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PubMed2000

Use of 16S-rRNA to investigate microbial population dynamics during biodegradation of toluene and phenol by a binary culture.

Interspecies interactions and changes in the rate and extent of biodegradation in mixed culture-mixed substrate studies were investigated. A binary mixed culture of Pseudomonas putida F1 and Burkholderia sp. JS150 degraded toluene, phenol, and their mixture. Both toluene and phenol can serve as sole sources of carbon and energy for both P. putida F1 and strain JS150. To investigate the population dynamics of this system, a fluorescent in-situ hybridization method was chosen because of its ability to produce quantitative data, its low standard error, and the ease of use of this method. When the binary mixed culture was grown on toluene or phenol alone, significant interactions between the species were observed. These interactions could not be explained by a pure-and-simple competition model and were substrate dependent. Strain JS150 growth was slightly inhibited when grown with P. putida F1 on phenol, and P. putida F1 grew more rapidly than expected. Conversely, when the two species were grown together on toluene alone, P. putida F1 was inhibited while strain JS150 was unaffected. During growth of the mixed culture on a combination of toluene and phenol, the interactions were similar to that observed during growth on phenol alone; P. putida F1 growth was enhanced while strain JS150 was unaffected. Because of the observed interspecies interactions, monoculture kinetic parameters were not sufficient to describe the mixed culture kinetics in any experiment. This is one of the first reports of microbial population dynamics in which molecular microbial ecology and mathematical modeling have been combined. The use of the 16S-rRNA-based method allowed for observation and understanding of interspecies interactions that were not observable with standard culture-based methods. These results suggest the need for more investigations that account for both substrate and microbial interactions when predicting the fate of organic pollutants in real systems.

Biodegradation, Environmental↗

PubMed2006

Prediction of glycolipid-binding domains from the amino acid sequence of lipid raft-associated proteins: application to HpaA, a protein involved in the adhesion of Helicobacter pylori to gastrointestinal cells.

Protein-glycolipid interactions mediate the attachment of various pathogens to the host cell surface as well as the association of numerous cellular proteins with lipid rafts. Thus, it is of primary importance to identify the protein domains involved in glycolipid recognition. Using structure similarity searches, we could identify a common glycolipid-binding domain in the three-dimensional structure of several proteins known to interact with lipid rafts. Yet the three-dimensional structure of most raft-targeted proteins is still unknown. In the present study, we have identified a glycolipid-binding domain in the amino acid sequence of a bacterial adhesin (Helicobacter pylori adhesin A, HpaA). The prediction was based on the major properties of the glycolipid-binding domains previously characterized by structural searches. A short (15-mer) synthetic peptide corresponding to this putative glycolipid-binding domain was synthesized, and we studied its interaction with glycolipid monolayers at the air-water interface. The synthetic HpaA peptide recognized LacCer but not Gb3. This glycolipid specificity was in line with that of the whole bacterium. Molecular modeling studies gave some insights into this high selectivity of interaction. It also suggested that Phe147 in HpaA played a key role in LacCer recognition, through sugar-aromatic CH-pi stacking interactions with the hydrophobic side of the galactose ring of LacCer. Correspondingly, the replacement of Phe147 with Ala strongly affected LacCer recognition, whereas substitution with Trp did not. Our method could be used to identify glycolipid-binding domains in microbial and cellular proteins interacting with lipid shells, rafts, and other specialized membrane microdomains.

Adhesins, Bacterial↗

PubMed2002

Toll-like receptors.

The ability of a host to sense invasion by pathogenic organisms and to respond appropriately to control infection is paramount to survival. In the case of sepsis and septic shock, however, an exaggerated systemic response may, in fact, contribute to the morbidity and mortality associated with overwhelming infections. The innate immune system has evolved as the first line of defense against invading microorganisms. The Toll-like receptors (TLRs) are a part of this innate immune defense, recognizing conserved patterns on microorganisms. These TLRs and their signaling pathways are represented in such diverse creatures as mammals, fruit flies, and plants. Ten members of the TLR family have been identified in humans, and several of them appear to recognize specific microbial products, including lipopolysaccharide, bacterial lipoproteins, peptidoglycan, and bacterial DNA. Signals initiated by the interaction of TLRs with specific microbial patterns direct the subsequent inflammatory response. Thus, TLR signaling represents a key component of the innate immune response to microbial infection.

Animals↗

PubMed2002

Toll-like receptors.

Journal Article↗

PubMed2006

An indigenous drought-tolerant strain of Glomus intraradices associated with a native bacterium improves water transport and root development in Retama sphaerocarpa.

The effects of interactions between Bacillus thuringiensis, a drought-adapted bacterium, and two isolates of Glomus intraradices, an arbuscular mycorrhizal (AM) fungus, on Retama sphaerocarpa, a drought-adapted legume, were investigated. The fungal isolates were an indigenous drought-tolerant and a nonindigenous drought-sensitive isolate. Shoot length and root growth, symbiotic parameters, water transport (in terms of percent relative plant water uptake), and volumetric soil moisture and soil enzymatic activities in response to microbial inoculations were evaluated. Retama plants colonized by G. intraradices plus Bacillus possessed similar shoot length after 30 days from sowing compared with noninoculated Retama plants after 150 days. Inoculation with drought-adapted bacterium increased root growth by 201%, but maximum root development was obtained by co-inoculation of B. thuringiensis and the indigenous G. intraradices. Nodules were formed only in plants colonized by autochthonous AM fungi. Relative water uptake was higher in inoculated than in noninoculated Retama plants, and these inoculants depleted soil water content concomitantly. G. intraradices-colonized Retama reached similar shoot length irrespective of the fungal origin, but there were strong differences in relative water uptake by plants colonized by each one of the fungi. Indigenous G. intraradices-colonized roots (evaluated as functional alkaline phosphatase staining) showed the highest intensity and arbuscule richness when associated with B. thuringiensis. The interactive microbial effects on Retama plants were more relevant when indigenous microorganisms were involved. Co-inoculation of autochthonous microorganisms reduced by 42% the water required to produce 1 mg of shoot biomass. This is the first evidence of the effectiveness of rhizosphere bacterium, singly or associated with AM fungus, in increasing plant water uptake, which represents a positive microbial effect on plants grown under drought environments.

Bacillus↗

PubMed2024

Genetic modification of the shikimate pathway to reduce lignin content in switchgrass (Panicum virgatum L.) significantly impacts plant microbiomes.

UNLABELLED: Switchgrass (Panicum virgatum L.) is considered a sustainable biofuel feedstock, given its fast-impact growth, low input requirements, and high biomass yields. Improvements in bioenergy conversion efficiency of switchgrass could be made by reducing its lignin content. Engineered switchgrass that expresses a bacterial 3-dehydroshikimate dehydratase (QsuB) has reduced lignin content and improved biomass saccharification due to the rerouting of the shikimate pathway towards the simple aromatic protocatechuate at the expense of lignin biosynthesis. However, the impacts of this QsuB trait on switchgrass microbiome structure and function remain unclear. To address this, wild-type and QsuB-engineered switchgrass were grown in switchgrass field soils, and samples were collected from inflorescences, leaves, roots, rhizospheres, and bulk soils for microbiome analysis. We investigated how QsuB expression influenced switchgrass-associated fungal and bacterial communities using high-throughput Illumina MiSeq amplicon sequencing of ITS and 16S rDNA. Compared to wild-type, QsuB-engineered switchgrass hosted different microbial communities in roots, rhizosphere, and leaves. Specifically, QsuB-engineered plants had a lower relative abundance of arbuscular mycorrhizal fungi (AMF). Additionally, QsuB-engineered plants had fewer Actinobacteriota in root and rhizosphere samples. These findings may indicate that changes in the plant metabolism impact both AMF and Actinobacteriota similarly or potential interactions between AMF and the bacterial community. This study enhances understanding of plant-microbiome interactions by providing baseline microbial data for developing beneficial bioengineering strategies and by assessing nontarget impacts of engineered plant traits on the plant microbiome. IMPORTANCE: Bioenergy crops provide an important strategy for mitigating climate change. Reducing the lignin in bioenergy crops could improve fermentable sugar yields for more efficient conversion into bioenergy and bioproducts. In this study, we assessed how switchgrass engineered for low lignin impacted aboveground and belowground switchgrass microbiome. Our results show unexpected reductions in mycorrhizas and actinobacteria in belowground tissues, raising questions on the resilience and function of genetically engineered plants in agricultural systems.

Panicum↗

PubMed2005

Bacteriophages--potential for application in wastewater treatment processes.

Bacteriophages are viruses that infect and lyse bacteria. Interest in the ability of phages to control bacterial populations has extended from medical applications into the fields of agriculture, aquaculture and the food industry. Here, the potential application of phage techniques in wastewater treatment systems to improve effluent and sludge emissions into the environment is discussed. Phage-mediated bacterial mortality has the potential to influence treatment performance by controlling the abundance of key functional groups. Phage treatments have the potential to control environmental wastewater process problems such as: foaming in activated sludge plants; sludge dewaterability and digestibility; pathogenic bacteria; and to reduce competition between nuisance bacteria and functionally important microbial populations. Successful application of phage therapy to wastewater treatment does though require a fuller understanding of wastewater microbial community dynamics and interactions. Strategies to counter host specificity and host cell resistance must also be developed, as should safety considerations regarding pathogen emergence through transduction.

Bacteria↗

PubMed2002

Introduction: innate recognition of bacteria and protozoan parasites.

Major advances have recently been achieved in the area of microbial recognition by the innate immune system. In this Forum, we discuss important issues related to innate recognition of bacteria and protozoan parasites. In particular, we highlight the structural characterization of pathogen-associated molecular patterns (PAMPs); the definition of the receptors required for recognition of PAMPs, especially the Toll-like receptors (TLRs); the signaling pathways triggered by PAMPs/PAMPs receptor interaction; and the functional consequences of these interactions for pathogenesis during microbial infection.

Animals↗

PubMed2001

Combined effects of two sulfonylurea herbicides on soil microbial biomass and N-mineralization.

The interaction effect of two sulfonylurea herbicides, bensulfuron-methyl (B) and metsulfuron-methyl(M), were tested on microbial biomass C, N, N-mineralization and C/N ratio in a loamy sand soil. The herbicides were applied at various levels of: control (B0M0), 0.01 and 0.01 (B1M1), 0.01 and 0.1 (B1M2), and 0.01 and 1.0 (B1M3) microgram/g soil. Determinations of soil microbial biomass-C, N and N-mineralization contents were carried out at 1, 3, 5, 7, 10, 15, 25 and 45 days after herbicides application. The results showed that the soil microbial biomass-C (Cmic) and microbial biomass-N (Nmic) decreased consistently with the increasing rates of herbicides. The results further indicated that B1M1 and B1M2 caused a significant reduction in Cmic and Nmic within first 10 and 7 days of incubation, respectively, as compared with the control. These reductions in Cmic and Nmic were also significant (P = 0.05) with B1M3 application especially within first 15 days of incubation. A significant reduction in N-mineralization (N-min) was observed with high doses (B1M2, B1M3) of herbicides within first 5 days of incubation, while low rate (B1M1) failed to produce any significant effect. An increase in the soil microbial biomass C:N ratio was also noted.

Analysis of Variance↗

PubMed2002

[Microbial resistance to antibiotics from the clinical viewpoint. Paradoxical therapy failures and the significance of pharmacokinetic-pharmacodynamic interactions in vivo].

Antimicrobial efficacy in vivo is not exclusively defined by the activity of an antibiotic as determined in the in vitro susceptibility test where constant inocula of log-phase bacteria are exposed to a steady concentration of drug for short periods of time in a well defined biochemical environment tailored to optimize antimicrobial activity. In vivo, many additional factors come into play which may be decisive for the clinical result of antimicrobial therapy. Clinically, high numbers of bacteria in various states of growth and metabolic activity are to be eliminated in an environment which may be unfavorable for antimicrobial activity. Low pH, low pO2, the presence of proteins, cell debris and biofilms, are all known to interfere with antimicrobial activity in vivo. Moreover, drug concentrations in vivo are fluctuating and vary greatly depending on a variety of pharmacokinetic factors. Finally, clinical treatment usually lasts for days. The impact of the pharmacokinetic profile of an antibiotic on its in vivo activity may be decisive. It is dependent on both, the type of drug in question and its target organisms. The time of supra-MIC concentrations appears to be the main parameter for the activity of macrolides, clindamycin, linezolide and betalactam antibiotics, particularly against gram-negative organisms. In contrast, peak concentrations and the AUC/MIC relationship are key parameters for the activity of aminoglycosides and fluoroquinolones. Regarding upcoming resistant bacterial strains, an as yet ill-defined mutation-prevention concentration has recently been discussed. Thus, in addition to the aim of eliminating target bacteria at the site of infection, in vivo concentrations of antibiotics may be important for both, the suppression of antimicrobial resistance and its epidemic spread. We are looking forward to prospective clinical trials to better understand the clinical relevance of the discussed phenomena.

Anti-Bacterial Agents↗

PubMed2003

The dendritic cell: its role in intestinal inflammation and relationship with gut bacteria.

Dendritic cells are antigen presenting cells that are likely to be pivotal in the balance between tolerance and active immunity to commensal microorganisms that is fundamental to inflammatory conditions, including Crohn's disease and ulcerative colitis. Interactions between dendritic cells and microbial products are discussed and how they contribute to regulation of immune responses. The concept that interactions between dendritic cells and commensal organisms may be responsible for maintaining intestinal immune homeostasis is also explored.

Bacterial Infections↗

PubMed1998

Different mathematical approaches to estimating microbial protein supply in ruminants.

Many of the amino acids that are available for absorption in ruminants are derived from microbial protein that has been synthesized in the reticulorumen. This paper focuses on the prediction of the microbial protein supply and evaluates different approaches to represent mathematically the process of microbial protein synthesis. In current protein evaluation systems for ruminants, the microbial protein supply is predicted using empirical equations that relate microbial protein production to the amounts of ruminally available energy and nitrogen. In contrast, mechanistic models of rumen function endeavor to describe quantitatively the microbial protein production that is based on underlying identifiable processes. A brief description is presented of two culture techniques used to examine microbial ecosystems, namely, batch culture and chemostat culture. The mathematical equations describing these cultures are helpful in understanding key parameters of microbial production for inclusion in models, including specific growth rate, growth yield, and substrate affinity. The availability of carbohydrates is a primary determinant of microbial protein production in the rumen, and the adequacy of mathematical representations of this relationship in empirical and mechanistic models is assessed. The representation of substrate utilization for nongrowth processes and the relationship between microbial protein production and the availability of various nitrogen sources are discussed. A variable part of the synthesized microbial protein does not reach the duodenum but is degraded in the rumen, and its representation is examined. The prediction of microbial protein supply should be based on a sound representation of the underlying mechanisms, including the interactions among microbes and between microbial activity and substrate degradation.

Animals↗

PubMed1990

Raman spectroscopy of synthetic antimicrobial frog peptides magainin 2a and PGLa.

Magainin and PGLa are 23- and 21-residue peptides isolated from the skin of the African clawed frog Xenopus laevis. They protect the frog from infection and exhibit a broad-spectrum antimicrobial activity in vitro. The mechanism of this activity involves the interaction of magainin with microbial membranes. We have measured the secondary structure and membrane-perturbing ability of these peptides to obtain information about this mechanism. Our results show that mgn2a forms a helix with an average length of less than 20 A upon binding to liposomes. At high concentrations (50 mg/mL) mgn2a spontaneously solubilizes phosphatidylcholine liposomes at temperatures above the gel-liquid-crystalline phase transition. Mgn2a appears to bind to the surface of liposomes made of negatively charged lipids without spontaneously penetrating the bilayer. Finally, mgn2a and PGLa interact together with liposomes in a synergistic way that enhances the helix content of one or both of the peptides and allows the peptides to more easily penetrate the bilayer. PGLa mixed with a small nonperturbing amount of magainin 2 amide is 25-43 times as potent as PGLa alone at inducing the release of carboxyfluorescein from liposomes. The results suggest that the mechanism of antimicrobial activity does not involve a channel formed by transmembrane helical peptides.

1,2-Dipalmitoylphosphatidylcholine↗

PubMed2002

Comparative ecology of H2 cycling in sedimentary and phototrophic ecosystems.

The simple biochemistry of H2 is critical to a large number of microbial processes, affecting the interaction of organisms with each other and with the environment. The sensitivity of each of these processes to H2 can be described collectively, through the quantitative language of thermodynamics. A necessary prerequisite is to understand the factors that, in turn, control H2 partial pressures. These factors are assessed for two distinctly different ecosystems. In anoxic sediments from Cape Lookout Bight (North Carolina, USA), H2 partial pressures are strictly maintained at low, steady-state levels by H2-consuming organisms, in a fashion that can be quantitatively predicted by simple thermodynamic calculations. In phototrophic microbial mats from Baja California (Mexico), H2 partial pressures are controlled by the activity of light-sensitive H2-producing organisms, and consequently fluctuate over orders of magnitude on a daily basis. The differences in H2 cycling can subsequently impact any of the H2-sensitive microbial processes in these systems. In one example, methanogenesis in Cape Lookout Bight sediments is completely suppressed through the efficient consumption of H2 by sulfate-reducing bacteria; in contrast, elevated levels of H2 prevail in the producer-controlled phototrophic system, and methanogenesis occurs readily in the presence of 40 mM sulfate.

Bacteria↗

PubMed1999

Escherichia coli and Porphyromonas gingivalis lipopolysaccharide interactions with CD14: implications for myeloid and nonmyeloid cell activation.

Porphyromonas gingivalis, a gram-negative bacterium, is an etiologic agent for adult periodontitis. Lipopolysaccharide (LPS) released from this bacterium can react with numerous host cell types. P. gingivalis LPS stimulates tumor necrosis factor alpha and interleukin-1beta secretion from monocytes (myeloid) but does not elicit E-selectin expression from human endothelial cells (nonmyeloid). In contrast, Escherichia coli LPS facilitates expression of these inflammatory mediators through CD14-dependent pathways on both myeloid and nonmyeloid cells. LPS binding studies have revealed that although P. gingivalis and E. coli LPSs bind to CD14 differently, this fact does not adequately explain the lack of endothelial cell activation by P. gingivalis LPS. Rather, LPS binding site and blocking monoclonal antibody epitope mapping studies have suggested that CD14 presents a charged surface that captures different microbial ligands by electrostatic interactions. We propose that human endothelial cells do not respond to P. gingivalis LPS because of their inability to "recognize" CD14-P. gingivalis LPS complexes.

Amino Acid Sequence↗

PubMed2025

The Streptococcus pyogenes mannose phosphotransferase system (Man-PTS) influences antimicrobial activity and niche-specific nasopharyngeal infection.

Streptococcus pyogenes is a human-adapted pathogen that can cause multiple diseases, including pharyngitis and skin infections. Although this bacterium produces many virulence factors, how S. pyogenes competes with the host microbiota is not well understood. Here, we detected antimicrobial activity from S. pyogenes MGAS8232 that prevented the growth of Micrococcus luteus. This activity was produced when cells were grown in 5% CO2 in M17 media supplemented with galactose; however, the addition of alternative sugars coupled with genome sequencing experiments revealed that the antimicrobial phenotype was not related to classical bacteriocins. To further determine genes involved in the production of this activity, a transposon mutant library in S. pyogenes MGAS8232 identified the mannose phosphotransferase system (Man-PTS), a major sugar transporter, as important for the antimicrobial phenotype. Loss-of-function transposon mutants linked to the antimicrobial activity were identified to also be involved in alternative sugar utilization, and additionally, the Man-PTS was further identified from an inadvertent secondary mutation in a bacteriocin operon mutant. Sugar utilization in the Man-PTS mutants demonstrated that galactose, mannose, and N-acetylglucosamine utilization was impaired. RNA-seq experiments in high and low glucose concentrations further characterized the Man-PTS as a glucose transporter; however, transcriptional regulators or virulence factors were not affected with the loss of the Man-PTS. Deletion of Man-PTS demonstrated defects in a mouse model of nasopharyngeal infection but not skin infection. This work suggests that the ability of S. pyogenes to utilize alternative sugars presented by glycans may play a role in acute infection and interactions with the endogenous microbial population existing in the nasopharynx.IMPORTANCEStreptococcus pyogenes is responsible for over 500,000 deaths per year primarily due to invasive infections and post-infection sequelae, although the most common manifestations include pharyngitis and impetigo. S. pyogenes can adapt to its environment through alternative sugar metabolism. Here, we identified an antimicrobial phenotype that was not bacteriocin-related but a by-product of alternative sugar metabolism. The mannose phosphotransferase system was involved in the production of the antimicrobial and was also important for S. pyogenes to utilize alternative sugars and establish nasopharyngeal infection but not skin infection. Overall, this study identified potential strategies used by S. pyogenes for interactions with the endogenous microbiota and further elucidated the importance of sugar metabolism in acute upper respiratory tract infection.

Streptococcus pyogenes↗

PubMed1994

Antimicrobial peptides of lactoferrin.

Lactoferrin was found to contain an antimicrobial sequence near its N-terminus which appears to function by a mechanism distinct from iron chelation. Antimicrobial peptides representing this domain were isolated following pepsin cleavage of human lactoferrin and bovine lactoferrin. The antimicrobial sequence was found to consist mainly of a loop of 18 amino acid residues formed by a disulfide bond between cysteine residues 20 and 37 of human lactoferrin, or 19 and 36 of bovine lactoferrin. The identified domain contains a high proportion of basic residues, like various other antimicrobial peptides known to target microbial membranes and it appears to be located on the surface of the folded protein allowing its interaction with surface components of microbial cells. The isolated domain, "lactoferrin", was shown to have potent broad spectrum antimicrobial properties and its effect was lethal causing a rapid loss of colony-forming capability. Such evidence points to the conclusion that this domain is the structural region responsible for the microbicidal properties of lactoferrin. The evidence also suggests the possibility that active peptides produced by enzymatic digestion of lactoferrin may contribute to the host defense against microbial disease.

Amino Acid Sequence↗

PubMed2006

Preferential feeding by an aquatic consumer mediates non-additive decomposition of speciose leaf litter.

Forest soils and streams receive substantial inputs of detritus from deciduous vegetation. Decay of this material is a critical ecosystem process, recycling nutrients and supporting detrital-based food webs, and has been attributed, in part, to leaf litter species composition. However, research on why speciose leaf litter should degrade differently has relied on a bottom-up approach, embracing interspecific variation in litter chemistry. We hypothesized that preferential feeding by an aquatic detritivore interacts with species-specific leaf palatability and slows decay of speciose leaf litter. We addressed this by offering four single- and mixed-species leaf resources to field densities of a leaf-shredding consumer. Mixing leaf species resulted in slower total leaf decomposition. Decreases in mixed-species decomposition was partly explained by preferential feeding by the consumers in one case, but the lack of preferential feeding in other mixtures suggested an interactive effect of feeding and microbial degradation. Loss of riparian tree biodiversity may have implications for in-stream consumer-resource interactions.

Analysis of Variance↗