Alexander V. Semenov

Survival of Escherichia coli in the environment: fundamental and public health aspects.

In this review, our current understanding of the species Escherichia coli and its persistence in the open environment is examined. E. coli consists of six different subgroups, which are separable by genomic analyses. Strains within each subgroup occupy various ecological niches, and can be broadly characterized by either commensalistic or different pathogenic behaviour. In relevant cases, genomic islands can be pinpointed that underpin the behaviour. Thus, genomic islands of, on the one hand, broad environmental significance, and, on the other hand, virulence, are highlighted in the context of E. coli survival in its niches. A focus is further placed on experimental studies on the survival of the different types of E. coli in soil, manure and water. Overall, the data suggest that E. coli can persist, for varying periods of time, in such terrestrial and aquatic habitats. In particular, the considerable persistence of the pathogenic E. coli O157:H7 is of importance, as its acid tolerance may be expected to confer a fitness asset in the more acidic environments. In this context, the extent to which E. coli interacts with its human/animal host and the organisms survivability in natural environments are compared. In addition, the effect of the diversity and community structure of the indigenous microbiota on the fate of invading E. coli populations in the open environment is discussed. Such a relationship is of importance to our knowledge of both public and environmental health.

Percolation and Survival of Escherichia coli O157:H7 and Salmonella enterica Serovar Typhimurium in Soil Amended with Contaminated Dairy Manure or Slurry

ABSTRACT The effect of cattle manure and slurry application on percolation and survival of Escherichia coli O157:H7 and Salmonella enterica serovar Typhimurium was investigated for different soil depths after the addition of water. Four treatments were chosen for the first set of experiments: (i) addition of inoculated farmyard manure on the soil surface, (ii) mixing of inoculated farmyard manure with the top 10 cm of soil, (iii) addition of inoculated slurry on the soil surface, and (iv) injection of inoculated slurry into the top 10 cm of the soil. Homogeneity of water distribution in the soil profile was confirmed by a nondestructive nuclear magnetic resonance method. Survival data were fitted to a modified logistic model, and estimated survival times were compared. In the second set of experiments, pathogen-inoculated farmyard manure or slurry was applied to soil columns with 1-month-old lettuce plants. More pathogen cells percolated to greater depths after slurry than after manure application. Survival of E. coli O157:H7 was significantly longer in soil with slurry than in that with manure, while survival of Salmonella serovar Typhimurium was equally high with manure and slurry. The densities of the pathogens were not different in the rhizosphere compared to the bulk soil with manure, while the densities were higher by 0.88 ± 0.11 and 0.71 ± 0.23 log CFU per g (dry weight), respectively, in the rhizosphere than in bulk soil after slurry application. Our results suggest that surface application of manure may decrease the risk of contamination of groundwater and lettuce roots compared to injection of slurry.

Comparative Analysis of Bacterial Communities in a Potato Field as Determined by Pyrosequencing

Background Plants selectively attract particular soil microorganisms, in particular consumers of root-excreted compounds. It is unclear to what extent cultivar type and/or growth stage affect this process. Methodology/Principal Findings DNA-based pyrosequencing was used to characterize the structure of bacterial communities in a field cropped with potato. The rhizospheres of six cultivars denoted Aveka, Aventra, Karnico, Modena, Premiere and Desiree, at three growth stages (young, flowering and senescence) were examined, in addition to corresponding bulk soils. Around 350,000 sequences were obtained (5,700 to 38,000 per sample). Across all samples, rank abundance distributions best fitted the power law model, which indicates a community composed of a few highly dominant species next to numerous rare species. Grouping of the sequences showed that members of the Actinobacteria, Alphaproteobacteria, next to as-yet-unclassified bacteria, dominated. Other groups that were consistently found, albeit at lower abundance, were Beta-, Gamma- and Deltaproteobacteria and Acidobacteria. Principal components analyses revealed that rhizosphere samples were significantly different from corresponding bulk soil in each growth stage. Furthermore, cultivar effects were found in the young plant stage, whereas these became insignificant in the flowering and senescence stages. Besides, an effect of time of season was observed for both rhizosphere and bulk soils. The analyzed rhizosphere samples of the potato cultivars were grouped into two groups, in accordance with the allocation of carbon to starch in their tubers, i.e. Aveka, Aventra and Karnico (high) versus Premiere and Desiree (low) and thus replicates per group were established. Conclusions Across all potato cultivars, the young plant stages revealed cultivar-dependent bacterial community structures, which disappeared in the flowering and senescence stages. Furthermore, Pseudomonas, Beta-, Alpha- and Deltaproteobacteria flourished under different ecological conditions than the Acidobacteria.

Seasonal variations in the diversity and abundance of diazotrophic communities across soils

The nitrogen (N)-fixing community is a key functional community in soil, as it replenishes the pool of biologically available N that is lost to the atmosphere via anaerobic ammonium oxidation and denitrification. We characterized the structure and dynamic changes in diazotrophic communities, based on the nifH gene, across eight different representative Dutch soils during one complete growing season, to evaluate the amplitude of the natural variation in abundance and diversity, and identify possible relationships with abiotic factors. Overall, our results indicate that soil type is the main factor influencing the N-fixing communities, which were more abundant and diverse in the clay soils (n=4) than in the sandy soils (n=4). On average, the amplitude of variation in community size as well as the range-weighted richness were also found to be higher in the clay soils. These results indicate that N-fixing communities associated with sandy and clay soil show a distinct amplitude of variation under field conditions, and suggest that the diazotrophic communities associated with clay soil might be more sensitive to fluctuations associated with the season and agricultural practices. Moreover, soil characteristics such as ammonium content, pH and texture most strongly correlated with the variations observed in the diversity, size and structure of N-fixing communities, whose relative importance was determined across a temporal and spatial scale.

Bacterial chitinolytic communities respond to chitin and pH alteration in soil

ABSTRACT Chitin amendment is a promising soil management strategy that may enhance the suppressiveness of soil toward plant pathogens. However, we understand very little of the effects of added chitin, including the putative successions that take place in the degradative process. We performed an experiment in moderately acid soil in which the level of chitin, next to the pH, was altered. Examination of chitinase activities revealed fast responses to the added crude chitin, with peaks of enzymatic activity occurring on day 7. PCR-denaturing gradient gel electrophoresis (DGGE)-based analyses of 16S rRNA and chiA genes showed structural changes of the phylogenetically and functionally based bacterial communities following chitin addition and pH alteration. Pyrosequencing analysis indicated (i) that the diversity of chiA gene types in soil is enormous and (i) that different chiA gene types are selected by the addition of chitin at different prevailing soil pH values. Interestingly, a major role of Gram-negative bacteria versus a minor one of Actinobacteria in the immediate response to the added chitin (based on 16S rRNA gene abundance and chiA gene types) was indicated. The results of this study enhance our understanding of the response of the soil bacterial communities to chitin and are of use for both the understanding of soil suppressiveness and the possible mining of soil for novel enzymes.

Influence of aerobic and anaerobic conditions on survival of Escherichia coli O157:H7 and Salmonella enterica serovar Typhimurium in Luria–Bertani broth, farm-yard manure and slurry

The influence of aerobic and anaerobic conditions on the survival of the enteropathogens Escherichia coli O157:H7 and Salmonella serovar Typhimurium was investigated in microcosms with broth, cattle manure or slurry. These substrates were inoculated with a green fluorescent protein transformed strain of the enteropathogens at 10(7) cells g(-1) dry weight. Survival data was fitted to the Weibull model. The survival curves in aerobic conditions generally showed a concave curvature, while the curvature was convex in anaerobic conditions. The estimated survival times showed that E. coli O157:H7 survived significantly longer under anaerobic than under aerobic conditions. Survival ranged from approximately. 2 weeks for aerobic manure and slurry to more than six months for anaerobic manure at 16 °C. On average, in 56.3% of the samplings, the number of recovered E. coli O157:H7 cells by anaerobic incubation of Petri plates was significantly (p < 0.05) higher in comparison with aerobic incubation. Survival of Salmonella serovar Typhimurium was not different between aerobic and anaerobic storage of LB broth or manure as well as between aerobic and anaerobic incubation of Petri dishes. The importance of changes in microbial community and chemical composition of manure and slurry was distinguished for the survival of E. coli O157:H7 in different oxygen conditions.

The effect of the native bacterial community structure on the predictability of E. coli O157:H7 survival in manure-amended soil

Aims:  The survival capability of pathogens like Escherichia coli O157:H7 in manure‐amended soil is considered to be an important factor for the likelihood of crop contamination. The aim of this study was to reveal the effects of the diversity and composition of soil bacterial community structure on the survival time (ttd) and stability (irregularity, defined as the intensity of irregular dynamic changes in a population over time) of an introduced E. coli O157:H7 gfp‐strain were investigated for 36 different soils by means of bacterial PCR‐DGGE fingerprints.

The use of statistical tools in field testing of putative effects of genetically modified plants on nontarget organisms

Abstract To fulfill existing guidelines, applicants that aim to place their genetically modified (GM) insect-resistant crop plants on the market are required to provide data from field experiments that address the potential impacts of the GM plants on nontarget organisms (NTOs). Such data may be based on varied experimental designs. The recent EFSA guidance document for environmental risk assessment (2010) does not provide clear and structured suggestions that address the statistics of field trials on effects on NTOs. This review examines existing practices in GM plant field testing such as the way of randomization, replication, and pseudoreplication. Emphasis is placed on the importance of design features used for the field trials in which effects on NTOs are assessed. The importance of statistical power and the positive and negative aspects of various statistical models are discussed. Equivalence and difference testing are compared, and the importance of checking the distribution of experimental data is stressed to decide on the selection of the proper statistical model. While for continuous data (e.g., pH and temperature) classical statistical approaches – for example, analysis of variance (ANOVA) – are appropriate, for discontinuous data (counts) only generalized linear models (GLM) are shown to be efficient. There is no golden rule as to which statistical test is the most appropriate for any experimental situation. In particular, in experiments in which block designs are used and covariates play a role GLMs should be used. Generic advice is offered that will help in both the setting up of field testing and the interpretation and data analysis of the data obtained in this testing. The combination of decision trees and a checklist for field trials, which are provided, will help in the interpretation of the statistical analyses of field trials and to assess whether such analyses were correctly applied. We offer generic advice to risk assessors and applicants that will help in both the setting up of field testing and the interpretation and data analysis of the data obtained in field testing.

Bacterial community establishment in native and non-native soils and the effect of fungal colonization

Abstract Bacterial communities are essential parts of living soils. However, we understand very little of how soil matrices govern the structure of the local microbiota. Here, we report on experiments that address such assembly rules