Abstract: In this work, based on morphological and molecular characterization Thrips origani Priesner (Thysanoptera: Thripidae) is recorded for the first time from the Americas, causing damage to cultivated Origanum vulgare L. in La Consulta, Mendoza, Argentina. An illustrated key is provided to aid morphological identification of the five species of the genus Thrips L. now known to be present in Argentina and partial sequences of the cytochrome oxidase subunit I gene from specimens of three species of Thrips from Argentina, including T. origani, have been added to an international data bank.
Keywords: COI, Morphological Key, Oregano, Thrips Pest.
Resumen: En este trabajo, basado en la caracterización morfológica y molecular, se registra por primera vez a Thrips origani Priesner (Thysanoptera: Thripidae) en América, causando daños a cultivos de Origanum vulgare L. en La Consulta, Mendoza, Argentina. Se proporciona una clave ilustrada para ayudar a la identificación morfológica de las cinco especies del género Thrips L. que se sabe que están presentes en Argentina y se han agregado secuencias parciales del gen de la subunidad I de citocromo oxidasa de especímenes de tres especies de Thrips de Argentina, incluido T. origani, a un banco de datos internacional.
Palabras clave: Clave Morfológica, COI, Orégano, Trips Plaga.
Notas
First record of Thrips origani (Thysanoptera: Thripidae), from the Americas and illustrated key for the known species of Thrips from Argentina
Primer registro de Thrips origani (Thysanoptera: Thripidae) para las Américas y clave ilustrada para las especies de Thrips conocidas de Argentina
Recepción: 24 Julio 2024
Aprobación: 04 Noviembre 2024
The genus Thrips L. (Thysanoptera: Thripidae) has 303 valid species (Thrips Wiki, 2023), of which 62 species (43 endemic, 18 introduced, and one Holarctic) were recorded from the Americas by Nakahara (1994). It can be distinguished from other genera of Thripidae by the following features: antennae 7- or 8-segmented, segment I without median dorso-apical setae, III and IV each with forked sense cones (Fig. 2i-k); ocellar setae pair I absent, ocellar setae III subequal to or longer than ocellar setae II; median pair of setae on posterior margin of pronotum subequal to or longer than submedian pair (Fig. 1d); ferna entire, undivided; mesosternum with spinula, metasternum without spinula; presence of paired ctenidia laterally on abdominal tergites V-VIII, terminating laterally at tergal setae S3 on tergites VI-VII; ctenidia on tergite VIII posteromesad of the spiracles; tergite X split longitudinally in both sexes (Bhatti, 1980; Mound, 2002; Masumoto & Okajima, 2013; Tyagi & Kumar, 2015).
Only four species from the genus Thrips have been recorded from Argentina: Thrips tabaci Lindeman, Thrips simplex (Morison), Thrips australis (Bagnall) and Thrips trehernei Priesner (de Borbón, 2009). The onion thrips (T. tabaci), a widely distributed, polyphagous pest species, has been characterized as containing at least three biotypes (leek-associated arrhenotokous L1-biotype, leek-associated thelytokous L2-biotype and tobacco-associated arrhenotokous T-biotype); however, the adults are morphologically indistinguishable (Iftikhar et al., 2016; Farkas et al., 2020). In 2016, Panonto & Bauzá reported for the first time a species of the genus Thrips that damage oregano (Origanum vulgare L.) crop in the province of Mendoza, without confirming the identity of the species.
The objectives of this work were to identify the species of thrips present on the oregano and then provide an illustrated morphological key to the Thrips species present in Argentina, and also to obtain and analyze a partial sequence of cytochrome oxidase I for these species. As a result of this work, we record the presence of Thrips origani for the first time anywhere in the Americas.
Specimens were collected from oregano being grown in Mendoza, Argentina, and these were then cleared with 10 % KOH, dehydrated with increasing concentrations of ethanol, transferred to clove oil and mounted onto slides in Canada Balsam (Mound & Marullo, 1996). The specimens were observed under a microscope with phase contrast at 400x and identified as T. origani using keys to European species of Thrips (zur Strassen, 2003; Mound et al., 2018). To confirm the identification, images were taken (Fig. 1 c-d; Fig. 2 a, e, f, i) and matched with slide-mounted specimens of T. origani that had been collected from wild-growing Origanum vulgare in England. As a result, T. origani is here recorded for the first time from both Argentina and the Americas.
Thrips origani is found across western, central, and eastern continental Europe, plus the United Kingdom, Norway, the Azores and Madeira (zur Strassen, 2003; Mound et al., 2018). It is host specific to O. vulgare, a woody perennial with a native range across the Palaearctic from Macaronesia to China (POWO, 2024). It is widely cultivated as a culinary herb, but not only has T. origani not previously been reported as a pest of cultivated oregano (in Europe or elsewhere), the thrips is very rarely mentioned in the literature at all. A problem with thrips causing damage to cultivated oregano in Mendoza dates back to at least 2011, with 40 % of growers in one survey reporting a problem (Panonto & Bauzá, 2016). The damage is manifested in spring (from September onwards); new shoots are attacked leaving the top of the stems brownish with only a few deformed flowers appearing.
La Consulta, Mendoza, is a locality situated in the centre-west of Argentina. The climate semi-arid and cold corresponds to the BSK type according to Köppen-Geiger classification (Peel et al., 2007). It has a great thermal amplitude, with cold nights and warm days. It is separated from Chile and the Pacific Ocean by a high part of the Los Andes range. By contrast, for example, in the maritime climate of England, T. origani is a mid-summer species, its visibility coinciding with the flowering season of wild O. vulgare on dry grasslands, particularly those on chalk geology.
This species is morphologically similar to two other species found in its home range in Europe, Thrips tabaci and a rarely collected host-specialist of the woodland plant Euphorbia amygdaloides, Thrips euphorbiicola Bagnall. These three species all have closely spaced rows of microtrichia on the abdominal pleurotergites (Fig. 2e); the posteromarginal comb on tergite VIII complete with long, fine, microtrichia; the anterior pairs of pores on tergite IX not developed; and second-instar larvae with the lateral spiracles on tergite II absent (this is unusual within the genus Thrips). However, T. origani is morphologically distinct from these other two species, with 1-5 discal setae present on each of sternites III-VI (Fig. 2a), strong dark antecostal ridges on the sternites (Fig. 2a) and tergites, and variable brown and yellow mottling on the tergites. Males are unknown, the species reproducing by thelytokous parthenogenesis.
NEW RECORDS: Argentina, Mendoza, La Consulta (-33.7625, -69.1153): 9 females on Origanum vulgare, 21.x.2014, and 7 females on Origanum vulgare, 13.iii.2023 (coll. S. Panonto). The specimens are deposited in the Thysanoptera collection of the Estación Experimental Agropecuaria, Mendoza, INTA.
1. Abdominal pleurotergites with closely spaced rows of microtrichias (Fig. 2e); abdominal sternite VII without discal setae (Fig. 2a-b) 2
-. Abdominal pleurotergites without closely spaced rows of microtrichia (Fig. 2d); abdominal sternite VII with many discal setae (Fig. 2c) 3
2. Abdominal sternites with a few discal setae (Fig. 2a); antennal segments III-V uniformly brown, III slightly paler than IV-V (Fig. 2i); on Origanum vulgare T. origani
-. Abdominal sternites without discal setae (Fig. 2b); antennal segments III-V, usually slightly bicoloured, darker at apex than base; polyphagous T. tabaci
3. Forewing first vein with continuous row of setae (Fig. 2g); female and male yellow to brown, typically yellow with brown post-occipital ridge on head and brown spots on tergites (Fig. 1a); metanotum with campaniform sensilla (Fig. 1e); on Eucalyptus spp....................... T. australis
-. Forewing first vein with discontinuous row of setae (Fig. 2h); female and male dark brown (Fig. 1b); metanotum without campaniform sensilla 4
4. Antenna 8-segmented; antenna brown, except segment III yellow (Fig. 2k); reticulate metanotum, many of the reticles with internal markings (Fig. 1f) ; on Gladiolus spp T. simplex
-. Antenna 7-segmented, antennal segments III to base of VI paler than the rest of the antennal segments (Fig. 2j); metanotum, with reticles lacking internal markings; on Asteraceae T. trehernei

a-c female adults’ full body (a) T. australis (b) T. trehernei (c) T. origani; (d) head with ocellar setae pair I absent, ocellar setae III subequal to or longer than ocellar setae II and pronotum with median pair of setae on posterior margin longer than submedian pair of T. origani; e-f metanotum (e) T. australis, campaniform sensilla present (f) T. simplex, reticles with internal markings.
Four specimens were individually examined under a stereoscopic microscope with 160x magnification and placed in identifiable vials (labeled with a “sample number”). Specimens originally collected from La Consulta, Mendoza, were reared on oregano plants and collected for morphological and molecular characterization (INTA CdB 147 and INTA CdB 148). Additionally, DNA was also extracted from one specimen collected on Senecio subulatus D. Don ex Hook. & Arn. from El Carrizal, Mendoza, and previously identified under a stereoscopic microscope as T. tabaci (INTA CdB 35) and one specimen collected from an unidentified Asteraceae from Mar del Plata, Buenos Aires, and classified as T. trehernei (INTA CdB 131).

a-c sternites VI-VIII (a) T. origani, sternites VI with few discal setae and VI without discal setae (b) T. tabaci, sternites VI-VII without discal setae (c) T. trehernei, sternites VI -VII with many discal setae; d-e. pleurotergites (d) T. trehernei, rows without microtrichias (e) T. origani, rows with microtrichias, f-h forewings setae distribution (f) T. origani, (g) T. australis (h) T. trehernei; i-k antenna, number of segments and colour (i) T. origani (j) T. trehernei (k) T, simplex, antenna 8-segmented.
Specimens of thrips were collected in tubes containing 100 % ethanol. The species were recognized using morphological characters observable under a stereoscopic microscope at 160x magnification. Specimens were preserved in a freezer at -20 ºC until the moment of their use. DNA extraction was performed following the protocol proposed by Rugman et al. (2006) and modified by Kadirvel et al. (2013). A whole specimen was placed in a
1.5 ml Eppendorf tube with 100 µl of TNES (50 mM Tris [pH: 7.5], 400 mM NaCl, 20 mM EDTA, and 0.5 % SDS) and ground with the help of a plastic tip. The ground-down tissue was transferred to a 0.5 ml Eppendorf tube and 0.85 µl of proteinase K (20 mg/ml) was added. The sample was then incubated in a bath at 37 ºC for 18 h. The protein was precipitated with the addition of 28 µl of 5 M NaCl and centrifuged for 5 min at 13,000 rpm. The supernatant was transferred to a new 0.5 ml Eppendorf tube, one volume of
100 % ethanol previously chilled in a freezer at -20 ºC was added, and the tube incubated for one hour at room temperature. A precipitation step was done by centrifugation for 5 min at 13,000 rpm. The supernatant was discarded, 200 µl of 70 % ethanol was added to wash, and another centrifugation was carried out for 5 min at 13,000 rpm. The supernatant was carefully discarded, and the pellet was allowed to air dry. Once dry, the DNA was resuspended in 20 µl of sterile water and stored in a freezer at -20 ºC until PCR was carried out. The following primers were used: forward C1-J-1718 GGAGGATTTGGAAATTGATTAGTTCC and reverse C1-N-2329: ACTGTAAATATATGATGAGCTCA, cited by Simon et al. (1994), to obtain the partial sequence of the mitochondrial COI gene. A total volume of 40 µl PCR reagents was used, containing a 4 µl sample with 10 ng/µl DNA, 4 µl 1X PCR buffer, 1µl MgCl2 [50 mM], 0.6 µl [10 µM], dNTPS, 0.4 µl [100 µM] of forward and reverse primers and 0.3 µl Platinum Taq polymerase, and 29 µl sterile distilled water. The PCR reaction was performed in an Eppendorf Mastercycler nexus model thermocycler. The program consisted of 94 ºC for 10 minutes, followed by 4 cycles of 94 ºC for 40 seconds, 45 ºC for 40 seconds, 72 ºC for 1 minute, then 35 cycles of 94 ºC for 40 seconds, 51 ºC for 40 seconds, 72 ºC for 1 minute, and finally 15 minutes at 72 ºC. A 1 % agarose gel (TBE) was run at 90 volts for 30 minutes to confirm the amplification of the fragment sought and bands of approximately 650 bp were visualized, using a 100 bp ladder (InvitrogenTM) as marker. The positive samples were purified according to the manufacturer's protocol with a Qiagen purification kit (QIAquick PCR Purification Kit), and they were sent to the laboratory of the Genomics Unit, Institute of Biotechnology, CICVyA - CNIA - INTA, where they were sequenced by capillary electrophoresis.


Four partial sequences contributed by us and deposited in the European Nucleotide Archives (ENA) and 25 obtained from GenBank (including three sequences for each biotype of T. tabaci , all available sequences of the genus Thrips tabaci filtered as cytochrome oxidase subunit I and Argentina, three sequences for T. simplex, three for T. trehernei and the only one available for T. australis) (Table I) were edited using BioEdit Sequence Alignment Editor (Hall, 1999) to conserve only the regions with good sequencing quality and that showed consensus in both directions. A BLASTN analysis (Zhang et al., 2000) was carried out using the consensus sequences of COI amplicon as query. Target sequences with the greatest similarity were retrieved from the GenBank database, and all taxa were aligned with ClustalW. Phylogenetic trees were constructed with MEGA11 software (Tamura et al., 2011). The evolutionary history was inferred using the Maximum Likelihood method based on the Tamura-Nei model (Tamura & Nei, 1993). The sequences were deposited in the EMBL-EBI database (PRJEB59814).
The phylogenetic tree for a partial sequence of cytochrome oxidase I, obtained for species of Thrips including those identified in Argentina up to May 23rd 2024 (Fig. 3) shows that both isolates of T. origani locate on the same clade distinct from T. tabaci (biotypes L1, L2 and T), T. australis, T. simplex and T. trehernei sequences, but show similarity (0.99) with one specimen classified as T. tabaci (sequence code in GenBank, MG334456). This sequence differs by more than 10 % from other T. tabaci COI sequences (598 sequences available in GenBank), which suggests a possible misidentification of a T. origani specimen labelled as having been collected in Canada.
The presence of T. origani in Argentina was reported to Sistema Nacional de Vigilancia y Monitoreo (SINAVIMO) of Servicio Nacional de Sanidad y Calidad Agroalimentaria (SENASA), Identificaction number 1306.
redalyc-journal-id: 3220
This work was funded by the Mendoza and La Consulta Agricultural Experimental Stations of the Instituto Nacional de Technología Agropecuaria (INTA). We thank the anonymous reviewers and the editor of this journal.
deborbon.carlos@inta.gob.ar

a-c female adults’ full body (a) T. australis (b) T. trehernei (c) T. origani; (d) head with ocellar setae pair I absent, ocellar setae III subequal to or longer than ocellar setae II and pronotum with median pair of setae on posterior margin longer than submedian pair of T. origani; e-f metanotum (e) T. australis, campaniform sensilla present (f) T. simplex, reticles with internal markings.

a-c sternites VI-VIII (a) T. origani, sternites VI with few discal setae and VI without discal setae (b) T. tabaci, sternites VI-VII without discal setae (c) T. trehernei, sternites VI -VII with many discal setae; d-e. pleurotergites (d) T. trehernei, rows without microtrichias (e) T. origani, rows with microtrichias, f-h forewings setae distribution (f) T. origani, (g) T. australis (h) T. trehernei; i-k antenna, number of segments and colour (i) T. origani (j) T. trehernei (k) T, simplex, antenna 8-segmented.

