TY - JOUR
AU - Walton, Vaughn, M
AB - Abstract This research aimed to more clearly describe the interactions of Drosophila suzukii (Matsumura; Diptera: Drosophilidae) with microorganisms that may contribute to spoilage or quality loss of wine grapes during harvest. Experiments were conducted in controlled laboratory experiments and under field conditions to determine these effects. Laboratory trials determined the role of insect contact and oviposition to vector spoilage bacteria onto wine grapes. In the field, the roles of key organoleptic parameters in grape fruit ripening were assessed to determine their relative contribution to oviposition potential as fruit ripened. Finally, field trials determined the relationships of egg and larval infestation to sour rot levels. Non-ovipositional trials indicated elevated levels of microbiota when D. suzukii was present. D. suzukii oviposition exponentially increased the concentration of acetic acid bacteria. Both incised and sound berries showed a significant increase in concentrations of acetic acid bacteria exposed to D. suzukii. Volatile acidity was higher in treatments infested with D. suzukii. Fruit with only eggs did not develop a significant increase of volatile acidity. Larva-infested grape berries in 9.5% of samples developed higher volatile acidity after 14 d. Sound grape berries were less susceptible to the development of microbiota associated with sour rot and spoilage. D. suzukii oviposition and larval development increase risk of spoilage bacteria vectored by D. suzukii adults. Acetic acid bacteria induced fermentation and produced several volatile compounds contributing to spoilage. Spoilage bacteria may create a positive feedback loop that attracts both D. suzukii and other drosophilids, which may contribute to additional spoilage. Spotted Wing Drosophila, invasive species, susceptibility, grapevine, spoilage microorganisms Drosophila suzukii (Matsumura; Diptera: Drosophilidae), also known as Spotted Wing Drosophila, is a threat for fruit crop production systems worldwide (Cini et al. 2012). The serrated ovipositor of female D. suzukii allows oviposition in suitable ripening intact fruits, conferring the ability to exploit otherwise undamaged fruits that are inaccessible to the larvae of other Drosophila species (Atallah et al. 2015). Small fruits and cherries are the most susceptible crops to D. suzukii infestation. The economic impacts of this pest on the susceptible crops are staggering and sustainable management is challenging (Asplen et al. 2015, Del Fava et al. 2017). There is currently no clear understanding of the poor host performance observed in wine grapes. D. suzukii was considered a commercial pest of grapes [Vitis vinifera (L.; Vitales: Vitaceae)] (Kanzawa 1935, 1939), but its actual economic impact on this crop was not fully described in early literature. Laboratory studies and field observations on grape susceptibility to D. suzukii oviposition report that very few eggs are laid on intact grape berries compared to small fruits and cherries (Bellamy et al. 2013, Ioriatti et al. 2015). Moreover, significant differences in susceptibility to oviposition are reported among grape varieties (Linder et al. 2014, Ioriatti et al. 2015, Andreazza et al. 2016) and physiological conditions (Ioriatti et al. 2015, Pelton et al. 2017). The combined properties of susceptible fruit skins (e.g., thickness or texture) allow successful insertion of D. suzukii eggs (Atallah et al. 2015, Karageorgi et al. 2017). When multiple characteristics were evaluated simultaneously, oviposition consistently increased as penetration force decreased, even though other physiological changes occurred during harvest. These additional factors included increased sugar content and decreased acidity levels (Burrack et al. 2013, Ioriatti et al. 2015, Lee et al. 2016). Wine grapes are typically a less suitable host for D. suzukii than are other thin-skinned fruits, as demonstrated by the fact that larvae in wine grapes develop at a slower rate and survive at lower levels compared to small fruits (Maiguashca et al. 2010, Lee et al. 2011, Linder et al. 2014, Ioriatti et al. 2015). Wine grape ranked last among seven types of fruits in a study assessing host potential index (Bellamy et al. 2013). Despite poor overall host suitability, a direct effect of grape berry infestation by D. suzukii is caused by puncturing of the fruit skin due to oviposition (Ioriatti et al. 2015, Pelton et al. 2017), likely providing a gateway for secondary infections with bacterial and fungal pathogens (Atallah et al. 2015). Previous studies have demonstrated that certain bacterial communities are closely associated with D. suzukii adults from cherry. Such species include a high frequency of Tatumella, Gluconobacter, and Acetobacter. These microorganisms are believed to improve the host fitness and the metabolic potential for D. suzukii on nutrient-scarce fruit (Chandler et al. 2014, Jaramillo et al. 2015). It is well known that the successful development of D. suzukii is closely linked to the presence of yeasts. Such yeasts are therefore most probably dispersed and cultivated by D. suzukii to improve life table parameters (Hamby et al. 2012, Hamby and Becher 2016). On grape, mixed microbial populations of yeast and acetic acid bacteria (Gluconobacter and Acetobacter spp.) (Barata et al. 2012a, 2012b) are responsible for fermenting processes involved in sour rot disease. These microorganisms principally produce acetic acid with the development of pungent flavors that can negatively affect grape quality. In addition, bacterial contamination associated with sour rot can modify the chemical composition of wine grapes, reducing their enological potential and favoring the microbial spoilage of grape must and wine that are produced from them (Gravot et al. 2001). At higher frequency, sour rot affects late-ripening, soft-berry, and tightly clustered cultivars (Ioriatti et al. 2015). Drosophilids, including D. melanogaster, are recognized as vectors of the microbial community responsible for these fermenting processes (Fermaud et al. 2002). D. suzukii can feed on cracked or damaged wine grapes during the harvest period (Saguez et al. 2013, Van Timmeren and Isaacs 2013), and such compromised berry skins can create a positive feedback loop resulting in increased D. suzukii feeding and egg-laying compared to fully intact berries, a phenomenon that holds true even for less susceptible cultivars (Ioriatti et al. 2015). Increased berry contact due to D. suzukii feeding and oviposition may therefore elevate the probability of vectoring spoilage bacteria such as Acetobacter spp. as demonstrated for D. melanogaster (Barata et al. 2012b). Considering the importance of wine grape cultivation in some European countries and regions of the United States where D. suzukii is found, the goal of this study was to determine if the presence of this insect on wine grape is an additional risk factor that contributes to sour rot development. Materials and Methods The study was conducted using controlled exclusion and inclusion techniques under both laboratory and field conditions. The field experimental design was set up in such a way to determine the role of D. suzukii as a possible sour rot vector organism. Preparation of Insects and Microorganisms A colony of D. suzukii was established using adults collected from multiple locations in Trentino, Italy. Collected adults were released into plastic cages, their offspring reared on a yeast-based standard artificial medium diet for drosophilids (Dalton et al. 2011), and maintained at the Fondazione Edmund Mach (FEM, San Michele all’Adige, Trentino, Italy, 200 m a.s.l., 46° 12′ N, 11° 8′ E). Wild flies were periodically introduced into the colony in order to minimize genetic drift. Reference cultures of acetic acid bacteria Gluconobacter oxydans ATCC 621 and Acetobacter aceti NRRL B-999 were cultured on GYC medium (OIV 2016). Oxoid (Basingstoke, United Kingdom) provided both components of synthetic media. After inoculation with the above microorganisms, GYC was incubated at 30°C under continuous agitation for 24 h to allow microbial growth up to stationary phase. The cellular concentration of acetic acid bacterial cultures was assayed using standard plate counts (OIV 2016). After growth, microbial cultures were concentrated using a centrifuge (5804 Centrifuge, Eppendorf, Hamburg, Germany) at 4,000 rpm for 10 min. The residues of GYC media were eliminated and cells were suspended in a peptone-based water buffer (mycological peptone, Oxoid, 1 g/liter) to reach a nominal concentration of 107 CFU/ml, and stored at 4°C until use (maximum 6 h). For the purposes of this study, the spoilage bacteria included a 1:1 suspension of G. oxydans ATCC 621, and A. aceti NRRL B-999. D. suzukii adults within rearing cages were infected with cultures of the above microorganisms by placing a cotton pad (15 × 10 × 0.5 cm, cotton wool, AF000641-2014, Luigi Salvadori S.p.A. Scandicci, FI, Italy) impregnated with 15 ml of suspension of the bacterial culture in the rearing cage for 48 h. The bacteria-infected flies were subsequently used in the experiments. Wild-collected individuals were caught in traps (Droso-trap, Biobest, Westerlo, Belgium) baited with a cotton ball soaked in Droskidrink (Prantil, Vervò di Priò, Trento, Italy) and installed on the bottom of the trap (Rossi-Stacconi et al. 2016). The interior of the trap was divided into two sectors by a frame of fine mesh tulle attached to the trap wall above the cotton ball; it allowed volatile release while avoiding contact between the captured flies and the cotton ball. Traps were installed in a vineyard at FEM, where rows of grapevines (cv. Schiava) were planted 3 m apart with a plant-to-plant distance of 1 m. Microbiological, Chemical and Texture Analysis Microbiological analyses of wine grape and D. suzukii were performed according to the OIV methods (OIV 2016) for the quantification of Saccharomyces and non-Saccharomyces yeasts, G. oxydans, and A. aceti. For this purpose, 1 g of D. suzukii (approximately 35–40 insects) was put in a sterile 50 ml Falcon test tube (Sarsted, Nümbrecht, Germany) containing 9 ml of sterile peptone water (1 g/liter of Mycological Peptone, Oxoid). The sample was homogenized for 60 s by an Ultra-turrax T18 disperser (IKA, Staufen im Breisgau, Germany), and further analyzed. Wine grape samples (25 g for each sample) were homogenized by a Stomacher blender (Seward, Worthing, United Kingdom) with the addition of 1:10 w/w of peptone water. Samples were diluted to the appropriate decimal dilution in 9 ml of peptone water, and then spread onto Petri plates containing the following synthetic media: WL agar (Oxoid) for the enumeration of yeast; Lysine agar (Oxoid) for non-Saccharomyces yeast; Act/s agar: Glucose (Sigma Aldrich, St. Louis, MO), 10 g/liter, yeast extract (Oxoid), 5 g/liter, hydrolyzed Casein (Oxoid), 5 g/liter, Cycloheximide 1% aqueous solution (Oxoid), 10 ml/liter, bacteriological agar (Oxoid), 15 g/liter for the quantification of acetic acid bacteria; MRS Agar (Oxoid) added with 15% apple juice (Santal, Parma, Italy) for the quantification of lactic acid bacteria. All media were incubated at 25°C for a time between 4 (Yeast) and 10 d (Acetic and Lactic acid bacteria) (OIV 2016). Lactic acid bacteria were incubated under anaerobic conditions (Anaerogen KIT, Oxoid). Plate count data were expressed according to the (ISO 2013). Chemical analyses of grape samples were performed using a WineScan (FOSS, Hillerød, Denmark) FT-IR apparatus to determine the levels of acetic acid (AAc), relative density (D), sugar content (Sc), total acidity (TotAc), malic acid (MAc), tartaric acid (TAc), pH, and potassium (K). Grape berries were examined under a stereo-microscope to check for their integrity before their use in the bioassays. The firmness levels of intact berries were evaluated using the Universal Testing Machine (TAxT2i Texture Analyzer, Stable Micro System, Godalming, Surrey, United Kingdom) by measuring the penetration force (cN) exerted by a 2-mm blunted needle probe to the point of penetration (Ioriatti et al. 2015). D. suzukii Non-Ovipositional Activity and its Relationship to Sour Rot The tests were performed in a plastic cage (30 × 30 × 30 cm) (BugDorm, Megaview Science Education Co., Taichung, Taiwan) partially divided into two sectors (here called ‘A’ and ‘B’) by placing a plastic wall (30 × 15 × 0.3 cm) in the middle of the cage. Store-bought grape berries of cv. Italia were washed and sterilized by immersion in 96% ethanol (Sigma-Aldrich) for 5 min, rinsed by distilled water and then dried under a fume hood. In order to simulate field damage, a sample of grape berries was incised with a scalpel prior to sterilization. The cv. Italia is a table grape variety known to be resistant to D. suzukii oviposition (Andreazza et al. 2016). This cultivar resisted D. suzukii oviposition during the experimental period and was verified to be free of egg infestation by checking berries under the stereo-microscope at the end of the period of insect exposure. Contact made between D. suzukii and grape berries during the experimental period would therefore be the only cause of contamination rather than oviposition. Sector ‘A’ contained either sound (‘a’, ‘b’, ‘c’) or incised (‘d’, ‘e’) sterile berries (n = 10) whose microbial load was quantified to verify whether D. suzukii was vectoring microorganisms developed on berries placed in sector ‘B’, which acted as a source of contamination. The source of contamination was represented by either sound (treatments ‘a’, ‘d’) or incised berries (treatments ‘b’, ‘c’, ‘e’) (n = 10), which were expected to develop a higher level of microorganisms. Moreover, to stimulate microorganism development, in treatments ‘c’ and ‘e’, incised berries placed in sector ‘B’ were also inoculated with G. oxydans ATCC 621 and A. aceti NRRL B-999 by dipping the incised berries in a suspension of peptone water containing bacteria of 107 CFU/ml at a ratio of 1:1. Treatment ‘a’, having sterilized sound berries on both sides of the cage, was considered as the control. The five treatments (Table 1) were replicated four times. D. suzukii adults (4 males, 6 females) were released in each cage. Each cage was equipped with a 30-ml cup fitted with a water-soaked sponge, allowing adequate hydration of insects. All adults were 1–3 d old and the cages were maintained at 25°C under continuous light. After 7 d, berries in sector A were retrieved and analyzed for the presence of microorganisms, as described previously. Mean data concerning microorganism load (CFU/g) in the five treatments were considered significantly different when separated by one order of magnitude, considering a confidence interval of plate counts of 0.5 log units, estimated as suggested by the (ISO 2013). Table 1. Microbial contamination of tested berries (column A) after 7 d of exposure to potential infection by contact with non-ovipositing D. suzukii flying from the source of contamination (column B) Experiment Acetic acid bacteria Lactic acid bacteria Saccharomyces yeast Non-Saccharomyces yeast Treatment A B CFU/g Tested berries Source of contamination A Sound Sound 5.0 ± 0.0E+01 5.0 ± 0.0E+01 1.8 ± 2.5E+04 2.2 ± 3.0E+04 B Sound Incised 6.0 ± 1.1E+03 1.6 ± 2.3E+02 6.8 ± 7.1E+04 2.6 ± 2.4E+04 C Sound Incised + AAB 6.8 ± 1.2E+03 2.8 ± 2.6E+02 4.7 ± 6.3E+03 5.8 ± 11E+04 D Incised Sound 4.8 ± 4.6E+06 2.0 ± 3.3E+0.6 1.3 ± 1.2 + 06 3.1 ± 5.3E+07 E Incised Incised + AAB 9.2 ± 9.2E+06 2.6 ± 4.6E+0.6 1.1 ± 0.9E+05 2.0 ± 3.2E+07 Experiment Acetic acid bacteria Lactic acid bacteria Saccharomyces yeast Non-Saccharomyces yeast Treatment A B CFU/g Tested berries Source of contamination A Sound Sound 5.0 ± 0.0E+01 5.0 ± 0.0E+01 1.8 ± 2.5E+04 2.2 ± 3.0E+04 B Sound Incised 6.0 ± 1.1E+03 1.6 ± 2.3E+02 6.8 ± 7.1E+04 2.6 ± 2.4E+04 C Sound Incised + AAB 6.8 ± 1.2E+03 2.8 ± 2.6E+02 4.7 ± 6.3E+03 5.8 ± 11E+04 D Incised Sound 4.8 ± 4.6E+06 2.0 ± 3.3E+0.6 1.3 ± 1.2 + 06 3.1 ± 5.3E+07 E Incised Incised + AAB 9.2 ± 9.2E+06 2.6 ± 4.6E+0.6 1.1 ± 0.9E+05 2.0 ± 3.2E+07 Mean data (n = 10) ± SD. AAB: Acetic acid bacteria. Data are considered significant different when separated by > 1 log unit. Open in new tab Table 1. Microbial contamination of tested berries (column A) after 7 d of exposure to potential infection by contact with non-ovipositing D. suzukii flying from the source of contamination (column B) Experiment Acetic acid bacteria Lactic acid bacteria Saccharomyces yeast Non-Saccharomyces yeast Treatment A B CFU/g Tested berries Source of contamination A Sound Sound 5.0 ± 0.0E+01 5.0 ± 0.0E+01 1.8 ± 2.5E+04 2.2 ± 3.0E+04 B Sound Incised 6.0 ± 1.1E+03 1.6 ± 2.3E+02 6.8 ± 7.1E+04 2.6 ± 2.4E+04 C Sound Incised + AAB 6.8 ± 1.2E+03 2.8 ± 2.6E+02 4.7 ± 6.3E+03 5.8 ± 11E+04 D Incised Sound 4.8 ± 4.6E+06 2.0 ± 3.3E+0.6 1.3 ± 1.2 + 06 3.1 ± 5.3E+07 E Incised Incised + AAB 9.2 ± 9.2E+06 2.6 ± 4.6E+0.6 1.1 ± 0.9E+05 2.0 ± 3.2E+07 Experiment Acetic acid bacteria Lactic acid bacteria Saccharomyces yeast Non-Saccharomyces yeast Treatment A B CFU/g Tested berries Source of contamination A Sound Sound 5.0 ± 0.0E+01 5.0 ± 0.0E+01 1.8 ± 2.5E+04 2.2 ± 3.0E+04 B Sound Incised 6.0 ± 1.1E+03 1.6 ± 2.3E+02 6.8 ± 7.1E+04 2.6 ± 2.4E+04 C Sound Incised + AAB 6.8 ± 1.2E+03 2.8 ± 2.6E+02 4.7 ± 6.3E+03 5.8 ± 11E+04 D Incised Sound 4.8 ± 4.6E+06 2.0 ± 3.3E+0.6 1.3 ± 1.2 + 06 3.1 ± 5.3E+07 E Incised Incised + AAB 9.2 ± 9.2E+06 2.6 ± 4.6E+0.6 1.1 ± 0.9E+05 2.0 ± 3.2E+07 Mean data (n = 10) ± SD. AAB: Acetic acid bacteria. Data are considered significant different when separated by > 1 log unit. Open in new tab D. suzukii Oviposition and its Relationship to Sour Rot To test if oviposition and subsequent larval development of D. suzukii could promote grape sour rot in fully intact, healthy and ripe grape berries of cvs. Pinot noir (late harvested, 2014), Schiava, and Cabernet sauvignon (2015) were sterilized in a 70% ethanol (Sigma-Aldrich) aqueous solution by dipping for 15 min, then rinsing with distilled water (100 ml for each sample of 5 berries), and subsequently allowed to dry for 12 h at 5°C before use. Fruit ripeness of the three varieties was assessed by measuring the penetration force on an additional sample of berries. We trialled two berry statuses, i.e., intact and incised, for each cultivar. Each treatment replicate consisted of five berries. Half of the treatments were not exposed to D. suzukii during the experimental period while in the remaining treatments four adult male and six female D. suzukii were released into the plastic cages. The insects were infected with bacteria prior to the test, using the method described above. Both cultivar (n = 3) and berry status (n = 2, i.e., intact and incised) were either positively or negatively subjected to D. suzukii infestation (12 treatments total) and were placed into a sterile 120 ml volume plastic bottle (Doctorpoint S.r.l. Borgomanero, NO, Italy). All treatments were replicated 10 times. Plastic boxes were equipped with a 2.5-ml plastic tube (Sarsted AG & Co., Nümbrecht, Germany) filled with cotton impregnated with an aqueous solution of 10% glucose (Sigma-Aldrich), and kept in a climatic chamber (23°C, 75% RH, 16:8 (L:D) h photoperiod). After 4 d, D. suzukii adults were removed from the plastic boxes and adult survival and oviposition levels on both intact and incised berries were assessed. Berries were maintained in the plastic boxes in the climatic chamber for 12 d, allowing eclosion of D. suzukii eggs as well as sour rot development. Grapes were submitted to microbiological analysis to quantify the bacterial load and chemical analysis to determine volatile acidity at the end of the experimental period. Values concerning oviposition and bacterial load in the three grape varieties were submitted to full factorial 2-way ANOVA to assess possible differences between berry status (i.e., intact vs incised; primary treatment) and grapevine varieties (Pinot noir, Schiava and Cabernet sauvignon; secondary treatment), followed by a post hoc Tukey’s multiple comparison test. To satisfy the conditions of normality, bacterial load data were log-transformed. Volatile acidity values of the assembled grape samples of each variety were submitted to the unilateral Wilcoxon signed-rank test to assess significant differences between berry status (sound vs incised) and D. suzukii infestation status (with and without flies). Artificial Infestation in Vineyard Oviposition Behavior of the Released D. suzukii Flies In 2016, three cv. Schiava blocks located in the same climatic area (Rotaliana plain, Trentino, Italy) and within a radius of 10 km were selected to investigate the contribution of D. suzukii to the development of sour rot under field scenarios. The vineyard blocks were regularly monitored for D. suzukii flight activity, oviposition and the fruit ripeness parameters. Forty-five healthy grape clusters (3 clusters/vine) were selected and enclosed in organza bags on 10 August in each of the three vineyards. On 7, 17, and 27 September (commercial harvest), 15 grape clusters/vineyard were artificially infested with four male and six female bacteria-infected D. suzukii adults. After 4 d, the flies were removed and grape clusters were collected and brought to the laboratory. All berries of each cluster were checked under the dissecting microscope and categorized in berries with (g+) and without (g−) eggs. The berries of the two groups were submitted to chemical analyses to quantify seven chemical parameters including MAc, TAc, TotAc, pH, D, Sc, and K. The fruit ripening parameters change during the ripening process. In order to determine the role of these changing parameters in relation to D. suzukii oviposition, a principal component analysis (PCA) was performed in order to determine the relative contributing value of each of these seven organoleptic parameters. A Wilcoxon signed-rank test was conducted between factor coordinates of cases for each of the three selected periods to assess significantly different positions of the centroids given by g+ and g−, respectively. Similarly, a Wilcoxon signed-rank test was conducted to assess significant differences in the contents of each organoleptic parameter between g+ and g−. Development of Sour Rot in Vineyards In each vineyard block, 30 grape bunches (3/vine) were selected and bagged with organza on 10 August 2016, before any D. suzukii flight activity was recorded. Two weeks before the expected harvest (7 September = T0) all the bagged grape clusters were infested with four male and six female bacteria-infected D. suzukii for 4 d. In each block, 15 bagged grape clusters were collected 7 d after infestation (T7). All berries of each cluster were checked under a dissecting microscope and divided into three classes: berries without any eggs (sound), berries with one or more eggs, and berries with developing larvae. The groups of berries of each class were weighed and chemical analyses were performed by WineScan (FOSS) FT-IR apparatus to quantify volatile acidity. A grape cluster with a detectable level of volatile acidity is considered affected by sour rot. Berries damaged during manipulation were excluded from the analyses. The same procedure was applied to the remaining 15 bagged clusters 14 d after infestation (T14) corresponding to the harvest date of the grape variety. The vineyard blocks are representative of the differences of cultural practices applied in the area, which cause different grape cluster weight. Grape cluster and infested berry weights in the three vineyards were compared by non-parametric ANOVA (Kruskal-Wallis test). Results and Discussion Insects Artificial contamination of the laboratory colony allowed generation of a population whose microorganism load was comparable to that of the wild population sampled in the vineyard. The concentration of acetic acid bacteria on the laboratory-infected flies was 2.2 × 107 CFU/g compared to the 7.5 × 106 CFU/g measured on the wild D. suzukii specimens. This difference appeared inconsequential considering the large variability of plate count method, typically on the order of 0.2–0.5 log units (ISO 2013). D. suzukii Non-Ovipositional Activity and its Relationship to Sour Rot Results of the microflora analysis showed the presence of acetic acid bacteria at very low levels (1 log unit) in treatment a) (control) where sterilized sound berries were installed in both sides of the cage (Table 1). Significantly (OIV 2016) higher levels of acetic acid bacteria (3 log units) were detected on sound berries when contamination was introduced into the cage by incised berries (6.0 × 103 and 6.8 × 103 CFU/g in treatments b and c, respectively). The highest bacterial load (6 log units) was detected on incised berries (4.8 × 106 and 9.2 × 106 CFU/g in treatments d and e, respectively) regardless of the status of the source of contamination (sound or incised). Inoculation of incised berries (treatments c and e) did not significantly increase the effect of the contaminants. The results concerning lactic acid bacteria were similar; an increase in bacterial load was found on the tested sound berries (1.6 × 102 and 2.8 × 102 CFU/g in treatments b and c, respectively) when the source of contamination included incised berries, compared to the control, but still far below the lactic acid bacterial load found on incised berries (2.0 × 106 and 2.6 × 106/g, respectively in treatments ‘d’ and ‘e’). On the contrary, when Saccharomyces and non-Saccharomyces yeast contamination was considered, we did not find any significant increase on tested sound berries compared to the control, while significantly higher loads of yeast were detected on tested incised berries (Table 1). Ability of D. suzukii to Vector Sour Rot Bacteria to Fully Intact Sound Berries by Oviposition The ripeness of the grapes used in the bioassays was assessed by measuring the penetration force of fruits originating from the study vineyard blocks. The mean penetration forces were 38.6 ± 14.0, 56.6 ± 12.3, and 61.9 ± 15.8 (cN; mean ± SD) in Schiava, Pinot noir, and Cabernet sauvignon varieties, respectively. A significantly higher number of eggs (ANOVA: F2,54 = 110.2, P < 0.001, Table 2) were laid on incised berries (average 7.7 eggs/berry), compared to the sound ones (average 1.9 eggs/berry). There were differences in oviposition levels between cultivars (ANOVA: F1,54 = 58.9, P < 0.001). The interaction between treatments (berry status × variety) was also significant (ANOVA: F1,24 = 19.5, P < 0.001); the number of eggs/berry was significantly higher in both Pinot noir and Cabernet sauvignon when berries were incised, while no significant differences were found in Schiava between incised and sound grape berries (Table 2). Table 2. Oviposition and microbial contamination of different vine grapes after contact with D. suzukii Wine grape cv. Year Oviposition (number of eggs) Acetic acid bacteria (CFU/g) Incised Sound Incised Sound Cabernet sauvignon 2015 14.3 ± 2.4d 3.7 ± 1.7bc 7.9 ± 1.6E+06b 0.6 ± 0.2E+06a Pinot noir 2014 5.5 ± 3.3c 1.2 ± 1.4ab 3.4 ± 0.6E+08c 9.0 ± 1.9E+06b Schiava 2015 3.3 ± 2.2abc 0.8 ± 0.9a 1.5 ± 0.4E+06b 1.4 ± 0.3E+07b mean ± SD 7.7 ± 5.5 1.9 ± 1.9 1.2 ± 0.5E+07 7.9 ± 1.9E+06 Wine grape cv. Year Oviposition (number of eggs) Acetic acid bacteria (CFU/g) Incised Sound Incised Sound Cabernet sauvignon 2015 14.3 ± 2.4d 3.7 ± 1.7bc 7.9 ± 1.6E+06b 0.6 ± 0.2E+06a Pinot noir 2014 5.5 ± 3.3c 1.2 ± 1.4ab 3.4 ± 0.6E+08c 9.0 ± 1.9E+06b Schiava 2015 3.3 ± 2.2abc 0.8 ± 0.9a 1.5 ± 0.4E+06b 1.4 ± 0.3E+07b mean ± SD 7.7 ± 5.5 1.9 ± 1.9 1.2 ± 0.5E+07 7.9 ± 1.9E+06 Data are presented as mean number of eggs per berry ± SD (N = 50 berries). Different letters indicate significant differences among treatments after two-way ANOVA followed by Tukey’s HSD post hoc test (P < 0.05). Pinot noir grape berries were harvested on 17 October, 3 wk after commercial ripening in 2014, Schiava and Cabernet sauvignon grape berries were harvested at their respective commercial ripening dates on 25 September and 9 October 2015. Open in new tab Table 2. Oviposition and microbial contamination of different vine grapes after contact with D. suzukii Wine grape cv. Year Oviposition (number of eggs) Acetic acid bacteria (CFU/g) Incised Sound Incised Sound Cabernet sauvignon 2015 14.3 ± 2.4d 3.7 ± 1.7bc 7.9 ± 1.6E+06b 0.6 ± 0.2E+06a Pinot noir 2014 5.5 ± 3.3c 1.2 ± 1.4ab 3.4 ± 0.6E+08c 9.0 ± 1.9E+06b Schiava 2015 3.3 ± 2.2abc 0.8 ± 0.9a 1.5 ± 0.4E+06b 1.4 ± 0.3E+07b mean ± SD 7.7 ± 5.5 1.9 ± 1.9 1.2 ± 0.5E+07 7.9 ± 1.9E+06 Wine grape cv. Year Oviposition (number of eggs) Acetic acid bacteria (CFU/g) Incised Sound Incised Sound Cabernet sauvignon 2015 14.3 ± 2.4d 3.7 ± 1.7bc 7.9 ± 1.6E+06b 0.6 ± 0.2E+06a Pinot noir 2014 5.5 ± 3.3c 1.2 ± 1.4ab 3.4 ± 0.6E+08c 9.0 ± 1.9E+06b Schiava 2015 3.3 ± 2.2abc 0.8 ± 0.9a 1.5 ± 0.4E+06b 1.4 ± 0.3E+07b mean ± SD 7.7 ± 5.5 1.9 ± 1.9 1.2 ± 0.5E+07 7.9 ± 1.9E+06 Data are presented as mean number of eggs per berry ± SD (N = 50 berries). Different letters indicate significant differences among treatments after two-way ANOVA followed by Tukey’s HSD post hoc test (P < 0.05). Pinot noir grape berries were harvested on 17 October, 3 wk after commercial ripening in 2014, Schiava and Cabernet sauvignon grape berries were harvested at their respective commercial ripening dates on 25 September and 9 October 2015. Open in new tab When berries were analyzed for quantification of microbial contamination, acetic acid bacteria were not detected in the treatments where flies were not released, in neither sound nor incised berries (data not shown). On the contrary, exposure of both types of berries to D. suzukii exponentially increased the concentration of acetic acid bacteria; sound grapes showed significantly lower levels of acetic acid bacteria (M ± SD = 7.9 ± 1.9 × 106) compared to incised berries (1.2 ± 0.5 × 107) (ANOVA: F2,24 = 25.5, P < 0.001). Similarly, the varieties showed significant differences in terms of acetic acid bacterial load (ANOVA: F2,24 = 79.7, P < 0.001) (Table 2). The interaction between treatments (berry status × variety) was also significant (ANOVA: F1,24 = 97.4, P < 0.001) in that both Pinot noir and Cabernet sauvignon developed a significant increase in the number of bacterial CFU when incised, while no significant differences were found in Schiava between incised and sound grape berries (Table 2). As expected, the volatile acidity value was significantly higher in the incised treatments than in the sound berries (3.55 g/liter vs 1.03 g/liter; T = 14, P = 0.040). Volatile acidity was also significantly higher in the treatments infested with D. suzukii compared to those where flies were excluded (3.61 g/liter vs 0.36 g/liter; T = 21, P = 0.014) (Fig. 1). Fig. 1. Open in new tabDownload slide Acetic acid content of grapes at the end of a laboratory test performed on three grape varieties. The value of volatile acidity (g/liter) are given for of the whole mass of grapes used in each treatment and significance of the differences between incised vs sound and between infested vs non-infested treatments was tested (*P < 0.05). Fig. 1. Open in new tabDownload slide Acetic acid content of grapes at the end of a laboratory test performed on three grape varieties. The value of volatile acidity (g/liter) are given for of the whole mass of grapes used in each treatment and significance of the differences between incised vs sound and between infested vs non-infested treatments was tested (*P < 0.05). In more detail, when contact between D. suzukii and grape berries was absent, the acetic acid level remained negligible in sound berries (0.15; 0.18; 0.22 g/liter in Pinot noir, Schiava, and Cabernet sauvignon, respectively) and it increased above the legal limits of dry wine in Europe (Reg. EC n. 606, 2009) only in Pinot noir when the berries were incised. On the contrary, when the berries were incised, the presence of D. suzukii always resulted in a significant increase of acetic acid that reached far above the legal limits of 1 g/liter (9.48, 4.00, 4.00 g/liter in Pinot noir, Schiava, and Cabernet sauvignon, respectively), but it was particularly high, as well, when D. suzukii was in contact with sound berries: Pinot noir developed an acetic acid content up to 2.84 g/liter, while Cabernet sauvignon and Schiava showed diminished, but still commercially unacceptable, accumulation of 0.78 and 0.60 g/liter respectively (Fig. 1). Artificial Infestation in Vineyard Oviposition Behavior of the Released D. suzukii flies Flight activity and oviposition did not differ significantly in the three selected vineyards, nor was the progression of grape skin softening during ripening significantly different, as monitored by penetration force measurements (data not shown). The relationship between the selected chemical parameters and D. suzukii oviposition behavior was studied only on a limited number of initially bagged grape clusters: 13, 20, and 19 clusters on the 7, 17, and 27 September, respectively. In fact, a sufficient amount of grape juice could be produced only from clusters bearing a significant number of berries with D. suzukii eggs to perform the quantification of the seven chemical parameters. The first two components of the PCA, PC1 and PC2, accounted for 55.5% and 28.1% of variance, respectively (Fig. 2, Table 3). The PCA biplot (PC1xPC2) showed that pH, D, Sc, and K were more highly correlated with PC1, while TotAc, MAc, and TAc were more highly correlated with PC2. The analysis of factor coordinate cases showed that the centroid g+ has significantly higher values than the centroid g− in both PC1 (Wilcoxon signed-rank test: W = 2, P < 0.01) and PC2 (W = 17, P < 0.05) on 7 September (n = 13), only in PC1 (W = 41, P < 0.05; PC2: W = 87, P > 0.05) on 17 September (n = 20) and in neither of them (PC1: W = 87; P > 0.05; PC2: W = 85; P > 0.05) on 27 September (n = 19). Taking each sampling period separately, on 7 September (n = 13) we measured a significant difference in all organoleptic parameters (except for TAc, W = 21, P > 0.05), where g+ showed significantly higher Sc (W = 2, P < 0.01), pH (W = 5, P < 0.01), D (W = 1, P < 0.01), and K (W = 7, P < 0.01) and lower TotAc (W = 1, P < 0.01) and MAc (W = 3, P < 0.01), compared to g−. Grapes sampled on 17 September (n = 20) showed a significant difference only in terms of Sc (W = 10.5, P < 0.01) and D (W = 22, P < 0.01), which was higher in g+, while MAc was significantly higher (W = 24, P < 0.01) in g−. Finally, grapes collected on 27 September (n = 19) differed between g+ and g− for TotAc and TAc values, which were higher in g− (W = 25, P < 0.01 and W = 37.5, P < 0.05, respectively), and pH, which was higher in g+ (W = 29, P < 0.05; Fig. 3). Fig. 2. Open in new tabDownload slide PCA based on seven chemical parameters: malic acid (MAc), tartaric acid (TAc), total acidity (TotAc), pH, density (D), sugar content (Sc), and potassium content (K). In the center is the PCA biplot (PC1xPC2) with the factor coordinates based on correlations between the studied variables and factor axes. The four frames represent the plots separated per different period of sampling (circles, first period; squares, second period; triangles third period) and the sum of the plots for the whole period. Filled and empty shapes represent plots with (+) and without (−) D. suzukii infestation, respectively. The centroids (g+ and g−) refer to the mean coordinates of all filled and empty shapes per period. Only in the first period is the distance significantly different between g+ and g− centroids in both PC1 (t-test: P < 0.001) and PC2 (P = 0.002), while in the second period the distance is significantly different only in PC1 (P = 0.006; PC2 = 0.15), and in the third period there was no significant difference (PC1: P = 0.86; PC2: P = 0.72). Fig. 2. Open in new tabDownload slide PCA based on seven chemical parameters: malic acid (MAc), tartaric acid (TAc), total acidity (TotAc), pH, density (D), sugar content (Sc), and potassium content (K). In the center is the PCA biplot (PC1xPC2) with the factor coordinates based on correlations between the studied variables and factor axes. The four frames represent the plots separated per different period of sampling (circles, first period; squares, second period; triangles third period) and the sum of the plots for the whole period. Filled and empty shapes represent plots with (+) and without (−) D. suzukii infestation, respectively. The centroids (g+ and g−) refer to the mean coordinates of all filled and empty shapes per period. Only in the first period is the distance significantly different between g+ and g− centroids in both PC1 (t-test: P < 0.001) and PC2 (P = 0.002), while in the second period the distance is significantly different only in PC1 (P = 0.006; PC2 = 0.15), and in the third period there was no significant difference (PC1: P = 0.86; PC2: P = 0.72). Table 3. PCA factor coordinates based on correlations between the studied variables and factor axes Variable Factor coordinates of the variables, based on correlations Factor 1 Factor 2 Brix 0.934924 −0.226844 pH 0.866383 0.134927 Total acidity −0.693177 −0.694274 Density 0.935014 −0.233322 Tartaric acid 0.366516 −0.815621 Malic acid −0.724196 −0.530403 K 0.870626 −0.290700 Variable Factor coordinates of the variables, based on correlations Factor 1 Factor 2 Brix 0.934924 −0.226844 pH 0.866383 0.134927 Total acidity −0.693177 −0.694274 Density 0.935014 −0.233322 Tartaric acid 0.366516 −0.815621 Malic acid −0.724196 −0.530403 K 0.870626 −0.290700 Open in new tab Table 3. PCA factor coordinates based on correlations between the studied variables and factor axes Variable Factor coordinates of the variables, based on correlations Factor 1 Factor 2 Brix 0.934924 −0.226844 pH 0.866383 0.134927 Total acidity −0.693177 −0.694274 Density 0.935014 −0.233322 Tartaric acid 0.366516 −0.815621 Malic acid −0.724196 −0.530403 K 0.870626 −0.290700 Variable Factor coordinates of the variables, based on correlations Factor 1 Factor 2 Brix 0.934924 −0.226844 pH 0.866383 0.134927 Total acidity −0.693177 −0.694274 Density 0.935014 −0.233322 Tartaric acid 0.366516 −0.815621 Malic acid −0.724196 −0.530403 K 0.870626 −0.290700 Open in new tab Fig. 3. Open in new tabDownload slide Box plots of the seven studied berry chemical parameters divided between egg-infested (g+) and egg-free (g−) berries and grouped by first, second, and third sampling periods. Asterisks (*P < 0.05; **P < 0.01) indicate significant differences between datasets of the same sampling period after bilateral Wilcoxon signed-rank test for paired data. Fig. 3. Open in new tabDownload slide Box plots of the seven studied berry chemical parameters divided between egg-infested (g+) and egg-free (g−) berries and grouped by first, second, and third sampling periods. Asterisks (*P < 0.05; **P < 0.01) indicate significant differences between datasets of the same sampling period after bilateral Wilcoxon signed-rank test for paired data. Development of Sour Rot in Vineyards The average weight of the grape clusters was 237.1 g ± 118.1 but with significant differences between the three vineyards (Kruskal-Wallis test: H = 45.3, P < 0.001). Grape clusters were significantly lighter in Facchinelli vineyard (199.3 g ± 51.1) compared to Pedot (351.6 g ± 99.0) and Pergher vineyards (392.8 g ± 105.8). At the time of the release of D. suzukii flies, on 7 September, the average berry penetration force value was 33.7 cN, considerably below the susceptibility threshold of 40 cN (Ioriatti et al. 2015). At T7, 1 wk after the artificial infestation, one group of bagged clusters of the Facchinelli vineyard was lost, while at T14, 2 wk after artificial infestation, two bagged clusters in the Facchinelli vineyard and one bagged cluster in the Pergher vineyard were missing. All recovered clusters, both at T7 and T14, were infested with eggs and larvae. The weight of infested berries was significantly different between Facchinelli (64.8 g ± 25.7) and Pergher (103.4 g ± 55.3) vineyards, while neither of them differed from Pedot vineyard (71.7 g ± 43.6) (Kruskal-Wallis test: H = 10.2, P = 0.006). Irrespective of the vineyard or the weight of the grape clusters, no grape berry samples infested with eggs alone developed a significant increase of volatile acidity at either T7 or at T14. Concerning larval infestation, grape berry samples infested with larvae during T7 did not develop a detectable amount of volatile acidity; four samples (1 Facchinelli, 1 Pergher, 2 Pedot) out of 42 (9.5%) developed a volatile acidity greater than 0.1 g/liter, up to 1.2 g/liter by 14 d after infestation with D. suzukii. Discussion The objective of this study was to determine whether D. suzukii is capable of vectoring a microbial community responsible for fermenting processes, as recognized for D. melanogaster and other drosophilids (Fermaud et al. 2002). In fact, despite the observation that eggs were not laid in the grape berries in our first laboratory bioassay, spoilage bacteria contamination of the tested sound berries nonetheless occurred. The data suggest that this contamination was exclusively due to insect contact as opposed to ovipositional penetration of the berry skins. Sound berries are characterized by a native yeast microbiota on the order of 3–4 log units (Barata et al. 2012a, Liu et al. 2017). The controlled conditions employed in these trials suggest that the elevated yeast loads of sterilized sound berries were not due to D. suzukii-mediated vectoring. Physical damage to berries triggers the release of several compounds including sugars from inside berries. These sugars in turn stimulate an increase of yeast populations by up to 7 log units as soon as they come into contact with these resources (Barata et al. 2012b). It is therefore clear that incised berries act as a source of bacteria that can easily be vectored by D. suzukii as the insects move from one point to another during feeding and oviposition. Our data illustrate that bacterial loads in incised berries easily reached levels as high as 6 log units, which confirms that berry injury is the primary cause of sour rot development. Yeast populations of ruptured berry skins resulted in rapid growth of yeasts as is also demonstrated in earlier studies (Bisiach et al. 1986; Barata et al. 2012c) under our experimental conditions. The composition of spoilage yeasts was mainly represented by non-Saccharomyces species, known to be well adapted to the vineyard environment (Jolly et al. 2006, Liu et al. 2017, Renouf et al. 2007). The presence of a large population of yeast inside berries certainly favored the proliferation of acetic acid bacteria. Yeasts ferment sugars and release small amounts of ethanol, one of the most preferred carbon sources for Acetobacter spp. metabolism (Fermaud et al. 2002). When grape berries susceptible to oviposition were used, D. suzukii laid eggs in higher numbers on damaged, compared to sound, berries. Moreover, in both incised vs sound berries, the concentration of acetic acid bacteria increased significantly compared to the treatments where the flies were excluded. Pinot noir and Cabernet sauvignon both showed significantly greater oviposition and concentration of acetic acid bacteria, while Schiava did not show significant differences in the number of eggs/berry. Schiava berry acetic acid bacteria also did not differ. The description of these differences was not within the scope of this study. However, the differences were probably attributable to seasonal fluctuations and time of the year when the bioassays were carried out, due to the need to use fully mature grape berries of cultivars with different ripening periods. However, only the availability of fermentable substrates, typical of damaged berries, stimulated proliferation of acetic acid bacteria due to two different phenomena: the direct utilization of sugars, especially by the bacteria belonging to the genus Gluconobacter; and the oxidation of ethanol previously produced by the fermentation of yeast on account of Acetobacter bacteria (Bartowsky and Henschke 2008, Gullo et al. 2009). Even though on sound berries the legal limit of 1 g/liter was reached only in 2014 on Pinot noir, the increase in acetic acid content was consistent across the three cultivars when exposed to D. suzukii oviposition. As expected, the acetic acid content rapidly increased when the damaged berries were exposed to flies. These data are more relevant when considering that the accumulation of acetic acid is observed prior to enological fermentation that usually contributes 0.4–0.6 g/liter of acetic acid from grape must to wine. The introduction of D. suzukii causes a significant alteration of grape berries, making them unsuitable for the production of wine. In order to investigate the association of D. suzukii with grape sour rot disease in the vineyard, we studied the oviposition behavior of the fly in relation to the physical and chemical changes occurring during grape berry ripening. As grape berries develop, they display changes in size, composition, color, texture, flavor, and in pathogen and pest susceptibility (Conde et al. 2007). D. suzukii is able to infest fully intact grape berries in the vineyard (Kanzawa 1935, 1939; Sinn 2012; Saguez et al. 2013; Van Timmeren and Isaacs 2013; Linder et al. 2014). Grape susceptibility to D. suzukii oviposition increases during the grape ripening period in the more susceptible varieties (Ioriatti et al. 2015). Selection of appropriate oviposition sites is essential for progeny survival and fitness in all insect species, including D. suzukii. Yet, little is known about the mechanisms regulating how the conditions of the oviposition substrate are perceived by D. suzukii to determine the final choice for egg deposition (Crava et al. 2016; Ramasamy et al. 2016; Karageorgi et al. 2017). Berry firmness is a critical component in host selection and is negatively correlated to oviposition, but concomitant increases in oviposition have been demonstrated with an increase of sugar content and a decrease in acidity (Ioriatti et al. 2015). In the present study, 10 and 20 d before harvest, artificial infestations of bagged grape bunches demonstrated that D. suzukii preferred to lay eggs in those berries with a significantly higher sugar and relative density level and lower malic acidity. Malic acid varies greatly as berries develop and mature. This compound accumulates and reaches a maximum value just prior to véraison. Malic acid begins to break down at the onset of ripening due to the induction of malate oxidation, suggesting that malic acid is transformed to fructose and glucose (Conde et al. 2007). On the other hand, it is reported that during berry ripening, the berry cuticle of visually intact berries may contain microfissures (Barata et al. 2012a) through which sugar can reach the berry skin surface. Drosophila females are known to selectively choose egg-laying sites (Yang et al. 2008). Oviposition site selection could be driven by inputs arising from labella and tarsi, where gustatory sensilla likely can select preferred sucrose-rich substrates (Yang et al. 2008, Montell 2009). This same phenomenon is observed in European grapevine moth [Lobesia botrana (Denis & Schiffermüller; Lepidoptera: Tortricidae)] where female oviposition increases as glucose and fructose increase during the ripening process (Varandas et al. 2004). On the other hand, Biolchini (2016) reported that each valve of the D. suzukii ovipositor presents five putative gustatory sensilla arranged in a row. Electrophysiological recordings indicate that these sensilla are sensitive to different gustatory stimuli, including that of sugar. Behavioral observations confirmed that females preferred to lay eggs on substrates containing elevated levels of sucrose. The experimental setup used by Biolchini (2016) suggests that flies use the inputs arising by the piercing action with their ovipositor to determine the contact of the gustatory sensilla with the chemicals that are below the grape skin. So far, there are no reports about direct observations of explorative piercing carried out by D. suzukii in its selection of an oviposition site. It is more likely that D. suzukii is using gustatory sensilla to detect changes in sugar level and uses this information to select the most suitable available oviposition site. Ad hoc infestation set up in the vineyard 2 wk before harvest resulted in successful oviposition in all 86 bagged grape clusters, affecting 34.4% of the average weight of grape clusters. In all but five clusters, oviposition resulted in larval development, which affected 11% of the average weight of the grape clusters. Despite this generalized description of infestation and the high susceptibility of the tested cultivars, only 4 grape clusters produced a detectable level of volatile acidity after 14 d, considered a symptom of sour rot disease. The hot, dry weather experienced during the field trial period may explain this low level of disease development. Under such conditions, the contamination of the microbial consortium disseminated by D. suzukii during the initial 4 d of contact did not occur under optimal conditions for infection and development in the wounds made by oviposition and feeding by young larvae. As the contact between D. suzukii adults and berries was restricted to the initial 4 d, after which flies were removed and contact with other vector insects was excluded by bagging the grape cluster, there was no way to contaminate larger wounds caused by larval feeding activity. Moreover, the high mortality of larvae when growing on grape (Maiguashca et al. 2010, Lee et al. 2011, Linder et al. 2014, Ioriatti et al. 2015) allowed plant defense mechanisms to heal the wounds, preventing establishment of microbial infection and disease development. Finally, we cannot exclude that the lower levels of sour rot development following oviposition of D. suzukii in the field compared to our laboratory assays were due to presence of a more complex microorganism community. This possibly more complex community could have interfered with the development of spoilage bacteria present on the skin of the non-sterilized berries (Barata et al. 2012a). Conclusions A wide range of microorganisms may negatively affect grape quality and promote morbid infections. Among them, sour rot is an emergent grapevine disease affecting late-ripening cultivars with tightly packed, thin-skinned, and dense bunches close to harvesting. Sour rot may cause heavy crop losses and negatively affect juice and wine quality (Bisiach et al. 1986, Zoecklein et al. 2001, Barata et al. 2012c). However, the presence of D. suzukii eggs in the grape berries does not necessarily affect quality. Whether egg laying and successive larval development promote infection by microorganisms is the greater issue. Results of this study demonstrate that D. suzukii is vectoring spoilage bacteria through contact and feeding on damaged berries, as already demonstrated by other drosophilids inhabiting the vineyards (Fermaud et al. 2002). Sour rot developed in the current study because of contamination of wounded berries by microbiota vectored by contaminated D. suzukii. Moreover, while oviposition on intact, sound berries is not a sufficient condition for triggering sour rot development, the successive larval development constitutes an additional way for spreading spoilage bacteria such as Acetobacter spp. in wine grapes during the harvest period. We demonstrated, both under controlled laboratory settings and field conditions, that inoculative infestation with D. suzukii and subsequent larval development within grape berries can promote the increase of volatile acidity, as an indicator of increased sour rot levels. In conclusion, a number of factors affecting skin integrity (rainfall, wind, temperature, diseases, insect pests, viticultural practices, etc.) can influence grape microbiota that are primarily responsible for development of grape sour rot (Barata et al. 2012a). In fact, sound grape berries are less susceptible to the development of microbiota associated with sour rot and spoilage. In cases where D. suzukii eggs hatch and larvae develop, spoilage bacteria are vectored by contaminated D. suzukii adults. Furthermore, acetic acid bacteria induce fermentation and produce a number of volatile compounds as secondary metabolites in addition to acetic acid (Raspor and Goranovič 2008). The production of spoilage bacteria may create a positive feedback loop to attract both D. suzukii and other drosophilids (Mazzetto et al. 2016, Rombaut et al. 2017), which, by feeding on the infested grape berries, contribute to facilitating the spread of the disease. Acknowledgments This work was funded by the Autonomous Province of Trento (Italy), ‘Accordo di Programma Ricerca’, and the Oregon Wine Research Institute. Many thanks are due to Laura Turrin and Federico Micheli for collaboration in the field trial data collection. References Cited Andreazza , F. , Baronio C. A. , Botton M. , Valgas R. A. , Ritschel P. S. , Maia J. D. G. , and Nava D. E. . 2016 . 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TI - Drosophila suzukii (Diptera: Drosophilidae) Contributes to the Development of Sour Rot in Grape
JF - Journal of Economic Entomology
DO - 10.1093/jee/tox292
DA - 2018-02-09
UR - https://www.deepdyve.com/lp/oxford-university-press/drosophila-suzukii-diptera-drosophilidae-contributes-to-the-MqAY0sypDW
SP - 283
VL - 111
IS - 1
DP - DeepDyve
ER -