Alicyclobacillus Contamination in the Production Line of Kiwi Products in China

Alicyclobacillus are spoilage microbes of many juice products, but contamination of kiwi products by Alicyclobacillus is seldom reported. This study aims to investigate the whole production line of kiwi products in China to assess the potential risk of their contamination. A total of 401 samples from 18 commercial products, 1 processing plant and 16 raw material orchards were tested, and 76 samples were positive, from which 76 strains of microbes were isolated and identified as 4 species of Alicyclobacillus, including Alicyclobacillus acidoterrestris, Alicyclobacillus contaminans, Alicyclobacillus herbarius and Alicyclobacillus cycloheptanicus, and another 9 strains as 3 species of Bacillus by sequencing of their 16S rDNA. Through phylogenetic tree construction and RAPD-PCR amplification, it was found that there exist genotypic diversities to some extent among these isolates. Four test strains (each from one species of the 4 Alicyclobacillus species isolated in this study) could spoil pH adjusted kiwi fruit juice and some commercial kiwi fruit products with producing guaiacol (11–34 ppb).     

Detection of Alicyclobacillus spp. in Fruit Juice by Combination of Immunomagnetic Separation and a SYBR Green I Real-Time PCR Assay

An approach based on immunomagnetic separation (IMS) and SYBR Green I real-time PCR (real-time PCR) with species-specific primers and melting curve analysis was proposed as a rapid and effective method for detecting Alicyclobacillus spp. in fruit juices. Specific primers targeting the 16S rDNA sequences of Alicyclobacillus spp. were designed and then confirmed by the amplification of DNA extracted from standard strains and isolates. Spiked samples containing known amounts of target bacteria were used to obtain standard curves; the correlation coefficient was greater than 0.986 and the real-time PCR amplification efficiencies were 98.9%- 101.8%. The detection limit of the testing system was 2.8×10¹ CFU/mL. The coefficient of variation for intra-assay and inter-assay variability were all within the acceptable limit of 5%. Besides, the performance of the IMS-real-time PCR assay was further investigated by detecting naturally contaminated kiwi fruit juice; the sensitivity, specificity and accuracy were 91.7%, 95.9% and 95.3%, respectively. The established IMS-real-time PCR procedure provides a new method for identification and quantitative detection of Alicyclobacillus spp. in fruit juice.     

Influence of Different Filling,Cooling, and Storage Conditions on the Growth of Alicyclobacillus acidoterrestris CRA7152 in Orange Juice

The prevention of spoilage by Alicyclobacillus acidoterrestris is a current challenge for fruit juice and beverage industries worldwide due to the bacterium''s acidothermophilic growth capability, heat resistance, and spoilage potential. This study examined the effect of storage temperature on A. acidoterrestris growth in hot-filled orange juice. The evolution of the A. acidoterrestris population was monitored under six different storage conditions after pasteurization (at 92°C for 10 s), maintenance at 85°C for 150 s, and cooling with water spray to 35°C in about 30 min and using two inoculum levels: <10¹ and 10¹ spores/ml. Final cooling and storage conditions were as follows: treatment 1, 30°C for the bottle cold point and storage at 35°C; treatment 2, 30°C for 48 h and storage at 35°C; treatment 3, 25°C for the bottle cold point and storage at 35°C; treatment 4, 25°C for 48 h and storage at 35°C; treatment 5, storage at 20°C (control); and treatment 6, filling and storage at 25°C. It was found that only in treatment 5 did the population remain inhibited during the 6 months of orange juice shelf life. By examining treatments 1 to 4, it was observed that A. acidoterrestris predicted growth parameters were significantly influenced (P < 0.05) either by inoculum level or cooling and storage conditions. The time required to reach a 10⁴ CFU/ml population of A. acidoterrestris was considered to be an adequate parameter to indicate orange juice spoilage by A. acidoterrestris. Therefore, hot-filled orange juice should be stored at or below 20°C to avoid spoilage by this microorganism. This procedure can be considered a safe and inexpensive alternative to other treatments proposed earlier.The first Alicyclobacillus sp. discovered was isolated in 1982 from spoiled apple juice aseptically packed in Germany and was considered at that time strictly limited to thermophilic and acidic environments (5). The spoilage of fruit juices by Alicyclobacillus is characterized by “off” flavors (medicinal or phenolic) due to guaiacol, 2,6-dibromophenol, and 2,6-dichlorophenol (20, 32, 25). As the spoilage does not show any evident signs like swelling of the container or any overt changes in the fruit juice (e.g., pH or turbidity), it is often not recognized until the packages are opened, the product is tasted, and consumer complaints are received by the manufacturer (6).Alicyclobacillus acidoterrestris, Alicyclobacillus cycloheptanicus and Alicyclobacillus acidocaldarius were the first three species described when the Alicyclobacillus genus was created in 1992 (31). Although there are currently more than 15 species described (27), only four Alicyclobacillus species (A. acidoterrestris, A. pomorum, A. herbarius and A. acidophilus) have shown the ability to produce off flavors in fruit juices or beverages (5, 13, 9, 1). Of these, A. acidoterrestris is considered the most important spoilage species within the Alicyclobacillus genus either by its frequency of occurrence or by its linkage to the spoilage problems of fruit juices and beverages. The broad temperature range for A. acidoterrestris growth (25 to 60°C) (33, 21, 20, 30, 11), its ability to grow under acidic environments (pH 2.5 to 6.0) (20, 30, 19, 22, 6), and its high heat resistance in orange juice (D at 95°C of 2.7 min) (8) together provide adequate conditions for both survival through pasteurization and growth during juice storage.A. acidoterrestris growth and the consequent orange juice spoilage can lead to enormous economic losses; therefore, this microorganism is currently considered a major challenge for the fruit juice industries. It is known to be difficult, if not impossible, to guarantee the absence of Alicyclobacillus spores on the surface of fruits used to make juices since the soil is the primary niche of Alicyclobacillus spp. (7). Subsequently, control measures such as avoiding fruit contact with soil and the use of sanitizers during the fruit washing step before crushing have been studied (16, 12). In addition, fruit juice producers need to better control the pasteurization conditions and to redesign their thermal processes for targeting Alicyclobacillus spp. (28, 24). However, the limited effectiveness of sanitizers against Alicyclobacillus spores and the sensory and nutritional problems that may arise from increased time and temperature regimes in pasteurization are recognized. Since the complete inactivation of Alicyclobacillus spores from raw materials may not be feasible and since juice spoilage by this microorganism depends on the germination of spores and outgrowth, studies reporting conditions that avoid spoilage by controlling Alicyclobacillus spore germination are necessary. The best option to manage the challenge that Alicyclobacillus presents to the fruit juice industries will be one that results in the fewest alterations in processing and storage conditions, preserves the nutritional and sensory aspects of the final products, does not impact production costs and commercial practices, and at the same time ensures the control of the microorganism.In the fruit juice industry, two main types of thermal processes are commonly applied: pasteurization followed by a hot-fill process or pasteurization followed by a cold-fill process (6). In the former, after the product is heated to >90 to 95°C, it is held hot for 15 to 20 s. As the temperature decreases to 82 to 84°C, the product is filled into the package. Next, the product is held for approximately 2 min before the packages are cooled to room temperature. Hot filling has been extensively used in the manufacturing processes of fruit-based drinks and beverages, but problems due to spoilage caused by Alicyclobacillus may arise during fruit juice shelf life. This is due to the extended time that the product is maintained at temperature conditions adequate for the germination and outgrowth of acidothermophilic spore-forming microbes. Despite several studies regarding the factors that affect A. acidoterrestris growth and heat resistance (12, 18, 2, 29), there is a lack of research on the effects of hot-filled fruit juice storage conditions on A. acidoterrestris growth during juice storage. Thus, this study aimed at estimating and comparing the growth parameters (maximum population ratio, κ; lag time, λ; and maximum growth rate, μ) of A. acidoterrestris survival in hot-filled orange juice that was cooled and stored under several conditions that simulate industrial and commercial practices. Primary growth parameters were estimated by using the Baranyi predictive model (3). Additionally, orange juice cooling and storage conditions that avoided germination, growth, and guaiacol production by A. acidoterrestris CRA 7152 were determined.     

Purification and Characterization of Warnericin RB4, Anti-Alicyclobacillus Bacteriocin, Produced by Staphylococcus warneri RB4

In this study we characterized a bacteriocin, warnericin RB4, produced by Staphylococcus warneri RB4. Warnericin RB4 activity was completely inactivated by trypsin and actinase E. The activity was stable at 100°C for 15 min, and had a pH range of 2 to 6. S. warneri RB4 showed antibacterial activity against only Alicyclobacillus acidoterrestris, A. acidocaldarius, and Micrococcus luteus, among 34 bacterial species tested. The amino acid sequence of the purified bacteriocin contained 27 amino acid residues (K-K-K-S-G-V-I-P-X-V-X-H-D-X-H-M-N-X-F-Q-F-V-F-X-X-X-S). The molecular mass of the bacteriocin was estimated to be 2,958.2 Da by ESI-MS. These results show that the Warnericin RB4 exhibiting specific antibacterial activity against thermo-acidophiles, Alicyclobacillus spp., is a Nukacin ISK-1 or closely related bacteriocin, classified with class IA (Lacticin 481 types). This is the first report that Warnericin RB-4 is effective to inhibit the growth of causative microorganisms of spoilage in various acidic drinks. Warnericin RB4 might prove useful in fruit juices and fruit juice–containing drinks.     

Biogenic amine production by Lactobacillus

AIMS: The aim of this work was to demonstrate that strains of Lactobacillus may be able to produce putrescine and agmatine from one of the major amino acids present in fruit juices and wine, arginine, and from amino acid-derived ornithine. METHODS AND RESULTS: Biogenic amines were determined by HPLC. Their production in the culture medium was similar under both microaerophilic and anaerobic conditions. The presence of Mn2+ had a minimal influence on the results, whereas the addition of pyridoxal phosphate increased amine production 10-fold. Lactobacillus hilgardii X1B, isolated from wine, was able to degrade arginine by two pathways: arginine deiminase and arginine decarboxylase. The isolate was able to produce putrescine from ornithine and from agmatine. Lactobacillus plantarum strains N4 and N8, isolated from orange, utilized arginine via the arginine deiminase system. Only the N4 strain was able to produce putrescine from ornithine. CONCLUSION: It has been demonstrated that Lact. hilgardii X1B is able to produce the most important biogenic amine found in wine, putrescine, and also agmatine from arginine and ornithine, and that Lactobacillus plantarum, considered to be an innocuous spoilage micro-organism in fruit juices, is able to produce amines. SIGNIFICANCE AND IMPACT OF THE STUDY: The results have significance in relation to food poisoning caused by beverages that have been contaminated with biogenic amines.     

Isolation and identification of species of Alicyclobacillus from orchard soil in the Western Cape, South Africa

Alicyclobacilli were isolated from orchard soil collected from an apple and pear farm in Elgin, Western Cape, South Africa. Morphological, biochemical and physiological characteristics of the isolates were used to presumptively classify them as belonging to the genus Alicyclobacillus. Strains were identified to species level by polymerase chain reaction (PCR) with genus-specific primers, and 16S ribosomal RNA (rRNA) gene sequencing. To our knowledge this is the first report on the isolation of Alicyclobacillus acidoterrestris and Alicyclobacillus acidocaldarius from orchard soil. The presence of these organisms in the soil suggests a possible source of contamination for the final fruit juice, concentrate or pulp.     

Identification of thermo-acidophilic bacteria isolated from the soil of several Japanese fruit orchards

Aims: To investigate the occurrence and distribution of thermo‐acidophilic bacteria (TAB) associated with various commercial fruit crop soils in Japan and to assess their ability to produce the odorous phenolic compound, guaiacol. Methods and Results: Phylogenetic analysis based on the 5′ end of the 16S rRNA gene (approximately 500 bp), was performed on 62 TAB isolated from the soil of several Japanese fruit orchards. The results suggested that 60 of the bacterial strains analysed belonged to the genus Alicyclobacillus, while the remaining two belonged to the genus Bacillus. The majority of strains (58%) were identified as Alicyclobacillus acidoterrestris. This group partitioned into three phylogenetically distinct subgroups (A–C). Isolates identified as A. acidiphilus (two strains), A. acidoterrestris (36 strains), and A. hesperidum subsp. aigle (one strain), produced guaiacol from vanillic acid. Levels of guaiacol production varied significantly among strains. The guaiacol producing phenotype was conserved among certain species, however no correlation was observed between levels of guaiacol production and 16S rRNA gene‐based phylogenetic relatedness. Conclusions: Alicyclobacillus acidoterrestris and Alicyclobacillus contaminans were widely distributed among various fruit orchards in Japan. Guaiacol production was common at the species/subspecies level; however the amount of guaiacol produced by each strain varied significantly. Significance and Impact of the Study: This study provides a comprehensive phylogenetic survey of Alicyclobacillus species in Japanese fruit orchards. Quality control standards for guaiacol producing Alicyclobacillus have also been described.     

Role of Broiler Carcasses and Processing Plant Air in Contamination of Modified-Atmosphere-Packaged Broiler Products with Psychrotrophic Lactic Acid Bacteria

Some psychrotrophic lactic acid bacteria (LAB) are specific meat spoilage organisms in modified-atmosphere-packaged (MAP), cold-stored meat products. To determine if incoming broilers or the production plant environment is a source of spoilage LAB, a total of 86, 122, and 447 LAB isolates from broiler carcasses, production plant air, and MAP broiler products, respectively, were characterized using a library of HindIII restriction fragment length polymorphism (RFLP) patterns of the 16 and 23S rRNA genes as operational taxonomic units in numerical analyses. Six hundred thirteen LAB isolates from the total of 655 clustered in 29 groups considered to be species specific. Sixty-four percent of product isolates clustered either with Carnobacterium divergens or with Carnobacterium maltaromaticum type strains. The third major product-associated cluster (17% of isolates) was formed by unknown LAB. Representative strains from these three clusters were analyzed for the phylogeny of their 16S rRNA genes. This analysis verified that the two largest RFLP clusters consisted of carnobacteria and showed that the unknown LAB group consisted of Lactococcus spp. No product-associated LAB were detected in broiler carcasses sampled at the beginning of slaughter, whereas carnobacteria and lactococci, along with some other specific meat spoilage LAB, were recovered from processing plant air at many sites. This study reveals that incoming broiler chickens are not major sources of psychrotrophic spoilage LAB, whereas the detection of these organisms from the air of the processing environment highlights the role of processing facilities as sources of LAB contamination.     

Isolation of UVC-Tolerant Bacteria from the Hyperarid Atacama Desert, Chile

Martian surface microbial inhabitants would be challenged by a constant and unimpeded flux of UV radiation, and the study of analog model terrestrial environments may be of help to understand how such life forms could survive under this stressful condition. One of these environments is the Atacama Desert (Chile), a well-known Mars analog due to its extreme dryness and intense solar UV radiation. Here, we report the microbial diversity at five locations across this desert and the isolation of UVC-tolerant microbial strains found in these sites. Denaturing gradient gel electrophoresis (DGGE) of 16S rDNA sequences obtained from these sites showed banding patterns that suggest distinct and complex microbial communities. Analysis of 16S rDNA sequences obtained from UV-tolerant strains isolated from these sites revealed species related to the Bacillus and Pseudomonas genera. Vegetative cells of one of these isolates, Bacillus S3.300-2, showed the highest UV tolerance profile (LD₁₀?=?318 J?m²), tenfold higher than a wild-type strain of Escherichia coli. Thus, our results show that the Atacama Desert harbors a noteworthy microbial community that may be considered for future astrobiological-related research in terms of UV tolerance.     

Saccharomyces cerevisiae: Population Divergence and Resistance to Oxidative Stress in Clinical,Domesticated and Wild Isolates

Background

Saccharomyces cerevisiae has been associated with human life for millennia in the brewery and bakery. Recently it has been recognized as an emerging opportunistic pathogen. To study the evolutionary history of S. cerevisiae, the origin of clinical isolates and the importance of a virulence-associated trait, population genetics and phenotypic assays have been applied to an ecologically diverse set of 103 strains isolated from clinics, breweries, vineyards, fruits, soil, commercial supplements and insect guts.

Methodology/Principal Findings

DNA sequence data from five nuclear DNA loci were analyzed for population structure and haplotype distribution. Additionally, all strains were tested for survival of oxidative stress, a trait associated with microbial pathogenicity. DNA sequence analyses identified three genetic subgroups within the recombining S. cerevisiae strains that are associated with ecology, geography and virulence. Shared alleles suggest that the clinical isolates contain genetic contribution from the fruit isolates. Clinical and fruit isolates exhibit high levels of recombination, unlike the genetically homogenous soil isolates in which no recombination was detected. However, clinical and soil isolates were more resistant to oxidative stress than any other population, suggesting a correlation between survival in oxidative stress and yeast pathogenicity.

Conclusions/Significance

Population genetic analyses of S. cerevisiae delineated three distinct groups, comprising primarily the (i) human-associated brewery and vineyard strains, (ii) clinical and fruit isolates (iii) and wild soil isolates from eastern U.S. The interactions between S. cerevisiae and humans potentiate yeast evolution and the development of genetically, ecologically and geographically divergent groups.     

rpoB-Based Microbial Community Analysis Avoids Limitations Inherent in 16S rRNA Gene Intraspecies Heterogeneity

Contemporary microbial community analysis frequently involves PCR-amplified sequences of the 16S rRNA gene (rDNA). However, this technology carries the inherent problem of heterogeneity between copies of the 16S rDNA in many species. As an alternative to 16S rDNA sequences in community analysis, we employed the gene for the RNA polymerase beta subunit (rpoB), which appears to exist in one copy only in bacteria. In the present study, the frequency of 16S rDNA heterogeneity in bacteria isolated from the marine environment was assessed using bacterial isolates from the red alga Delisea pulchra and from the surface of a marine rock. Ten strains commonly used in our laboratory were also assessed for the degree of heterogeneity between the copies of 16S rDNA and were used to illustrate the effect of this heterogeneity on microbial community pattern analysis. The rock isolates and the laboratory strains were also used to confirm nonheterogeneity of rpoB, as well as to investigate the versatility of the primers. In addition, a comparison between 16S rDNA and rpoB PCR-DGGE (denaturing gradient gel electrophoresis)-based community analyses was performed using a DNA mixture of nine isolates from D. pulchra. Eight out of 14 isolates from D. pulchra, all rock isolates, and 6 of 10 laboratory strains displayed multiple bands for 16S rDNA when analyzed by DGGE. There was no indication of heterogeneity for either the rock isolates or the laboratory strains when rpoB was used for PCR-DGGE analysis. Microbial community pattern analysis using 16S rDNA PCR-DGGE showed an overestimation of the number of laboratory strains in the sample, while some strains were not represented. Therefore, the 16S rDNA PCR-DGGE-based community analysis was proven to be severely limited by 16S rDNA heterogeneity. The mixture of isolates from D. pulchra proved to be more accurately described using rpoB, compared to the 16S rDNA-based PCR-DGGE.     

大连海域沉积物中可培养海洋链霉菌活性及其多样性

以大肠杆菌、金黄色葡萄球菌和尖孢镰刀枯萎病菌作为测试靶目标,采用9种分离培养基从大连海域13个不同采样点的海洋沉积物样品中分离到165株海洋链霉菌。从165株海洋放线菌中筛选到对金黄色葡萄球菌具有抑制活性的菌株85株,占总菌株数的51.5%;对大肠杆菌具有抑制活性的菌株27株,占总菌株数的16.4%;对尖孢镰刀枯萎病菌具有抑制活性的菌株仅有6株,占总菌株数的3.6%。因此,海洋链霉菌的活性更多地表现为对细菌的抗性,尤其对革兰氏阳性细菌具有更高的抑制活性。对其中具有抑制活性或形态独特的菌株进行了16S r DNA序列分析,并构建系统发育树,显示活性海洋链霉菌具有丰富的种类多样性和广谱抗菌活性。同种海洋链霉菌与土壤链霉菌活性比较结果也表明,海洋链霉菌多表现抗革兰氏阳性细菌活性。     

Fate ofescherichia coli andE. coli O157∶H7 in apple juice treated with propolis extract

Fruit juices are targets of spoilage moulds, yeasts and acid tolerant bacteria. They might be contaminated with bacteria from raw materials, environment, packaging and during the handling of the product. These contaminations have frequently resulted in the spoilage of fruit juice and consequently commercial losses. The objective of this study was to determine the influence of propolis in apple juice againstEscherichia coli andE. coli O157:H7 strains of the spoilage and pathogenic bacteria. For this purpose, apple juice was obtained from fresh apples and then was pasteurised. The pH value, titrable acidity (as % malic acid) and Brix degree of this apple juice were 3.72±0.10, 0.67±0.05% and 12.1±0.01, respectively. Propolis extract at 1,2 and 5% concentrations were tested to determine ofE. coli andE. coli O157:H7 inhibition using paper disc diffusion method. The control treatment had no propolis extract. The apple juices were contaminated with these bacteria, and the activity of propolis was observed at first, 18^th, 24^th, 48^th and 72^nd hours at 4 and 25°C. The number of cells in the tubes was counted using serial dilution method. Results indicated that propolis extract at 2 and 5% concentrations had significant antimicrobial activity againstE. coli andE. coli O157:H7, therefore we can conclude that propolis extract is worthy of further study as a natural preservative for the foods prone to microbial spoilage.     

Characterization of culturable heterotrophic bacteria in hydrocarbon-contaminated soil from an alpine former military site

We characterized the culturable, heterotrophic bacterial community in soil collected from a former alpine military site contaminated with petroleum hydrocarbons. The physiologically active eubacterial community, as revealed by fluorescence-in situ-hybridization, accounted for 14.9 % of the total (DAPI-stained) bacterial community. 4.0 and 1.2 % of the DAPI-stained cells could be attributed to culturable, heterotrophic bacteria able to grow at 20 and 10 °C, respectively. The majority of culturable bacterial isolates (23/28 strains) belonged to the Proteobacteria with a predominance of Alphaproteobacteria. The remaining isolates were affiliated with the Firmicutes, Actinobacteria and Bacteroidetes. Five strains could be identified as representatives of novel species. Characterization of the 28 strains demonstrated their adaptation to the temperature and nutrient conditions prevailing in the studied soil. One-third of the strains was able to grow at subzero temperatures (?5 °C). Studies on the effect of temperature on growth and lipase production with two selected strains demonstrated their low-temperature adaptation.     

Short-term plant species impact on microbial community structure in soils with long-term agricultural history

The goal of our study was to capture the short-term effects of individual plant species on an established microbial community in a soil with a well-defined agricultural history. Using biochemical and molecular techniques we quantified the effects of plant species on changes in the soil microbial community over an 8-week time-course. We conducted a greenhouse experiment using field soil from a site that was managed as a Zea mays monoculture for over 50 years. The conditioned soil provided a baseline from which changes in microbial community composition through the effects of newly introduced plants could be determined. Within a short time frame (8 weeks), introduced plants influenced the soil microbial community in ways unique to each plant. Some species (Fagopyrm esculentum and xTriticosecale spp.) resulted in an increase of total microbial community richness, diversity and the stimulation of new microbial species not associated with the legacy vegetation. Other plants (Vicia villosa and Lolium multiflorum) tended to reduce community diversity. We suggest root surface area is good general predictor of rhizosphere microbial community diversity, but in some cases other plant traits may have dominant influence on plant-induced changes in microbial community composition.     

Endophytic Fungal Strains of Soybean for Lipid Production

Lipids created via microbial biosynthesis are a potential raw material to replace plant-based oil for biodiesel production. Oleaginous microbial species currently available are capable of accumulating high amount of lipids in their cell biomass, but rarely can directly utilize lignocellulosic biomass as substrates. Thus this research focused on the screening and selection of new fungal strains that generate both lipids and hydrolytic enzymes. To search for oleaginous fungal strains in the soybean plant, endophytic fungi and fungi close to the plant roots were studied as a microbial source. Among 33 endophytic fungal isolates screened from the soybean plant, 13 have high lipid content (>20 % dry biomass weight); among 38 fungal isolates screened from the soil surrounding the soybean roots, 14 have high lipid content. Also, five fungal isolates with both high lipid content and promising biomass production were selected for further studies on their cell growth, oil accumulation, lipid content and profile, utilization of various carbon sources, and cellulase production. The results indicate that most strains could utilize different types of carbon sources and some strains accumulated >40 % of the lipids based on the dry cell biomass weight. Among these promising strains, some Fusarium strains specifically showed considerable production of cellulase, which offers great potential for biodiesel production by directly utilizing inexpensive lignocellulosic material as feedstock.     

Why fruit rots: theoretical support for Janzen's theory of microbe–macrobe competition

We present a formal model of Janzen''s influential theory that competition for resources between microbes and vertebrates causes microbes to be selected to make these resources unpalatable to vertebrates. That is, fruit rots, seeds mould and meat spoils, in part, because microbes gain a selective advantage if they can alter the properties of these resources to avoid losing the resources to vertebrate consumers. A previous model had failed to find circumstances in which such a costly spoilage trait could flourish; here, we present a simple analytic model of a general situation where costly microbial spoilage is selected and persists. We argue that the key difference between the two models lies in their treatments of microbial dispersal. If microbial dispersal is sufficiently spatially constrained that different resource items can have differing microbial communities, then spoilage will be selected; however, if microbial dispersal has a strong homogenizing effect on the microbial community then spoilage will not be selected. We suspect that both regimes will exist in the natural world, and suggest how future empirical studies could explore the influence of microbial dispersal on spoilage.     

Phylogeny and ecophysiological features of prokaryotes isolated from temporary saline tidal pools

Although hypersaline environments have been extensively examined, only a limited number of microbial community studies have been performed in saline tide pools. We have studied a temporary salt-saturated tide pool and isolated prokaryotes from the water. Chlorinity measurements revealed that the tide pool brine could be characterized as one of the most hypersaline ecosystems on earth. Enumeration of microorganisms at different salinities showed that the tide pool was dominated by moderate halophiles. Based on 16S rRNA gene sequence analysis, the prokaryotic strains isolated were related to the bacterial genera Rhodovibrio, Halovibrio, Aquisalimonas, Bacillus and Staphylococcus and to the haloarchaeal species Haloferax alexandrinus. Four bacterial isolates were distantly related to their closest validly described species Aquisalimonas asiatica (96.5 % similarity), representing a novel phylogenetic linkage. Ecophysiological analysis also revealed distinct phenotypic profiles for the prokaryotic strains analyzed. The herbicide 2,4-dichlorophenoxyacetate could be effectively utilized by selected strains as the sole carbon source, but phenolic compounds could not be utilized by any of the halophilic isolates examined. None of the halophilic strains were able to grow without the presence of sea salt or seawater. Based on these results, we conclude that moderate halophilic bacteria rather than extremely halophilic archaea dominate in such a hypersaline environment.     

Spatial and Resource Factors Influencing High Microbial Diversity in Soil

To begin defining the key determinants that drive microbial community structure in soil, we examined 29 soil samples from four geographically distinct locations taken from the surface, vadose zone, and saturated subsurface using a small-subunit rRNA-based cloning approach. While microbial communities in low-carbon, saturated, subsurface soils showed dominance, microbial communities in low-carbon surface soils showed remarkably uniform distributions, and all species were equally abundant. Two diversity indices, the reciprocal of Simpson’s index (1/D) and the log series index, effectively distinguished between the dominant and uniform diversity patterns. For example, the uniform profiles characteristic of the surface communities had diversity index values that were 2 to 3 orders of magnitude greater than those for the high-dominance, saturated, subsurface communities. In a site richer in organic carbon, microbial communities consistently exhibited the uniform distribution pattern regardless of soil water content and depth. The uniform distribution implies that competition does not shape the structure of these microbial communities. Theoretical studies based on mathematical modeling suggested that spatial isolation could limit competition in surface soils, thereby supporting the high diversity and a uniform community structure. Carbon resource heterogeneity may explain the uniform diversity patterns observed in the high-carbon samples even in the saturated zone. Very high levels of chromium contamination (e.g., >20%) in the high-organic-matter soils did not greatly reduce the diversity. Understanding mechanisms that may control community structure, such as spatial isolation, has important implications for preservation of biodiversity, management of microbial communities for bioremediation, biocontrol of root diseases, and improved soil fertility.     

Native soil fungi associated with compostable plastics in three contrasting agricultural settings

Plastics are used widely as agricultural mulches to suppress weeds and retain soil moisture. Disposal of conventional plastic mulches requires physical removal for disposal in a landfill or incineration. Biodegradable plastic mulches that could be tilled into the soil at the end of a growing season represent an attractive alternative to conventional plastic mulches. In this study, three commercially available mulches labeled as “biodegradable” and one experimental, potentially biodegradable mulch were used during a tomato growing season, and then buried in field soil at three locations for approximately 6 months, as would occur typically in an agricultural setting. Degradation after 6 months in soil was minimal for all but the cellulosic mulch. After removal of mulches from soil, fungi were isolated from the mulch surfaces and tested for their ability to colonize and degrade the same mulches in pure culture. The majority of culturable soil fungi that colonized biodegradable mulches were within the family Trichocomaceae (which includes beneficial, pathogenic, and mycotoxigenic species of Aspergillus and Penicillium). These isolates were phylogenetically similar to fungi previously reported to degrade both conventional and biodegradable plastics. Under pure culture conditions, only a subset of fungal isolates achieved detectable mulch degradation. No isolate substantially degraded any mulch. Additionally, DNA was extracted from bulk soil surrounding buried mulches and ribosomal DNA was used to assess the soil microbial community. Soil microbial community structure was significantly affected by geographical location, but not by mulch treatments.