Endogenous Microbial Community Research Articles

Enhancement of microbial community dynamics and metabolism in compost through ammonifying cultures inoculation

The efficient use of livestock and poultry manure waste has become a global challenge, with microorganisms playing an important role. To investigate the impact of novel ammonifying microorganism cultures (NAMC) on microbial community dynamics and carbon and nitrogen metabolism, five treatments [5% (v/w) sterilized distilled water, Amm-1, Amm-2, Amm-3, and Amm-4] were applied to cow manure compost. Inoculation with NAMC improved the structure of bacterial and fungal communities, enriched the populations of the functional microorganisms, enhanced the role of specific microorganisms, and promoted the formation of tight modularity within the microbial network. Further functional predictions indicated a significant increase in both carbon metabolism (CMB) and nitrogen metabolism (NMB). During the thermophilic phase, inoculated NAMC treatments boosted carbon metabolism annotation by 10.55%–33.87% and nitrogen metabolism annotation by 26.69%–63.11. Structural equation modeling supported the NAMC-mediated enhancement of NMB and CMB. In conclusion, NAMC inoculation, particularly with Amm-4, enhanced the synergistic interaction between bacteria and fungi. This collaboration promoted enzymatic catabolic and synthetic processes, resultng in positive feedback loops with the endogenous microbial community. Understanding these mechanisms not only unravels how ammonifying microorganisms influence microbial communities but also paves the way for the development of the composting industry and global waste management practices.

Role of endogenous soil microorganisms in controlling antimicrobial resistance after the exposure to treated wastewater

The reuse of treated wastewater (TWW) for irrigation appears to be a relevant solution to the challenges of growing water demand and scarcity. However, TWW contains not only micro-pollutants including pharmaceutical residues but also antibiotic resistant bacteria. The reuse of TWW could contribute to the dissemination of antimicrobial resistance in the environment. The purpose of this study was to assess if exogenous bacteria from irrigation waters (TWW or tap water-TP) affect endogenous soil microbial communities (from 2 soils with distinct irrigation history) and key antibiotic resistance gene sul1 and mobile genetic elements intl1 and IS613. Experiments were conducted in microcosms, irrigated in one-shot, and monitored for three months. Results showed that TP or TWW exposure induced a dynamic response of soil microbial communities but with no significant increase of resistance and mobile gene abundances. However, no significant differences were observed between the two water types in the current experimental design. Despite this, the 16S rDNA analysis of the two soils irrigated for two years either with tap water or TWW resulted in soil microbial community differentiation and the identification of biomarkers from Xanthomonadaceae and Planctomycetes families for soils irrigated with TWW. Low-diversity soils were more sensitive to the addition of TWW. Indeed, TWW exposure stimulated the growth of bacterial genera known to be pathogenic, correlating with a sharp increase in the copy number of selected resistance genes (up to 3 logs). These low-diversity soils could thus enable the establishment of exogenous bacteria from TWW which was not observed with native soils. In particular, the emergence of Planctomyces, previously suggested as a biomarker of soil irrigated by TWW, was here demonstrated. Finally, this study showed that water input frequency, initial soil microbial diversity and soil history drive changes within soil endogenous communities and the antibiotic resistance gene pool.

Cerium oxide and neodymium oxide phytoextraction by ryegrass in bioenhanced hydroponic environments

Sustainable technologies for the recovery of rare earth elements (REE) from waste need to be developed to decrease the volume of ore mining extractions and its negative environmental consequences, while simultaneously restoring previously impacted lands. This is critical due to the extensive application of REE in everyday life from electronic devices to energy and medical technologies, and the dispersed distribution of REE resources in the world. REE recovery by plants has been previously studied but the feasibility of REE phytoextraction from a poorly soluble solid phase (i.e., nanoparticles) by different plant species has been rarely investigated. In this study, the effect of biostimulation and bioaugmentation on phytorecovery of REE nanoparticles (REE-NP) was investigated by exposing ryegrass seeds to REE-NP in hydroponic environments. This was studied in two sets of experiments: bioaugmentation (using CeO2 nanoparticles and Methylobacterium extorquens AM1 pure culture), and biostimulation (using CeO2 or Nd2O3 nanoparticles and endogenous microorganisms). Addition of M. extorquens AM1 in bioaugmentation experiment including 500 mg/L CeO2 nanoparticles could not promote the nanoparticles accumulation in both natural and surface-sterilized treatments. However, it enhanced the translocation of Ce from roots to shoots in sterile samples. Moreover, another REE-utilizing bacterium, Bacillus subtilis, was enriched more than M. extorquens in control samples (no M. extorquens AM1), and associated with 52% and 14% higher Ce extraction in both natural (165 μg/gdried-plant) and surface-sterilized samples (136 μg/gdried-plant), respectively; showing the superior effect of endogenous microorganisms’ enrichment over bioaugmentation in this experiment. In the biostimulation experiments, up to 705 μg/gdried-plant Ce and 19,641 μg/gdried-plant Nd could be extracted when 500 mg/L REE-NP were added. Furthermore, SEM-EDS analysis of the surface and longitudinal cross-sections of roots in Nd2O3 treatments confirmed surface and intracellular accumulation of Nd2O3-NP. These results demonstrate stimulation of endogenous microbial community can lead to an enhanced REE phytoaccumulation.

Screening and field application of microbial-flooding activator systems

This study aims to further enhance the oil recovery of reservoirs in the Zhong-2 Block of the Gudao Oilfield by identifying the most effective microbial-flooding activator systems and applying them in the field. We began by analyzing the structure of the reservoirs' endogenous microbial communities to understand the potential impact of microbial flooding. This was followed by determining commonly used activator systems based on their abilities to stimulate oil-displacement functional bacteria. Through laboratory experiments on oil displacement efficiency and sweep characteristics, we determined the optimal activator injection method (injection ratio) and the requisite bacterial concentration for maximal microbial-flooding efficacy. Finally, we selected the optimal activator systems and applied them to field tests. Our findings suggest the target block is highly receptive to microbial-flooding. In terms of performance, the activator systems ranked as No. 3 > No. 4 > No. 1 > No. 2. Interestingly, a deep activator system, when compared to the top-performing No. 3 system, exhibited a higher bacterial concentration peak and longer peaking duration. Optimal oil displacement effects were observed at a 1:4 vol ratio between the No. 3 activator and deep activator systems, with bacterial concentrations of up to 106 cells/mL or above. Field tests with the selected activator systems, following a specific injection protocol, demonstrated a notable increase in oil production and a reduction in water cut.

Assessing the impacts of fine sediment removal on endogenous pollution release and microbial community structure in the shallow lakes

Resuspension is a crucial process for releasing endogenous pollution from shallow lakes into the overlying water. Fine particle sediment, which has a higher contamination risk and longer residence time, is the primary target for controlling endogenous pollution. To this end, a study coupling aqueous biogeochemistry, electrochemistry, and DNA sequencing was conducted to investigate the remediation effect and microbial mechanism of sediment elution in shallow eutrophic water. The results indicated that sediment elution can effectively remove some fine particles in situ. Furthermore, sediment elution can inhibit the release of ammonium nitrogen and total dissolved phosphorous into the overlying water from sediment resuspension in the early stage, resulting in reductions of 41.44 %–50.45 % and 67.81 %–72.41 %, respectively. Additionally, sediment elution greatly decreased the concentration of nitrogen and phosphorus pollutants in pore water. The microbial community structure was also substantially altered, with an increase in the relative abundance of aerobic and facultative aerobic microorganisms. Redundancy analysis, PICRUSt function prediction, and the correlation analysis revealed that loss on ignition was the primary factor responsible for driving changes in microbial community structure and function in sediment. Overall, the findings provide novel insights into treating endogenous pollution in shallow eutrophication water.

Characterizing the blood microbiota of omnivorous and frugivorous bats (Chiroptera: Phyllostomidae) in Casanare, eastern Colombia.

Bats are known reservoirs of seemingly-innocuous pathogenic microorganisms (including viruses, bacteria, fungi, and protozoa), which are associated with triggering disease in other zoonotic groups. The taxonomic diversity of the bats' microbiome is likely associated with species-specific phenotypic, metabolic, and immunogenic capacities. To date, few studies have described the diversity of bat blood microbial communities. Then, this study used amplicon-based next generation sequencing of the V4 hypervariable region of the 16S-rRNA gene in blood samples from omnivorous (n = 16) and frugivorous (n = 9) bats from the department of Casanare in eastern Colombia. We found the blood microbiota in bats to be composed of, among others, Bartonella and Mycoplasma bacterial genera which are associated with various disease phenotypes in other mammals. Furthermore, our results suggest that the bats' dietary habits might determine the composition and the persistence of some pathogens over others in their bloodstream. This study is among the first to describe the blood microbiota in bats, to reflect on co-infection rates of multiple pathogens in the same individual, and to consider the influence of diet as a factor affecting the animal's endogenous microbial community.

Endogenous small intestinal microbiome determinants of transient colonisation efficiency by bacteria from fermented dairy products: a randomised controlled trial

BackgroundThe effects of fermented food consumption on the small intestine microbiome and its role on host homeostasis are largely uncharacterised as our knowledge on intestinal microbiota relies mainly on faecal samples analysis. We investigated changes in small intestinal microbial composition and functionality, short chain fatty acid (SCFA) profiles, and on gastro-intestinal (GI) permeability in ileostomy subjects upon the consumption of fermented milk products.ResultsWe report the results from a randomised, cross-over, explorative study where 16 ileostomy subjects underwent 3, 2-week intervention periods. In each period, they consumed either milk fermented by Lacticaseibacillus rhamnosus CNCM I-3690, or milk fermented by Streptococcus thermophilus CNCM I-1630 and Lactobacillus delbrueckii subsp. bulgaricus CNCM I-1519, or a chemically acidified milk (placebo) daily. We performed metataxonomic, metatranscriptomic analysis, and SCFA profiling of ileostomy effluents as well as a sugar permeability test to investigate the microbiome impact of these interventions and their potential effect on mucosal barrier function. Consumption of the intervention products impacted the overall small intestinal microbiome composition and functionality, mainly due to the introduction of the product-derived bacteria that reach in several samples 50% of the total microbial community. The interventions did not affect the SCFA levels in ileostoma effluent, or gastro-intestinal permeability and the effects on the endogenous microbial community were negligible. The impact on microbiome composition was highly personalised, and we identified the poorly characterised bacterial family, Peptostreptococcaceae, to be positively associated with a low abundance of the ingested bacteria. Activity profiling of the microbiota revealed that carbon- versus amino acid-derived energy metabolism of the endogenous microbiome could be responsible for the individual-specific intervention effects on the small intestine microbiome composition and function, reflected also on urine microbial metabolites generated through proteolytic fermentation.ConclusionsThe ingested bacteria are the main drivers of the intervention effect on the small intestinal microbiota composition. Their transient abundance level is highly personalised and influenced by the energy metabolism of the ecosystem that is reflected by its microbial composition (http://www.clinicaltrials.gov, ID NCT NCT02920294).3z8GaJs8-4AmNdyX8busiEVideo

Mining the landfill soil metagenome for denitrifying methanotrophic taxa and validation of methane oxidation in microcosm

In the present study, the microbial community residing at different depths of the landfill was characterized to assess their roles in serving as a methane sink. Physico-chemical characterization revealed the characteristic signatures of anaerobic degradation of organic matter in the bottom soil (50–60 cm) and, active process of aerobic denitrification in the top soil (0–10 cm). This was also reflected from the higher abundance of bacterial domain in the top soil metagenome represented by dominant phyla Proteobacteria and Actinobacteria which are prime decomposers of organic matter in landfill soils. The multiple fold higher relative abundances of the two most abundant genera; Streptomyces and Intrasporangium in the top soil depicted greater denitrifying taxa in top soil than the bottom soil. Amongst the aerobic methanotrophs, the genera Methylomonas, Methylococcus, Methylocella, and Methylacidiphilum were abundantly found in the top soil metagenome that were essential for oxidizing methane generated in the landfill. On the other hand, the dominance of archaeal domain represented by Methanosarcina and Methanoculleus in the bottom soil highlighted the complete anaerobic digestion of organic components via acetoclasty, carboxydotrophy, hydrogenotrophy, methylotrophy. Functional characterization revealed a higher abundance of methane monooxygenase gene in the top soil and methyl coenzyme M reductase gene in the bottom soil that correlated with the higher relative abundance of aerobic methanotrophs in the top soil while methane generation being the active process in the highly anaerobic bottom soil in the landfill. The activity dependent abundance of endogenous microbial communities in the different zones of the landfill was further validated by microcosm studies in serum bottles which established the ability of the methanotrophic community for methane metabolism in the top soil and their potential to serve as sink for methane. The study provides a better understanding about the methanotrophs in correlation with their endogenous environment, so that these bacteria can be used in resolving the environmental issues related to methane and nitrogen management at landfill site.

Niches and Seasonal Changes, Rather Than Transgenic Events, Affect the Microbial Community of Populus × euramericana 'Neva'.

Exploring the complex spatiotemporal changes and colonization mechanism of microbial communities will enable microbial communities to be better used to serve agricultural and ecological operations. In addition, evaluating the impact of transgenic plants on endogenous microbial communities is necessary for their commercial application. In this study, microbial communities of Populus × euramericana ‘Neva’ carrying Cry1Ac-Cry3A-BADH genes (ECAA1 line), Populus × euramericana ‘Neva’ carrying Cry1Ac-Cry3A-NTHK1 genes (ECAB1 line), and non-transgenic Populus × euramericana ‘Neva’ from rhizosphere soil, roots, and phloem collected in different seasons were compared and analyzed. Our analyses indicate that the richness and diversity of bacterial communities were higher in the three Populus × euramericana ‘Neva’ habitats than in those of fungi. Bacterial and fungal genetic-distance-clustering results were similar; rhizosphere soil clustered in one category, with roots and phloem in another. The diversity and evenness values of the microbial community were: rhizosphere soil > phloem > root system. The bacterial communities in the three habitats were dominated by the Proteobacteria, and fungal communities were dominated by the Ascomycota. The community composition and abundance of each part were quite different; those of Populus × euramericana ‘Neva’ were similar among seasons, but community abundance fluctuated. Seasonal fluctuation in the bacterial community was greatest in rhizosphere soil, while that of the fungal community was greatest in phloem. The transgenic lines ECAA1 and ECAB1 had a bacterial and fungal community composition similar to that of the control samples, with no significant differences in community structure or diversity among the lines. The abundances of operational taxonomic units (OTUs) were low, and differed significantly among the lines. These differences did not affect the functioning of the whole specific community. Sampling time and location were the main driving factors of changes in the Populus × euramericana ‘Neva’ microbial community. Transgenic events did not affect the Populus × euramericana ‘Neva’ rhizosphere or endophytic microbial communities. This study provides a reference for the safety evaluation of transgenic plants and the internal colonization mechanism of microorganisms in plants.

Insight Into Host-Microbe Interactions Using Microbial Flow Cytometry Coupled to Next-Generation Sequencing.

Antibody-based assays have been a cornerstone of infectious disease diagnostics for over 100 years [1]. These assays rely on the exquisite sensitivity and specificity of humoral response to almost all infections. While next-generation sequencing (NGS) has tremendous potential to improve diagnostics and uncover host-microbial relationships by directly identifying nucleic acids from infectious microbes, challenges and opportunities for new approaches remain. Here, we review a group of cutting-edge techniques that couple antibody responses with flow cytometry of antibody tagged microbes and NGS. These studies are bringing into focus the dynamic relationship between our immune systems and endogenous microbial communities, which are an important source of pathogens. For simplicity, we use the umbrella term mFLOW-Seq (microbial flow cytometry coupled to NGS) to describe these approaches.

Mucosal Bacteria Modulate Candida albicans Virulence in Oropharyngeal Candidiasis.

ABSTRACTOropharyngeal candidiasis (OPC) is the most prevalent oral infection in immunocompromised patients, primarily associated with Candida albicans. Increasing evidence points to a significant role of mucosal bacteria on the transition of C. albicans from commensal to pathogenic. In this work, we hypothesized that changes in the abundance or composition of the mucosal bacterial microbiota induced by dietary sucrose during the development of OPC can modulate C. albicans virulence. C. albicans burdens and mucosal lesions were evaluated in a mouse cortisone immunosuppression model amended with sucrose. We also analyzed the mucosal bacterial composition using 16S rRNA gene sequencing and culture methods. In immunocompetent mice, sucrose significantly increased total bacterial burdens and reduced alpha diversity, by increasing the relative abundance of mitis group streptococci. In immunocompromised mice, C. albicans infection was associated with a significantly reduced bacterial alpha diversity due to an increase in the relative abundance of enterococci. When exposed to dietary sucrose, these mice had reduced C. albicans burdens and reduced bacterial alpha diversity, associated with an increase in the relative abundance of Lactobacillus. SparCC correlation networks showed a significant negative correlation between Lactobacillus and Enterococcus in all Candida-infected mice. Depletion of lactobacilli with antibiotic treatment partially restored C. albicans burdens in mice receiving sucrose. In coculture in vitro experiments, mouse oral Lactobacillus johnsonii isolates inhibited growth of Enterococcus faecalis isolates and C. albicans. These results support the hypothesis that the sucrose-induced attenuation of C. albicans virulence was a result of changes in the mucosal bacterial microbiome characterized by a reduction in enterococci and an increase in lactobacilli.

1281. Longitudinal and Spatial Variation in the Human Microbiome in a Phase 2a Clinical Study of Gepotidacin in Adult Females with Uncomplicated Urinary Tract Infection

BackgroundGepotidacin (GSK2140944) is a first in class novel oral triazaacenaphthylene bacterial topoisomerase inhibitor. In this study, we evaluated the potential impact of orally administered gepotidacin on the human microbiome, across three body-sites and at three specific time-points, as an exploratory endpoint in a Phase 2a clinical trial for the treatment of uncomplicated Urinary Tract Infection (uUTI) (ClinicalTrials.gov: NCT03568942).MethodsThrough DNA sequencing of the 16S rRNA variable region 4, we analyzed samples collected with consent from study subjects from the gastrointestinal tract or GIT (stool), pharyngeal cavity (saliva swabs) and vagina (vaginal swabs). Samples were taken at three time points which were pre-dosing (Day 1), end of dosing (Day 5) and follow-up visit (Day 28 ±3 days). A total of 156 samples were collected and 141 samples passed quality control criteria for DNA sequence analyses. Using a rigorous computational work-flow, changes in microbiome diversity and relative abundances of microbial species were quantified.ResultsTime series analyses showed that microbiota alpha diversity dropped, relative to pre-dose, by the end of gepotidacin dosing but trended a return trajectory to original pre-dose levels by the follow-up visit, for all body sites (Figure). However, the character and extent of the microbiota changes varied by location. The relative ordering from least to greatest changes in microbiota diversity of body sites is vaginal, pharyngeal and GIT. We found no statistically significant occurrences of pathogen related taxa, such as Clostridioides or Enterobacterales spp., at the final timepoints.ConclusionSince gepotidacin is both orally dosed and elimination includes the biliary route, it was predicted to affect the GIT microbiome, however changes in the distal pharyngeal and vaginal microbiota were also observed. Gepotidacin alteration of the endogenous microbial community appears to be temporary and reversible as microbiota diversity rebounded to near pre-dosing status within a period of several weeks. Our study illustrates how microbiome analyses in antibiotic clinical studies can quantify patterns of microbiota disruption and recovery.DisclosuresAndrea Nuzzo, PHD, GlaxoSmithKline (Employee) Stephanie Van Horn, B.Sc., GlaxoSmithKline (Employee) Christopher Traini, PHD, GlaxoSmithKline (Employee) Caroline R. Perry, PhD, GlaxoSmithKline (Employee) Etienne Dumont, MD, GlaxoSmithKline (Employee) Nicole Scangarella-Oman, MS, GlaxoSmithKline plc. (Employee, Shareholder) David Gardiner, MD, GlaxoSmithKline (Employee) James R. Brown, PhD, GlaxoSmithKline (Employee)

Specific enrichment of hydrocarbonclastic bacteria from diesel-amended soil on biochar particles

Biochar has been proposed as a suitable biostimulant for the remediation of hydrocarbon contamination, and also has the potential to act as a carrier for hydrocarbonoclastic microorganisms which could bioaugment endogenous microbial communities. However, the evidence regarding the biostimulatory effects of biochars on hydrocarbon bioremediation is somewhat equivocal, possibly due to variability of the physicochemical properties of biochar and soil across studies. Here, we use standard biochars with defined properties produced from softwood pellets (SWP) and rice husk (RH) at pyrolysis temperatures of 550 °C or 700 °C to test the effects of biochar amendment on microbial community composition and hydrocarbon degradation in soil microcosms contaminated with diesel oil. Combining this approach for the first time with specific analysis of microbial community composition using amplicon sequence variants (ASVs), we find that oil contamination causes extreme short-term loss of soil microbial diversity, and highly-specific selection of a limited set of genera defined by 13 ASVs. Biochar ameliorates the short-term loss of diversity, and in the longer term (9 weeks), changes community composition in a type-specific manner. The majority of the 13 selected ASVs are further enriched on biochar particles, although SWP biochars perform better than RH biochar in enrichment of putative hydrocarbonoclastic Aquabacterium spp. However, complete degradation of normal (n) alkanes from the aliphatic hydrocarbon fraction is prevented in the presence of biochar amendment, possibly due to their adsorption onto the char surface. Furthermore, we show that putative hydrocarbon degraders released from diesel-amended soil can subsequently be enriched to high levels on SWP biochar particles in growth medium supplemented with diesel oil as the sole carbon source; these include selected ASVs representing the genera Rhodococcus, Aquabacterium, and Cavicella. This work suggests that use of biochar pre-enriched with endogenous, conditionally-rare hydrocarbon degrading bacteria is a promising strategy for bioaugmentation of diesel-contaminated soils.

Exogenous and endogenous microbiomes of wild-caught Phormia regina (Diptera: Calliphoridae) flies from a suburban farm by 16S rRNA gene sequencing

The black blow fly, Phormia regina (Meigen) (Diptera: Calliphoridae) is one of the most abundant carrion flies in North America. Calliphorids are important in agriculture and animal production, veterinary sciences, forensics and medical entomology. While the role of flies in the epidemiology of human and animal diseases is an active area of research, little is known about the microorganisms associated with these insects. We examined the diversity of wild-caught black blow fly endogenous (internal body) and exogenous (external body) microbial communities using 16S rRNA gene sequencing. Overall, 27 phyla, 171 families and 533 genera were detected, and diversity was significantly higher (P < 0.05) on external body surfaces. At the genus level, Dysgonomonas, Ignatzschineria, Acinetobacter, Vagococcus, Myroides, and Wohlfahrtiimonas were predominant. Cloning and sequencing of nearly full-length fragments of the 16S rRNA gene showed that some of the species identified are known to be pathogenic to humans, animals, and plants. Myroides odoratimimus and Acinetobacter radioresistens are well-known, multi-drug resistant bacteria. These results provide a snapshot of the microbial communities harbored by adult black blow flies and call for more comprehensive studies to better characterize the role these flies may play in the transmission of pathogenic microorganisms.

Mammary microbiome of lactating organic dairy cows varies by time, tissue site, and infection status.

Infections of the cow udder leading to mastitis and reducing milk quality are a critical challenge facing all dairy farmers. Mastitis may be linked to the ecological disruption of an endogenous mammary microbial community, suggesting an ecosystems approach to management and prevention of this disease. The teat end skin represents a first point of host contact with mastitis pathogens and may offer an opportunity for microbially mediated resistance to infection, yet we know little about the microbial community of teat end skin or its potential interaction with the microbial community of intramammary milk of organic dairy cattle. High-throughput sequencing of marker genes for bacterial and fungal communities was used to characterize the skin and milk microbiome of cows with both a healthy and infected gland (i.e., udder quarter) and to assess the sharing of microbial DNA between these tissue habitat sites. The mammary microbiome varied among cows, through time, and between skin and milk. Microbiomes of milk from healthy and infected quarters reflected a diverse group of microbial DNA sequences, though milk had far fewer operational taxonomic units (OTUs) than skin. Milk microbiomes of infected quarters were generally more variable than healthy quarters and were frequently dominated by a single OTU; teat end skin microbiomes were relatively similar between healthy and infected quarters. Commonly occurring genera that were shared between skin and milk of infected glands included Staphylococcus spp. bacteria and Debaryomyces spp. fungi. Commonly occurring genera that were shared between skin and milk of healthy glands included bacteria SMB53 (Clostridiaceae) and Penicillium spp. fungi. Results support an ecological interpretation of the mammary gland and the notion that mastitis can be described as a dysbiosis, an imbalance of the healthy mammary gland microbiome.

1490 Ursodeoxycholic Acid for the Management of Recurrent Clostridioides difficile Infection

INTRODUCTION: Fecal microbiota transplant (FMT) is an effective treatment for recurrent Clostridioides difficile infection (rCDI), but its precise mechanism remains unknown (1-2). Recent studies suggest that the restoration of gut bacteria, which carry necessary enzymes to convert primary bile acids (PBAs) to secondary bile acids (SBAs), may play a key role in rCDI risk (3, Figure 1). For example, in vitro studies have demonstrated that PBAs stimulate C. difficile spore germination and that deoxycholic acid, a SBA, inhibits toxin-producing C. difficile (4). As normal fecal bile acid pools consist predominantly of SBAs, we hypothesize that a contributor of rCDI risk is the decrease in SBAs from enteric bacteria loss following antibiotic use (5-8). We present the first case enrolled in our clinical trial using ursodeoxycholic acid (UDCA), a SBA, as an adjunct treatment to prevent future rCDI. CASE DESCRIPTION/METHODS: An otherwise healthy 46-year-old man presented with S. aureusbacteremia secondary to septic knee arthritis. While on an extended course of cefazolin, he developed profuse, watery diarrhea and stool C. difficile toxin PCR was positive. He was then treated with intravenous metronidazole for one week and oral vancomycin for two weeks, with resolution of symptoms. Six weeks later, CDI recurred and was successfully treated with oral vancomycin for two weeks, in conjunction with concurrent UDCA 300 mg twice daily for eight weeks. Stool samples were collected at two-week intervals and fecal bile acid compositions were assessed via liquid chromatography - mass spectrometry. Fecal bile acid analyses demonstrated a shift towards protective SBAs by the end of the 8-week course of ursodiol (Figure 2). This profile of increased SBAs remained consistent despite subsequent retreatment with cefazolin at 5 months for recurrent septic arthritis. The patient has been without rCDI now for almost 2 years. DISCUSSION: The findings in this case are consistent with our hypothesis that increasing SBAs through adjunct treatment with UDCA can be effective in breaking the rCDI cycle. While some CDI patients have adequate enteric bacteria to produce SBAs after antibiotic treatment, others may have a delayed normalization of their gut microbiome, and may therefore require “bridge therapy” with supplementary SBAs (e.g., UDCA) until their endogenous microbial community is restored. Treatment and analysis of additional patients with rCDI in this clinical trial are in progress.

Mineralization and Biotransformation of Estrone in Simulated Poultry Litter and Cow Manure Runoff Water.

Application of animal manure on agricultural lands is one of the main sources of estrogen contamination in the environment. Poultry and cow manure contain free and conjugated forms of the natural estrogens (e.g., estrone [E1] and estradiol [E2]) that can enter surface waters during runoff events. Estrone has been identified as the major form of estrogen in the environment; therefore, this study is focused on the evaluation of the degree of mineralization and fate of E1 in a simulated poultry litter and cow manure runoff water. A time-course study was conducted using simulated runoff water that consisted of 0.5 mg cow manure or poultry litter dissolved in 1 L of water spiked with radiolabeled E1 (C-E1). Samples were analyzed for estrogen concentrations at Day 0, 0.5, 1, 2, 3, 5, and 7 using liquid chromatography with tandem mass spectrometry. In the poultry litter simulated runoff water, E1 was biotransformed to 17β-estrone-3-sulfate (E1-3S) but was eventually mineralized to CO; a total E1 mineralization of 92.2% occurred after 7 d of aerobic incubation. In contrast, the concentrations of E1 and other forms of endogenous estrogens detected in the cow manure simulated runoff water, such as E1-3S, 17α-estradiol (α-E2), and 17β-estradiol (β-E2), remained relatively constant and persisted over the 7 d of aerobic incubation. Results of this study demonstrate the differences in the fate of estrone in the simulated poultry litter and cow manure runoff water, highlighting the ability of the endogenous microbial community from poultry litter to mineralize estrogens to CO.

Growth promotion of greenhouse tomatoes with Pseudomonas sp. and Bacillus sp. biofilms and planktonic cells

The effects on plants of commercial applications of biofertilizer and biocontrol agents containing planktonic rhizobacteria cells are inconsistent mainly due to an unsuccessful competition with existing endogenous microbial communities. The present study investigated rhizobacteria inoculants in the form of biofilms, which are dense colonies of single or multi-species microbial cells, adherent to either biotic or abiotic surfaces, and encased in a self-produced matrix composed of extracellular polymeric substances. Bacillus sp., Pseudomonas sp., or a mixed inoculum of both bacteria at the planktonic or biofilm stage of growth were applied to greenhouse-grown tomato (Solanum lycopersicum L. cv Trust) plants fertilized with a half strength Hoagland solution or a modified solution lacking phosphate or iron. Based on multivariate analysis of variance, availability of nutrients, bacteria, and stages of bacterial growth significantly affected plant variables known to be contributing factors to greenhouse tomato yield. Results suggest that Bacillus sp. and Pseudomonas sp. biofilms are more effective inoculants than planktonic bacteria cells. Pseudomonas sp. applied as a biofilm increased the height and root dry weight of tomato plants, but it inhibited the positive effects of inoculation on tomato leaf number and tomato root length. Path analysis identified positive causal paths from (Pseudomonas) biofilm and planktonic cells, to the tomato plant growth variables. Presence of Bacillus sp. alone or in a mixture inhibited the positive effects of inoculation on tomato plant leaf area and root dry weight. Nevertheless, the effect of Bacillus sp. on root length was relatively positive. Results suggest that the use of innovative rhizobacteria biofilm technologies could prove advantageous as biofertilizer agents, and alleviate the dependence of greenhouse tomato producers on non-biological agro-chemicals.

Effects of endogenous inhibitors on the evolution of antibiotic resistance genes during high solid anaerobic digestion of swine manure

Livestock manure is a reservoir of antibiotic resistance genes (ARGs). The endogenous inhibitors that emerge during high solid anaerobic digestion (HSAD) greatly influence the bioprocess performance and the ARGs. The effect of endogenous inhibitors including volatile fatty acids (VFAs) and free ammonia (FA) on the ARG occurrences during HSAD of swine manure was investigated in this study. The ARG properties during HSAD (8%–14% total solids (TS)) largely differed from the low TS control (4%). The endogenous inhibitors and microbial communities greatly contributed to the three-phase changes in ARGs. The concentrations of VFAs and abundances of Proteobacteria and intI1 were correlated with the changes in ARGs. FA inhibition and VFA accumulation (especially propionate) delayed and restricted the decline of ARGs. The relatively slow rate of changes in ARGs and high ARGs in the end product suggested the high risk of the HSAD digestate for land application.

Revelation of mechanism for aqueous saponins content decrease during storage of Dioscorea zingiberensis C. H. Wright tubers: An essential prerequisite to ensure clean production of diosgenin

Releasing saponins to aqueous phase from Dioscorea zingiberensis C. H. Wright tubers is an effective way to decrease water pollution in diosgenin industry. Aqueous saponins (AS) are unstable during storage and could be hydrolyzed under certain conditions. To preserve the maximum AS content, different storage methods were evaluated, and it was found that storage at −20 °C and storage after 80 °C drying could well maintain AS content stability. AS model solution system was then constructed and utilized to simulate storage process. Adding sodium dodecyl sulphate and boiling can both effectively ensure the stability of AS content, while acid shock deferred the decrease of AS content. AS content decreased after sequential effects of endogenous enzymes and microbial community from tubers in different phase during storage. Myroides, Enterococcus, Alcaligenes and Stenotrophomonas were enriched as storage proceeded. It was also concluded that multiple glycosidase and surfactants secreted by selectively enriched microbial community decreased AS content stability at late stage of storage. This study also provides a guideline for preventing the loss of glycoside bioactive constituents in other industrial crops during storage.