoculated 2-week-old seedlings were cold stressed and evaluated for survival and other phenotypes such as electrolyte leakage (EL) and necessary elements for cold tolerance. The results of this study showed that of the five bacteria, Pseudomonas mosselii, improved both indica and japonica varietal plants' survival and decreased EL, indicating increased membrane integrity. We observed different possible cold tolerance mechanisms in japonica and indica plants such as increases in proline and reduced glutathione levels, respectively. This bacterium also improved the shoot growth of cold exposed indica plants during the recovery period. This study confirmed the host genotype dependent activity of P. mosselii and indicated that there is an interaction between specific plant genes and bacterial genes that causes different plant responses to cold stress.

Dynamic shifts within volatile fatty acid-degrading microbial communities indicate process imbalance in anaerobic digesters.

Buildup of volatile fatty acids (VFAs) in anaerobic digesters (ADs) often results in acidification and process failure. Understanding the dynamics of microbial communities involved in VFA degradation under stable and overload conditions may help optimize anaerobic digestion processes. In this study, five triplicate mesophilic completely mixed AD sets were operated at different organic loading rates (OLRs; 1-6 g chemical oxygen demand [COD] LR-1day-1), and changes in the composition and abundance of VFA-degrading microbial communities were monitored using amplicon sequencing and taxon-specific quantitative PCRs, respectively. AD sets operated at OLRs of 1-4 g COD LR-1day-1 were functionally stable throughout the operational period (120 days) whereas process instability (characterized by VFA buildup, pH decline, and decreased methane production rate) occurred in digesters operated at ≥ 5 g COD LR-1day-1. Though microbial taxa involved in propionate (Syntrophobacter and Pelotomaculum) and butyrate (Syntrophomonas) degradation were detected across all ADs, their abundance decreased with increasing OLR. The overload conditions also inhibited the proliferation of the acetoclastic methanogen, Methanosaeta, and caused a microbial community shift to acetate oxidizers (Tepidanaerobacter acetatoxydans) and hydrogenotrophic methanogens (Methanoculleus). This study's results highlight the importance of operating ADs with conditions that promote the maintenance of microbial communities involved in VFA degradation.

A microfluidic platform for the simultaneous quantification of methanogen populations in anaerobic digestion processes.

Methanogens are a diverse group of archaea that play a critical role in the global carbon cycle. The lack of appropriate molecular tools to simultaneously quantify numerous methanogenic taxa, however, has largely limited our ability to study these communities in a wide variety of habitats, such as anaerobic digesters (ADs). In this study, 34 probe-based quantitative PCR (qPCR) assays were designed to target all known methanogenic genera within the archaeal phylum Euryarchaeota. These qPCR assays were adapted to a high-throughput microfluidic platform, which allowed for the simultaneous detection and absolute quantification of numerous taxa in a single run. The resulting microfluidic qPCR (MFQPCR) platform was successfully used to decipher structure-function relationships among methanogenic communities in four laboratory-scale digesters exposed to a transient organic overload. Twelve of the 34 genera targeted in the MFQPCR were detected in the ADs, similar to results obtained using high-throughput sequencing. The MFQPCR platform and conventional qPCR assays also generated similar quantitative results. The MFQPCR tool developed here will help optimize AD technologies for efficient waste treatment and enhanced biogas production and can facilitate studies that will increase our understanding of methanogenic communities in other environments.

Quantitative detection of syntrophic fatty acid-degrading bacterial communities in methanogenic environments.

In methanogenic habitats, volatile fatty acids (VFA), such as propionate and butyrate, are major intermediates in organic matter degradation. VFA are further metabolized to H(2), acetate and CO(2) by syntrophic fatty acid-degrading bacteria (SFAB) in association with methanogenic archaea. Despite their indispensable role in VFA degradation, little is known about SFAB abundance and their environmental distribution. To facilitate ecological studies, we developed four novel genus-specific quantitative PCR (qPCR) assays, with primer sets targeting known SFAB: Syntrophobacter, Smithella, Pelotomaculum and Syntrophomonas. Primer set specificity was confirmed using in silico and experimental (target controls, clone libraries and melt-curve analysis) approaches. These qPCR assays were applied to quantify SFAB in a variety of mesophilic methanogenic habitats, including a laboratory propionate enrichment culture, pilot- and full-scale anaerobic reactors, cow rumen, horse faeces, an experimental rice paddy soil, a bog stream and swamp sediments. The highest SFAB 16S rRNA gene copy numbers were found in the propionate enrichment culture and anaerobic reactors, followed by the bog stream and swamp sediment samples. In addition, it was observed that SFAB and methanogen abundance varied with reactor configuration and substrate identity. To our knowledge, this research represents the first comprehensive study to quantify SFAB in methanogenic habitats using qPCR-based methods. These molecular tools will help investigators better understand syntrophic microbial communities in engineered and natural environments.

Plant Growth-Promoting Activity of Bacteria Isolated from Asian Rice (Oryza sativa L.) Depends on Rice Genotype.

Asian rice is one of the most important crops because it is a staple food for almost half of the world's population. To have production of rice keep pace with a growing world population, it is anticipated that the use of fertilizers will also need to increase, which may cause environmental damage through runoff impacts. An alternative strategy to increase crop yield is the use of plant growth-promoting bacteria. Thousands of microbial species can exist in association with plant roots and shoots, and some are critical to the plant's survival. We isolated 140 bacteria from two distantly related rice accessions and investigated whether their impact on the growth of four different rice accessions. The bacterial isolates were screened for their ability to solubilize phosphate, a known plant growth-promoting characteristic, and 25 isolates were selected for further analysis. These 25 phosphate-solubilizing isolates were also able to produce other potentially growth-promoting factors. Five of the most promising bacterial isolates were chosen for whole-genome sequencing. Four of these bacteria, isolates related to Pseudomonas mosselii, a Microvirga sp., Paenibacillus rigui, and Paenibacillus graminis, improved root and shoot growth in a rice genotype-dependent manner. This indicates that while bacteria have several known plant growth-promoting functions, their effects on growth parameters are rice genotype dependent and suggest a close relationship between plants and their microbial partners. IMPORTANCE In this study, endophytic bacterial isolates from roots and shoots of two distantly related rice accessions were characterized phenotypically and genotypically. From the isolated bacterial species, five of the most promising plant growth-promoting bacteria were selected to test their abilities to enhance growth of the four rice accessions. Interestingly, plant growth enhancement was both bacterial isolate specific and plant genotype specific. However, the positive interactions between plant and bacteria could not easily be predicted because rice growth-promoting bacteria isolated from their host plants did not necessarily stimulate growth of their own host.

Redirecting marine antibiofouling innovations from sustainable horizons.

Biofouling has great environmental, economic, and societal impacts. Emerging and promising strategies for antibiofouling require incorporation of sustainability concepts. To this end, key research priorities should be given to disrupting attachment of organisms or engineering innovative surfaces to slough off fouling organisms from the surfaces, with more holistic considerations of other viable options, including eco-friendly antifouling chemicals.

Dominance of epiphytic filamentous Thiothrix spp. on an aquatic macrophyte in a hydrothermal vent flume in Sedge Bay, Yellowstone Lake, Wyoming.

Sublacustrine hydrothermal vents, geysers, and fumaroles impart regions of Yellowstone Lake with distinctive chemical compositions that generate unique freshwater habitats and support diverse microbial life. Some microbial communities within Sedge Bay manifest themselves as accumulations of white-colored films on the surfaces of aquatic macrophytes located within the hydrothermal flow of vents. It was hypothesized that the white films were the product of microbial growth, particularly sulfur-oxidizing bacteria. An investigation of the relevant biological compounds in the vent waters was conducted. Microscopy, non-culture molecular techniques, and phylogenetic analysis were used to assay the bacterial diversity associated with the films. Microscopic analysis of the white films revealed the presence of long filaments (>200 µm) that contained sulfur granules. Filaments with these characteristics were not detected on the normal macrophyte samples. Nucleic acids were extracted from the surface of macrophyte coated with the white film (SB1, SB2) and from the surface of an uncoated macrophyte (SC). 16S ribosomal (rRNA) genes were amplified with the polymerase chain reaction (PCR) and cloned. Amplified ribosomal DNA restriction analysis (ARDRA) was used to examine 100 clones from each library and identify unique phylotypes. S(Chao1) and the Shannon Index, mathematical measures of richness and heterogeneity, were employed to assess the ARDRA pattern diversity of each sample. The SC community contained 50 unique phylotypes, predominantly cyanobacteria and proteobacteria, and was the most heterogeneous. SB1 and SB2 communities were less heterogeneous and dominated by Thiothrix. Dilution to extinction PCR conducted with specific primers indicated that the relative abundance of Thiothrix 16S rRNA gene copies in all three samples were similar. However, reduced sulfur compounds from the vent resulted in a more narrow habitat that supported the sulfur-oxidizing Thiothrix in the white film to the exclusion of cyanobacteria and other proteobacteria found on the normal macrophyte. The majority of 16S rRNA gene sequences obtained in this study displayed similarities ≤98% to any known sequence in public data bases which suggests an abundance of new bacterial species in Sedge Bay.

An in vitro comparison of bacterial leakage of three common restorative materials used as an intracoronal barrier.

The purpose of this in vitro study was to evaluate quantitatively the effectiveness of three different restorative materials used as an intracoronal barrier to prevent microleakage of endodontically treated teeth. Fifty-five extracted human single-canal teeth were used in this study. The teeth were endodontically prepared and obturated. Forty-five teeth were randomly assigned to three experimental groups: group 1: sealed with Ketac-Cem (3M ESPE, St Paul, MN) (n = 15), group 2: sealed with Clearfil Protect Bond/Clearfil AP-X (Kuraray, New York, NY) (n = 15), and group 3: sealed with Maxcem (Kerr, Orange, CA) (n = 15). Ten teeth were also randomly assigned to a negative control group (n = 5) and a positive control group (n = 5). Microleakage was tested by using a sterile two-chamber bacterial method and Enterococcus faecalis was used as a microbial marker. Samples were incubated aerobically at 37 degrees C for 120 days. Bacterial leakage was determined by change in turbidity in the medium. Statistical analysis was performed using a Wald chi-square test. No significant difference (p > 0.05) in bacterial leakage was found between the three experimental groups tested. All positive controls leaked within 60 days and broth of the negative control group remained clear throughout the entire experimental period.

Can jua be a weapon in combating oral diseases?

Jua (or Joazeiro) is a plant that has been used in Brazil as a folk remedy for many diseases, but mostly as a dentifrice. Several scientific investigations have been conducted in Brazil to examine the therapeutic use of Jua for oral health. This article presents information about the Ziziphus joazeiro plant, its medicinal use, and its chemical components while also reviewing the literature concerning Jua and especially its anti-plaque activity. Although Joazeiro has been investigated, more studies must be performed to better understand these Jua compounds, their antimicrobial activity, and their mechanism of action. Such studies would confirm whether Joazeiro has the potential to fight or prevent diseases like dental caries and periodontitis.

In vitro evaluation of the antimicrobial effects of a root canal sealer-antibiotic combination against Enterococcus faecalis.

The purpose of this in vitro study was to evaluate the antimicrobial effects of five antibiotics when added to Kerr Pulp Canal Sealer EWT against Enterococcus faecalis. Five antibiotics: amoxicillin, penicillin, clindamycin, metronidazole, and doxycycline, were added separately to Kerr sealer. Thirty brain heart infusion agar plates were inoculated with E. faecalis and sterile paper discs containing a sealer-antibiotic combination were randomly assigned to the inoculated plates. Kerr sealer on a sterile paper disc and a blank sterile paper disc served as the controls. Fifteen plates were incubated aerobically and the remaining 15 plates were incubated anaerobically at 37 degrees C. The zones of inhibition were measured at 48 hours. No significant differences (p > 0.05) were found between aerobic and anaerobic groups. The groups were combined and data analysis using a one-way ANOVA and Tukey's post hoc test was performed. Results revealed that sealer-antibiotic combinations containing amoxicillin, penicillin, clindamycin, and doxycycline had a significant difference (p < 0.001) in the mean zones of inhibition when compared to Kerr EWT sealer alone. No significant differences (p > 0.05) were found between the metronidazole-sealer combinations and Kerr EWT sealer alone.

Methyl coenzyme M reductase (mcrA) gene abundance correlates with activity measurements of methanogenic H2 /CO2 -enriched anaerobic biomass.

Biologically produced methane (CH4) from anaerobic digesters is a renewable alternative to fossil fuels, but digester failure can be a serious problem. Monitoring the microbial community within the digester could provide valuable information about process stability because this technology is dependent upon the metabolic processes of microorganisms. A healthy methanogenic community is critical for digester function and CH4 production. Methanogens can be surveyed and monitored using genes and transcripts of mcrA, which encodes the α subunit of methyl coenzyme M reductase - the enzyme that catalyses the final step in methanogenesis. Using clone libraries and quantitative polymerase chain reaction, we compared the diversity and abundance of mcrA genes and transcripts in four different methanogenic hydrogen/CO2 enrichment cultures to function, as measured by specific methanogenic activity (SMA) assays using H2 /CO2 . The mcrA gene copy number significantly correlated with CH4 production rates using H2 /CO2 , while correlations between mcrA transcript number and SMA were not significant. The DNA and cDNA clone libraries from all enrichments were distinctive but community diversity also did not correlate with SMA. Although hydrogenotrophic methanogens dominated these enrichments, the results indicate that this methodology should be applicable to monitoring other methanogenic communities in anaerobic digesters. Ultimately, this could lead to the engineering of digester microbial communities to produce more CH4 for use as renewable fuel.

Bacterial intracellular sulfur globules: structure and function.

Bacteria that oxidize reduced sulfur compounds like H2S often transiently store sulfur in protein membrane-bounded intracellular sulfur globules; intracellular in this case meaning found inside the cell wall. The cultured bacteria that form these globules are primarily phylogenetically classified in the Proteobacteria and are chemotrophic or photoautotrophic. The current model organism is the purple sulfur bacterium Allochromatium vinosum. Research on this bacterium has provided the groundwork for understanding the protein membranes and the sulfur contents of globules. In addition, it has demonstrated the importance of different genes (e.g. sulfur oxidizing, sox) in their formation and in the final oxidation of sulfur in the globules to sulfate (e.g. dissimilatory sulfite reductase, dsr). Pursuing the characteristics of other intracellular sulfur globule-forming bacteria through genomics, transcriptomics and proteomics will eventually lead to a complete picture of their formation and breakdown. There will be commonality to some of the genetic, physiological and morphological characteristics involved in intracellular sulfur globules of different bacteria, but there will likely be some surprises as well.

In vitro evaluation of the antimicrobial effect of three endodontic sealers mixed with amoxicillin.

INTRODUCTION: The purpose of this in vitro study was to evaluate the antimicrobial effect of mixing amoxicillin with three different sealers when freshly mixed and set. METHODS: Using a direct contact test, Pulp Canal Sealer EWT (SybronEndo Corporation, Orange, CA), AH Plus (Dentsply International Inc, York, PA), and RealSeal SE (Pentron Clinical Technologies LLC, Wallingford, CT) were freshly mixed with amoxicillin and placed on the side wall of the microtiter plate. A 10-microL bacterial suspension of Enterococcus faecalis was placed directly onto the fresh sealers, and sealers set 1 day, 3 days, and 7 days after mixing. The bacteria were allowed to dry in direct contact with the sealer sample. Fresh media was then added, and growth of the bacteria was measured by spectrophotometry over an 8-hour period. One-way analysis of variance (ANOVA), two-way ANOVA, and Tukey multiple comparison were used for statistical significance. RESULTS: All sealers mixed with amoxicillin showed complete inhibition of the growth of E. faecalis. Sealers mixed with amoxicillin had no statistical difference in inhibiting growth between freshly mixed samples and samples set for 1 day, 3 days, or 7 days (p > 0.05). Sealers without amoxicillin did not inhibit growth of E. faecalis, and no statistical difference was found between freshly mixed and set samples (p > 0.05). Sealers with amoxicillin were statistically different than sealers without amoxicillin when freshly mixed and set (p < 0.001). CONCLUSIONS: Sealers mixed with amoxicillin inhibited the growth of E. faecalis significantly greater than sealers without amoxicillin (p < 0.001).

Bioaugmentation for improved recovery of anaerobic digesters after toxicant exposure.

Bioaugmentation was investigated as a method to decrease the recovery period of anaerobic digesters exposed to a transient toxic event. Two sets of laboratory-scale digesters (SRT = 10 days, OLR = 2 g COD/L-day), started with inoculum from a digester stabilizing synthetic municipal wastewater solids (MW) and synthetic industrial wastewater (WW), respectively, were transiently exposed to the model toxicant, oxygen. Bioaugmented digesters received 1.2 g VSS/L-day of an H2-utilizing culture for which the archaeal community was analyzed. Soon after oxygen exposure, the bioaugmented digesters produced 25-60% more methane than non-bioaugmented controls (p < 0.05). One set of digesters produced lingering high propionate concentrations, and bioaugmentation resulted in significantly shorter recovery periods. The second set of digesters did not display lingering propionate, and bioaugmented digesters recovered at the same time as non-bioaugmented controls. The difference in the effect of bioaugmentation on recovery may be due to differences between microbial communities of the digester inocula originally employed. In conclusion, bioaugmentation with an H(2)-utilizing culture is a potential tool to decrease the recovery period, decrease propionate concentration, and increase biogas production of some anaerobic digesters after a toxic event. Digesters already containing rapidly adaptable microbial communities may not benefit from bioaugmentation, whereas other digesters with poorly adaptable microbial communities may benefit greatly.

The effects of natural biofilms on the reattachment of young adult zebra mussels to artificial substrata.

This laboratory study examined the effects of natural biofilms on the reattachment of young adult zebra mussels, Dreissena polymorpha, in Petri dishes. Natural biofilms were developed in glass and polystyrene Petri dishes using water samples collected at various times of the year. Biofilms were developed over 1, 3, 8, and 14 d. Controls were clean glass and polystyrene Petri dishes. Zebra mussels collected from the field (< or = 10 mm, ventral length) were placed in the dishes and their reattachment by byssal threads was recorded after 1 d. Zebra mussels reattached to the dish surface or the shells of other mussels in the dish, or remained unattached. The data indicate that reattachment to clean glass was greater than to clean polystyrene (p < or = 0.05, ANOVA), but there were no consistent differences between reattachment to filmed polystyrene and filmed glass dish surfaces. Zebra mussels in control and filmed glass dishes reattached in higher percentages to the dish surface compared to the shells of other mussels (p < or = 0.05, ANOVA). There was no difference in mussel of reattachment between the dish surface and the shells of other mussels in most control polystyrene dishes (p > 0.05, ANOVA), whereas in filmed polystyrene the percentage of reattachment to the dish surface was greater than to the shells of other mussels (p < or = 0.05, ANOVA). These results indicate that substratum wettability and the presence of biofilms on some types of substrata can be factors in the reattachment of young adult zebra mussels.