Proposals for new spray drift exposure values in orchards and vineyards for residents and bystanders.

In the EU, predicted exposure to spray drift for residents and bystanders from applications in orchards and vineyards is based on data from one study published in 1987, where one downwind distance (8 m) was considered. CropLife Europe conducted sixteen new GLP compliant studies in 4 EU countries, 8 in orchards, 8 in vineyards with early and late season applications, using adult and child mannequins located 5, 10 and 15 m downwind from the last row to measure dermal and inhalation exposures. The resulting "Bystander Resident Orchard Vineyard (BROV)" database comprises 288 observations and offers a more comprehensive option for exposure prediction. There were differences between adult and child, crop type, leaf cover and distance from the sprayer, supporting the derivation of mean, median, 75th and 95th percentile exposures for each subset. Exposures did not generally correlate with wind speed, wind direction, sprayer type, spray quality, spray concentration or amount applied. Dermal and inhalation exposure were lower in vineyards than in orchards and further analysis is required to understand why.

Terroir Influence on Polyphenol Metabolism from Grape Canes: A Spatial Metabolomic Study at Parcel Scale.

The composition of bioactive polyphenols from grape canes, an important viticultural byproduct, was shown to be varietal-dependent; however, the influence of soil-related terroir factors remains unexplored. Using spatial metabolomics and correlation-based networks, we investigated how continuous changes in soil features and topography may impact the polyphenol composition in grape canes. Soil properties, topography, and grape cane extracts were analyzed at georeferenced points over 3 consecutive years, followed by UPLC-DAD-MS-based metabolomic analysis targeting 42 metabolites. Principal component analyses on intra-vintage metabolomic data presented a good reproducibility in relation to geographic coordinates. A correlation-driven approach was used to explore the combined influence of soil and topographic variables on metabolomic responses. As a result, a metabolic cluster including flavonoids was correlated with elevation and curvature. Spatial metabolomics driven by correlation-based networks represents a powerful approach to spatialize field-omics data and may serve as new field-phenotyping tool in precision agriculture.

Linkages between traits and decomposition of weed communities along a soil management and pedoclimate gradient in Mediterranean vineyards.

BACKGROUND AND AIMS: Decomposition is a major ecosystem process which improves soil quality. Despite that, only a few studies have analysed decomposition in an agricultural context, while most agrosystems (e.g. vineyards) are facing decreasing soil quality. The objective of this study is to understand the impacts of both pedoclimate and weed management on the mass loss of vineyard weed communities during the early stages of the decomposition process through their functional properties. METHODS: In 16 Mediterranean vineyards representing both a pedoclimate and a soil management gradient, we measured the mass loss of green above-ground biomass of 50 weed communities during decomposition in standard conditions and key leaf traits of dominant species [e.g. leaf dry matter content (LDMC) and leaf lignin to nitrogen ratio (lignin:N)]. Both the mean [i.e. community-weighted mean (CWM)] and diversity (i.e. Rao index) were computed at the community level. Path analysis was used to quantify the effects of agro-environmental filters on the mass loss of weed communities through their functional properties. KEY RESULTS: Tillage and mowing filtered more decomposable communities than chemical weeding (16 and 8 % of higher mass loss after 2 months of decomposition). Path analysis selected weed management practice type as the main factor determining mass loss through its effect on functional properties, while soil and climate had minor and no effects, respectively. Chemical weeding favoured communities with higher investment in resistant leaves (e.g. 38 % higher lignin:N, 22 % lower leaf nitrogen content) which resulted in lower mass loss compared with tilled and mowed communities. Mowing favoured communities with 47 % higher biomass and with 46 % higher nitrogen content. CONCLUSIONS: Weed management significantly influenced weed mass loss, while the pedoclimate had little effect. Our results suggest that mowing is a promising alternative to herbicide use, favouring higher biomass, nitrogen content and decomposability potential of weeds.

Diversity of soil fungi in the vineyards of Changli region in China.

This study aimed to elucidate the fungal diversity of Changli vineyard soil in China. High-throughput sequencing technology was used to investigate the diversity and composition of soil fungi in five vineyards from different geographical locations in Changli. Although the five vineyards had similar fungal communities, the diversity, composition, and distribution of the high-abundance species differed. Ascomycota, Basidiomycota, and Mortierellomycota were dominant phyla. Among the 14 high-abundance genera of fungi, Odiodendron, Pleotrichocladium, and Plectosephalella have rarely been reported in other vineyards and are unique to the Changli region. In addition, Solicoccozyma aeria and Solicoccozyma terrea were the dominant species in the five vineyards and have rarely been reported in domestic vineyards. Additionally, Rhizophagus, Wardomyces, Mortierella, Volutella, and Cryptococcus were significantly different among the five vineyard soils. Among these species, Mortierella was highly abundant in each vineyard, but its contents were significantly different across vineyards. These findings enrich the information on the composition and diversity of soil fungi in the vineyard of the Changli region, which helps to explore the regional or distinctive sensorial attributes of wine from the perspective of microbial biogeography.

Experimenting Agriculture 4.0 with Sensors: A Data Fusion Approach between Remote Sensing, UAVs and Self-Driving Tractors.

Geomatics is important for agriculture 4.0; in fact, it uses different types of data (remote sensing from satellites, Unmanned Aerial Vehicles-UAVs, GNSS, photogrammetry, laser scanners and other types of data) and therefore it uses data fusion techniques depending on the different applications to be carried out. This work aims to present on a study area concerning the integration of data acquired (using data fusion techniques) from remote sensing techniques, UAVs, autonomous driving machines and data fusion, all reprocessed and visualised in terms of results obtained through GIS (Geographic Information System). In this work we emphasize the importance of the integration of different methodologies and data fusion techniques, managing data of a different nature acquired with different methodologies to optimise vineyard cultivation and production. In particular, in this note we applied (focusing on a vineyard) geomatics-type methodologies developed in other works and integrated here to be used and optimised in order to make a contribution to agriculture 4.0. More specifically, we used the NDVI (Normalized Difference Vegetation Index) applied to multispectral satellite images and drone images (suitably combined) to identify the vigour of the plants. We then used an autonomous guided vehicle (equipped with sensors and monitoring systems) which, by estimating the optimal path, allows us to optimise fertilisation, irrigation, etc., by data fusion techniques using various types of sensors. Everything is visualised on a GIS to improve the management of the field according to its potential, also using historical data on the environmental, climatic and socioeconomic characteristics of the area. For this purpose, experiments of different types of Geomatics carried out individually on other application cases have been integrated into this work and are coordinated and integrated here in order to provide research/application cues for Agriculture 4.0.

Learning from 80 years of studies: a comprehensive catalogue of non-Saccharomyces yeasts associated with viticulture and winemaking.

Non-Saccharomyces yeast species are nowadays recognized for their impact on wine´s chemical composition and sensorial properties. In addition, new interest has been given to the commercial exploitation of non-Saccharomyces starter cultures in the wine sector. However, over many years, these yeast species were considered sources of contamination in wine production and conservation, mainly due to the high levels of volatile acidity obtained. The present manuscript systematizes 80 years of literature describing non-Saccharomyces yeast species isolated from grapes and/or grape musts. A link between each reference, the accepted taxonomic name of each species and their geographical occurrence is presented, compiling information for 293 species, in a total of 231 citations. One major focus of this work relates to the isolation of non-Saccharomyces yeasts from grapevines usually ignored in most sampling studies, also as isolation from damaged grapes. These particular niches are sources of specific yeast species, which are not identified in most other explored environments. These yeasts have high potential to be explored for important and diversified biotechnological applications.

Year, Location, and Variety Impact on Grape-, Soil-, and Leaf-Associated Fungal Microbiota of Arkansas-Grown Table Grapes.

With the recent advancement of next-generation sequencing methods, there has been an increase in studies on identification of vineyard microbiota, winery-associated microbiota, and microbiota in wine fermentation. However, there have been few studies investigating the fungal microbiota of table grapes which present distinct spoilage and food safety challenges. The aims of this study were to identify and compare the impact of year, variety, and vineyard location on grape, leaf, and soil fungal communities of two varieties of table grapes, Faith and Gratitude, grown in two open-air vineyards and one high tunnel vineyard. The grape, leaf, and soil mycobiota were analyzed using high throughput amplicon sequencing of the ITS region. The sampling year and location of table grapes had an impact on grape, leaf, and soil mycobiota. Fungal diversity of grape, leaf, and soil was greater in 2017 than in 2016. Grape and leaf samples presented strong similarities in fungal communities with abundance of Sporidiobolaceae and Filobasidium in two vineyards and Cladosporium in another one. The high tunnel structure had distinct grape and leaf fungal communities compared to the two other vineyard locations. Mortierella was the predominant genus (27%) in soil samples for the three locations; however, genera of lower abundance varied between locations. These results provide extensive description of fungal communities in less-studied table grape vineyards and high tunnels, providing useful insight of potential threats and preventive strategies to help improve the production and marketability of table grapes.

First report of Seimatosporium vitifusiforme causing trunk disease in Chilean grapevines (Vitis vinifera).

Grapevine is one of the most important fruit crops in Chile and trunk diseases reduce the productivity, quality, and longevity of the vineyards. A survey was conducted in ancient (> 50 years) vineyards of Cauquenes (35°57´14´´S 72°17´07´´W) and Itata valleys (36°38´13´´S 72°30´57´´W), located in the central area of Chile, during 2019. Trunks and cordons showing dieback and dark brown to black wood discoloration were collected from 50 to 200-year-old plants of six cultivars: País, Moscatel, Torontel Amarilla, Carignan, Aliatica and Aligote. The bark was removed and 0.5-cm sections were cut from the edges of necrotic wood lesions. Subsequently, pieces were surface disinfected using 10% v/v sodium hypochlorite bleach (4.9% chlorine), plated on acidified quarter-strength potato dextrose agar (APDA) (25% PDA, acidified with 0.1% v/v 85% lactic acid) and incubated at 25°C, for 14 to 28 days. Hyphal tips were excised and transferred to PDA to obtain pure cultures. Along with the conidiomata and conidia produced, growth rate, color and shape of the colonies on PDA, after 7 and 14 days of incubation at 25°C (n=17), were recorded. DNA was extracted from pure cultures of three isolates on PDA: HMV3, HMV64 and HMV81. The internal transcribed spacer region and partial ß-tubulin genes were amplified, using ITS1/ITS4 (White et al. 1990) and bt2A/bt2B (Glass & Donaldson 1995) primers, respectively. Sequences were subjected to NCBI BLAST search and compared to the published sequences. Isolated colonies were whitish to light-brown, cottony with a smooth margin (n=37). Their mycelium grew 1.9 cm after 7-days and 3.2 cm after 14-days of incubation on PDA, at 25°C. Colonies produced black globose pycnidia and curved, slightly-pigmentated, three-septated conidia 22.3-(29.8)-32.2 x 3.9-(4.8)-5.3 µm (n=30), with apical and basal flexuous appendages 4.3-(12.7)-21.5 µm (n=20). When compared to type sequences of Seimatosporium vitifusiforme (Lawrence et al. 2018), ITS and ßtub sequences identity of these isolates were 99 to 100% identical. To produce uniform healthy plants for pathogenicity tests, Petit Syrah canes (1-year old) were rooted in tap water amended with 500 ppm of indole-butyric acid, for 30 days. Plants were inoculated with 0.5-cm diameter mycelial plugs of actively growing colonies of the isolates HMV3, HMV64 and HMV81 (GenBank accessions no. MW026664, MW048518; MW026665, MW048519, and MW026666, MW048520, respectively). Sterile agar plugs were used for controls. Five plants per pathogen isolate were incubated at 25°C, in a humid chamber, for 25 days, and seven additional plants per isolate were incubated in aerated tap water, for 55 days. After the incubation period, the bark was removed and the lesions were measured. Dark necrotic lesions identical to the original observations were reproduced, both in the high humidity chamber (6% length) and water (10% length). There were no differences in lesion length among the isolates (P < 0.05). Control vines remained asymptomatic. To fulfill Koch´s postulates, isolations were made from symptomatic vines and compared to the ones used for inoculation, and found to be identical. Seimatosporium vitifusiforme was previously reported as a pathogen of Vitis vinifera in California, USA (Lawrence et al. 2018). Consequently, this is the second report of this fungus as a grapevine pathogen and the first one affecting Latin-American grapevines.

Habitat amount mediates the effect of fragmentation on a pollinator's reproductive performance, but not on its foraging behaviour.

Agricultural intensification, with its associated habitat loss and fragmentation, is among the most important drivers of the ongoing pollination crisis. In this quasi-experimental study, conducted in intensively managed vineyards in southwestern Switzerland, we tested the separate and interdependent effects of habitat amount and fragmentation on the foraging activity and reproductive performance of bumblebee Bombus t. terrestris colonies. Based on a factorial design, we selected a series of spatially replicated study sites across a dual gradient of habitat amount (area of ground-vegetated vineyards) and fragmentation (density of ground-vegetated vineyard fields) in a landscape predominantly consisting of vineyards with bare grounds. The foraging activity of individual bumblebees was measured using the radio frequency identification (RFID) technology, and we assessed final colony size to measure reproductive performance. We found an interactive effect of habitat amount and fragmentation on colony size. More specifically, the degree of fragmentation had a negative effect on bumblebee colony size when the amount of habitat was low, while it had a weak positive effect on colony size in landscapes with high amounts of habitat. At the level of individual vineyard fields, ground vegetation cover exerted a positive effect on bumblebee colony size. Fragmentation, but not habitat amount, significantly influenced foraging activity, with more foraging trips in sites with lower degrees of fragmentation. Our results emphasise the importance of studying the separate and interdependent effects of habitat amount and fragmentation to understand their influence on pollinators, providing guidance for optimising the spatial configuration of agricultural landscapes from a biodiversity viewpoint.

Evaluation of Vineyard Cropping Systems Using On-Board RGB-Depth Perception.

A non-destructive measuring technique was applied to test major vine geometric traits on measurements collected by a contactless sensor. Three-dimensional optical sensors have evolved over the past decade, and these advancements may be useful in improving phenomics technologies for other crops, such as woody perennials. Red, green and blue-depth (RGB-D) cameras, namely Microsoft Kinect, have a significant influence on recent computer vision and robotics research. In this experiment an adaptable mobile platform was used for the acquisition of depth images for the non-destructive assessment of branch volume (pruning weight) and related to grape yield in vineyard crops. Vineyard yield prediction provides useful insights about the anticipated yield to the winegrower, guiding strategic decisions to accomplish optimal quantity and efficiency, and supporting the winegrower with decision-making. A Kinect v2 system on-board to an on-ground electric vehicle was capable of producing precise 3D point clouds of vine rows under six different management cropping systems. The generated models demonstrated strong consistency between 3D images and vine structures from the actual physical parameters when average values were calculated. Correlations of Kinect branch volume with pruning weight (dry biomass) resulted in high coefficients of determination (R2 = 0.80). In the study of vineyard yield correlations, the measured volume was found to have a good power law relationship (R2 = 0.87). However due to low capability of most depth cameras to properly build 3-D shapes of small details the results for each treatment when calculated separately were not consistent. Nonetheless, Kinect v2 has a tremendous potential as a 3D sensor in agricultural applications for proximal sensing operations, benefiting from its high frame rate, low price in comparison with other depth cameras, and high robustness.

Local Perceptions, Vulnerability and Adaptive Responses to Climate Change and Variability in a Winegrowing Region in Uruguay.

Understanding climate variability in a winegrowing region is fundamental to understanding how its vineyards can adapt to climate change. For Uruguay, studying the vulnerability and adaptive responses of vineyards to climate change and climate variability is relevant due to its winegrowing region's economic importance and cultural heritage. Winegrowers and technical advisors were interviewed to evaluate their perceptions of climate change, vulnerability of their vineyards and how to adapt them. The main results showed that winegrowers had a clear perception of annual climate variability. The respondents highlighted the extreme climate events that had occurred over the previous few years and 71% of them believed that they had increased in frequency. Despite the perception of increase in climate variability in the region, they did not associate it with climate change. Overall, 43% of respondents agreed that changes in certain viticulture practices in recent years could have been due to climate change, especially those related to the preventive use of pesticides. The respondents identified climate risks that resulted in "bad" years for yield and quality (increase in extreme events (e.g., storms, hail), decrease in "cold" units in winter (i.e., temperatures <0 °C), increase in "hot" hours (i.e., >35 °C), increase in precipitation during the growing season and ripening period) as well as their impacts on vineyards. An adaptation matrix was developed from the viticulture practices that the winegrowers used in response to climate variability. Medium- and long-term adaptive responses to climate change can be based on the knowledge of winegrowers and their advisors.

Organic farming at local and landscape scales fosters biological pest control in vineyards.

While organic farming practices, which are often promoted as models of ecological intensification, generally enhance biodiversity, their effects on the delivery of ecosystem services, such as biological pest control, are still unknown. Here, using a multi-scale hierarchical design in southwestern France, we examined the effects of organic farming and seminatural habitats at the local and landscape scales on biological control services of three pests, including weeds and insects, in 42 vineyards. Organic farming at the local and landscape scales was beneficial to the mean and temporal stability of biological control services, while the proportion of seminatural habitats in the landscape reduced the level of biological pest control potential. The effects of organic farming and seminatural habitats across spatial scales varied with the type of prey considered and with time. Egg moth removal rates were higher in fields under organic management compared to conventional management while weed seed removal rates increased with the proportion of organic farming in the landscape. Larval removal rates as well as seed removal rates were always more stable within time in organic fields than in conventional fields. Moreover, independently of farming system type, local variables describing the agricultural management intensity, such as pesticide use or crop productivity, were also found to be important variables explaining levels of biological control services. Pesticide use tended to reduce biological control potential, while crop productivity was associated with contrasting biological control responses depending on the pest type. Our study demonstrates the need to target multiple spatial scales and to consider farming practices, as well as the proportion of seminatural habitats, to design functional landscapes that optimize biological pest control services.

Changes in rhizosphere bacterial communities associated with tree decline: grapevine esca syndrome case study.

An investigation was carried out on rhizosphere bacteria to determine if they may be associated with perennial crops affected by nonspecific decline, a phenomenon that is difficult to diagnose and prevent. Esca disease of grapevine was chosen for this case study because of its easy foliar symptom identification. Ribosomal DNA fingerprint analysis by polymerase chain reaction - denaturing gradient gel electrophoresis (PCR-DGGE), quantitative PCR (qPCR), and rDNA amplicon sequencing by next-generation sequencing (NGS) were adopted to investigate the bacterial communities associated with grapevines, which were selected for the presence and absence of external foliar symptoms in 11 vineyards. According to PCR-DGGE and qPCR, bacterial communities differed in site of origin (vineyards), but not between symptomatic and asymptomatic plants, whereas qPCR gave a significantly higher presence of total bacteria and Pseudomonas spp. in asymptomatic plants. NGS confirmed no difference between symptomatic and asymptomatic plants, apart from a few minor genera (<0.5%) such as Salinibacterium, Flavobacterium, Nocardia, and Janthinobacterium, which were, in all cases, higher in asymptomatic plants and whose functional role should be the object of further investigation. The fact that total bacteria and Pseudomonas were more abundant in the rhizosphere of asymptomatic grapevines and that some bacterial genera were associated with the latter, represents a new element when investigating the multiple-origin phenomenon such as esca disease of grapevine.

Effects of rows arrangement, soil management, and rainfall characteristics on water and soil losses in Italian sloping vineyards.

Erosional processes are highly affected by seasonal climatic fluctuations and soil management practices. Controlled grass cover is one of the most used soil conservation practices adopted in temperate climates, even if the protective effect of grass cover may decrease according to seasonal pattern. This technique is effective and, thus, widely adopted in the inter-rows of orchards such as olives, citrus or vineyards. This study reports the erosive events recorded in two different rain-fed hillslope vineyards with different rows orientation located in the Monferrato region, NW Italy. The study is addressed at compare the effects of different inter-row managements and rainfall characteristics on runoff and soil loss in hillslope vineyards (average slope from 15% to 35%). Rainfall, runoff and erosion variables were monitored in hydraulically bounded vineyard plots, where the inter-rows were managed with tillage and grass cover. Seventy-two erosive events were recorded in the period 1992-1996 in two vineyard plots with rows along the contour lines while 86 erosive events were recorded in two plots with rows up-and-down the slope from 2000 to 2014 (158 erosive events and four plots in total). Events were classified according to rainfall characteristics as "long-lasting", "intense" and "normal". In plots with rows along the contour lines, "intense" events were responsible for the highest mean soil loss in tilled plots (0.7 Mg ha-1) with very high erosion rates (12.3 Mg ha-1) observed during a single storm. In plots with rows up-and-down the slope the highest erosion rates, 21.2 and 3.4 Mg ha-1, were recorded during fall "long-lasting" events in the tilled and grass cover plots respectively. The grass cover proved to be effective in decreasing runoff and soil losses during most of the events (at least 68% and 61% of the occurrences, respectively) reducing soil losses especially during summer storms when most of the "intense" events occured. Furthermore, the results show the fundamental role of contour-slope row orientation in reducing runoff and soil losses, disregarding the inter-rows soil management that is adopted.

Differentiation of Saccharomyces cerevisiae populations from vineyards of the Azores Archipelago: Geography vs Ecology.

Aiming to elucidate the roles that ecology and geography play in shaping the differentiation of fermentative grape-associated Saccharomyces cerevisiae populations, several locations on six islands of the Azores Archipelago were surveyed. A total of 249 strains were isolated from spontaneous fermentations of grape samples from several varieties of two distinct grapevine species (Vitis vinifera L. and Vitis labrusca L.), in vineyards that are under regular cultivation or in abandoned vineyards. Strains were genetically analyzed using a set of nine microsatellite loci, and also phenotypically characterized using relevant physiological/biotechnological tests. Results showed that genetic divergence among populations of the same island was lower than from populations from different islands. Phenotypic comparison of the populations from each of the islands revealed significant differences between them. Strains isolated from the islands with more intensive viticultural activity - Pico, Terceira and Graciosa - showed higher levels of SO2 tolerance, possibly resulting from selection by human activity. The percentage of strains producing low levels of H2S was higher in S. Jorge (60%). Our findings were supported both by genetic and phenotypic data and provide clear evidence for the prevailing role of the geography over ecology in the differentiation of S. cerevisiae populations in the Azores Archipelago.

Spatio-temporal analysis of land use/land cover change and its effects on soil erosion (Case study in the Oplenac wine-producing area, Serbia).

In this paper, various spatial modelling techniques were applied to analyse changes in soil cover and their impact on soil erosion in the Oplenac wine-producing area in Serbia in the past (1985 and 2013) and in the future (with predictions for 2041). The Integrated Valuation of Ecosystem Services and Trade-offs Sediment Delivery Ratio (InVEST SDR) model and the Modules for Land Use Change Evaluation (MOLUSCE) model, integrated with methods of remote sensing, were successfully applied and were shown to be valid tools for predicting the impact of Land Use Land Cover (LULC) changes when estimating soil loss. The results revealed that the greatest impact of land use changes between 1985 and 2013 was on a reduction in areas under vineyards and an extension of meadow and pasturelands as an individual and social response to economic conditions during the research period. The forecast for 2041 reflected the trends observed in the previous period, with the greatest changes witnessing an increase in urban areas and a decrease in areas of arable land. It was also found that the effect of LULC changes on spatio-temporal patterns in the Oplenac wine-producing area did not have a major impact on soil loss, meaning this area, with its good agro-climatic characteristics, is suitable for the intensification of agricultural production.

VineSens: An Eco-Smart Decision-Support Viticulture System.

This article presents VineSens, a hardware and software platform for supporting the decision-making of the vine grower. VineSens is based on a wireless sensor network system composed by autonomous and self-powered nodes that are deployed throughout a vineyard. Such nodes include sensors that allow us to obtain detailed knowledge on different viticulture processes. Thanks to the use of epidemiological models, VineSens is able to propose a custom control plan to prevent diseases like one of the most feared by vine growers: downy mildew. VineSens generates alerts that warn farmers about the measures that have to be taken and stores the historical weather data collected from different spots of the vineyard. Such data can then be accessed through a user-friendly web-based interface that can be accessed through the Internet by using desktop or mobile devices. VineSens was deployed at the beginning in 2016 in a vineyard in the Ribeira Sacra area (Galicia, Spain) and, since then, its hardware and software have been tested to prevent the development of downy mildew, showing during its first season that the system can led to substantial savings, to decrease the amount of phytosanitary products applied, and, as a consequence, to obtain a more ecologically sustainable and healthy wine.

Environmental yeast communities in vineyards in the mountains of Santa Catarina State, Brazil.

Yeasts were isolated from three vineyards located in the South Region of Brazil. A cross evaluation was carried out at the oldest vineyard of the study in Pinheiro Preto. Samples of grape berries, grapevine leaves and the soil, along with samples of the winery equipment and effluent, were collected. In the Serra do Marari and Campos Novos vineyards only samples of grape clusters were obtained. The 106 yeast isolates were identified by sequencing the D1/D2 domain of LSU rDNA or ITS1-5.8S-ITS2 region in 22 species. The values for the richness indices varied between the vineyards. A comparison of the taxonomic diversity of the yeasts from these regions using the reciprocal Simpson index showed a significant difference between the Serra do Marari and Campos Novos vineyards (5.72 ± 0.36 and 2.92 ± 0.36, respectively, p < 0.0001). The functional diversity was assessed in relation to the use of carbon and nitrogen sources by the yeasts isolated from each location. In general, we observed that the Pinheiro Preto and Campos Novos vineyards differed consistently from the Serra do Marari vineyard according to these indices (FAD2, FDc and Rao, p < 0.0001). The possible spreading of Saccharomyces cerevisiae from the winery to the vineyard in Pinheiro Preto was observed.

Characterization of Aspergillus section Nigri species populations in vineyard soil using droplet digital PCR.

Identification of populations of Aspergillus section Nigri species in environmental samples using traditional methods is laborious and impractical for large numbers of samples. We developed species-specific primers and probes for quantitative droplet digital PCR (ddPCR) to improve sample throughput and simultaneously detect multiple species in each sample. The ddPCR method was used to distinguish Aspergillus niger, Aspergillus welwitschiae, Aspergillus tubingensis and Aspergillus carbonarius in mixed samples of total DNA. Relative abundance of each species measured by ddPCR agreed with input ratios of template DNAs. Soil samples were collected at six time points over two growing seasons from two raisin vineyards in Fresno County, California. Aspergillus section Nigri strains were detected in these soils in the range of 102 -105  CFU g-1 . Relative abundance of each species varied widely among samples, but in 52 of 60 samples, A. niger was the most abundant species, ranging from 38 to 88% of the total population. In combination with total plate counts, this ddPCR method provides a high-throughput method for describing population dynamics of important potential mycotoxin-producing species in environmental samples. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first study to demonstrate the utility of ddPCR as a means to quantify species of Aspergillus section Nigri in soil. This method eliminates the need for isolation and sequence identification of individual fungal isolates, and allows for greater throughput in measuring relative population sizes of important (i.e. mycotoxigenic) Aspergillus species within a population of morphologically indistinguishable species.