Long-term effects of neonicotinoid insecticides on ants

The widespread prophylactic usage of neonicotinoid insecticides has a clear impact on non-target organisms. However, the possible effects of long-term exposure on soil-dwelling organisms are still poorly understood especially for social insects with long-living queens. Here, we show that effects of chronic exposure to the neonicotinoid thiamethoxam on black garden ant colonies, Lasius niger, become visible before the second overwintering. Queens and workers differed in the residue-ratio of thiamethoxam to its metabolite clothianidin, suggesting that queens may have a superior detoxification system. Even though thiamethoxam did not affect queen mortality, neonicotinoid-exposed colonies showed a reduced number of workers and larvae indicating a trade-off between detoxification and fertility. Since colony size is a key for fitness, our data suggest long-term impacts of neonicotinoids on these organisms. This should be accounted for in future environmental and ecological risk assessments of neonicotinoid applications to prevent irreparable damages to ecosystems.

Nature Research wishes to improve the reproducibility of the work that we publish. This form provides structure for consistency and transparency in reporting. For further information on Nature Research policies, seeAuthors & Referees and theEditorial Policy Checklist .

Statistics
For all statistical analyses, confirm that the following items are present in the figure legend, table legend, main text, or Methods section.
n/a Confirmed The exact sample size (n) for each experimental group/condition, given as a discrete number and unit of measurement A statement on whether measurements were taken from distinct samples or whether the same sample was measured repeatedly The statistical test(s) used AND whether they are one-or two-sided Only common tests should be described solely by name; describe more complex techniques in the Methods section.
A description of all covariates tested A description of any assumptions or corrections, such as tests of normality and adjustment for multiple comparisons A full description of the statistical parameters including central tendency (e.g. means) or other basic estimates (e.g. regression coefficient) AND variation (e.g. standard deviation) or associated estimates of uncertainty (e.g. confidence intervals) For null hypothesis testing, the test statistic (e.g. F, t, r) with confidence intervals, effect sizes, degrees of freedom and P value noted Give P values as exact values whenever suitable.

For Bayesian analysis, information on the choice of priors and Markov chain Monte Carlo settings
For hierarchical and complex designs, identification of the appropriate level for tests and full reporting of outcomes Estimates of effect sizes (e.g. Cohen's d, Pearson's r), indicating how they were calculated Our web collection on statistics for biologists contains articles on many of the points above.

Software and code
Policy information about availability of computer code Data collection

Data analysis
For manuscripts utilizing custom algorithms or software that are central to the research but not yet described in published literature, software must be made available to editors/reviewers. We strongly encourage code deposition in a community repository (e.g. GitHub). See the Nature Research guidelines for submitting code & software for further information.

Data
Policy information about availability of data All manuscripts must include a data availability statement. This statement should provide the following information, where applicable: -Accession codes, unique identifiers, or web links for publicly available datasets -A list of figures that have associated raw data -A description of any restrictions on data availability Field-specific reporting Please select the one below that is the best fit for your research. If you are not sure, read the appropriate sections before making your selection. No computer code or software was used to generate data. The data is based on observations of laboratory Lasius niger colonies.
Only publicly available software (Custom code in R + associated packages; cited and referenced in the manuscript) were used for the data analyses (Standard statistical approaches described in material and methods).
Data supporting the findings of this study are available fin the supplementary materials.

nature research | reporting summary
October 2018 For a reference copy of the document with all sections, see nature.com/documents/nr-reporting-summary-flat.pdf

Ecological, evolutionary & environmental sciences study design
All studies must disclose on these points even when the disclosure is negative. Most studies on sublethal effects have focused on pollinators due to their economic importance. However, little research exists on long-term consequences of chronical insecticide exposure to long-lived soil-dwelling organisms. Regarding long life spans, ant-queens are outstanding, especially black garden ant queens that can live up to 30 years. L. niger is endemic to Europe and colonies can be regarded as sedentary, potentially leading to exposure over decades. Thus, L. niger was chosen as study species.
Gynes of L. niger were captured after their nuptial flight and then exposed to field-realistic sublethal levels of thiamethoxam for for two developmental seasons until the second overwintering. Due to logistical reasons involved in keeping colonies for such a long duration, 30 queens were assigned to three thiamethoxam treatments (control = 0 !g/L, low = 4.5 !g/L, high = 30 !g/L; N=10 each). No data were excluded.
The experimental set up is described in the material and methods section and could be reproduced by other research groups. Data supporting the findings of this study are available from the corresponding author in order to reproduce the results from the collected raw data.
Colonies were randomly assigned to the three treatments based on a random number generator created as custom code in R.

N/A
The fieldwork consisted of sampling queens after mating flight. Nuptial flights always take place at the same time of year (late June-early July) and the same time of day (late afternoon), and occur during periods of hot weather just before or following rainfall.

Bern , Switzerland
The ants were collected on a concrete place just outside a supermarket (accessible to general public).
Apart from the collection stress induced on captured queens no further disturbance was caused.