Metabolic Status of Bacteria and Fungi in the Rhizosphere of Ponderosa Pine Seedlings

We determined the quantity and metabolic status of bacteria and fungi in rhizosphere and nonrhizosphere soil from microcosms containing ponderosa pine seedlings. Rhizosphere soil was sampled adjacent to coarse, fine, or young roots. The biovolume and metabolic status of bacterial and fungal cells was determined microscopically and converted to total and active biomass values. Cells were considered active if they possessed the ability to reduce the artificial electron acceptor 2-(4-iodophenyl)-3-(4-nitrophenyl)-5-phenyltetrazolium chloride (INT) to visible intracellular deposits of INT formazan. A colorimetric assay of INT formazan production was also used to assess dehydrogenase activity. INT-active microorganisms made up 44 to 55% of the microbial biomass in the soils studied. The proportion of fungal biomass that exhibited INT-reducing activity (40 to 50%) was higher than previous estimates of the active proportion of soil fungi determined by using fluorescein diacetate. Comparison between soils from different root zones revealed that the highest total and INT-active fungal biomass was adjacent to fine mycorrhizal roots, whereas the highest total and active bacterial biomass was adjacent to the young growing root tips. These observations suggest that fungi are enhanced adjacent to the fine roots compared with the nonrhizosphere soil, whereas bacteria are more responsive than fungi to labile carbon inputs in the young root zone. Colorimetric dehydrogenase assays detected gross differences between bulk and rhizosphere soil activity but were unable to detect more subtle differences due to root types. Determination of total and INT-active biomass has increased our understanding of the role of spatial compartmentalization of bacteria and fungi in rhizosphere carbon flow.     

Importance of Saprotrophic Freshwater Fungi for Pollen Degradation

Fungi and bacteria are the major organic matter (OM) decomposers in aquatic ecosystems. While bacteria are regarded as primary mineralizers in the pelagic zone of lakes and oceans, fungi dominate OM decomposition in streams and wetlands. Recent findings indicate that fungal communities are also active in lakes, but little is known about their diversity and interactions with bacteria. Therefore, the decomposer niche overlap of saprotrophic fungi and bacteria was studied on pollen (as a seasonally recurring source of fine particulate OM) by performing microcosm experiments with three different lake types. Special emphasis was placed on analysis of fungal community composition and diversity. We hypothesized that (I) pollen select for small saprotrophic fungi and at the same time for typical particle-associated bacteria; (II) fungal communities form specific free-living and attached sub-communities in each lake type; (III) the ratio between fungi or bacteria on pollen is controlled by the lake''s chemistry. Bacteria-to-fungi ratios were determined by quantitative PCR (qPCR), and bacterial and fungal diversity were studied by clone libraries and denaturing gradient gel electrophoresis (DGGE) fingerprints. A protease assay was used to identify functional differences between treatments. For generalization, systematic differences in bacteria-to-fungi ratios were analyzed with a dataset from the nearby Baltic Sea rivers. High abundances of Chytridiomycota as well as occurrences of Cryptomycota and yeast-like fungi confirm the decomposer niche overlap of saprotrophic fungi and bacteria on pollen. As hypothesized, microbial communities consistently differed between the lake types and exhibited functional differences. Bacteria-to-fungi ratios correlated well with parameters such as organic carbon and pH. The importance of dissolved organic carbon and nitrogen for bacteria-to-fungi ratios was supported by the Baltic Sea river dataset. Our findings highlight the fact that carbon-to-nitrogen ratios may also control fungal contributions to OM decomposition in aquatic ecosystems.     

The Ability of Saprotrophic Bacteria Isolated from Natural Habitats to Lyse Yeasts

More than 600 bacterial strains isolated from different horizons of steppe biogeocenoses and zoogenous loci (diplopod intestines and feces) were tested for the ability to lyse yeast cell walls. About half of the strains that were isolated from biotopes with active degradation of plant debris (steppe litters and diplopod intestines and feces) were found to possess yeast-lytic activity. Most of the yeast-lytic strains belonged to the genera Streptomyces, Promicromonospora, Oerskovia, and Agromyces. The yeast-lytic activity of actinobacteria from the genera Agromyces, Mycobacterium, and Micrococcus has not previously been reported.     

Interaction of Rhizosphere Bacteria, Fertilizer, and Vesicular-Arbuscular Mycorrhizal Fungi with Sea Oats

Plants must be established quickly on replenished beaches in order to stabilize the sand and begin the dune-building process. The objective of this research was to determine whether inoculation of sea oats (Uniola paniculata L.) with bacteria (indigenous rhizosphere bacteria and N₂ fixers) alone or in combination with vesicular-arbuscular mycorrhizal fungi would enhance plant growth in beach sand. At two fertilizer-N levels, Klebsiella pneumoniae and two Azospirillum spp. did not provide the plants with fixed atmospheric N; however, K. pneumoniae increased root and shoot growth. When a sparingly soluble P source (CaHPO₄) was added to two sands, K. pneumoniae increased plant growth in sand with a high P content. The phosphorus content of shoots was not affected by bacterial inoculation, indicating that a mechanism other than bacterially enhanced P availability to plants was responsible for the growth increases. When sea oats were inoculated with either K. pneumoniae or Acaligenes denitrificans and a mixed Glomus inoculum, there was no consistent evidence of a synergistic effect on plant growth. Nonetheless, bacterial inoculation increased root colonization by vesicular-arbuscular mycorrhizal fungi when the fungal inoculum consisted of colonized roots but had no effect on colonization when the inoculum consisted of spores alone. K. pneumoniae was found to increase spore germination and hyphal growth of Glomus deserticola compared with the control. The use of bacterial inoculants to enhance establishment of pioneer dune plants warrants further study.     

Diversification of Fungal Specific Class A Glutathione Transferases in Saprotrophic Fungi

Glutathione transferases (GSTs) form a superfamily of multifunctional proteins with essential roles in cellular detoxification processes and endogenous metabolism. The distribution of fungal-specific class A GSTs was investigated in saprotrophic fungi revealing a recent diversification within this class. Biochemical characterization of eight GSTFuA isoforms from Phanerochaete chrysosporium and Coprinus cinereus demonstrated functional diversity in saprotrophic fungi. The three-dimensional structures of three P. chrysosporium isoforms feature structural differences explaining the functional diversity of these enzymes. Competition experiments between fluorescent probes, and various molecules, showed that these GSTs function as ligandins with various small aromatic compounds, derived from lignin degradation or not, at a L-site overlapping the glutathione binding pocket. By combining genomic data with structural and biochemical determinations, we propose that this class of GST has evolved in response to environmental constraints induced by wood chemistry.     

Fungi as Eukaryotes: Understanding the Antifungal Effects of Immunosuppressive Drugs

The immunosuppressive agents cyclosporine, tacrolimus, and sirolimus are naturally occurring products of environmental fungi or bacteria, so the fact that they possess intrinsic antifungal activity is not surprising. Both calcineurin and the target of rapamycin (TOR) are conserved across eukaryotes and share a common function, regulating the organism’s ability to react to environmental changes and response to stress. In the medically important fungi Candida, Cryptococcus, and Aspergillus, mutations in the calcineurin gene affect in vitro patterns of growth and serum sensitivity, and attenuate virulence in animal models. Notably, cyclosporine, tacrolimus, and sirolimus exhibit strong synergy with many classes of antifungal drugs including azoles, amphotericin B, and the echinocandins, with potentiation of fungicidal effects even against drug-resistant strains. Hopefully, future studies will realize the promise of exploiting the antifungal properties of the immunosuppressive drugs to help decrease the burden of these clinically important infections on patient survival.     

Phylogenetic Diversity of Aerobic Saprotrophic Bacteria Isolated from the Daqing Oil Field

A diverse and active microbial community in the stratal waters of the Daqing oil field (China), which is exploited with the use of water-flooding, was found to contain aerobic chemoheterotrophic bacteria (including hydrocarbon-oxidizing ones) and anaerobic fermentative, sulfate-reducing, and methanogenic bacteria. The aerobic bacteria were most abundant in the near-bottom zones of injection wells. Twenty pure cultures of aerobic saprotrophic bacteria were isolated from the stratal waters. Under laboratory conditions, they grew at temperatures, pH, and salinity values typical of the stratal water from which they were isolated. These isolates were found to be able to utilize crude oil and a wide range of hydrocarbons, fatty acids, and alcohols. Phylogenetic analysis carried out with the use of complete 16S rRNA sequences showed that the isolates could be divided into three major groups: gram-positive bacteria with a high and a low G+C content of DNA and gram-negative bacteria of the -subclass of the Proteobacteria. Gram-positive isolates belonged to the genera Bacillus, Brevibacillus, Rhodococcus, Dietzia, Kocuria, Gordonia, Cellulomonas, and Clavibacter. Gram-negative isolates belonged to the genera Pseudomonas and Acinetobacter. In their 16S rRNA sequences, many isolates were similar to the known microbial species and some probably represented new species.     

Antifungal Susceptibility Testing of Filamentous Fungi

Methods developed for testing filamentous fungi (molds) include standardized broth microdilution (Clinical and Laboratory Standards Institute [CLSI] and European Committee for Antimicrobial Susceptibility Testing [AFST-EUCAST]) methods and disk diffusion (CLSI) methods. Quality control limits also are available from CLSI for MIC (minimal inhibitory concentration), MEC (minimal effective concentration), and zone diameters. Although clinical breakpoints based on correlations of in vitro results with clinical outcome have not been established, epidemiologic cutoff values have been defined for six Aspergillus species and the triazoles, caspofungin, and amphotericin B. The link between resistance molecular mechanisms, elevated MICs, and clinical treatment failure has also been documented, especially for Aspergillus and the triazoles. Other insights into the potential clinical value of high MICs have also been reported. Various commercial methods (e.g., YeastOne, Etest, and Neo-Sensitabs) have been evaluated in comparison with reference methods. This review summarizes and discusses these developments.     

霉克舒对致病性浅部真菌的体外抑菌作用研究

从湛江地区 15 4例癣病患者分离真菌 ,评价一种新的抗真菌药—霉克舒对上述真菌菌株的体外抑菌活性。应用琼脂稀释法测定霉克舒对浅部真菌的最小抑菌浓度 (MIC) ,同时以兰美抒作为对照药物。从 15 4例癣病患者中分离出 14 1株真菌 ,其中以红色毛癣菌和须癣毛癣菌为主 ,分别占 6 6 .7%和 14 .2 %。霉克舒的抑菌作用与兰美抒相比略强或相当 ;对霉克舒各单一成分的初步抑菌效果进行比较 ,复合物的抑菌作用明显强于水杨酸或特比萘芬等单一成分。上述结果显示 ,霉克舒在体外对常见致病性浅部真菌具有较强的抗菌活性。     

Sequential and Structural Aspects of Antifungal Peptides from Animals,Bacteria and Fungi Based on Bioinformatics Tools

Emerging drug resistance varieties and hyper-virulent strains of microorganisms have compelled the scientific fraternity to develop more potent and less harmful therapeutics. Antimicrobial peptides could be one of such therapeutics. This review is an attempt to explore antifungal peptides naturally produced by prokaryotes as well as eukaryotes. They are components of innate immune system providing first line of defence against microbial attacks, especially in eukaryotes. The present article concentrates on types, structures, sources and mode of action of gene-encoded antifungal peptides such as mammalian defensins, protegrins, tritrpticins, histatins, lactoferricins, antifungal peptides derived from birds, amphibians, insects, fungi, bacteria and their synthetic analogues such as pexiganan, omiganan, echinocandins and Novexatin. In silico drug designing, a major revolution in the area of therapeutics, facilitates drug development by exploiting different bioinformatics tools. With this view, bioinformatics tools were used to visualize the structural details of antifungal peptides and to predict their level of similarity. Current practices and recent developments in this area have also been discussed briefly.     

Violet-Pigmented Pseudomonads With Antifungal Activity From the Rhizosphere of Beans

Bean rhizosphere bacteria antagonistic to four root-infecting fungi and an antibiotic produced by these bacteria were studied. The bacteria were violet-pigmented gram-negative rods, probably belonging to the genus Pseudomonas. The antibiotic, which was localized largely in the bacterial cell mass, was easily extracted with acetone. It was selectively active against a wide variety of plant-pathogenic and saprophytic fungi tested in vitro but was relatively inactive against bacteria. The compound, partially purified by chromatography, was soluble in all organic solvents tried, but nearly insoluble in water. It demonstrated no characteristic ultraviolet- or visible-absorption spectrum and was chemically unidentified. The antagonistic bacteria or crude antibiotic applied to buried buckwheat segments suppressed the colonization of this substrate by Rhizoctonia spp. The data suggested that the bacteria or the antibiotic may play a role in the suppression of root-infecting fungi in soil.     

Methanotrophic Bacteria in the Rhizosphere of Trichloroethylene-Degrading Plants

The presence and density of methanotrophic bacteria has been shown to play an important role in the bioremediation of trichloroethylene (TCE). This article describes the methanotrophic bacterial densities in rhizosphere soils from two areas of the Department of Energy's Savannah River Site in Aiken, South Carolina. A direct fluorescent antibody (DFA) technique was evaluated to determine the presence of methanotrophic bacteria in roots and rhizospheres from vascular plants. The first site, the Miscellaneous Chemical Basin (MCB), was contaminated with a mixture of chemicals, including chlorinated solvents. The second site will be potentially affected by outcropping of TCE-contaminated groundwater. Significantly higher numbers of methanotrophic bacteria were observed with DFA in rhizosphere soils and on roots of Lespedeza cuneata and Pinus taeda (that previously showed higher rates of ¹⁴C-TCE mineralization) compared with nonvegetated soils. In addition, viable and heterotrophic microbial counts were consistently higher in rhizosphere soils and on plant roots compared with nonvegetated soils. Therefore, the presence of these plant species may enhance ¹⁴C-TCE mineralization by selectively increasing the microbial population in the root zone. Methanotrophic bacteria were directly observed by DFA in soils, on the surface of root hairs, within plant roots, and in higher densities associated with mycorrhizal fungi. These experiments provide further evidence that specific types of bacterial interactions with vegetation in the rhizosphere may play an important role in remediation of TCE-contaminated soils and groundwater.     

单叶蔓荆根际土壤真菌多样性

对江西鄱阳湖区5个不同地区单叶蔓荆(Vitex trifolia)的根际土壤真菌种类和多样性进行了研究。从100份单叶蔓荆植物根际土壤样品中分离获得128株真菌,隶属于无性型菌和接合菌,共15个属,其中青霉属(Penicillium)为单叶蔓荆根际土壤真菌的优势属,占总菌株数的14.06%,其次为单端孢属(Trichothecium),占总菌株数的11.72%,葡萄孢属(Botrytis)、卷头霉属(Helicocephalum)、茎点霉属(Phoma)、根霉属(Rhizopus)占总菌株数的7.81%,可见单叶蔓荆根际真菌具有较丰富的多样性。研究还发现不同地区单叶蔓荆根际土壤真菌的种群组成和结构存在一定的差异,各属真菌在不同地区的单叶蔓荆根际土壤中所占的优势度也不同。     

Response of Bacteria to Mechanical Stimuli

Microbiology - Bacteria adapt rapidly to changes in ambient conditions, constantly inspecting their surroundings by means of their sensor systems. These systems are often thought to respond only to...     

Reactions of Pinus sylvestris (Scots Pine) Root Tissues to the Presence of Mutualistic, Saprotrophic and Necrotrophic Micro-organisms

Using Scots pine seedlings as experimental hosts, the general reactions and recognition patterns of living conifer tissues challenged in vitro with either parasitic, saprotrophic, epiphytic or mutualistic microbial species were investigated. Three distinct reaction and recognition patterns were observed. In one, a necrosis type of browning reaction occurred that restricted vascular penetration of the seedlings when inoculated with a saprotroph (Marasmius androsaceus) or a mycorrhizal-associated bacterium (Pseudomonas fluorescens). The second pattern, a kind of necrosis in which the host defence system was ultimately ineffective, was typically observed in seedlings challenged with the S or P types of the root rot fungus (Heterobasidion annosum), the fine root parasite (Fusarium avenaceum) or the saprotrophic wood decay fungus (Phlebia gigantea). The third type, described as non-recognition, was observed on seedlings showing no necrosis and was particularly common with seedlings inoculated with either mycorrhizal fungi (Suillus granulatus, Piloderma croceum) or saprotrophs (Trichoderma aureoviride, Coriolus versicolor), or an unidentified rod-shaped bacterial isolate. Since necrosis is often linked to rapid accumulation of H₂O₂, increases in peroxidase activity were determined. Only seedlings challenged with the S-type of H. annosum had a systemic increase in peroxidase levels of needles, shoot and roots. Similarly, only H. annosum, F. avenaceum, P. gigantea and the Pseudomonas bacterium caused a two- to four-fold increase in the peroxidase level of inoculated roots, results suggesting that a relationship exists between the degree of necrosis and the ratio of peroxidase induction. But the higher levels of peroxidase activity was not correlated with increased resistance to disease development.     

Can Insects Develop Resistance to Insect Pathogenic Fungi?

Microevolutionary adaptations and mechanisms of fungal pathogen resistance were explored in a melanic population of the Greater wax moth, Galleria mellonella. Under constant selective pressure from the insect pathogenic fungus Beauveria bassiana, 25^th generation larvae exhibited significantly enhanced resistance, which was specific to this pathogen and not to another insect pathogenic fungus, Metarhizium anisopliae. Defense and stress management strategies of selected (resistant) and non-selected (susceptible) insect lines were compared to uncover mechanisms underpinning resistance, and the possible cost of those survival strategies. We hypothesize that the insects developed a transgenerationally primed resistance to the fungus B. bassiana, a costly trait that was achieved not by compromising life-history traits but rather by prioritizing and re-allocating pathogen-species-specific augmentations to integumental front-line defenses that are most likely to be encountered by invading fungi. Specifically during B. bassiana infection, systemic immune defenses are suppressed in favour of a more limited but targeted repertoire of enhanced responses in the cuticle and epidermis of the integument (e.g. expression of the fungal enzyme inhibitor IMPI, and cuticular phenoloxidase activity). A range of putative stress-management factors (e.g. antioxidants) is also activated during the specific response of selected insects to B. bassiana but not M. anisopliae. This too occurs primarily in the integument, and probably contributes to antifungal defense and/or helps ameliorate the damage inflicted by the fungus or the host’s own immune responses.     

C型凝集素LSECtin识别细菌的研究

目的:通过sLSECtin-Fc蛋白与细菌及细胞的黏附,寻找能与LSECtin结合的细菌。方法:通过ELISA检测sLSECtin-Fc与细菌的黏附,同时构建CHO-pDsRed1-N1-LSECtin稳定细胞株进行与细菌的黏附实验,荧光显微镜进行镜鉴拍照。结果:获得能够与sLSECtin-Fc蛋白黏附的细菌,并且通过过表达LSECtin的稳定株与细菌的黏附,发现LSECtin可以结合大肠杆菌和肺炎链球菌,而不能结合金黄色葡萄球菌和肺炎克雷伯杆菌。结论:发现LSECtin可以结合细菌,为进一步研究LSECtin在先天免疫中的作用奠定了基础。     

Plant Reactions to Inoculation of Roots with Fungi and Bacteria

The potential of 120 isolates of fungi and bacteria from plant rhizospheres to interfere with plant development and growth was studied in greenhouse experiments. The pure cultured isolates were obtained from plant roots in the field and applied as suspensions to the roots of eight test plant species. 10–20% of the isolates caused distinct symptoms on shoots, growth retardations without other symptoms or growth promotions. Responses of treated plants ranged from death of plants soon after treatment to up to about 40% higher shoot fresh weight than in control plants. Two bacterial isolates induced strong reactions in most of the plant species tested while other isolates showed a more or less pronounced specificity by giving reactions in only some of the plant species tested.     

Rhizosphere Competitiveness of Trichloroethylene-Degrading, Poplar-Colonizing Recombinant Bacteria

Indigenous bacteria from poplar tree (Populus canadensis var. eugenei ‘Imperial Carolina’) and southern California shrub rhizospheres, as well as two tree-colonizing Rhizobium strains (ATCC 10320 and ATCC 35645), were engineered to express constitutively and stably toluene o-monooxygenase (TOM) from Burkholderia cepacia G4 by integrating the tom locus into the chromosome. The poplar and Rhizobium recombinant bacteria degraded trichloroethylene at a rate of 0.8 to 2.1 nmol/min/mg of protein and were competitive against the unengineered hosts in wheat and barley rhizospheres for 1 month (colonization occurred at a level of 1.0 × 10⁵ to 23 × 10⁵ CFU/cm of root). In addition, six of these recombinants colonized poplar roots stably and competitively with populations as large as 79% ± 12% of all rhizosphere bacteria after 28 days (0.2 × 10⁵ to 31 × 10⁵ CFU/cm of root). Furthermore, five of the most competitive poplar recombinants (e.g., Pb3-1 and Pb5-1, which were identified as Pseudomonas sp. strain PsK recombinants) retained the ability to express TOM for 29 days as 100% ± 0% of the recombinants detected in the poplar rhizosphere expressed TOM constitutively.