Modelling the monthly abundance of Culicoides biting midges in nine European countries using Random Forests machine learning.

BACKGROUND: Culicoides biting midges transmit viruses resulting in disease in ruminants and equids such as bluetongue, Schmallenberg disease and African horse sickness. In the past decades, these diseases have led to important economic losses for farmers in Europe. Vector abundance is a key factor in determining the risk of vector-borne disease spread and it is, therefore, important to predict the abundance of Culicoides species involved in the transmission of these pathogens. The objectives of this study were to model and map the monthly abundances of Culicoides in Europe. METHODS: We obtained entomological data from 904 farms in nine European countries (Spain, France, Germany, Switzerland, Austria, Poland, Denmark, Sweden and Norway) from 2007 to 2013. Using environmental and climatic predictors from satellite imagery and the machine learning technique Random Forests, we predicted the monthly average abundance at a 1 km2 resolution. We used independent test sets for validation and to assess model performance. RESULTS: The predictive power of the resulting models varied according to month and the Culicoides species/ensembles predicted. Model performance was lower for winter months. Performance was higher for the Obsoletus ensemble, followed by the Pulicaris ensemble, while the model for Culicoides imicola showed a poor performance. Distribution and abundance patterns corresponded well with the known distributions in Europe. The Random Forests model approach was able to distinguish differences in abundance between countries but was not able to predict vector abundance at individual farm level. CONCLUSIONS: The models and maps presented here represent an initial attempt to capture large scale geographical and temporal variations in Culicoides abundance. The models are a first step towards producing abundance inputs for R0 modelling of Culicoides-borne infections at a continental scale.

Modeling the Influence of Ambient Temperature on the Interactions Between the Stable Fly (Diptera: Muscidae) and Its Natural Enemy Spalangia cameroni (Hymenoptera: Pteromalidae) to Assess Consequences of Climate Change.

A simulation model was used to predict how temperature influences biological control of stable flies (Stomoxys calcitrans (L.)) by the pupal parasitoid Spalangia cameroni. Temperature, which was either constant or fluctuated due to seasonal variation and/or environmental stochasticity, was modeled as a first order autocorrelation process. The simulations showed that stable flies could tolerate a wider temperature interval than expected from their thermal performance curve (TPC). This was attributed to the fact that immature flies develop in manure, which protects them against low air temperatures. In contrast, the parasitoids were found to have a narrower thermal tolerance range than expected from their TPC. This was attributed to the temperature-dependent functional response of S. cameroni, which was a limiting factor for the parasitoid's development and survival when host densities were low at suboptimal temperatures. The effects of seasonal variation on critical thermal limits were studied by means of thermal performance diagrams (TPDs). Fluctuating temperatures narrowed the thermal tolerance range of both species. At constant temperatures, the simulations showed that the optimal temperature for using S. cameroni in control of stable flies is ~20°C and that the parasitoid can persist in environments with yearly average temperatures between 18 and 29°C. However, if temperature variation was taken into consideration, it changed both the optimal temperature and the temperature interval at which biological control will be possible. This indicates that climate change causing increasing temperatures compounded with greater fluctuations may have serious consequences for biological control of pests.

Integrated genome-wide investigations of the housefly, a global vector of diseases reveal unique dispersal patterns and bacterial communities across farms.

BACKGROUND: Houseflies (Musca domestica L.) live in intimate association with numerous microorganisms and is a vector of human pathogens. In temperate areas, houseflies will overwinter in environments constructed by humans and recolonize surrounding areas in early summer. However, the dispersal patterns and associated bacteria across season and location are unclear. We used genotyping-by-sequencing (GBS) for the simultaneous identification and genotyping of thousands of Single Nucleotide Polymorphisms (SNPs) to establish dispersal patterns of houseflies across farms. Secondly, we used 16S rRNA gene amplicon sequencing to establish the variation and association between bacterial communities and the housefly across farms. RESULTS: Using GBS we identified 18,000 SNPs across 400 individuals sampled within and between 11 dairy farms in Denmark. There was evidence for sub-structuring of Danish housefly populations and with genetic structure that differed across season and sex. Further, there was a strong isolation by distance (IBD) effect, but with large variation suggesting that other hidden geographic barriers are important. Large individual variations were observed in the community structure of the microbiome and it was found to be dependent on location, sex, and collection time. Furthermore, the relative prevalence of putative pathogens was highly dependent on location and collection time. CONCLUSION: We were able to identify SNPs for the determination of the spatiotemporal housefly genetic structure, and to establish the variation and association between bacterial communities and the housefly across farms using novel next-generation sequencing (NGS) techniques. These results are important for disease prevention given the fine-scale population structure and IBD for the housefly, and that individual houseflies carry location specific bacteria including putative pathogens.

Quantifying the potential for bluetongue virus transmission in Danish cattle farms.

We used a mechanistic transmission model to estimate the number of infectious bites (IBs) generated per bluetongue virus (BTV) infected host (cattle) using estimated hourly microclimatic temperatures at 22,004 Danish cattle farms for the period 2000-2016, and Culicoides midge abundance based on 1,453 light-trap collections during 2007-2016. We used a range of published estimates of the duration of the hosts' infectious period and equations for the relationship between temperature and four key transmission parameters: extrinsic incubation period, daily vector survival rate, daily vector biting rate and host-to-vector transmission rate resulting in 147,456 combinations of daily IBs. More than 82% combinations of the parameter values predicted > 1 IBs per host. The mean IBs (10-90th percentiles) for BTV per infectious host were 59 (0-73) during the transmission period. We estimated a maximum of 14,954 IBs per infectious host at some farms, while a best-case scenario suggested transmission was never possible at some farms. The use of different equations for the vector survival rate and host-to-vector transmission rates resulted in large uncertainty in the predictions. If BTV is introduced in Denmark, local transmission is very likely to occur. Vectors infected as late as mid-September (early autumn) can successfully transmit BTV to a new host until mid-November (late autumn).

Monthly variation in the probability of presence of adult Culicoides populations in nine European countries and the implications for targeted surveillance.

BACKGROUND: Biting midges of the genus Culicoides (Diptera: Ceratopogonidae) are small hematophagous insects responsible for the transmission of bluetongue virus, Schmallenberg virus and African horse sickness virus to wild and domestic ruminants and equids. Outbreaks of these viruses have caused economic damage within the European Union. The spatio-temporal distribution of biting midges is a key factor in identifying areas with the potential for disease spread. The aim of this study was to identify and map areas of neglectable adult activity for each month in an average year. Average monthly risk maps can be used as a tool when allocating resources for surveillance and control programs within Europe. METHODS: We modelled the occurrence of C. imicola and the Obsoletus and Pulicaris ensembles using existing entomological surveillance data from Spain, France, Germany, Switzerland, Austria, Denmark, Sweden, Norway and Poland. The monthly probability of each vector species and ensembles being present in Europe based on climatic and environmental input variables was estimated with the machine learning technique Random Forest. Subsequently, the monthly probability was classified into three classes: Absence, Presence and Uncertain status. These three classes are useful for mapping areas of no risk, areas of high-risk targeted for animal movement restrictions, and areas with an uncertain status that need active entomological surveillance to determine whether or not vectors are present. RESULTS: The distribution of Culicoides species ensembles were in agreement with their previously reported distribution in Europe. The Random Forest models were very accurate in predicting the probability of presence for C. imicola (mean AUC = 0.95), less accurate for the Obsoletus ensemble (mean AUC = 0.84), while the lowest accuracy was found for the Pulicaris ensemble (mean AUC = 0.71). The most important environmental variables in the models were related to temperature and precipitation for all three groups. CONCLUSIONS: The duration periods with low or null adult activity can be derived from the associated monthly distribution maps, and it was also possible to identify and map areas with uncertain predictions. In the absence of ongoing vector surveillance, these maps can be used by veterinary authorities to classify areas as likely vector-free or as likely risk areas from southern Spain to northern Sweden with acceptable precision. The maps can also focus costly entomological surveillance to seasons and areas where the predictions and vector-free status remain uncertain.

Detection of live larvae in cocoons of Bathyplectes curculionis (Hymenoptera: Ichneumonidae) using visible/near-infrared multispectral imaging.

BACKGROUND: The multispectral (MS) imaging system is a non-destructive method with potential to reduce the labour and time required for quality control in the production of beneficial arthropods such as the parasitoid Bathyplectes curculionis. In Denmark, a project is being undertaken that focuses on the possible use of B. curculionis in augmentative control of Hypera weevil pests in white clover seed production where cocoons of the parasitoid remain as a by-product of seed processing. Only a fraction of the by-product contains live parasitoid larvae and an effective method is required detect live cocoons for later augmentative control of the pest. Therefore, this study aims to identify live larval cocoons of B. curculionis using the MS imaging system. RESULTS: Live and dead cocoons were identified using the canonical discriminant analysis (CDA) model with an accuracy of 91% and 80% (error rate 14%) in the training set, and a predicted accuracy of 89% and 81% (error rate 15%) in the test set. Reflectance from the near-infrared region was valuable in identifying live cocoons compared with that from the visible region. CONCLUSION: The MS imaging system is a rapid method for the separation of live and dead cocoons of B. curculionis. This study shows the potential of developing an MS imaging system to facilitate sorting of live and dead cocoons and optimize augmentative control of Hypera weevil pests. © 2018 Society of Chemical Industry.

Spatial and temporal variation in the abundance of Culicoides biting midges (Diptera: Ceratopogonidae) in nine European countries.

BACKGROUND: Biting midges of the genus Culicoides (Diptera: Ceratopogonidae) are vectors of bluetongue virus (BTV), African horse sickness virus and Schmallenberg virus (SBV). Outbreaks of both BTV and SBV have affected large parts of Europe. The spread of these diseases depends largely on vector distribution and abundance. The aim of this analysis was to identify and quantify major spatial patterns and temporal trends in the distribution and seasonal variation of observed Culicoides abundance in nine countries in Europe. METHODS: We gathered existing Culicoides data from Spain, France, Germany, Switzerland, Austria, Denmark, Sweden, Norway and Poland. In total, 31,429 Culicoides trap collections were available from 904 ruminant farms across these countries between 2007 and 2013. RESULTS: The Obsoletus ensemble was distributed widely in Europe and accounted for 83% of all 8,842,998 Culicoides specimens in the dataset, with the highest mean monthly abundance recorded in France, Germany and southern Norway. The Pulicaris ensemble accounted for only 12% of the specimens and had a relatively southerly and easterly spatial distribution compared to the Obsoletus ensemble. Culicoides imicola Kieffer was only found in Spain and the southernmost part of France. There was a clear spatial trend in the accumulated annual abundance from southern to northern Europe, with the Obsoletus ensemble steadily increasing from 4000 per year in southern Europe to 500,000 in Scandinavia. The Pulicaris ensemble showed a very different pattern, with an increase in the accumulated annual abundance from 1600 in Spain, peaking at 41,000 in northern Germany and then decreasing again toward northern latitudes. For the two species ensembles and C. imicola, the season began between January and April, with later start dates and increasingly shorter vector seasons at more northerly latitudes. CONCLUSION: We present the first maps of seasonal Culicoides abundance in large parts of Europe covering a gradient from southern Spain to northern Scandinavia. The identified temporal trends and spatial patterns are useful for planning the allocation of resources for international prevention and surveillance programmes in the European Union.

Modeling the Temperature- and Age-Dependent Survival, Development, and Oviposition Rates of Stable Flies (Stomoxys calcitrans) (Diptera: Muscidae).

Stable flies (Stomoxys calcitrans (L.)) can be a serious pest associated with cattle facilities. In Denmark, they occur most abundantly at organic farms, where they cannot be controlled by means of insecticides. On traditional farms, where chemical control is widely used, development of resistance is of increasing concern. Therefore, interest in biological control or other alternative methods has been growing during the recent years. In order to understand the complex relationships between a pest and its natural enemies in a variable environment, it is necessary to know how temperature affects the dynamics of the involved species. In this paper, we apply data derived from several existing sources to investigate the influence of temperature on development and survival of eggs, larvae, pupae, and adult stable flies, as well as on the fecundity of adult females. We demonstrate that the same modeling framework (called SANDY), previously applied to lifetable data of the pteromalid pupal parasitoid (Spalangia cameroni Perkins), a biological control agent used against stable flies, can also be used to model S. calcitrans. However, the predicted temperature responses depend on the data sources used to parameterize the model, which is reflected by differences in estimated population growth rates obtained from American and non-American studies. Elasticity analysis shows that growth rates are more sensitive to changes in viability, in particular of adult flies, than in fecundity, which may have implications for the management of stable fly populations.

Role of the aphid species and their feeding locations in parasitization behavior of Aphelinus abdominalis, a parasitoid of the lettuce aphid Nasonovia ribisnigri.

Aphid species feeding on lettuce occupy distinct feeding sites: the lettuce aphid Nasonovia ribisnigri prefers to feed on heart leaves, whereas the potato aphid Macrosiphum euphorbiae feeds only on outer leaves. The aphid parasitoid Aphelinus abdominalis, known to be able to regulate M. euphorbiae on many crops, has recently been indicated as a promising biocontrol candidate also for use against N. ribisnigri, a major pest of lettuce. This study therefore examined A. abdominalis parasitization preference between N. ribisnigri and M. euphorbiae and its ability to parasitize aphids feeding on different parts of lettuce plants. In addition, life history traits of A. abdominalis on these aphid species were investigated. In no-choice laboratory experiments on leaf discs and 24 h exposure, A. abdominalis successfully parasitized 54% and 60% of the offered N. ribisnigri and M. euphorbiae, respectively, with no significant difference. In the corresponding choice experiment, however, A. abdominalis had a tendency for a significantly higher preference for M. euphorbiae (38%) compared to N. ribisnigri (30%). Growth chamber experiments on whole plants demonstrated that A. abdominalis was able to parasitize aphids, regardless of their feeding locations on lettuce plants. However, aphid feeding behavior had a significant effect on the parasitization rate. A. abdominalis parasitized significantly higher percentages of M. euphorbiae or N. ribisnigri when aphids were exposed separately to parasitoids on whole lettuce plants as compared with N. ribisnigri exposed only on heart leaf. A significant preference of A. abdominalis for M. euphorbiae compared to N. ribisnigri was also observed in the growth chamber choice experiment. A high percentage of adult emergence (> 84%) and female-biased sex ratio (> 83%) were found irrespective of the aphid species.

Genome-wide fitness analyses of the foodborne pathogen Campylobacter jejuni in in vitro and in vivo models.

Campylobacter is the most common cause of foodborne bacterial illness worldwide. Faecal contamination of meat, especially chicken, during processing represents a key route of transmission to humans. There is a lack of insight into the mechanisms driving C. jejuni growth and survival within hosts and the environment. Here, we report a detailed analysis of C. jejuni fitness across models reflecting stages in its life cycle. Transposon (Tn) gene-inactivation libraries were generated in three C. jejuni strains and the impact on fitness during chicken colonisation, survival in houseflies and under nutrient-rich and -poor conditions at 4 °C and infection of human gut epithelial cells was assessed by Tn-insertion site sequencing (Tn-seq). A total of 331 homologous gene clusters were essential for fitness during in vitro growth in three C. jejuni strains, revealing that a large part of its genome is dedicated to growth. We report novel C. jejuni factors essential throughout its life cycle. Importantly, we identified genes that fulfil important roles across multiple conditions. Our comprehensive screens showed which flagella elements are essential for growth and which are vital to the interaction with host organisms. Future efforts should focus on how to exploit this knowledge to effectively control infections caused by C. jejuni.

Bacterial Communities Associated with Houseflies (Musca domestica L.) Sampled within and between Farms.

The housefly feeds and reproduces in animal manure and decaying organic substances and thus lives in intimate association with various microorganisms including human pathogens. In order to understand the variation and association between bacteria and the housefly, we used 16S rRNA gene amplicon sequencing to describe bacterial communities of 90 individual houseflies collected within and between ten dairy farms in Denmark. Analysis of gene sequences showed that the most abundant classes of bacteria found across all sites included Bacilli, Clostridia, Actinobacteria, Flavobacteria, and all classes of Proteobacteria and at the genus level the most abundant genera included Corynebacterium, Lactobacillus, Staphylococcus, Vagococcus, Weissella, Lactococcus, and Aerococcus. Comparison of the microbiota of houseflies revealed a highly diverse microbiota compared to other insect species and with most variation in species richness and diversity found between individuals, but not locations. Our study is the first in-depth amplicon sequencing study of the housefly microbiota, and collectively shows that the microbiota of single houseflies is highly diverse and differs between individuals likely to reflect the lifestyle of the housefly. We suggest that these results should be taken into account when addressing the transmission of pathogens by the housefly and assessing the vector competence variation under natural conditions.

Temperature- and Age-Dependent Survival, Development, and Oviposition Rates of the Pupal Parasitoid Spalangia cameroni (Hymenoptera: Pteromalidae).

The combined effect of temperature and age on development, survival, attack rate, and oviposition of the parasitoid Spalangia cameroni (Perkins) (Hymenoptera: Pteromalidae) exploiting house fly pupae was investigated by conducting life-table experiments at 15, 20, 25, 30 and 35°C. Temperature had a pronounced effect on survival and development of the immature stages. Survival was highest at 25°C, where 88.5% of the parasitized host pupae resulted in adult parasitoids, and lowest at 35°C when only 3.78% emerged. Females constituted between 50% (at 20°C) and 100% (at 35°C) of the surviving immatures. Males developed faster than females, with the shortest developmental times at 30°C (18.18 d for males and 19.41 d for females). Longevity of adult females decreased with temperature from 80 d at 15°C to 18 d at 35°C. Total attack rate of female parasitoids was highest at 20°C (106 hosts per female), and life-time reproduction highest at 20°C and 25°C (about 60 offspring per female). Sex ratio was female biased (65% females). A generic model was used to estimate and predict the temperature effect on the intrinsic rate of increase (rm), the net reproduction rate (R0), and the generation time (G). The model predicted that rm peaks at 33.5°C (rm = 0.182 d(-1)), that maximum R0 is reached at 27.2°C (R0 = 50.2), and that the shortest generation time occurs at 34.5°C (G = 21.1 d). Doubling time was 4.19 d at 33°C. In the temperature range between 20°C and 30°C, S. cameroni has the potential to be an efficient control agent against nuisance flies.

Persistence of low-pathogenic avian influenza H5N7 and H7N1 subtypes in house flies (Diptera: Muscidae).

Avian influenza caused by avian influenza virus (AIV) has a negative impact on poultry production. Low-pathogenic AIV (LPAIV) is naturally present in wild birds, and the introduction of the virus into domestic poultry is assumed to occur through contact with wild birds and by human activity, including the movement of live and dead poultry, and fomites such as clothing and vehicles. At present, the possible role of insects in the spread of AIV is dubious. The objective of the present work was to investigate the potential transmission of LPAIV by persistence of the virus in the alimentary tract of house flies, Musca domestica L. (Diptera: Muscidae). Flies were fed three virus concentrations of two AIV strains and then incubated at different temperatures for up to 24 h. The persistence of the two virus strains in the flies declined with increasing incubation temperatures and incubation periods. Similarly, increased virus uptake by the flies increased the persistence of virus. Persistence of infective AIV in flies differed significantly between the two virus strains. The laboratory experiments of the present study indicate that the house fly can be a potential carrier of AIV.

Molecular identification of bloodmeals from biting midges (Diptera: Ceratopogonidae: Culicoides Latreille) in Denmark.

Culicoides vectors are critical to the survival and transmission of bluetongue virus as infection only occurs in areas or regions where competent vectors are present. The success of Culicoides biting midges as vectors is mainly related to their vast population sizes and to their means of dispersal. Their choice of host for blood feeding is sparsely described. The aim of the present study was to establish methods for the identification of bloodmeal hosts and determine the identity and diversity of bloodmeals of vertebrate hosts from wild-caught biting midges near livestock farms. The study includes some of the most common and abundant species of biting midges in Denmark: Culicoides obsoletus, Culicoides scoticus, Culicoides pulicaris and Culicoides punctatus. We collected 8,378 biting midges including nine species of Culicoides of which blood-fed specimens were found from six species. We identified 251 blood engorged biting midges, and hosts were identified in 115 of 125 analysed specimens (90%). Cow, roe deer, horse, mallard and wood pigeon were identified as hosts. The most abundant host species was cow, which constituted 73.9% of the total identified bloodmeals, but the common wood pigeon was found with a frequency as high as 18.3%. In conclusion, the molecular methods applied were proven useful in identifying bloodmeal hosts from different Culicoides species. The results indicate that Culicoides species are opportunistic in their choice of bloodmeal host with a preference for cattle when present, which is important to have in mind for epidemiologist when making predictive models. Accordingly, the results of this study will add useful parameters for modelling bluetongue virus transmission and in the development of veterinary contingency plans.

Use of fly screens to reduce Campylobacter spp. introduction in broiler houses.

Fly screens that prevented influx of flies in 20 broiler houses during the summer of 2006 in Denmark caused a decrease in Campylobacter spp.-positive flocks from 51.4% in control houses to 15.4% in case houses. A proportional reduction in the incidence of chicken-borne campylobacteriosis can be expected by comprehensive intervention against flies in broiler production houses.

Flies and Campylobacter infection of broiler flocks.

A total of 8.2% of flies caught outside a broiler house in Denmark had the potential to transmit Campylobacter jejuni to chickens, and hundreds of flies per day passed through the ventilation system into the broiler house. Our study suggests that flies may be an important source of Campylobacter infection of broiler flocks in summer.

Life table study of Sitotroga cerealella (Lepidoptera: Gelichiidae), a strain from West Africa.

Life table studies for the Angoumois grain moth, Sitotroga cerealella (Olivier), a pest on stored maize, Zea mays L., in West Africa, were conducted as part of the expansion of a mathematical simulation model that has been developed for two pests of stored maize. The effects of four temperatures (20, 25, 30, and 35 degrees C) and two relative humidity levels (44 and 80%) on developmental time, age-specific survivorship and fecundity, sex ratio, and intrinsic rate of natural increase (r(m)) of S. cerealella were investigated. Sex ratio was close to 1:1 at all temperatures and humidity. Minimum development time occurred close to 32 degrees C and 80% RH for both males and females, and developmental time of females was significantly shorter than that of males. Immature survivorship was highest between 25 and 30 degrees C and 80% RH and lowest at 35 degrees C under both humidity conditions. A similar low level was found at 20 degrees C and 44% RH. The greatest fecundity (124 eggs per female) occurred at 20 degrees C, 80% RH. The maximum r(m) value was 0.086 d(-1) at 30 degrees C and 80% RH, but the growth rate declined dramatically at 35 degrees C. If compared with the few other life table studies conducted on this species on maize in India and North America, some variation among the strains becomes evident. A common conclusion for the current study and previous ones is that optimal population development for S. cerealella occurs at approximately 30 degrees C and at high humidity.