Analysis of insecticide resistance and de novo transcriptome assembly of resistance associated genes in the European grapevine moth, Lobesia botrana (Lepidoptera: Tortricidae).

The European grapevine moth Lobesia botrana (Denis & Shiffermüller 1776) is an economically important pest of the vine-growing areas worldwide. Chemical insecticides have been used for its control; however, its resistance status is largely unknown in many regions. We monitored the susceptibility of several L. botrana populations from Greece and Turkey. In addition, based on RNAseq transcriptome analysis, we identified and phylogenetically classify the cytochrome P450 genes of L. botrana, as well as analysed target site sequences and looked for the presence of known resistance mutations. Resistance against chlorantraniliprole, alpha-cypermethrin, spinetoram, etofenprox, and acetamiprid was very low (below 2.5-fold in all cases, compared to a reference strain from Greece) in all populations from Greece that were included in the study. However, resistance against indoxacarb (4-30-fold), spinosad (5-59-fold), and deltamethrin (18-30 fold) was detected in the L. botrana populations from Turkey, compared to a reference population from Turkey. De novo transcriptome assembly and manual annotation, and subsequent PCR-based analysis of insecticide target sequences (i.e. voltage-gated sodium channel - VGSC: target of pyrethroids and oxadiazines; nicotinic acetylcholine receptor subunit a6 - nAChR_α6: target of spinosad; ryanodine receptor - RyR: target of diamides; glutamate-gated chloride channel - GluCl: target of avermectins and; acetylcholinesterase - AChE: target of organophosphates) showed the absence of known resistance mutations in all specimens from both countries. Finally, the L. botrana CYPome (116 genes) was manually analysed and phylogenetically characterised, to provide resources for future studies that will aim the analysis of metabolic resistance.

First Report of the Pepper Fruit Fly Atherigona orientalis (Schiner 1968) (Diptera: Muscidae) Infesting Commercial Pepper Crops in Greece.

The pepper fruit fly Atherigona orientalis (Schiner 1968) (Diptera: Muscidae) is a cosmopolitan tropical pest which has been recently recorded in several European countries. The biology of the pest has been primarily associated not only with decomposing fruits and vegetables, but even vertebrate and invertebrate carrion, dung and faeces. Relatively recently, A. orientalis has been reported as a primary pest of pepper fruits as well. In this short communication, we report, for the first time in Greece and, to the best of our knowledge, in Europe, cases of pepper fruit fly damage to pepper fruits in commercial greenhouse crops (in Crete in 2022). In this direction, possible implications and concerns regarding the occurrence of this pest in Crete are discussed.

Fish farming, metals and antibiotics in the eastern Mediterranean Sea: Is there a threat to sediment wildlife?

Chemical residues released from aquaculture farms may persist in the environment and may pose adverse ecological effects. The aim of this study was to assess the pollution status of marine sediments underneath or close to aquaculture farms in the Eastern Mediterranean Sea, the factors that affect the elemental distribution, and the exceedance of environmental quality standards and factors. To this end, surface sediment samples were collected (underneath fish cages, at 25 m distance and from a reference station) from 48 fish farms in Greece with variable environmental and geochemical characteristics. The content of 29 metals and major and trace elements, and three antibiotics (oxytetracycline, florfenicol, and flumequine), was determined in the sediment samples. Most of the elements found in the sediments originated from geological sources and their concentrations were close to Earth's Crust content. Below and close to fish farm cages, the sediment was enriched with P, Cu, Zn, Mo, and Cd mainly due to the aquaculture biosolid deposition, and changes in environmental conditions (increased organic matter, low redox potential). Cr and As were found to exceed twice the upper threshold limit of the available sediment quality guidelines in 13.5% and 7.3% of sampling stations, respectively. The potential ecological risk of V, Cr, Mn, Co, Ni, Cu, Zn, As and Pb was found to be low in the sediment close to fish cages except for Cd, which may pose a moderate to considerable risk in 12.5% of sampled fish farms. However, the estimated risks for this metal may be influenced by the high background levels found in the investigated areas. This study also shows that the occurrence of antibiotics in sediments below fish farms in the Eastern Mediterranean Sea is very sparse (i.e., only flumequine was found in two farms) and concentration levels are relatively low.

Metals and elements in sardine and anchovy: Species specific differences and correlations with proximate composition and size.

Species - specific differences in the levels of 26 metals and elements in sardine and anchovy are investigated and the factors of proximate composition (proteins, lipids, ash) and body size (length, weight) that may affect the metal and elemental concentrations in fish are explored. Statistical analysis revealed that levels of metals and elements in fish seem to strongly depend on species. Significantly higher levels of most of the metals and elements studied have been observed in anchovy compared to sardine at each of six different sites. The observed species-specific differences in metal content could be attributed to different proximate composition and size among other factors. The significant (positive or negative) correlations found between lipids, proteins, ash, weight, length and metals or elements, suggest that higher levels of most of the metals and elements studied can be associated with lower lipid content, greater protein and ash content and smaller size.

Arsenic speciation in fish from Greek coastal areas.

Arsenic speciation analysis was conducted on fish samples (sardine and anchovy) collected from six areas along the Greek coastline, i.e. Artemisium Straits, Thermaikos Gulf, Amvrakikos Gulf, Strymonian Gulf, Thracian Sea, and Elefsina Gulf. Total arsenic levels ranging from 11.8 to 62.6mg As/kg dry weight were determined. Arsenobetaine, a non-toxic form of arsenic, was found to be the main arsenic species, present at 8.6 to 58.8mg As/kg dry weight, accounting for 67-95% of the total arsenic. Also detected in all fish samples was dimethylarsinic acid, although at considerably lower concentrations, ranging from 0.072-0.956mg As/kg dry weight. Monomethylarsonic acid was detected at low levels in all anchovy samples, and only in sardines from one area. Finally, inorganic arsenic in the form of arsenate was detected only in fish at one area, indicating the possible effect of an environmental parameter on its presence at detectable amounts. Statistical analysis revealed the environmental variables, such as salinity, total organic carbon and nitrogen, ammonium, phosphate, total phosphorus, dissolved oxygen and pressure index, are potentially correlated to As species concentrations. Furthermore, based on factor analysis, the biological parameters, such as fish weight, lipids, protein and ash content, that are correlated to As species concentrations of fish were also identified. The interrelationship of arsenobetaine and dimethylarsinic acid concentrations within each fish species was evaluated.

The impact of silver nanoparticles on marine plankton dynamics: Dependence on coating, size and concentration.

During this study, three microcosm experiments were carried out with natural coastal seawater, collected in the Eastern Mediterranean Sea, in order to assess the effect of silver nanoparticle (AgNP) exposure to natural plankton communities. The impact of coating (branched-polyethyleneimine: BPEI vs. poly-vinylpyrrolidone: PVP), size (40 vs. 60nm), concentration (200, 500, 2000, 5000 and 10,000ng Ag L-1) and silver form (dissolved Ag+ vs. AgNPs) were tested. The results of chlorophyll a concentration revealed that PVP AgNPs caused a higher toxicity than BPEI AgNPs, and this was possibly related to the measured higher dissolution rate. Additionally, toxicity of BPEI AgNPs was size-dependent, with 40 being more toxic than 60 nm AgNPs, which was nevertheless not seen clearly for PVP AgNPs. Interestingly, community composition altered in response to AgNP exposure: cyanobacterial abundance was negatively affected at concentrations ≥200ng Ag L-1, and dinoflagellate abundance and composition were altered at a 2000ng Ag L-1 concentration. Specifically, dinoflagellate (Gymnodinium, Prorocentrum and Gyrodinium) and diatom (Nitzschia, Navicula and Climacosphenia) genera either increased or decreased, highlighting taxa-specific effects, with some of them being able to tolerate, compensate or even benefit from AgNPs. Silver in either form (dissolved Ag+ or in NPs) caused almost identical results in the plankton community, further indicating that Ag+ release is the primary cause of AgNP toxicity. This study employed for the first time environmentally relevant AgNP concentrations (minimum 200ng Ag L-1) in natural seawater without pre-filtration steps and showed that community changes were driven by the exposure but were largely dependent on ambient physico-chemical characteristics and should be further investigated.

Silver nanoparticles in seawater: A dynamic mass balance at part per trillion silver concentrations.

This study investigates the dynamic processes affecting silver (Ag) nanoparticles that have been spiked into seawater at environmentally relevant concentrations (200 and 2000ngAgL-1). Seawater samples were taken at regular time intervals from multiple microcosm tanks and analysed rapidly, without any sample preparation, using a recently developed flow injection on-line dilution single particle inductively coupled plasma mass spectrometry method. Dissolution was found to be the predominant process of Ag nanoparticle transformation, with its rate being influenced by the type and thickness of the nanoparticle organic coating. More specifically the branched poly(ethyleneimine) coating provided additional stability to the 40 and 60nmAg nanoparticles that were tested, compared to the poly(vinylpyrrolidone) coated ones. At high Ag nanoparticle spiking levels and after 24h of exposure an extra Ag-containing nanoparticle peak appeared at the low range of the NP size distribution histogram. This peak corresponds to Ag-containing particles that contain Ag mass equivalent to 25-30nm Ag nanoparticles (assuming spherical shape). However, the composition and the "real" size of these particles remains unknown as the particles may have formed from the in-situ reduction of dissolved silver or they originate from other processes involving nanocrystal formation, as has been shown to occur in sewage sludge, or interaction with natural organic matter. Overall, this study provides additional insight into the physicochemical mechanisms behind Ag nanoparticle behavior in marine media.