Morphomolecular characterization of invasive fruit borer infesting apple.

Apple, a vital fruit crop worldwide and a major crop grown in northern parts of India, acts as a backbone for the survival and livelihood of the farming community. However, it is prone to severe damage from insect pests and diseases. In the past few years, due to erratic weather patterns, there has been an alarmingly increased infestation of different insect pests, both invasive and non-invasive, resulting in substantial economic losses to this industry. One similar case was seen in the Himalayan region of India, where the apple crop in Karewas was heavily damaged by some invasive fruit borer, feeding on pulp and making galleries to reach the seed locules, thereby destroying the seed material. To ascertain the true identity of this pest, a study based on morpho-molecular characterization of this pest was carried out in the Faculty of Agriculture, Wadura, SKUAST Kashmir, India, during the years 2021 and 2022. The invasive fruit samples were collected from apple orchards at different locations (experimental sites) in North India by installing delta sticky traps @ 5 traps/ha for moth collection. The fruit-boring larvae and pupa were also collected and reared in confined chambers of the laboratory with controlled temperature and humidity. All the laboratory investigations were conducted at the Division of Entomology, FoA, Wadura, SKUAST Kashmir. During the investigation, it was confirmed that the invasive borer is a codling moth (Cydia pomonella L.), a member of the family Tortricidae, order Lepidoptera, which was earlier having quarantine importance in India, as it was only present in the Ladakh region of India. From the phylogenetic analysis of sample sequences, the species of codling moth present at all experimental sites was more identical to the codling moth sequence from Leh (the northernmost arid region), India. Further, the study of life cycle and voltinism revealed that codling moth completes three generations per year in Kashmir, with a single cycle lasting up to 2.5 months. However, the timing of various generations varies, depending on prevailing weather conditions at specific locations and times. The number of generations completed by codling moth in Kashmir conditions during apple growing season was three full and a half overwintering generations. The study of the life cycle of this important pest was necessary to know the weak link for its effective management and to prevent the economic loss in apple fruit.

The Root-Lesion Nematode, Pratylenchus penetrans, Affects Early Growth and Physiology of Grafted M.9, G.41, and G.935 Apple Rootstocks Similarly Under Field Microplot Conditions.

The root-lesion nematode, Pratylenchus penetrans, is a ubiquitous parasite of roots of temperate fruit trees. It affects early growth of trees replanted into former orchard sites where populations have built up and may contribute to decline complexes of older trees. Most British Columbia, Canada, apple acreage is planted with M.9 rootstock, but growers are increasingly considering Geneva-series rootstocks such as G.41 and G.935. Among these rootstocks, responses to P. penetrans, specifically, are poorly known. To compare the resistance and tolerance to P. penetrans of G.41, G.935, and M.9 rootstocks ('Ambrosia' scion), a field microplot experiment was established in spring of 2020 at the Summerland Research and Development Centre. The experimental design was a two by three factorial combination of: P. penetrans inoculation (+/-) and rootstock (G.41, G.935, and M.9), with 20 replicate microplots of each of the six treatment combinations arranged in a randomized complete block design. The P. penetrans inoculum was 5,400 nematodes per microplot (54 P. penetrans liter-1 soil), which is below commonly accepted damage thresholds. Though P. penetrans population densities were lower for the G.41 rootstock by the end of the 2021 growing season, the effects of P. penetrans were similar among rootstocks. In the establishment year (2020), P. penetrans caused significant reductions in aboveground growth. In 2021, shoot growth and root weight were reduced by P. penetrans. The nematode also reduced rates of leaf gas exchange and stem water potential. These data suggest that while G.41 and G.935 may have other horticultural benefits over M.9, they are equally susceptible to P. penetrans at the early stages of tree growth.

EPG analysis of stylet penetration preference of woolly apple aphid on different parts of apple trees.

Woolly apple aphid (WAA), Eriosoma lanigerum (Hausmann), is an important global pest that feeds on Malus species. We studied the feeding preference of WAA on apple trees in the field for two consecutive years and in the laboratory we used electronic penetration graphs (EPG) to record the stylet penetration behavior of WAA on different parts of apple trees. We found that in the field WAA fed primarily on twigs and branches, not on leaves and fruits. Six EPG waveforms were produced during WAA probing on shoots, trunks and leaves of apple trees, including the non-penetration wave (np), the stylet pathway phase wave (C), the intracellular feeding wave (pd), the xylem feeding wave (G), waves indicative of release of saliva into the phloem (E1), and a wave indicative of ingestion from phloem (E2). In the laboratory, aphids only successfully fed on shoots, trunks and leaves, not on fruits. The EPG parameters on the phloem of shoots were significantly higher than those on trunks, indicating WAA prefer to feed on shoots. These laboratory findings explain the relative field feeding preference of WAA on different parts of apple trees, which occurs primarily on branches, barks, and young twigs in orchards, especially on young twigs.

Genetic structure of Malus sylvestris and potential link with preference/performance by the rosy apple aphid pest Dysaphis plantaginea.

The European crabapple Malus sylvestris, a crop wild relative of Malus domestica, is a major contributor to the cultivated apple genome and represents a potential source of interesting alleles or genes, particularly pest resistance traits. An original approach was used to explore the trophic interaction between M. sylvestris populations and its pest, the rosy apple aphid (Dysaphis plantaginea). Using 13 microsatellite markers, population genetic structure and level of crop-to-wild introgressions were inferred between M. sylvestris seedlings from three sites in Europe (Denmark, France, Romania), and M. domestica seedlings. Genetically characterized plants were also used to analyze aphid feeding behavior and fitness parameters. First, aphids submitted to two genetically close M. sylvestris populations (the Danish and French) exhibited similar behavioral parameters, suggesting similar patterns of resistance in these host plants. Second, the Romanian M. sylvestris population was most closely genetically related to M. domestica. Although the two plant genetic backgrounds were significantly differentiated, they showed comparable levels of sensitivity to D. plantaginea infestation. Third, aphid fitness parameters were not significantly impacted by the host plant's genetic background. Finally, crop-to-wild introgression seemed to significantly drive resistance to D. plantaginea independent of host plant population genetic structure, with hybrids being less suitable hosts.

Developmental stages of peach, plum, and apple fruit influence development and fecundity of Grapholita molesta (Lepidoptera: Tortricidae).

Host plant attributes are essential factors determining the population dynamics of herbivorous insects. The developmental stage of host plants, in particular, may affect the biology of Grapholita molesta (Busck), a possibility that has rarely been examined. Here we assessed the effect of developmental stage of plum, peach, and apple fruits on the development and fecundity performance of G. molesta, along with an examination of the firmness and sugar content of the fruits. Among the fruits collected earliest (May 31), plum and apple were better food sources for G. molesta compared to peach in terms of development, reproduction, and life table parameters. However, despite the higher sugar content in peach, G. molesta larvae showed a lower rate of fruit penetration in peach, probably due to fruit firmness. In the later-collected fruit (June 25), both peach and apple were better than plum, as peach and apple were softer and had higher sugar content. Nevertheless, the penetration rate of larva was still low in peach probably due to pubescence on the fruit surface. Although the plum fruits in the later collection date were softer with higher sugar content, there was a negative impact on the development and reproduction because fruits started to liquefy earlier. In conclusion, the developmental stage of fruits with changes in fruit firmness or sugar content affected the development and reproduction of G. molesta, and apple would be the best food source.

Large-scale genetic admixture suggests high dispersal in an insect pest, the apple fruit moth.

Knowledge about population genetic structure and dispersal capabilities is important for the development of targeted management strategies for agricultural pest species. The apple fruit moth, Argyresthia conjugella (Lepidoptera, Yponomeutidae), is a pre-dispersal seed predator. Larvae feed on rowanberries (Sorbus aucuparia), and when rowanberry seed production is low (i.e., inter-masting), the moth switches from laying eggs in rowanberries to apples (Malus domestica), resulting in devastating losses in apple crops. Using genetic methods, we investigated if this small moth expresses any local genetic structure, or alternatively if gene flow may be high within the Scandinavian Peninsula (~850.000 km2, 55o - 69o N). Genetic diversity was found to be high (n = 669, mean He = 0.71). For three out of ten tetranucleotide STRs, we detected heterozygote deficiency caused by null alleles, but tests showed little impact on the overall results. Genetic differentiation between the 28 sampling locations was very low (average FST = 0.016, P < 0.000). Surprisingly, we found that all individuals could be assigned to one of two non-geographic genetic clusters, and that a third, geographic cluster was found to be associated with 30% of the sampling locations, with weak but significant signals of isolation-by-distance. Conclusively, our findings suggest wind-aided dispersal and spatial synchrony of both sexes of the apple fruit moth over large areas and across very different climatic zones. We speculate that the species may recently have had two separate genetic origins caused by a genetic bottleneck after inter-masting, followed by rapid dispersal and homogenization of the gene pool across the landscape. We suggest further investigations of spatial genetic similarities and differences of the apple fruit moth at larger geographical scales, through life-stages, across inter-masting, and during attacks by the parasitoid wasp (Microgaster politus).

Aphid-infested beans divert ant attendance from the rosy apple aphid in apple-bean intercropping.

Ecological intensification of cropping systems aims at restoring multi-functionality while supporting current productivity levels. Intercropping is a form of ecological intensification involving ecological processes beneficial to farmers that do not take place in monocultures. Thus, it represents a practical approach to decrease the use of synthetic inputs such as insecticides in cultivated systems. Whereas insecticide reduction via intercropping-facilitated suppression of aphids is reported in literature, the majority of published studies focussed on herbaceous crops. Thus, the effect of intercropping on aphid populations of cultivated trees remains largely unaddressed. In this study we hypothesized that intercropping a specific companion plant within perennial crops would divert ant attendance from an aphid attacking the crop to another aphid feeding on the newly introduced plant, reducing aphid damage on the crop. We tested our hypothesis in the system of apple (Malus domestica Borkhausen), the rosy apple aphid (Dysaphis plantaginea Passerini) and the black garden ant (Lasius niger L.). Bean plants (Vicia faba) with the black bean aphid (Aphis fabae Scopoli) were intercropped within apple trees inoculated with D. plantaginea. We measured ant attendance, aphid development and survival as well as honeydew composition on both plant species through semi-field and field experiments. The majority of ants chose to attend A. fabae over D. plantaginea in the semi-field experiment with potted plants. In the orchard, a larger majority of scouts were scored on A. fabae over D. plantaginea. A higher number of D. plantaginea colonies remained active in the apple control, whilst they were almost eradicated by intercropping. Although chemical analyses of honeydew disclosed differences in the carbohydrate and amino acid profiles between aphid species, the difference in honeydew composition did not explain the preference for A. fabae. Ants did not discriminate between the two honeydew mimics both in laboratory and field bioassays. Our results showed the potential of intercropping apple trees with beans as a method to reduce ant attendance and thus colony survival. We propose that intercropping represents a bottom-up approach towards ecological intensification of perennial crops. Together with other ecosystem-based measures such as habitat management, intercropping should be considered when planning ecosystem redesign to increase biological control of pests.

Honeybee Pollinators Use Visual and Floral Scent Cues to Find Apple (Malus domestica) Flowers.

Apple flowers of most varieties require pollinator-mediated cross-pollination. However, little is known about the cues used by pollinators to find the flowers. We used bioassays to investigate the importance of visual and olfactory cues for the attraction of honeybee pollinators to apple flowers. Chemical-analytical and electrophysiological approaches were used to determine floral scents and investigate antennal responses of honeybees to scents from flowering twigs. Bioassays showed that visual and olfactory cues were equally important for the attraction of honeybees to apple flowers. Floral scents were dominated by aromatic components, mainly benzyl alcohol, and the antennae of honeybees responded to a large number of components, among them to benzyl alcohol, linalool, and indole. Our study aims to better understand how this important fruit crop communicates with its main pollinators. This knowledge might be used to improve the attractiveness of apple flowers to pollinators to optimize fruit sets.

'Candidatus Phytoplasma mali' Genome Encodes a Protein that Functions as an E3 Ubiquitin Ligase and Could Inhibit Plant Basal Defense.

Phytoplasmas are the causative agent of numerous diseases of plant species all over the world, including important food crops. The mode by which phytoplasmas multiply and behave in their host is poorly understood and often based on genomic data. We used yeast two-hybrid screening to find new protein-protein interactions between the causal agent of apple proliferation 'Candidatus Phytoplasma mali' and its host plant. Here, we report that the 'Ca. P. mali' strain PM19 genome encodes a protein PM19_00185 that interacts with at least six different ubiquitin-conjugating enzymes (UBC; E2) of Arabidopsis thaliana. An in vitro ubiquitination assay showed that PM19_00185 is enzymatically active as E3 ligase with A. thaliana E2 UBC09 and Malus domestica E2 UBC10. We show that a nonhost bacteria (Pseudomonas syringae pv. tabaci) can grow in transgenic A. thaliana plant lines expressing PM19_00185. A connection of phytoplasma effector proteins with the proteasome proteolytic pathway has been reported before. However, this is, to our knowledge, the first time that a phytoplasma effector protein with E3 ligase activity has been reported.

Development and fecundity performance of Grapholita molesta and Grapholita dimorpha (Lepidoptera: Tortricidae) on different immature fruits.

Both Grapholita molesta (Busck) and Grapholita dimorpha Komai (Lepidoptera: Tortricidae) are important pests of pome and stone fruits. Mature fruits of peach and plum have been tested as food sources for G. molesta, but no studies have examined the suitability of immature fruits, which are the fruit stage more likely to be available for the spring populations of both G. molesta and G. dimorpha. Thus, we evaluated immature fruits of peach, plum, and apple as food sources by assessing their effects on biological and behavioral attributes of these moths in the laboratory. Immature fruits were collected in May and June of 2016 and again in 2017. The first-instar larvae of G. molesta preferred either peach or plum, which showed exit rates of mature larvae of 81.0 and 100.0% for the two fruits, respectively. On peach, development time was shorter, and fecundity was lower than plum. However, G. dimorpha preferred plum and apple, showing the lowest fruit-boring rate and lowest mature larval exit rate on peach, from which only one female moth emerged but produced no eggs. In conclusion, it seems that at the immature fruit stage, plum and apple are better food sources for both G. molesta and G. dimorpha than is peach.

Effects of restricting movement between root and canopy populations of woolly apple aphid.

Movement of insect pests between spatially subdivided populations can allow them to recolonize areas where local extinction has occurred, increasing pest persistence. Populations of woolly apple aphid (Eriosoma lanigerum [Hausmann]; Hemiptera: Aphididae), a worldwide pest of apple (Malus domestica [Borkhausen]), occur both below- and aboveground. These spatially subdivided subpopulations encounter different abiotic conditions, natural enemies, and control tactics. Restricting movement between them might be an effective management tactic to decrease woolly apple aphid persistence and abundance. We examined this possibility in the field, using sticky barriers to restrict upward woolly apple aphid movement to tree canopies, and in the greenhouse, using mulches and sand amendments to restrict downward movement to roots. In the field, blocking aphid movement up tree trunks did not decrease the number of colonies in tree canopies. Instead, sticky-banded apple trees had higher aphid colony counts late in the study. Earwigs, which are woolly apple aphid predators, were excluded from tree canopies by sticky bands. In the greenhouse, fewer root galls (indicative of aphid feeding) occurred on trees in sandy potting media and on those with mulch (wood chips or paper slurry). Our results suggest that upward movement is less important than other factors that affect aboveground aerial woolly apple aphid population dynamics. In addition, apple orchards planted in sandier soils or with mulches may be partially protected from woolly apple aphid root feeding.

The Efficacy of Semiselective Chemicals and Chloropicrin/1,3-Dichloropropene-Containing Fumigants in Managing Apple Replant Disease in South Africa.

Apple replant disease (ARD) is a biological phenomenon that is encountered when old apple orchards are replanted, resulting in tree growth and yield reductions in young trees. Three ARD orchard trials were conducted, which showed that semiselective chemicals (fenamiphos, metalaxyl, imidacloprid, and phosphonates) used independently, two fumigant formulations (33.3% chloropicrin and 60.8% 1,3-dichloropropene [Pic33-1,3D] and 57.% chloropicrin and 38% 1,3 dichloropropene [Pic57-1,3D]), and semiselective chemicals combined with Pic33-1,3D or Pic57-1,3D all contributed to significant increases in tree growth (trunk diameter and shoot length) relative to the untreated control 3 to 4 years postplanting. The treatments did not differ significantly from each other in improving tree growth. Yield was more indicative of treatment efficacy, but this varied between the three orchards. The Pic33-1,3D fumigant in combination with semiselective chemistries was the most consistent in significantly increasing cumulative yields. The Pic57-1,3D treatment was superior in increasing yields relative to the Pic33-1,3D treatment, because (i) it significantly increased cumulative yields in comparison with the Pic33-1,3D treatment in one orchard and (ii) in another orchard, a significant increase in yield was obtained with Pic57-1,3D relative to the control treatment but not with the Pic33-1,3D treatment. The quantification of ARD causative agents 20 months postplant showed that Phytophthora cactorum contributed to disease development in all three orchards; significant negative correlations existed between the quantity of P. cactorum DNA detected in tree roots and tree growth and less often, yield. In two orchards, only some of the treatments that significantly reduced the quantity of P. cactorum DNA in tree roots relative to the control also resulted in a significant increase in tree growth. Some of the aforementioned trends were also evident for Pratylenchus spp. root densities in two of the orchards. There was a significant positive correlation between P. cactorum root DNA quantities and Pratylenchus spp. root densities. Pythium spp. and "Cylindrocarpon"-like DNA quantities detected in tree roots typically were not indicative of treatment efficacy. However, a significant positive correlation existed between these two pathogen groups, suggesting complex interactions not associated with pathogen quantities per se.

An apple a day: an outbreak of cryptosporidiosis in Norway associated with self-pressed apple juice.

In the autumn of 2018, an outbreak of cryptosporidiosis affected adult employees from the same company in Western Norway. The organism was Cryptosporidium parvum, GP60 subtype IIaA14G1R1. All those infected had drunk from the same container of self-pressed apple juice. Incubation period (1 week) and clinical signs were similar among those infected, although some experienced a more prolonged duration of symptoms (up to 2-3 weeks) than others. The infections resulted after consumption from only one of 40 containers of juice and not from any of the other containers. It seems that although Cryptosporidium oocysts were detected in a sample from another container, the contamination did not affect the whole batch. This is perhaps indicative of a restricted contamination event, either from contaminated ground in the orchard, or during collection of the fruit, or during processing. Although outbreaks of food-borne cryptosporidiosis have previously been associated with consumption of contaminated apple juice, most of the more recent outbreaks of food-borne cryptosporidiosis have been associated with salad vegetables or herbs. This outbreak, the first outside USA reported to be associated with apple juice, is a timely reminder that such juice is a suitable transmission vehicle for Cryptosporidium oocysts, and that appropriate hygienic measures are essential in the production of such juice, including artisanal (non-commercial) production.

Field Evaluation of Reduced Rate Brassicaceae Seed Meal Amendment and Rootstock Genotype on the Microbiome and Control of Apple Replant Disease.

An orchard field trial was conducted to assess the utility of reduced rate Brassicaceae seed meal (SM) amendment in concert with specific rootstock genotypes for effective control of apple replant disease. Three amendment rates of a 1:1 formulation of Brassica juncea-Sinapis alba SM were compared with preplant 1,3-dichloropropene/chloropicrin soil fumigation for disease control efficacy. When applied at the highest rate (6.6 t ha-1) in the spring of planting, SM caused significant phytotoxicity and tree mortality, which was higher for Gala/M.26 than for Gala/G.41 but was not observed at SM application rates of 2.2 or 4.4 t ha-1. SM treatment resulted in growth and yield increases of Gala/M.26 and Gala/G.41 trees in a manner similar to the fumigation treatment and significantly greater than the no treatment control. Tree growth in soils treated with SM at 4.4 t ha-1 was similar or superior to that obtained with SM at 6.6 t ha-1 and superior to that attained at an SM application rate of 2.2 t ha-1. Soil fumigation and all SM treatments reduced Pratylenchus penetrans root infestation relative to the control treatment at the end of the initial growing season. Lesion nematode root densities in the fumigation treatment, but not SM treatments, rapidly recovered and were indistinguishable from the control at the end of the second growing season. Soil fumigation and all SM treatments significantly suppressed Pythium spp. root infection relative to the control. Trees grafted to rootstock G.41 possessed lower P. penetrans root densities relative to trees grafted to rootstock M.26. One year after planting, composition of microbial communities from SM-amended soils was distinct from those detected in control and fumigated soils, and the differences were amplified with increasing SM application rate. Specific fungal and bacterial phyla associated with suppression of plant pathogens were more abundant in SM-treated soil relative to the control, and they were similar in abundance in 4.4- and 6.6-t ha-1 SM treatments. Findings from this study demonstrated that use of the appropriate apple rootstock genotype will allow for effective replant disease control at SM application rates significantly less than that utilized previously (6.6 t ha-1).

Characterization of the gut microbiota of invasive Agrilus mali Matsumara (Coleoptera: Buprestidae) using high-throughput sequencing: uncovering plant cell-wall degrading bacteria.

The genus Agrilus comprises diverse exotic and agriculturally important wood-boring insects that have evolved efficient digestive systems. Agrilus mali Matsumara, an invasive insect, is causing extensive mortality to endangered wild apple trees in Tianshan. In this study, we present an in-depth characterization of the gut microbiota of A. mali based on high-throughput sequencing of the 16S rRNA gene and report the presence of lignocellulose-degrading bacteria. Thirty-nine operational taxonomic units (OTUs) were characterized from the larval gut. OTUs represented 6 phyla, 10 classes, 16 orders, 20 families, and 20 genera. The majority of bacterial OTUs belonged to the order Enterobacteriales which was the most abundant taxa in the larval gut. Cultivable bacteria revealed 9 OTUs that all belonged to Gammaproteobacteria. Subsequently, we examined the breakdown of plant cell-wall compounds by bacterial isolates. Among the isolates, the highest efficiency was observed in Pantoea sp., which was able to synthesize four out of the six enzymes (cellulase, cellobiase, ß-xylanase, and ß-gluconase) responsible for plant-cell wall degradation. One isolate identified as Pseudomonas orientalis exhibited lignin peroxidase activity. Our study provides the first characterization of the gut microbial diversity of A. mali larvae and shows that some cultivable bacteria play a significant role in the digestive tracts of larvae by providing nutritional needs.

Leafroller-induced phenylacetonitrile and acetic acid attract adult Lobesia botrana in European vineyards.

We recently identified unique caterpillar-induced plant volatile compounds emitted from apple leaves infested with the larvae of various leafroller species. In subsequent field tests, binary blends of phenylacetonitrile+acetic acid and 2-phenylethanol+acetic acid were found to be attractive to a range of tortricid leafroller species (Tortricidae: Tortricinae) in both the Southern and Northern Hemispheres. In this work, the caterpillar-induced plant volatiles from the apple-leafroller system were tested in two vineyards in Spain and Hungary for their attractiveness to the grape frugivore Lobesia botrana (Tortricidae: Olethreutinae). As seen for Tortricinae species, a binary blend of phenylacetonitrile+acetic acid attracted significantly more male and female L. botrana to traps than acetic acid or blank lures. Traps baited with other caterpillar-induced plant volatile compounds (benzyl alcohol, 2-phenylethanol, indole, and (E)-nerolidol, each as a binary blend with acetic acid) did not catch significantly more moths than traps containing acetic acid alone. The catches of male and female moths support an optimistic future for new products in female tortricid surveillance and control that are based on combinations of kairomone compounds released from larval-damaged foliage.

Interaction of Brassicaceae Seed Meal Soil Amendment and Apple Rootstock Genotype on Microbiome Structure and Replant Disease Suppression.

Preplant soil application of a Brassica juncea-Sinapis alba seed meal formulation (SM) at a rate of 6.6 t ha-1 alters composition of the orchard soil microbiome in a manner that yields sustainable long-term suppression of soilborne pathogens in apple production systems. However, the cost of SM amendment has hindered the adoption of this tactic to manage apple replant disease in commercial orchards. Greenhouse trials were conducted to assess the effect of reduced SM application rates in concert with apple rootstock genotype on structure of the rhizosphere microbiome and associated disease control outcomes. At all application rates assessed, SM treatment increased tree growth and reduced disease development relative to the control. In general, total tree biomass and leader shoot length were similar in soils treated with SM at 4.4 or 6.6 t ha-1 regardless of rootstock genotype. Equivalent increase in tree biomass when cultivated in soil treated at the lowest and highest SM amendment rate was attained when used in conjunction with G.41 or G.210 apple rootstocks. Suppression of Pythium spp. or Pratylenchus penetrans root densities was similar at all SM application rates. When cultivated in nontreated replant orchard soil, Geneva rootstocks (G.41 and G.210) exhibited lower levels of Pythium spp. and P. penetrans root colonization relative to Malling rootstocks (M.9 and MM.106). For a given rootstock, structure of the rhizosphere microbiome was similar in soils treated with SM at 4.4 and 6.6 t ha-1. G.41 and G.210 rootstocks but not M.9 or MM.106 cultivated in soil treated with SM at 2.2 t ha-1 possessed a rhizosphere bacterial community structure that differed significantly from the control. Findings indicate that effective control of apple replant disease may be attained at lower SM amendment rates than employed previously, with lower effective rates possible when integrated with tolerant rootstock genotypes such as G.41 or G.210.

Extract of Nicotiana tabacum as a potential control agent of Grapholita molesta (Lepidoptera: Tortricidae).

Oriental fruit moth, Grapholita molesta (Busck) (Lepidoptera: Tortricidae), is an important pest of stone and pome fruits. Growers usually depend on chemical insecticides to control this pest, but demand for more environmentally-friendly means of controlling pests is increasing. At least 91 plant extracts have been reported to be effective against other lepidopterans, but their acute toxicity against G. molesta has rarely been studied. Among these 91 materials, we assessed the residual toxicity of 32 extracts against first instar larvae (< 5 h old) of G. molesta in the laboratory. Nicotiana tabacum L., used at the concentration of 2 mg/ml, showed the highest corrected mortality (92.0%) with a lethal time (LT50) value of 12.9 h. The extract was followed in its efficacy by Allium sativum L. (88.0%), Zanthoxylum piperitum (L.) De Candolle (70.0%), and Sapindus mukorossi Gaertner (65.0%), when mortality was assessed at 20 h after exposure. Against adult fruit moths (< 5 d old), N. tabacum also showed the highest corrected mortality among tested extracts, being 85 and 100% in adult females and males, respectively, at 168 h after exposure. However, there was no synergistic effect of the combined application of any of the top four extracts in either laboratory or greenhouse assays. Oviposition by G. molesta on peach twigs was reduced 85-90% when N. tabacum was applied at 4 ml/ twig compared to control (methanol), demonstrating that N. tabacum may have potential for use as a botanical insecticide against G. molesta.

Effects of Temperature and Moisture on the Infection and Development of Apple Fruit Rot Caused by Phytophthora cactorum.

Phytophthora fruit rot, caused by Phytophthora cactorum, is an important disease of apple in China, often causing more than 50% fruit rot in rainy years. We examined the effects of temperature and moisture on the development of the disease and effects of the variables on zoospore release and germination, infection, and lesion development. In vitro, a temperature range of 5 to 20°C had no significant effects on zoospore release dynamics but did significantly affect the quantities of released zoospores. The largest quantity of zoospores was released at 9.9°C according to a fitted model. Zoosporangia released zoospores within 15 min at the test temperatures (0 to 20°C), which peaked at the fourth hour. Zoospores germinated in vitro, requiring free water, at temperatures from 5 to 35°C. The optimum germination temperature was 25.1°C according to a fitted model. The minimum wetness duration required for zoospores to complete the infection process and induce visible lesions on Fuji fruit was 0.40 h at the optimal temperature of 23.0°C according to the fitted model, whereas observed values were 4.5, 1.5, 0.5, 1.5 and 8.5 h at 10, 15, 20, 25, and 30°C, respectively. The number of zoospore infections on fruit at various temperatures and wetness durations were well fitted by the modified Weibull model; based on the model, the optimal temperature for zoospore infections was 23.0°C. Young apple fruit infected by zoospores developed visible lesions from 10 to 30°C, with a predicted optimum of 23.5°C; no lesions developed at 5 or 35°C. The shortest incubation period of the disease was 4 days. These results can be used to develop disease forecasting models for improved fungicide control.

Comparative mitogenome analysis on species of four apple snails (Ampullariidae: Pomacea).

The genus Pomacea contains most of the economically important Ampullariid apple snails. Five Pomacea species were reported introduced out of their native ranges, including three highly invasive species, i.e. P. maculata, P. canaliculata and an unidentified species, Pomacea sp. In this study, the mitogenome of P. maculata was determined, which carried typical gene set of metazoan mitogenomes and shared the same gene rearrangement of Ampullariidae mitogenomes. The control region of the P. maculata has a 13-bp inverted repeat unit. We compared the mitogenome of P. maculata with P. canaliculata, Pomacea sp. and the aquatic pet species, P. diffusa. The three highly invasive species showed high sequence similarity of PCGs and RNAs. atp8 and nad2 showed the lowest similarity and the highest Ka/Ks ratios, indicating that both genes have potential for studying species identification and populations genetics in apple snails. All PCGs have the Ka/Ks ratios <1, indicating the existence of purifying selection in Pomacea species. We reconstructed phylogenetic trees using 14 Caenogastropda species based on sequences of PCGs and rRNAs using Bayesian inference. Pomacea maculata grouped with other Ampullariids and was most closely related to Pomacea sp. The mitogenome of P. maculata provides useful genetic resource for exploring the genetics and evolution of P. maculata and other apple snails.