In-Grove Spatiotemporal Spread of Citrus Huanglongbing and Its Psyllid Vector in Relation to Weather.

Reports of spatial patterns of 'Candidatus Liberibacter asiaticus'-infected asymptomatic citrus trees and 'Ca. L. asiaticus'-positive Asian citrus psyllids (ACP) are rare, as are published relationships between huanglongbing (HLB), ACP, and weather. Here, spatial patterns of 'Ca. L. asiaticus'-positive asymptomatic and symptomatic trees were determined every half year in a small grove over 2.5 years, and of HLB-symptomatic trees and ('Ca. L. asiaticus'-positive) ACP populations every month in two commercial groves for 1 year. Spread of symptomatic trees followed that of asymptomatic 'Ca. L. asiaticus'-positive trees with <6 months' delay. 'Ca. L. asiaticus'-positive asymptomatic and symptomatic fronts moved at 2.5 to 3.6 m month-1. No spatial relationship was detected between ACP populations and HLB-infected trees. HLB incidence and 'Ca. L. asiaticus'-positive ACP dynamics were tentatively positively correlated with monthly rainfall data and, to a lesser extent, with average minimum temperature.

Regional Spatial-Temporal Spread of Citrus Huanglongbing Is Affected by Rain in Florida.

Citrus huanglongbing (HLB), associated with 'Candidatus Liberibacter asiaticus' (Las), disseminated by Asian citrus psyllid (ACP), has devastated citrus in Florida since 2005. Data on HLB occurrence were stored in databases (2005 to 2012). Cumulative HLB-positive citrus blocks were subjected to kernel density analysis and kriging. Relative disease incidence per county was calculated by dividing HLB numbers by relative tree numbers and maximum incidence. Spatiotemporal HLB distributions were correlated with weather. Relative HLB incidence correlated positively with rainfall. The focus expansion rate was 1626 m month-1, similar to that in Brazil. Relative HLB incidence in counties with primarily large groves increased at a lower rate (0.24 year-1) than in counties with smaller groves in hotspot areas (0.67 year-1), confirming reports that large-scale HLB management may slow epidemic progress.

Environmental and health effects of the herbicide glyphosate.

The herbicide glyphosate, N-(phosphonomethyl) glycine, has been used extensively in the past 40years, under the assumption that side effects were minimal. However, in recent years, concerns have increased worldwide about the potential wide ranging direct and indirect health effects of the large scale use of glyphosate. In 2015, the World Health Organization reclassified glyphosate as probably carcinogenic to humans. A detailed overview is given of the scientific literature on the movement and residues of glyphosate and its breakdown product aminomethyl phosphonic acid (AMPA) in soil and water, their toxicity to macro- and microorganisms, their effects on microbial compositions and potential indirect effects on plant, animal and human health. Although the acute toxic effects of glyphosate and AMPA on mammals are low, there are animal data raising the possibility of health effects associated with chronic, ultra-low doses related to accumulation of these compounds in the environment. Intensive glyphosate use has led to the selection of glyphosate-resistant weeds and microorganisms. Shifts in microbial compositions due to selective pressure by glyphosate may have contributed to the proliferation of plant and animal pathogens. Research on a link between glyphosate and antibiotic resistance is still scarce but we hypothesize that the selection pressure for glyphosate-resistance in bacteria could lead to shifts in microbiome composition and increases in antibiotic resistance to clinically important antimicrobial agents. We recommend interdisciplinary research on the associations between low level chronic glyphosate exposure, distortions in microbial communities, expansion of antibiotic resistance and the emergence of animal, human and plant diseases. Independent research is needed to revisit the tolerance thresholds for glyphosate residues in water, food and animal feed taking all possible health risks into account.

Plant Diseases and Management Approaches in Organic Farming Systems.

Organic agriculture has expanded worldwide. Numerous papers were published in the past 20 years comparing plant diseases in organic and conventional crops. Root diseases are generally less severe owing to greater soil health, whereas some foliar diseases can be problematic in organic agriculture. The soil microbial community and nitrogen availability play an important role in disease development and yield. Recently, the focus has shifted to optimizing organic crop production by improving plant nutrition, weed control, and plant health. Crop-loss assessment relating productivity to all yield-forming and -reducing factors would benefit organic production and sustainability evaluation.

Potential effects of diurnal temperature oscillations on potato late blight with special reference to climate change.

Global climate change will have effects on diurnal temperature oscillations as well as on average temperatures. Studies on potato late blight (Phytophthora infestans) development have not considered daily temperature oscillations. We hypothesize that growth and development rates of P. infestans would be less influenced by change in average temperature as the magnitude of fluctuations in daily temperatures increases. We investigated the effects of seven constant (10, 12, 15, 17, 20, 23, and 27°C) and diurnally oscillating (±5 and ±10°C) temperatures around the same means on number of lesions, incubation period, latent period, radial lesion growth rate, and sporulation intensity on detached potato leaves inoculated with two P. infestans isolates from clonal lineages US-8 and US-23. A four-parameter thermodynamic model was used to describe relationships between temperature and disease development measurements. Incubation and latency progression accelerated with increasing oscillations at low mean temperatures but slowed down with increasing oscillations at high mean temperatures (P < 0.005), as hypothesized. Infection efficiency, lesion growth rate, and sporulation increased under small temperature oscillations compared with constant temperatures but decreased when temperature oscillations were large. Thus, diurnal amplitude in temperature should be considered in models of potato late blight, particularly when predicting effects of global climate change on disease development.

[Transition of entheropathogenic and saprotrophic bacteria in the eco-niche cycle: animals-excrement-soil-plants-animals].

The possibility of transition of saprotrophic and enteropathohenic bacterial populations following the chain of naturally related habitats--fodder-animal gastrointestinal tract (GIT)-animals excrement-soil-plants and again animals with a cyclic formation--has been investigated quantitatively. All bacteria used in the experiments have been shown to successfully overcome all the mechanical, physical-chemical, and biological barriers in the food chain and to come out into the environment with a quite high number. It has been demonstrated that the same bacterial population can pass the whole cycle without additional introduction of similar populations from the outside.

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. METHODS AND RESULTS: Bacterial PCR-DGGE fingerprints made with DNA extracts from the different soils using bacterial 16S-rRNA-gene-based primers were grouped by cluster analysis into two clusters consisting of six and 29 soils and one single soil at a cross-correlation level of 16% among samples per cluster. Average irregularity values for E. coli O157:H7 survival in the same soils differed significantly between clusters (P = 0.05), whereas no significant difference was found for the corresponding average ttd values (P = 0.20). The irregularity was higher for cluster 1, which consisted primarily of soils that had received liquid manure and artificial fertilizer and had a significant higher bacterial diversity and evenness values (P < 0.001). CONCLUSIONS: Bacterial PCR-DGGE fingerprints of 36 manure-amended soils revealed two clusters which differed significantly in the stability (irregularity) of E. coli O157 decline. The cluster with the higher irregularity was characterized by higher bacterial diversity and evenness. SIGNIFICANCE AND IMPACT OF THE STUDY: The consequence of a high temporal irregularity is a lower accuracy of predictions of population behaviour, which results in higher levels of uncertainty associated with the estimates of model parameters when modelling the behaviour of E. coli O157:H7 in the framework of risk assessments. Soil community structure parameters like species diversity and evenness can be indicative for the reliability of predictive models describing the fate of pathogens in (agricultural) soil ecosystems.

Modelling the contamination of lettuce with Escherichia coli O157:H7 from manure-amended soil and the effect of intervention strategies.

AIMS: A growing number of foodborne illnesses has been associated with the consumption of fresh produce. In this study, the probability of lettuce contamination with Escherichia coli O157:H7 from manure-amended soil and the effect of intervention strategies was determined. METHODS AND RESULTS: Pathogen prevalence and densities were modelled probabilistically through the primary production chain of lettuce (manure, manure-amended soil and lettuce). The model estimated an average of 0.34 contaminated heads per hectare. A minimum manure storage time of 30 days and a minimum fertilization-to-planting interval of 60 days was most successful in reducing the risk. Some specific organic farming practices concerning manure and soil management were found to be risk reducing. CONCLUSIONS: Certain specific organic farming practices reduced the likelihood of contamination. This cannot be generalized to organic production as a whole. However, the conclusion is relevant for areas like the Netherlands where there is high use of manure in both organic and conventional vegetable production. SIGNIFICANCE AND IMPACT OF THE STUDY: Recent vegetable-associated disease outbreaks stress the importance of a safe vegetable production chain. The present study contributed to this by providing a first estimate of the likelihood of lettuce contamination with E. coli O157:H7 and the effectiveness of risk mitigation strategies.

Physiological and molecular responses of Lactuca sativa to colonization by Salmonella enterica serovar Dublin.

This paper describes the physiological and molecular interactions between the human-pathogenic organism Salmonella enterica serovar Dublin and the commercially available mini Roman lettuce cv. Tamburo. The association of S. enterica serovar Dublin with lettuce plants was first determined, which indicated the presence of significant populations outside and inside the plants. The latter was evidenced from significant residual concentrations after highly efficient surface disinfection (99.81%) and fluorescence microscopy of S. enterica serovar Dublin in cross sections of lettuce at the root-shoot transition region. The plant biomass was reduced significantly compared to that of noncolonized plants upon colonization with S. enterica serovar Dublin. In addition to the physiological response, transcriptome analysis by cDNA amplified fragment length polymorphism analysis also provided clear differential gene expression profiles between noncolonized and colonized lettuce plants. From these, generally and differentially expressed genes were selected and identified by sequence analysis, followed by reverse transcription-PCR displaying the specific gene expression profiles in time. Functional grouping of the expressed genes indicated a correlation between colonization of the plants and an increase in expressed pathogenicity-related genes. This study indicates that lettuce plants respond to the presence of S. enterica serovar Dublin at physiological and molecular levels, as shown by the reduction in growth and the concurrent expression of pathogenicity-related genes. In addition, it was confirmed that Salmonella spp. can colonize the interior of lettuce plants, thus potentially imposing a human health risk when processed and consumed.

Comparison of methods of extracting Salmonella enterica serovar Enteritidis DNA from environmental substrates and quantification of organisms by using a general internal procedural control.

This paper compares five commercially available DNA extraction methods with respect to DNA extraction efficiency of Salmonella enterica serovar Enteritidis from soil, manure, and compost and uses an Escherichia coli strain harboring a plasmid expressing green fluorescent protein as a general internal procedural control. Inclusion of this general internal procedural control permitted more accurate quantification of extraction and amplification of S. enterica serovar Enteritidis in these samples and reduced the possibility of false negatives. With this protocol it was found that the optimal extraction method differed for soil (Mobio soil DNA extraction kit), manure (Bio101 soil DNA extraction kit), and compost (Mobio fecal DNA extraction kit). With each method, as little as 1.2 x 10(3) to 1.8 x 10(3) CFU of added serovar Enteritidis per 100 mg of substrate could be detected by direct DNA extraction and subsequent S. enterica-specific TaqMan PCR. After bacterial enrichment, as little as 1 CFU/100 mg of original substrate was detected. Finally, the study presents a more accurate molecular analysis for quantification of serovar Enteritidis initially present in soil or manure using DNA extraction and TaqMan PCR.

Short-term wavelike dynamics of bacterial populations in response to nutrient input from fresh plant residues.

The objectives of the research were to investigate short-term dynamics of bacterial populations in soil after a disturbance in the form of fresh organic matter incorporation and to investigate how these dynamics are linked to those of some environmental parameters. To reach these objectives, soil bacterial populations, mineral nitrogen, pH, and redox potential (ROP) were monitored daily for 1 month after incorporation of clover-grass (CG) plant material in microcosm experiments. Colony-forming units (CFUs) and direct microscopic counts of FDA-stained and FTTC-stained bacteria increased immediately after incorporation of the plant material, dropped within 2 days, and fluctuated thereafter. Harmonics analysis demonstrated that there were significant wavelike fluctuations with three or four significant peaks within 1 month after incorporation of clover-grass material. Peaks in CFUs were 1-2 days ahead of those in direct counts. Ammonium (NH4) concentrations increased from the start of the experiments until nitrification commenced. Nitrate (NO3) concentrations dropped immediately after plant incorporation, and then rose monotonically until the end of the experiments. There were no wavelike fluctuations in NH4 and NO3 concentrations, so that bacterial fluctuations could not be attributed to alternating mineral N shortages and sufficiencies. pH levels rose and declined with NH4 levels. ROP dropped shortly before NH4 concentrations rose, and increased before NH4 concentrations decreased; there were no regular fluctuations in ROP, so that temporary oxygen shortages may not have been responsible for the observed fluctuations in bacterial populations. Thus, for the first time, regular wavelike dynamics were demonstrated for bacterial populations after perturbation by addition of fresh organic matter to soil, and several potential reasons for the death phase of the fluctuations could be excluded from further consideration.

Analyses of the Relationships Between Lettuce Downy Mildew and Weather Variables Using Geographic Information System Techniques.

Previous studies in coastal California suggested that morning leaf wetness duration and temperature immediately after the prolonged leaf wetness period affect infection of lettuce by the downy mildew pathogen, Bremia lactucae. In this study, spatial analysis tools in a geographic information system were used to interpolate disease assessment data and then relate them to weather variables measured in 1995 and 1997 at weather stations in the Salinas Valley. Among the variables monitored at these weather stations, midday temperature (10:00 A.M. to 2:00 P.M.) was related most strongly to the interpolated downy mildew incidence in a circular area (radius = 5 km) around each station (r = 0.52, P < 0.0001); the higher the midday temperature, the lower the disease incidence. High humidity and prolonged morning leaf wetness duration also were associated with high downy mildew incidence. Cluster analysis resulted in distinct regions with different midday temperatures, which overlapped well (92.2% of the total area) with regions distinguished in previous cluster analyses of downy mildew incidence. Clusters of morning relative humidity showed similar patterns, although they overlapped less well with clusters of disease incidence. These results confirmed that midday temperature is an important determining factor for lettuce downy mildew, and its effects should be incorporated into a disease warning system for coastal California. Cluster analyses based on the effects of temperature have great potential for use in regional downy mildew risk assessment.

Comparison of real-time PCR methods for detection of Salmonella enterica and Escherichia coli O157:H7, and introduction of a general internal amplification control.

The objectives of this study were to compare different real-time PCR-based methods for detection of either Salmonella spp. or E. coli O157:H7 with respect to sensitivity, precision and accuracy. In addition, a general internal amplification control (IAC) is presented, allowing prevention of false negative results. The IAC allows insight in amplification efficiency and enables a more accurate quantification with the evaluated real-time PCR methods. Implementation of the IAC with the different PCR methods did not affect the precision of the methods, but the sensitivity was reduced 10-fold. Introduction of an IAC with the Salmonella enterica specific detection method showed a shift in Ct-value (increase of target Ct-value with 0.45+/-0.17 cycles), while with the method to detect E. coli O157:H7 no influence of IAC co-amplification was observed. The quantification threshold of the methods in which the IAC was included was determined at 1 pg of target DNA (equal to 200 CFU) per reaction. Qualitative detection was feasible down to 10 fg of target DNA per reaction using both methods in which the IAC was incorporated. The adjusted methods have the potential to provide fast and sensitive detection of Salmonella spp. or E. coli O157:H7, enabling accurate quantification and preventing false negative results by using the general IAC.

Sporulation of Bremia lactucae Affected by Temperature, Relative Humidity, and Wind in Controlled Conditions.

ABSTRACT The effects of temperature (5 to 25 degrees C), relative humidity (81 to 100%), wind speed (0 to 1.0 m s(-1)), and their interactions on sporulation of Bremia lactucae on lettuce cotyledons were investigated in controlled conditions. Sporulation was affected significantly (P < 0.0001) by temperature, with an optimum at 15 degrees C, and by relative humidity (RH), with sporulation increasing markedly at RH >/= 90%. There was a significant effect of exposure time in relation to temperature (P = 0.0007) but not to RH. In separate experiments, both RH and wind speed significantly (P < 0.0001) affected the number of cotyledons with sporulation and the number of sporangia produced per cotyledon. No sporulation was observed at wind speeds of >0.5 m s(-1), regardless of RH. In still air, the number of sporangiophores produced per cotyledon increased linearly with RH from 81 to 100% (P = 0.0001, r = 0.98). Histological observations indicated that sporulation may be affected by stomatal aperture in response to RH, as more closed stomata and correspondingly fewer sporangiophores were present at lower RH. These results are important for understanding the mechanism of RH effects on sporulation and for predicting conditions conducive to downy mildew development.

Wavelike distributions of infections by an introduced and naturally occurring root pathogen along wheat roots.

Previously, we showed that copiotrophic and oligotrophic bacteria fluctuate as moving waves along roots. These waves probably originate as a result of growth and death cycles at any location where a moving nutrient source passed. In this study, we placed sclerotia of Rhizoctonia solani AG8 along growing roots of wheat and showed that the proportions of root sections from which R. solani was isolated fluctuated with distance from the root tip. Similarly, proportions of root sections from which naturally occurring Pythium spp. were isolated fluctuated with distance from the root tip. Fourier analysis showed that these fluctuations constituted significant waves. Cross-correlation analyses demonstrated that there were negative correlations between R. solani infections and colony forming units of copiotrophic bacteria at the time of inoculation at the same locations on the root (lag = 0 cm), indicating that infection by R. solani could have been inhibited by these bacteria. There was a positive correlation between Pythium infections and copiotrophic bacteria at a lag of 6 cm along the roots. It therefore appears that Pythium infection took place shortly after the initial peak in copiotrophic bacteria following the passage of the root tip.

Incorporation of temperature and solar radiation thresholds to modify a lettuce downy mildew warning system.

ABSTRACT The effect of temperature on infection of lettuce by Bremia lactucae was investigated in controlled environment studies and in the field. In controlled conditions, lettuce seedlings inoculated with B. lactucae were incubated at 15, 20, 25, or 30 degrees C during a 4-h wet period immediately after inoculation or at the same temperatures during an 8-h dry period after the 4-h postinoculation wet period at 15 degrees C. High temperatures during wet and dry periods reduced subsequent disease incidence. Historical data from field studies in 1991 and 1992, in which days with or without infection had been identified, were analyzed by comparing average air temperatures during 0600 to 1000 and 1000 to 1400 Pacific standard time (PST) between the two groups of days. Days without infection had significantly higher temperatures (mean 21.4 degrees C) than days with infection (20.3 degrees C) during 1000 to 1400 PST (P < 0.01) but not during 0600 to 1000 PST. Therefore, temperature thresholds of 20 and 22 degrees C for the 3-h wet period after sunrise and the subsequent 4-h postpenetration period, respectively, were added to a previously developed disease warning system that predicts infection when morning leaf wetness lasts >/=4 h from 0600 PST. No infection was assumed to occur if average temperature during these periods exceeded the thresholds. Based on nonlinear regression and receiver operating characteristic curve analysis, the leaf wetness threshold of the previous warning system was also modified to >/=3-h leaf wetness (>/=0900 PST). Furthermore, by comparing solar radiation on days with infection and without infection, we determined that high solar radiation during 0500 to 0600 PST in conjunction with leaf wetness ending between 0900 and 1000 PST was associated with downy mildew infection. Therefore, instead of starting at 0600 PST, the calculation of the 3-h morning leaf wetness period was modified to start after sunrise, defined as the hour when measured solar radiation exceeded 8 W m(-2) (or 41 mumol m(-2) s(-1) for photon flux density). The modified warning system was compared with the previously developed system using historical weather and downy mildew data collected in coastal California. The modified system was more conservative when disease potential was high and recommended fewer fungicide applications when conditions were not conducive to downy mildew development.

Comparison of Three Fungicide Spray Advisories for Lettuce Downy Mildew.

Lettuce growers in coastal California have relied mainly on protective fungicide sprays to control downy mildew. Thus, timing of sprays before infection is critical for optimal results. A leaf-wetness-driven, infection-based advisory system, previously developed, did not always perform satisfactorily. In this study, the advisory system was modified by incorporating a pathogen survival component (system 1) or both survival and sporulation components (system 2). These systems were then evaluated in commercial lettuce fields in coastal California during 1996-1998. Three or four treatments were carried out in each field: (i) no spray; (ii) sprays as scheduled by the growers; (iii) sprays following modified system 1; and (iv) sprays following the original advisory system (1996) or modified system 2 (1998). Downy mildew incidence was evaluated every 2 to 9 days. In fields with drip irrigation, the number of fungicide applications was reduced by one or two regardless of the advisory system used compared to the grower's calendar-based schedule, although one unnecessary spray was recommended in 1996 at Soledad and 1997 at Salinas. Under all three systems, disease levels were low (incidence <25% and about 1 lesion per plant) for fields with drip irrigation, but not for fields with sprinklers (incidence up to 100% and 5 to 10 lesions per plant). For the first time, we established that survival and sporulation components are not needed for a lettuce downy mildew forecasting system. Instead, a threshold with a shorter period of morning leaf wetness and high temperatures were found to have potential for improving forecasting efficiency.

Effects of salinity on severity of infection by Phytophthora parasitica Dast., ion concentrations and growth of tomato, Lycopersicon esculentum Mill.

The response of tomato (Lycopersictm esculentum Mill., cv. UC82B) to salinity, alone and in combination with Phytophthora parasitica Dast. (a fungal pathogen causing root rot) was investigated in a field study. Three salinity regimes were established: 1) a low salinity control, 2 medium salinity, where 75 mm total salts (NaCl and CaCU in a 4:1 molar ratio of Na:Ca) were added to the irrigation water to give an electrical conductivity (EC) of approximately 8 dS m-1 , and 3) high salinity, where ISO mM total salts (4:1 molar ratio of Na: Ca) gave an EC of approximately 16 dS m-1 . Half of the plots were inoculated with P. parasitica. and the remainder were treated with a selective fungicide to inhibit the pathogen. Soil salinity markedly increased the incidence of Phytophthora root rot in both years of the study. The combination of salinity and enhanced disease severity led to significant reductions in fresh fruit yields, fruit size and, to a lesser extent, total above ground biomass. Fruit size was affected to the greatest extent and showed a strong interaction between the effects of disease and salinity, suggesting that the import of water by fruit was more sensitive than dry matter production to the combination of these stresses. Net root growth (0-50 cm depth) was greatly reduced (by 40-50%) in the presence of salinity, whereas P. parasitica had no discernable effect even when more than 50% of the root system showed severe root rot lesions. In spite of the reduced root system, leaf water potential was not affected by disease in the 1989 growing season. During the fruit-fill period in 1988, however, leaf water potential was more negative in inoculated plots. A marked degree of leaf ion homeostasis was maintained even under high salt and root rot stress. Excessive build up of Cl or Na concentrations in the leaves did not contribute substantially to the observed reductions in plant growth and yield. The results suggest that a reduced root growth rate or an enhanced root death rate may be at least partially responsible for the increased disease severity at high salinity.