Aerial Dispersal and Epiphytic Survival of Pseudomonas syringae during a Pretest for the Release of Genetically Engineered Strains into the Environment

Prospective experimental field evaluation of genetically engineered microorganisms, such as microbial pest control agents, raises issues of how to properly ascertain their fate and survival in the environment. Field trials with recombinant organisms must reflect requirements for sampling and monitoring. Field trials were conducted at Tulelake, Calif., to monitor the numbers of viable cells of a nonrecombinant strain of Pseudomonas syringae that entered the atmosphere and landed on plants and soil during and after an aerosol spray application. An exponential decrease in numbers of viable cells deposited at increasing distances from three sprayed plots was observed. The relative rate of survival of cells sprayed directly on plants was more than 10 times higher than that of cells dispersed through the air to similar adjacent plants. Results are being used to gain experience with the characteristics of a release site that influence containment or dispersal and to develop appropriate sampling methodologies for evaluating survival and dispersal characteristics of genetically engineered bacteria released into the environment. The ability to make predictions about microbial dispersal and survival will reduce the uncertainties associated with environmental releases of recombinant organisms.     

Model to predict aerial dispersal of bacteria during environmental release

Risk assessment for genetically engineered bacteria sprayed onto crops includes determination of off-site dispersal and deposition. The ability to predict microbial dispersal patterns is essential to characterize the uncertainty (risk) associated with environmental release of recombinant organisms. Toward this end, a particle dispersal model was developed to predict recovery of bacteria on fallout plates at various distances and directions about a test site. The microcomputer simulation incorporates particle size distribution, wind speed and direction, turbulence, evaporation, sedimentation, and mortality, with a time step of 0.5 s. The model was tested against data reported from three field applications of nonrecombinant bacteria and two applications of recombinant bacteria. Simulated dispersal of 10(5) particles was compared with reported deposition measurements. The model may be useful in defining appropriate populations of organisms for release, methods of release or application, characteristics of a release site that influence containment or dispersal, and in developing an appropriate sampling methodology for monitoring the dispersal of organisms such as genetically engineered bacteria.     

Model to predict aerial dispersal of bacteria during environmental release.

Risk assessment for genetically engineered bacteria sprayed onto crops includes determination of off-site dispersal and deposition. The ability to predict microbial dispersal patterns is essential to characterize the uncertainty (risk) associated with environmental release of recombinant organisms. Toward this end, a particle dispersal model was developed to predict recovery of bacteria on fallout plates at various distances and directions about a test site. The microcomputer simulation incorporates particle size distribution, wind speed and direction, turbulence, evaporation, sedimentation, and mortality, with a time step of 0.5 s. The model was tested against data reported from three field applications of nonrecombinant bacteria and two applications of recombinant bacteria. Simulated dispersal of 10(5) particles was compared with reported deposition measurements. The model may be useful in defining appropriate populations of organisms for release, methods of release or application, characteristics of a release site that influence containment or dispersal, and in developing an appropriate sampling methodology for monitoring the dispersal of organisms such as genetically engineered bacteria.     

受水杨酸盐浓度调控的细菌遏制系统

由于存在基因工程微生物(GEMs)不受控制地在环境中释放的风险, 利用GEMs的生物降解能力治理环境污染的方法受到了限制。在大肠杆菌JM109中构建了一个受环境污染物调控的细菌遏制系统, 该系统是由杀伤元件和调控元件组成的双质粒体系, 使细菌的存活受环境中水杨酸盐浓度的调控。当培养基含水杨酸盐时, 阻遏蛋白LacI合成, 阻止自杀基因gef表达, 细菌快速繁殖; 当水杨酸盐不存在时, 自杀基因gef的表达导致细胞杀伤, 菌体大量死亡。该遏制系统可作为模型用于具有生物修复功能的基因工程菌的构建。     

Containment in industrial biotechnology within wastewater treatment plants

Both physical and biological containment are considered to be essential parts in the risk analysis of industrial Good Industrial Large-Scale Practice (GILSP) processes using genetically modified organisms (GMOs). Biological containment of industrial microorganisms has become a more important issue since the introduction of recombinant DNA techniques. In the event of an accidental discharge in the production plant, a large amount of organisms could be released into the wastewater treatment (WWT) system. This WWT system should therefore be considered as a part of the containment. This study demonstrates both a hydrodynamic and a microbiological model for the containment aspects of industrial WWT plants. The models are verified by measurements using industrial hosts of GILSP GMOs at full scale. Both models describe the full-scale equipment accurately. The results are supplemented with microcosm studies on survival of GMOs in defined niches. It is shown that WWT plants can be considered as useful additional parts of the containment of microorganisms, in case of an accidental discharge. The effect of drainage of an enormous amount of microorganisms (several tons) through the WWT plant into the environment is shown to be comparable to the direct drainage of a small-scale fermenter. Microcosm experiments correlate well with the survival rates in the WWT and therefore can be of use to predict the behaviour of GMOs in this environment. Journal of Industrial Microbiology & Biotechnology (2002) 28, 65–69 DOI: 10.1038/sj/jim/7000210 Received 16 July 2001/ Accepted in revised form 05 September 2001     

Molecular methods for environmental monitoring and containment of genetically engineered microorganisms

Plans to introduce genetically engineered microorganisms into the environment has led to concerns over safety and has raised questions about how to detect and to contain such microorganisms. Specific gene sequences, such as lacZ, have been inserted into genetically engineered microorganisms to permit their phenotypic detection. Molecular methods have been developed based upon recovery of DNA from environmental samples and gene probe hybridization to specific diagnostic gene sequences for the specific detection of genetically engineered microorganisms. DNA amplification using the polymerase chain reaction has been applied to enhance detection sensitivity so that single gene targets can be detected. Detection of messenger RNA has permitted the monitoring of gene expression in the environment. The use of reporter genes, such as the lux gene for bioluminescence, likewise has permitted the observation of gene expression. Conditional lethal constructs have been developed as models for containment of genetically engineered microorganisms. Suicide vectors, based upon the hok gene have been developed as model containment systems.     

Implications of nonrandom seed abscission and global stilling for migration of wind‐dispersed plant species

Migration of plant populations is a potential survival response to climate change that depends critically on seed dispersal. Biological and physical factors determine dispersal and migration of wind‐dispersed species. Recent field and wind tunnel studies demonstrate biological adaptations that bias seed release toward conditions of higher wind velocity, promoting longer dispersal distances and faster migration. However, another suite of international studies also recently highlighted a global decrease in near‐surface wind speeds, or ‘global stilling’. This study assessed the implications of both factors on potential plant population migration rates, using a mechanistic modeling framework. Nonrandom abscission was investigated using models of three seed release mechanisms: (i) a simple drag model; (ii) a seed deflection model; and (iii) a ‘wear and tear’ model. The models generated a single functional relationship between the frequency of seed release and statistics of the near‐surface wind environment, independent of the abscission mechanism. An Inertial‐Particle, Coupled Eulerian‐Lagrangian Closure model (IP‐CELC) was used to investigate abscission effects on seed dispersal kernels and plant population migration rates under contemporary and potential future wind conditions (based on reported global stilling trends). The results confirm that nonrandom seed abscission increased dispersal distances, particularly for light seeds. The increases were mitigated by two physical feedbacks: (i) although nonrandom abscission increased the initial acceleration of seeds from rest, the sensitivity of the seed dispersal to this initial condition declined as the wind speed increased; and (ii) while nonrandom abscission increased the mean dispersal length, it reduced the kurtosis of seasonal dispersal kernels, and thus the chance of long‐distance dispersal. Wind stilling greatly reduced the modeled migration rates under biased seed release conditions. Thus, species that require high wind velocities for seed abscission could experience threshold‐like reductions in dispersal and migration potential if near‐surface wind speeds continue to decline.     

Seed release by invasive thistles: the impact of plant and environmental factors

Dispersal is a key process in biological studies of spatial dynamics, but the initiation of dispersal has often been neglected, despite strong indications that differential timing of dispersal can significantly affect dispersal distances. To investigate which plant and environmental factors determine the release of plumed seeds by the invasive thistles Carduus acanthoides and Carduus nutans, we exposed 192 flower heads of each species to increasing wind speeds in a full-factorial wind tunnel experiment with four air flow turbulence, three flower head wetness and two flower head temperature levels. The number of seed releases was highest under dry and turbulent conditions and from heads that had already lost a considerable number of seeds, but was not affected by flower head size, head angle or temperature. Inspection of the trials on video showed that higher wind speeds were needed to meet the seed release threshold in laminar flows and for C. acanthoides heads that had been wet for a longer time. Species differences were minimal, although seed release was more sensitive to lower levels of turbulence in the larger-headed and more open C. nutans heads. Knowledge of seed release biases towards weather conditions favourable for long-distance dispersal improves our understanding of the spread of invaders and allows managers to increase the efficiency of their containment strategies by applying them at crucial times.     

Laboratory Design for Microbiological Safety

Of the large amount of funds spent each year in this country on construction and remodeling of biomedical research facilities, a significant portion is directed to laboratories handling infectious microorganisms. This paper is intended for the scientific administrators, architects, and engineers concerned with the design of new microbiological facilities. It develops and explains the concept of primary and secondary barriers for the containment of microorganisms. The basic objectives of a microbiological research laboratory, (i) protection of the experimenter and staff, (ii) protection of the surrounding community, and (iii) maintenance of experimental validity, are defined. In the design of a new infectious-disease research laboratory, early identification should be made of the five functional zones of the facility and their relation to each other. The following five zones and design criteria applicable to each are discussed: clean and transition, research area, animal holding and research area, laboratory support, engineering support. The magnitude of equipment and design criteria which are necessary to integrate these five zones into an efficient and safe facility are delineated.     

Estimating dispersal rate of the silky cane weevil (Coleoptera: Curculionidae)*

Abstract: The objective of this study was to estimate the silky cane weevil rate of dispersal under near‐natural conditions inside a screened enclosure where an array of buckets was baited with cut sugarcane stalks. One hundred weevils were released and weevils inside the buckets were counted hourly for 8 h, and then 24 and 48 h after release. A passive diffusion model was used to estimate the weevil's dispersal and disappearance rates, within and between rows of buckets with sugarcane. The weevils concentrated around the release point and slowly moved towards the boundaries of the experimental plot over time with an overall average dispersal rate of 2.8 ± 3.58 cm²/h. Dispersal and disappearance rates within and between rows were not significantly different among the time intervals considered (1–8, 8–24 and 24–48 h after release) except for the 1–8 time interval on the array representing the release point when the dispersal rate, D, was significantly higher than those at other time intervals. Continuum of the substratum to disperse from one side of the array to another via a wooden bridge may explain the higher dispersal rate through this array. The number of buckets exposed to the sun during the morning hours was significantly higher on those rows exposed to the sun (south side of the screen enclosure) than on the shaded side. Longer times of bucket exposure to the sun may explain the predominant distribution of weevils in that area suggesting that the weevil population is constantly expanding and retracting according to micro environmental conditions.     

Dispersal behavior of the parasitic wasp Cotesia urabae (Hymenoptera: Braconidae): A recently introduced biocontrol agent for the control of Uraba lugens (Lepidoptera: Nolidae) in New Zealand

Estimates of the dispersal range of a recently introduced biocontrol agent in its new environment are vital to understanding its relative searching capacity, and to foresee the maximum area that could be covered in a release event. In New Zealand, the solitary endoparasitoid Cotesia urabae Austin and Allen (Hymenoptera: Braconidae) was first released in January 2011 as a biological control agent for the gum leaf skeletoniser, Uraba lugens Walker (Lepidoptera: Nolidae). The objective of this study was to utilize an experimental approach to quantify the dispersal behavior of one generation of C. urabae. In our experiment, which used sentinel larvae as target hosts, parasitoids dispersed up to 20 m away from the release point but parasitism was highest within 5 m of the release site. A high level of parasitism was observed at the release tree (87.6%) which suggests that most of the females released may have stayed there. According to the dispersal model developed from the data collected, Cotesia would be able to disperse up to 53 m in one release event. In addition, significant differences were found between the different directions tested for dispersal, showing a clear downwind effect on dispersal suggesting that wind has a direct effect on the dispersal behavior of C. urabae in the field.     

Timing of propagule release significantly alters the deposition area of resulting aerial dispersal

Aim The aim of this study is to determine whether changes to the seasonal and circadian timing of propagule release can a have a significant effect on the area covered by resulting aerial dispersal. Location Western Australia. Methods Using the atmospheric pollution model (TAPM), an existing meso‐scale dispersal model, a range of release patterns was simulated and the resulting deposition compared. Comparisons were based on observations of deposition patterns and the calculated area of deposition. Results Small changes to the timing of propagule release were shown to significantly impact on the area experiencing deposition from the resulting aerial dispersal. Main conclusions Simulations performed in this study show that, for small propagules, changes to the timing of release can lead to alternate, clearly differentiable dispersal events. Small changes in both the seasonal and circadian patterns of release can have significant effects on the area that experiences deposition during the resulting dispersal event. This effect is particularly important at the landscape scale and when there is a need to quantify individual dispersal events. Predictive modelling of aerial dispersal needs to be undertaken with an understanding of the manner in which biological and environmental factors that affect the timing of propagule release can influence results. Results presented highlight the need to characterize the epidemiology of pathogenic organisms of importance to biosecurity as much as possible before they arrive.     

EXPOSE, an Astrobiological Exposure Facility on the International Space Station - from Proposal to Flight

Following an European Space Agency announcement of opportunity in 1996 for ”Externally mounted payloads for 1st utilization phase” on the International Space Station (ISS), scientists working in the fields of astrobiology proposed experiments aiming at long-term exposure of a variety of chemical compounds and extremely resistant microorganisms to the hostile space environment. The ESA exposure facility EXPOSE was built and an operations´ concept was prepared. The EXPOSE experiments were developed through an intensive pre-flight experiment verification test program. 12 years later, two sets of astrobiological experiments in two EXPOSE facilities have been successfully launched to the ISS for external exposure for up to 1.5 years. EXPOSE-E, now installed at the balcony of the European Columbus module, was launched in February 2008, while EXPOSE-R took off to the ISS in November 2008 and was installed on the external URM-D platform of the Russian Zvezda module in March 2009.     

Mother's timing and duration of corticosterone exposure modulate offspring size and natal dispersal in the common lizard (Lacerta vivipara)

Although multiple condition dependence in dispersal is common, the proximate mechanisms that integrate information from multiple sources remain largely unknown. In the common lizard (Lacerta vivipara), earlier studies have shown that maternal plasma corticosterone level interacts with maternal phenotype to affect offspring phenotype and dispersal strategy, and may reflect the mother's external and/or internal environment. However, the mechanism by which these two types of environmental information are integrated has not been identified. We explored the possibility that the timing and duration of the corticosterone signal are the key factors by which the message is modulated. We found that the timing of corticosterone exposure affects the juvenile phenotype: an exposure to corticosterone early in development has negative effects on juvenile size, weight, and body condition, that can nevertheless be restored in the case of a prolonged exposure. The duration of corticosterone exposure affects the dispersal strategy, although the precise effect depends on the sex ratio of the clutch. That is, in female-biased clutches, a prolonged exposure during gestation promotes philopatry of juveniles, while a short exposure enhances their dispersal, a result that is consistent with kin competition theory. Therefore, our results demonstrate that while corticosterone titer may signal a mother's external and/or internal environment to her developing young, differences in the timing and duration of this endocrine signal produce offspring with specialized phenotypes that exhibit different dispersal strategies.     

Synthetic genetic circuits for programmable biological functionalities

Living organisms evolve complex genetic networks to interact with the environment. Due to the rapid development of synthetic biology, various modularized genetic parts and units have been identified from these networks. They have been employed to construct synthetic genetic circuits, including toggle switches, oscillators, feedback loops and Boolean logic gates. Building on these circuits, complex genetic machines with capabilities in programmable decision-making could be created. Consequently, these accomplishments have led to novel applications, such as dynamic and autonomous modulation of metabolic networks, directed evolution of biological units, remote and targeted diagnostics and therapies, as well as biological containment methods to prevent release of engineered microorganisms and genetic materials. Herein, we outline the principles in genetic circuit design that have initiated a new chapter in transforming concepts to realistic applications. The features of modularized building blocks and circuit architecture that facilitate realization of circuits for a variety of novel applications are discussed. Furthermore, recent advances and challenges in employing genetic circuits to impart microorganisms with distinct and programmable functionalities are highlighted. We envision that this review gives new insights into the design of synthetic genetic circuits and offers a guideline for the implementation of different circuits in various aspects of biotechnology and bioengineering.     

Biofilm dispersal: multiple elaborate strategies for dissemination of bacteria with unique properties

In most environments, microorganisms evolve in a sessile mode of growth, designated as biofilm, which is characterized by cells embedded in a self‐produced extracellular matrix. Although a biofilm is commonly described as a “cozy house” where resident bacteria are protected from aggression, bacteria are able to break their biofilm bonds and escape to colonize new environments. This regulated process is observed in a wide variety of species; it is referred to as biofilm dispersal, and is triggered in response to various environmental and biological signals. The first part of this review reports the main regulatory mechanisms and effectors involved in biofilm dispersal. There is some evidence that dispersal is a necessary step between the persistence of bacteria inside biofilm and their dissemination. In the second part, an overview of the main methods used so far to study the dispersal process and to harvest dispersed bacteria was provided. Then focus was on the properties of the biofilm‐dispersed bacteria and the fundamental role of the dispersal process in pathogen dissemination within a host organism. In light of the current body of knowledge, it was suggested that dispersal acts as a potent means of disseminating bacteria with enhanced colonization properties in the surrounding environment.     

Dispersal kernels of the invasive alien western corn rootworm and the effectiveness of buffer zones in eradication programmes in Europe

Europe is attempting to contain or, in some regions, to eradicate the invading and maize destroying western corn rootworm (WCR). Eradication and containment measures include crop rotation and insecticide treatments within different types of buffer zones surrounding new introduction points. However, quantitative estimates of the relationship between the probability of adult dispersal and distance from an introduction point have not been used to determine the width of buffer zones. We address this by fitting dispersal models of the negative exponential and negative power law families in logarithmic and non-logarithmic form to recapture data from nine mark-release-recapture experiments of marked WCR adults from habitats as typically found in the vicinity of airports in southern Hungary in 2003 and 2004. After each release of 4000–6300 marked WCR, recaptures were recorded three times using non-baited yellow sticky traps at 30–305 m from the release point and sex pheromone-baited transparent sticky traps placed at 500–3500 m. Both the negative exponential and negative power law models in non-log form presented the best overall fit to the numbers of recaptured adults (1% recapture rate). The negative exponential model in log form presented the best fit to the data in the tail. The models suggested that half of the dispersing WCR adults travelling along a given bearing will have travelled between 117 and 425 m and 1% of the adults between 775 and 8250 m after 1 day. An individual-based model of dispersal and mortality over a generation of WCR adults indicated that 9.7–45.3% of the adults would escape a focus zone (where maize is only grown once in 3 consecutive years) of 1 km radius and 0.6–21% a safety zone (where maize is only grown once in 2 consecutive years) of 5 km radius and consequently current European Commission (EC) measures are inadequate for the eradication of WCR in Europe. Although buffer zones large enough to allow eradication would be economically unpalatable, an increase of the minimum width of the focus zone from 1 to 5 km and the safety zone from 5 to 50 km would improve the management of local dispersal.     

应用细胞自动机对一年生植物种群扩散进行理论研究

应用细胞自动机方法构造了用于研究一年生植物扩散的理论模型并应用该模型模拟了一年生植物（杂草）种群在同质环境中的扩散。一年生杂草种子的扩散距离和分布是其种群扩散的主要方式,故本文将其种子扩散分布作为构建邻域细胞函数的基础。根据Howard(1991)^[1]提供的某一年生杂草种子的扩散数据,本文导出了一个25邻的邻域细胞函数和相关的转移函数。建立了一个受控的细胞自动机模型。通过模拟,发现在同质环境中聚集在一起的一年生植物杂草越多就需要越大的控制力才能限制它们扩散;生长于农田边缘的杂草更容易被控制。这些模拟结果表明该模型能较好地表现生态学中的两个众所周知的现象;生物的聚集效应和边缘效应。希望自动机方法和在本文获得的知识将有助于我们制订植物种群的最优管理策略。     

Testing a mechanistic dispersal model against a dispersal experiment with a wind‐dispersed moss

Wind is the main dispersal agent for a wide array of species and for these species the environmental conditions under which diaspores are released can potentially modify the dispersal kernel substantially. Little is known about how bryophytes regulate spore release, but conditions affecting peristome movements and vibration of the seta may be important. We modelled airborne spore dispersal of the bryophyte species Discelium nudum (spore diameter 25 μm), in four different release scenarios, using a Lagrangian stochastic dispersion model and meteorological data. We tested the model predictions against experimental data on colonization success at five distances (5, 10, 30, 50 and 100 m) and eight directions from a translocated point source during seven two‐day periods. The model predictions were generally successful in describing the observed colonization patterns, especially beyond 10 m. In the laboratory we established spore release thresholds; horizontal wind speed sd > 0.25 m s^?1 induced the seta to vibrate and in relative humidity < 75% the peristome was open. Our dispersal model predicts that the proportion of spores dispersing beyond 100 m is almost twice as large if the spores are released under turbulent conditions than under more stable conditions. However, including release thresholds improved the fit of the model to the colonization data only minimally, with roughly the same amount of variation explained by the most constrained scenario (assuming both vibration of the seta and an open peristome) and the scenario assuming random release. Model predictions under realised experimental conditions suggest that we had a low statistical power to rank the release scenarios due to the lack of measurements of the absolute rate of spore release. Our results hint at the importance of release conditions, but also highlight the challenges in dispersal experiments intended for validating mechanistic dispersal models.     

Interactions between spatial and temporal scales in the evolution of dispersal rate

The evolution of dispersal rate is studied with a model of several local populations linked by dispersal. Three dispersal strategies are considered where all, half or none of the offspring disperse. The spatial scale (number of patches) and the temporal scale (probability of local extinction) of the environment are critical in determining the selective advantage of the different dispersal strategies. The results from the simulations suggest that an interaction between group selection and individual selection results in a different outcome in relation to the spatial and temporal scales of the environment. Such an interaction is able to maintain a polymorphism in dispersal strategies. The maintenance of this polymorphism is also scale-dependent. This study suggests a mechanism for the short-term evolution of dispersal, and provides a testable prediction of this hypothesis, namely that loss of dispersal abilities should be more frequent in spatially more continuous environments, or in temporally more stable environments. This revised version was published online in July 2006 with corrections to the Cover Date.