Faecal indicator bacteria enumeration in beach sand: a comparison study of extraction methods in medium to coarse sands

Aims:  The absence of standardized methods for quantifying faecal indicator bacteria (FIB) in sand hinders comparison of results across studies. The purpose of the study was to compare methods for extraction of faecal bacteria from sands and recommend a standardized extraction technique.
Methods and Results:  Twenty-two methods of extracting enterococci and Escherichia coli from sand were evaluated, including multiple permutations of hand shaking, mechanical shaking, blending, sonication, number of rinses, settling time, eluant-to-sand ratio, eluant composition, prefiltration and type of decantation. Tests were performed on sands from California, Florida and Lake Michigan. Most extraction parameters did not significantly affect bacterial enumeration. anova revealed significant effects of eluant composition and blending; with both sodium metaphosphate buffer and blending producing reduced counts.
Conclusions:  The simplest extraction method that produced the highest FIB recoveries consisted of 2 min of hand shaking in phosphate-buffered saline or deionized water, a 30-s settling time, one-rinse step and a 10 : 1 eluant volume to sand weight ratio. This result was consistent across the sand compositions tested in this study but could vary for other sand types.
Significance and Impact of the Study:  Method standardization will improve the understanding of how sands affect surface water quality.     

Effects of Surface Area and Flow Rate on Marine Bacterial Growth in Activated Carbon Columns

The colonization of granular activated carbon columns by bacteria can have both beneficial and potentially detrimental consequences. Bacterial growth on the carbon surface can remove adsorbed organics and thus partially regenerate the carbon bed. However, growth can also increase the levels of bacteria in the column effluents, which can adversely affect downstream uses of the treated water. This study of a sand column and several activated carbon columns demonstrated that considerable marine bacterial growth occurred in both sand and carbon columns and that this growth increased the number of bacteria in column effluents. Activated carbon supported approximately 50% more bacteria than did sand. Bacterial growth on activated carbon was reduced by increasing the flow rate through a carbon column and increasing the carbon particle size. Scanning electron micrographs showed that bacteria preferred to attach in the protected crevices on both the sand and carbon surface. The results of this study indicated that the colonization of activated carbon by marine bacteria was enhanced because of carbon's high surface area, its rough surface texture, and its ability to absorb organic materials.     

腾格里沙漠东南缘可培养微生物群落数量与结构特征

以腾格里沙漠东南缘沙漠化土壤为研究对象,研究了不同沙漠化修复程度土壤结皮及结皮下微生物分布特征及多样性。结果表明:研究区域可培养细菌数量随沙漠化生态修复进程呈升高趋势,随采样深度呈下降趋势。数量以节杆菌属和芽孢杆菌属为主,其含量随沙漠化修复程度分别呈降低与升高趋势。修复过程中可培养土壤微生物数量与土壤碳、氮含量呈极显著正相关关系,与pH值呈极显著负相关关系,说明微生物数量与沙漠化土壤改良程度密切相关。通过16S rDNA基因测序及构建系统发育树,研究区域可培养细菌归类为18个属,分属于6个系统发育组:高G+C革兰氏阳性类群、低G+C革兰氏阳性类群、α-变形菌、β-变形菌、γ-变形菌和CFB类群,序列比对显示菌株功能多样。     

Measurement of Horizontal and Vertical Movement of Ralstonia solanacearum in Soil

Two model systems were constructed to measure horizontal and vertical movement of bacteria in soil. These systems were applied to measuring movement of Ralstonia solanacearum (race 1, biovar 3), a causal agent of bacterial wilt of tomato, in andosol and sand at 28°C. The first system was used to measure horizontal movement of the bacteria in soil packed in a narrow horizontal frame. Suspension of the pathogen was applied to soil at one end of the frame, and bacterial number per gram of soil was measured over distance from the inoculation point after 4 days. Horizontal movement of R. solanacearum in supersaturated soil, but without flow, was possibly due to diffusion and the front advanced at 2.2 cm/day in andosol, and at 8.1 cm/day in sand. Using the same experimental system, but applying water inflow to one end of the frame only, the bacterium was detected at the front of water in andosol and sand. The front of the distribution advanced at 20.4 cm/h in andosol and 66.3 cm/h in sand. In the second experimental system, a cylinder of soil packed in a short tube was soaked with water, and soil at the top of the tube was inoculated with bacterial suspension. Immediately, soil cylinders were turned upward, and the bacterial number per gram of soil was measured along vertical distance from the inoculation point after 7 days. Using the system with andosol, the capillary water front rose to 32.5 cm over 7 days after inoculation, and R. solanacearum reached to 18.8 cm height. In sand, capillary water rose to 20.0 cm and the bacteria reached to 16.3 cm height.     

Physical and chemical factors influencing transport of microorganisms through porous media.

Resting-cell suspensions of bacteria isolated from groundwater were added as a pulse to the tops of columns of clean quartz sand. An artificial groundwater solution (AGW) was pumped through the columns, and bacterial breakthrough curves were established and compared to test the effects of ionic strength of the AGW, cell size (by using strains of similar cell surface hydrophobicity but different size), mineral grain size, and presence of heterogeneities within the porous media on transport of the bacteria. The proportion of cells recovered in the effluent ranged from nearly 90% for AGW of a higher ionic strength (I = 0.0089 versus 0.00089 m), small cells (0.75-micron-diameter spheres versus 0.75 by 1.8-micron rods), and coarse-grained sand (1.0 versus 0.33 mm) to less than 1% for AGW of lower ionic strength, large cells, and fine-grained sand. Differences in the widths of peaks (an indicator of dispersion) were significant only for the cell size treatment. For treatments containing heterogeneities (a vein of coarse sand in the center of a bed of fine sand), doubly peaked breakthrough curves were obtained. The first peak represents movement of bacteria through the transmissive coarse-grained vein. The second peak is thought to be dominated by cells which have moved (due to dispersion) from the fine-grained matrix to the coarse-grained vein near the top of the column and thus had been retarded, but not retained, by the column. Strength of effects tests indicated that grain size was the most important factor controlling transport of bacteria over the range of values tested for all of the factors examined.(ABSTRACT TRUNCATED AT 250 WORDS)     

Physical and chemical factors influencing transport of microorganisms through porous media.

Resting-cell suspensions of bacteria isolated from groundwater were added as a pulse to the tops of columns of clean quartz sand. An artificial groundwater solution (AGW) was pumped through the columns, and bacterial breakthrough curves were established and compared to test the effects of ionic strength of the AGW, cell size (by using strains of similar cell surface hydrophobicity but different size), mineral grain size, and presence of heterogeneities within the porous media on transport of the bacteria. The proportion of cells recovered in the effluent ranged from nearly 90% for AGW of a higher ionic strength (I = 0.0089 versus 0.00089 m), small cells (0.75-micron-diameter spheres versus 0.75 by 1.8-micron rods), and coarse-grained sand (1.0 versus 0.33 mm) to less than 1% for AGW of lower ionic strength, large cells, and fine-grained sand. Differences in the widths of peaks (an indicator of dispersion) were significant only for the cell size treatment. For treatments containing heterogeneities (a vein of coarse sand in the center of a bed of fine sand), doubly peaked breakthrough curves were obtained. The first peak represents movement of bacteria through the transmissive coarse-grained vein. The second peak is thought to be dominated by cells which have moved (due to dispersion) from the fine-grained matrix to the coarse-grained vein near the top of the column and thus had been retarded, but not retained, by the column. Strength of effects tests indicated that grain size was the most important factor controlling transport of bacteria over the range of values tested for all of the factors examined.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects on microbial activity by extraction of indigenous cells from soil slurries

Abstract: Possible effects on the physiological activity and culturability of soil microorganisms by different soil dispersion procedures, and effects on activity caused by extracting bacteria from soil, were investigated. There was no apparent difference in cfu's with dispersion of a silty loam soil and a loamy sand soil with pyrophosphate as compared to dispersion in NaCl. Substrate-induced respiration was reduced in the silty loam soil, and methanol oxidation was reduced in the loamy sand soil with dispersion in pyrophosphate, and the soil pH was irreversibly increased by the treatment. Extracted bacterial fractions had lower numbers of culturable cells as percentage of the total number of bacteria in each fraction, lower respiration rates and no methanol oxidation activity as compared to the soil slurry both before and after extraction. The physiological activity was apparently not affected by the number of cells extracted. This indicates that the increased extraction rate of indigenous soil bacteria obtained by effective disruption of aggregates and detachment of cells from surfaces, only results in increased extraction of cells that have been physiologically changed as a result of the extraction process.     

Enhancing the growth of Vicia faba plants by microbial inoculation to improve their phytoremediation potential for oily desert areas

Two experiments were conducted to investigate the effect of inoculating Vicia faba plants (broad beens) raised in clean and oily sand with nodule-forming rhizobia and plant-growth-promoting rhizobacteria (PGPR) on growth of these plants in sand and to test whether this can improve the phytoremediation potential of this crop for oily desert areas. It was found that crude oil in sand at concentrations < 1.0% (w/w) enhanced the plant heights, their fresh and dry weights, the total nodule weights per plant, and the nitrogen contents of shoots and fruits. Similar enhancing effects were recorded when roots of the young plants were inoculated with nodule bacteria alone, PGPR alone, or a mixture of one strain of nodule bacteria and one of the PGPR. Such plant growth effects were associated with a better phytoremediation potential of V. faba plants for oily sand. The total numbers of oil-utilizing bacteria increased in the rhizosphere and more hydrocarbons were eliminated in sand close to the roots. The nodule bacteria tested were two strains of Rhizobium leguminosarum and the PGPR were Pseudomonas aeruginosa and Serratia liquefaciens. The four strains were found to use crude oil, n-octadecane, and phenanthrene as sole sources of carbon and energy. It was concluded that coinoculation of V. faba plant roots in oily sand with nodule bacteria and PGPR enhances the phytoremediation potential of this plant for oily desert sand through improving plant growth and nitrogen fixation.     

小叶锦鸡儿固沙林土壤G+与G-细菌碳代谢功能多样性

为探讨科尔沁沙地流动沙丘上建立小叶锦鸡儿固沙植被后,土壤微生物群落中革兰氏阴性菌和革兰氏阳性菌代谢功能的变化规律,采用Biolog-GN2和GP2微平板检测法,研究了9、16、26年生人工群落以及流动沙丘和天然小叶锦鸡儿群落土壤革兰氏阴性和革兰氏阳性菌碳源代谢的多样性变化特征。结果表明,流动沙丘建立小叶锦鸡儿固沙植被后,土壤革兰氏阴性菌多样性表现为持续缓慢增加的趋势,0～26年生群落Shannon指数由3.85逐渐增加到4.17,均匀度指数由0.85增加到0.94;而革兰氏阳性菌则表现为先显著降低,后逐渐增加的趋势。随着固沙群落年龄的增长,土壤微生物代谢糖类、羧酸类、双亲化合物类、聚合物类、胺/酰胺类和氨基酸类等碳源种类逐渐增加。26年生小叶锦鸡儿人工群落土壤革兰氏阴性和革兰氏阳性菌的碳源代谢模式已逐渐接近天然群落。     

An annual cycle,at constant temperature,in a model sandy beach. II. Microbial biology

A model sandy beach successfully mimicking the natural environment has been operated under controlled conditions at a constant temperature (5°C, representative of British winter coastal sea-water temperatures) for a year. During this time the bacterial population of the sand and inundating sea water has been enumerated. Bacterial numbers were highest near the high and low water marks and had a minimum in between on the beach slope; they decreased with depth in the sand profile and showed little variation with time. A number of the bacteria were isolated from the sand system and a preliminary classification attempted. The results are discussed in relation to the meagre literature pertaining to the microbial biology of natural marine sandy beaches.     

Microbial community composition but not diversity changes along succession in arctic sand dunes

The generality of increasing diversity of fungi and bacteria across arctic sand dune succession was tested. Microbial communities were examined by high‐throughput sequencing of 16S rRNA genes (bacteria) and internal transcribed spacer (ITS) regions (fungi). We studied four microbial compartments (inside leaf, inside root, rhizosphere and bulk soil) and characterized microbes associated with a single plant species (Deschampsia flexuosa) across two sand dune successional stages (early and late). Bacterial richness increased across succession in bulk soil and leaf endosphere. In contrast, soil fungal richness remained constant while root endosphere fungal richness increased across succession. There was, however, no significant difference in Shannon diversity indices between early and late successional stage in any compartment. There was a significant difference in the composition of microbial communities between early and late successional stage in all compartments, although the major microbial OTUs were shared between early and late successional stage. Co‐occurrence network analysis revealed successional stage‐specific microbial groups. There were more co‐occurring modules in early successional stage than in late stage. Altogether, these results emphasize that succession strongly affects distribution of microbial species, but not microbial diversity in arctic sand dune ecosystem and that fungi and bacteria may not follow the same successional trajectories.     

Influence of Bulking Agents,Fertilizers and Bacteria on the Removal of Diesel from a Newfoundland Soil

Laboratory experiments in culture flasks, containing diesel-contaminated Newfoundland soil samples, were undertaken to compare the influence of fertilizers, microorganisms and bulking agents on bioremediation. In Phase I experiments only one fertilizer (cow manure or poultry manure), one bulking agent (sand or hay), or one inoculum (cold-tolerant indigenous bacteria or exogenous commercial bacteria) was added to a soil sample. In Phase II experiments, Design-Expert® Version 6 design of experiment software determined the combinations of fertilizers, bulking agents and inocula to be mixed with the soil samples to study the interactions among the amendments. The maximum diesel removal at 90 days occurred in the sample with sand (Phase I) and in the sample with cow manure, an inoculum of cold-tolerant indigenous bacteria, and sand (Phase II). Diesel removal at 45 days for the same two samples was 85.4% (Phase I) and 91.9% (Phase II), suggesting the cow manure and/or cold-tolerant bacteria inoclum accelerated the process. The poultry manure, commercial bacteria and hay were less effective than their counterparts. The commercial bacteria were more sensitive to diesel concentration than the indigenous cold-tolerant bacteria. The addition of sand, cow manure, and poultry manure improved diesel removal.     

Influence of soil pH on the nitrate-reducing microbial populations and their potential to reduce nitrate to NO and N2O

Abstract After the introduction of Rhizobium leguminosarum biovar trifolii into natural loamy sand and silt loam, bacterial numbers increased only directly after inoculation. Thereafter, bacterial numbers decreased until an equilibrium was reached. This decrease was exponential on a log scale and could be described by the function Y = A + B − R ', where Y is the log number of rhizobial cells at time: T ; A represents the lgo of the final population size; B is the difference between the log (initial number of bacteria) and A ; R is the daily reduction factor of Y−A and t is time in days after inoculation. The final population sizes increased with increasing inoculum densities (10⁴−10⁸ bacteria/g soil). In sterilized soil, however, the populations increased up to an equilibrium, which was not affected by the inoculum density.
The final population sizes were higher in silt loam than in loamy sand in natural, as well as in sterilized soil. The final population size was reached earlier in natural silt loam than in loamy sand. Also the growth rate in sterilized soil was higher in silt loam than in loamy sand. The growth rate of low inoculum densities in silt loam was exponential and approximately the same as in yeast extract mannitol broth. The growth rate in loamy sand could be improved by incresing the bulk density of the soil from 1.0 to 1.4 g/cm³.     

Sampling Design and Enumeration Statistics for Bacteria Extracted from Marine Sediments

The spatial and temporal distributions of marine bacteria were studied at both a muddy and a sandy subtidal site in North Inlet, S.C. The sampling design was hierarchical, since subsampling (by a dilution series) of the sediments was necessary to count bacterial cells using acridine orange epifluoresence microscopy. The cell count data fit a log-normal distribution. The abundance of bacteria was 10¹¹ g⁻¹ (dry weight) of mud and 10⁹ g⁻¹ (dry weight) of sand. Variance component analyses demonstrated that variation due to the subsampling procedures was always statistically significant. Thus the common practice of counting 20 fields from one filter preparation is inadequate for estimating the true bacterial population variance in marine sediments. It is recommended that replication of the subsampling level be performed. Standardization of data (by dry weight of sediment) decreased sampling variance at the mud site but not at the sand site, implying that bacteria are more homogeneously distributed in sand than in mud.     

THE CONTROL OF DISEASES OF LETTUCE BY THE USE OF ANTAGONISTIC ORGANISMS II. THE CONTROL OF RHIZOCTONIA SOLANI KUHN.

The antagonism of a number of bacteria, actinomycetes and fungi in pure culture on soil extract agar has been investigated. A considerable number of each of the three groups were highly antagonistic at 25° C., fewer were active at 15° C. and only a very limited number were effective at 5° C.
In greenhouse experiments damping-off of lettuce seedlings was substantially controlled by selected antagonists in sterile sand, soil or in sand/soil mixtures. Under similar conditions in unsterile soil, forms highly antagonistic in pure culture were generally ineffective in controlling disease. In one experiment control of disease in natural soil was obtained by the use of soil amendments, the effect being enhanced by the inoculation of one of these amendments with suspensions of antagonists.
Attempts to control disease in the field by the use of soil amendments are described. In one year a substantial increase in the stand of healthy seedlings was obtained with certain treatments.     

荒漠草原区土壤粒径组成对柠条根际土壤微生物数量及酶活性的影响

柠条(Caragana korshinskii)是荒漠草原区主要的造林绿化树种,研究其根际土壤微生物和酶活性与不同土壤类型土壤粒径组成的关系有重要意义,然而土壤粒径对荒漠草原柠条根际土壤微生物数量和酶活性的影响知之甚少,探讨土壤颗粒组分与微生物数量、土壤酶活性之间的关系,以及土壤颗粒组成对荒漠草原区固沙灌木植物柠条根际土壤微生物数量及酶活性的影响,可为揭示荒漠草原土壤退化及生态修复提供参考。以宁夏荒漠草原区土壤粒径组成差异显著的灰钙土、红黏土、风沙土环境下栽植的柠条为研究对象,研究不同土壤颗粒组成对根际土壤微生物数量及酶活性的相互关系与影响。结果表明:土壤微生物的数量表现为细菌放线菌真菌。根际土壤中的细菌、真菌数量显著高于非根际,且在3种不同类型的土壤中随着细砂粒的增多,真菌和放线菌数量逐渐降低,而细菌数量呈先增大后减小的趋势;根际与非根际土壤的蔗糖酶、碱性磷酸酶及过氧化氢酶活性均呈现出灰钙土红黏土风沙土的趋势,红黏土根际土壤中的脲酶活性显著高于灰钙土与风沙土;除过氧化氢酶外,土壤酶活性表现为根际高于非根际,在3种不同类型的土壤中随着细砂含量的增加,土壤酶活性均呈递减趋势。土壤颗粒组成与微生物数量之间没有明显的相关性,而与土壤酶活性之间显著相关,土壤酶活性与黏粒、粉粒呈正相关,与细砂、中砂呈负相关关系,根际土壤中酶活性更高,能够为植物及微生物提供更多的营养。     

细菌解磷能力测定方法的研究

选择分解有机磷能力较强的3株细菌和溶解磷矿粉较强的4株细菌,砂培4周后,用不同的方法测定水浸提液中磷的含量。发现不同的细菌解磷能力差异很大,细菌在分解磷化合物的同时,一部分磷被细菌同化,一部分以无机磷酸盐状态贮藏在细菌细胞内。直接测定浸提液中无机磷酸盐的含量,将大大低估细菌的解磷能力,必须将浸提液消煮,才能比较正确地反映细菌分解磷的能力。     

Bacterial transport through porous solids: interactions betweenMicrococcus luteus cells and sand particles

Summary As part of an evaluation of microbial systems for potential application in enhanced oil recovery (EOR) in situ, the behaviour of bacteria within the labyrinths of porous structures was investigated. Sandpacks were utilised as models of reservoir formations. UsingMicrococcus luteus (NCIB 8553) cells under non-growing conditions, the interaction between the bacteria and sand particles was investigated by a simple shake flask system. The attachment of bacterial cells to sand was time-dependent and reversible. With increasing density of cell suspensions, competivive effects on binding to sand were observed, indicating a possible interaction between two subpopulations of cells. Similar effects occurred when suspensions ofM. luteus cells were pumped through sandpacks. Shake flask experiments indicated that the maximum binding of bacteria to sand was about 2×10⁹ cells · g^–1, representing a total coverage of approximately 20% of the surface of sand particles by bacterial cells.     

Relationship between Desiccation and Exopolysaccharide Production in a Soil Pseudomonas sp

The relationship between desiccation and the production of extracellular polysaccharides (EPS) by soil bacteria was investigated by using a Pseudomonas species isolated from soil. Cultures subjected to desiccation while growing in a sand matrix contained more EPS and less protein than those growing at high water potential, suggesting that resources were allocated to EPS production in response to desiccation. Desiccation did not have a significant effect on activity as measured by reduction of iodonitrotetrazolium. Purified EPS produced by the Pseudomonas culture contained several times its weight in water at low water potential. Sand amended with EPS held significantly more water and dried significantly more slowly than unamended sand, implying that an EPS matrix may buffer bacterial colonies from some effects of desiccation. We conclude that bacteria may use EPS production to alter their microenvironment to enhance survival of desiccation.     

Scanning electron microscopy and epifluorescence investigation of bacterial colonization of marine sand sediments

Scanning electron microscopy (SEM) was employed for the investigation of microorganisms living in marine sand sediments. Epifluorescence, as well as sediment analyses, gave further data on the parameters of the sediment samples.SEM revealed a correlation between the site and density of bacterial colonization and the microtopography of the individual sand grains.Sand grains with a medium roundness showed the greatest density of bacterial colonization. Protected surface sites were favored in the colonization process. The mode of bacterial attachment varied; mostly the barren sand grain surface was colonized. However, bacteria were also observed close to or within detritus or attached to diatoms. Many of the attaching bacteria observed were found to produce polymer strands.In some cases special structures were discovered which could serve bacterial attachment. Entire colonies attached by means of polymer nets, and disc-shaped bacteria were observed.