DOI: 10.5061/dryad.qbzkh18wx
title: Supplementary information: Integrative taxonomy unravels the true identity of Linnaeus’ <em>Termes fatalis</em> (Isoptera: Termitidae), the source of termites’ name

description: The identity of Termes fatalis Linnaeus, the species that gave termites
 their name, is still controversial since recent molecular and chemical
 studies showed that at least two different taxa matched its description.
 This paper aims to resolve this uncertainty and provide an unambiguous
 characterization of the type species of the genus Termes. We collected
 colonies matching the description of T. fatalis in French Guiana, compared
 the morphology of the different castes with type specimens of the Termes
 species reported from the northern Neotropics, sequenced their
 mitochondrial genomes, and analyzed the chemical composition of cuticular
 hydrocarbons in workers and defensive secretions in soldiers. Our
 phylogeny revealed that the current definition of T. fatalis encompasses
 three distinct taxa, distinguished by morphological details, the chemistry
 of cuticular hydrocarbons and defensive compounds. One species
 morphologically matches T. fatalis, while another one matches T.
 panamaensis Snyder and T. medioculatus Emerson, which cannot be
 distinguished and may be synonyms. The third species, herein described as
 T. incognitus sp. nov., is new to science.
description: # Supplementary information: Integrative taxonomy unravels the true
 identity of Linnaeus’ *Termes fatalis* (Isoptera: Termitidae), the source
 of termites’ name Original article accessible at:
 [https://doi.org/10.1111/syen.70016](https://doi.org/10.1111/syen.70016)
 ## Description of the data and file structure **Sample collection**
 Colonies of *Termes aff. fatalis* were collected in French Guiana and
 Trinidad. Specimens were preserved in 100% ethanol for genetic analyses,
 in 80% ethanol for morphological studies, and extracted in *n*-hexane for
 chemical analyses. Additional reference material from the University of
 Florida collection was used for genetic analyses of *T. hispaniolae* and
 *T. panamaensis*. Type material from the American Museum of Natural
 History and the Smithsonian National Museum of Natural History was
 examined for morphological comparisons with *T. fatalis*, *T.
 medioculatus*, and *T. panamaensis*. **DNA sequencing** DNA was extracted
 from the head of one soldier per colony collected in French Guiana and
 Trinidad (n = 236). The mitochondrial cytochrome c oxidase subunit II
 (COII) gene was sequenced using Sanger technology. Illumina sequencing was
 also performed to obtain the complete mitochondrial genome of at least one
 individual from each *T. aff. fatalis* lineage from French Guiana, as well
 as from the reference colonies of the University of Florida.
 **Phylogenetic analyses** Phylogenetic relationships were first inferred
 from all COII sequences, including those obtained from Illumina-sequenced
 mitogenomes and additional Termitinae sequences (for rooting and
 phylogenetic context). Sequences were aligned using MUSCLE in
 CodonCodeAligner, and the phylogenetic tree was reconstructed with
 MrBayes. A second phylogeny was generated using complete mitogenomes,
 including newly sequenced data and previously published mitogenomes of
 *Termes*, other Termitinae, and representative termite taxa. The
 concatenated alignment was partitioned into five categories: rRNA, tRNA,
 and the first, second, and third codon positions of protein-coding genes.
 Analyses were conducted with and without the third codon positions using
 BEAST. **Chemical analyses** For 22 colonies, 4–50 workers and soldiers
 were extracted in hexane and analyzed by gas chromatography. This allowed
 recovery of cuticular hydrocarbons from workers and defensive secretions
 from soldiers. **Morphological analyses** A total of 172 soldiers from 39
 colonies, 55 workers from 27 colonies, and 68 reproductives from 9
 colonies were measured using a micrometer mounted on a stereomicroscope.
 Workers’ mandibles were dissected and measured to calculate the mandibular
 index following Emerson (1960). A linear discriminant analysis (LDA) was
 performed on the measurements of soldiers and workers. ### Files and
 variables #### File: Aligned_COII_sequences.nex **Description:** This
 Nexus file contains the alignment of (1) newly generated COII sequences,
 (2) COII sequences extracted from newly assembled mitogenomes, and (3)
 COII sequences retrieved from GenBank. Sequences were aligned in
 CodonCodeAligner using the MUSCLE algorithm. #### File:
 Aligned_mitogenomes_with_3rd_codon.nexus **Description:** This Nexus file
 contains the whole mitogenome alignment, partitioned into rRNA, tRNA, and
 the first, second, and third codon positions. #### File:
 BEAST_configuration_without_3rd_codon.xml **Description:** This Nexus file
 contains the whole mitogenome alignment, partitioned into rRNA, tRNA, and
 the first and second codon positions (excluding third positions) ####
 File: BEAST_configuration_with_3rd_codon.xml **Description:** This XML
 file was generated with BEAUti and contains the parameter configuration
 for the BEAST analysis including third codon positions. #### File:
 Aligned_mitogenomes_without_3rd_codon.nexus **Description:** This XML file
 was generated with BEAUti and contains the parameter configuration for the
 BEAST analysis excluding third codon positions. #### File:
 BEAST_consensus_tree_with_3rd_codon.trees **Description:** This file
 contains the consensus tree from the BEAST analysis including third codon
 positions. The tree was summarized in TreeAnnotator as a maximum clade
 credibility tree. #### File: BEAST_consensus_tree_without_3rd_codon.trees
 **Description:** This file contains the consensus tree from the BEAST
 analysis excluding third codon positions. The tree was summarized in
 TreeAnnotator as a maximum clade credibility tree. #### File:
 MrBayes_consensus_tree.tre **Description:** This file contains the
 consensus tree generated with MrBayes using the sumt command. #### File:
 COII_sequences_(fasta).zip **Description:** This compressed archive
 contains the new cytochrome c oxidase subunit II (COII) sequences
 generated in this study, provided in FASTA format. Each sequence is
 labeled with its colony code, with corresponding details available in
 Samples_information.xlsx. Sequences were obtained from DNA extractions of
 soldier heads and sequenced using Sanger technology. COII sequences
 extracted from Illumina-sequenced mitogenomes are not included in this
 folder. #### File: BEAST_raw_trees_with_3rd_codon.trees
 **Description:** This file contains the raw BEAST output (10,000 trees)
 including third codon positions. #### File:
 BEAST_raw_trees_without_3rd_codon.trees.trees **Description:** This file
 contains the raw BEAST output (10,000 trees) excluding third codon
 positions. #### File: Raw_measurements.xlsx **Description:** This Excel
 file contains raw morphometric measurements of termites collected in
 French Guiana and of type specimens. Data are organized in three sheets,
 one for each caste (soldiers, workers, reproductives). Measurements are
 given in millimetres, except for the mandibular index (IM), which is
 unitless. Missing values are indicated with “NA”. **Variables**: *
 *Colony*: colony code of the measured specimen * *Species_name*:
 scientific name of identified type specimens (*T. fatalis*, *T.
 medioculatus*, *T. panamaensis*), and temporary name given to new
 specimens (*T.* aff.* fatalis* 1–3). * *Sex*: sex of the individual
 identified through the disposition of sternites. An additionnal (Q) or (K)
 indicate that the specimen is a queen or a king. * *Head lenght to the tip
 of the frontal process (HLT)*: measured according to Miller 1991 * *Head
 lenght to the anterior part of the genae (HLG)*: measured according to
 Miller 1991 * *Head depth (HD)*: measured according to Miller 1991 * *Head
 width (HW)*: measured according to Roonwal (1969), measure number 17 *
 *Pronotum width (PW)*: measured according to Roonwal (1969), measure
 number 68 * *Mandible lenght (ML)*: measured according to Miller 1991 *
 *Minimal width of the gula (Gwmin)*: measured according to Miller 1991 *
 *Maximal width of the gula (Gwmax)*: measured according to Miller 1991 *
 *Tibia lenght (TL)*: measured according to Roonwal (1969), measure number
 85 * *Mandibular index (IM)*: measured according to Emerson (1960) * *Head
 lenght (HL)*: measured according to Roonwal (1969), measure number 8 *
 *Pronotum lenght (PL)*: measured according to Roonwal (1969), measure
 number 66 * *Inter-eyes distance (EyesDist)*: measured according to
 Roonwal (1969), measure number 52 * *Diameter of the eye (EyeDiam)*:
 measured according to Roonwal (1969), measure number 48 * *Diameter of the
 ocellus (OcDiam)*: measured according to Roonwal (1969), measure number 55
 * *Eye-ocellus distance (EyeOcDist)*: measured according to Roonwal
 (1969), measure number 57 **Cited references:** Emerson, A.E. (1960) New
 genera of termites related to *Subulitermes* from the Oriental, Malagasy,
 and Australian regions (Isoptera, Termitidae, Nasutitermitinae). *American
 Museum Novitates*, 1986, 1–28. Miller, L.R. (1991) A revision of the
 *Termes-Capritermes* branch of the Termitinae in Australia (Isoptera:
 Termitidae). *Invertebrate Systematics*, 4 (6), 1147–282.
 [https://doi.org/10.1071/IT9901147](https://doi.org/10.1071/IT9901147)
 Roonwal, M.L. (1969) Measurements of termites for taxonomic purposes.
 *Journal of the Zoological Society of India*, 21, 9–66 #### File:
 Chemical_identities.xlsx **Description:** This Excel file contains raw
 results from chemical extractions of termite cuticular hydrocarbons (sheet
 “workers”) and defensive products (sheet “soldiers”). Results are
 expressed as relative abundances (%), and are colored using conditional
 formatting along a green (low values) to red (high values) gradient.
 Colonies G18-10c, G18-21, G18-22, G18-27, G18-28, G18-33, G18-34, G18-36A,
 G18-44, G18-45 are *T. fatalis*, G18-02, G18-03, G18-06, G18-16, G18-24,
 G18-37, G18-42 are *T. medioculatus* and G18-01, G18-11, G18-13, G18-19,
 G18-29 are *T. incognitus*. **Variables**: * *Peak label*: letter
 identifying the chromatographic peak of each compound * *Retention index*:
 retention index of the compound * *Identity*: chemical name of the
 compound identified from mass spectral analysis and retention index,
 confirmed by comparison with the NIST 20 library. * *Chemical class*:
 category of the identified compound * *Retention time (min)*: elution time
 in minutes of the compound #### File: LDA_soldiers.Rmd **Description:** R
 Markdown file containing the code used for the LDA analysis of the
 soldiers dataset. Results are presented in the file
 "LDA_soldiers_markdown.pdf". #### File:
 LDA_soldiers_markdown.pdf **Description:** This PDF was produced using the
 R Markdown file "LDA_soldiers.Rmd". It integrates the code used
 for the LDA analysis of the soldiers dataset with contextual text for
 explanation and organization, and displays the corresponding outputs
 alongside the code #### File: LDA_workers.Rmd **Description:** R Markdown
 file containing the code used for the LDA analysis of the workers dataset.
 Results are presented in the file "LDA_workers_markdown.pdf".
 #### File: LDA_workers_markdown.pdf **Description:** This PDF was produced
 using the R Markdown file "LDA_workers.Rmd". It integrates the
 code used for the LDA analysis of the worers dataset with contextual text
 for explanation and organization, and displays the corresponding outputs
 alongside the code. #### File: Samples_information.xlsx
 **Description:** This Excel file contains detailed data on all samples
 used in the study. Missing values are indicated by “NA”. **Variables**: *
 *Colony code*: unique identifier assigned to each colony by the collector
 or the institution of deposit * *Country*: country where the specimens
 were collected * *Locality*: region and eventual exact locality in which
 the specimens were collected * *GPS (North) / GPS (East)*: geographic
 coordinates (in decimal degrees) of the collection site * *Date of
 collect*: date when the specimens were collected, formatted as
 “yyyy-mm-dd” * *Collector(s)*: names of the person(s) who collected the
 specimens * *Collection*: institution where the specimens are deposited *
 *Species name*: final scientific name assigned to the specimens *
 *Temporary name*: provisional name used in the article or supplementary
 material (e.g., *T.* aff. *fatalis* 1) * *Type status*: type status of the
 specimens, if applicable * *Presence of alates*: an “x” indicates the
 presence of alates in the nest at the time of collection * *Soldiers
 measured*: an “x” indicates that soldiers from this colony were used for
 morphological analyses * *Workers measured and/or dissected*: an “x”
 indicates that workers from this colony were used for morphological
 analyses * *Alates measured*: an “x” indicates that reproductives (alates
 and eventually the queen/king) from this colony were used for
 morphological analyses * *Mitogenome accession*: GenBank accession numbers
 of mitogenome sequences * *COII accession*: GenBank accession numbers of
 COII gene sequences * *COII haplotype*: “Identical to …” indicates
 correspondence between the colony (if barcoded) and another haplotype
 whose sequence is either deposited in GenBank or listed in the previous
 column. All sequences are available in the file
 *COII_sequences_(fasta).zip*