Description of immature stages of Xestia brunneopicta ( Matsumura , 1925 ) , with a key to the mature larvae of the European species of Xestia ( Pachnobia ) ( Lepidoptera , Noctuidae )

Immature stages of Xestia brunneopicta (Matsumura, 1925) are described and illustrated from an ex ovo rearing. The female was collected during a Finnish-Russian expedition to the province of Chita in East Siberia in 2013. Eggs were laid in a plastic jar at Chara Sands on the 7th of July. Larvae hatched between the 20th and 21st of July. Rearing of larvae was undertaken in Finland by four lepidopterologists. A key is given that includes the known European larvae of the subgenus Pachnobia Guenée, 1852 sensu Lafontaine et al. (1998), mostly based on the morphology and larval chaetotaxy. The closest relatives on the basis of larval morphology are discussed.


Introduction
The early stages of Xestia brunneopicta (Matsumura, 1925) have remained undescribed in spite of its wide distribution from Magadan to the East-Siberian Tuva in Russia (Kononenko 2005) and rarely also in Kuusamo, Finland (Mikkola et al. 1989).Ahola and Silvonen (2011) described larvae found in nature in 1982 (Kuusamo) and 1992 (Kuusamo, Kuhmo) as X. brunneopicta on the basis of their differences in chaetotaxy and habitus compared to those of X. gelida (Sparre-Schneider, 1883).However, through the recent findings, we can now state that the larvae of X. gelida and the real X.brunneopicta are very distinct from each other, particularly in their outer appearance, as shown in this paper.The Finnish larvae formerly reported as Xestia brunneopicta are now re-identified as those of X. gelida even though some of their characters differ slightly from other individuals of X. gelida examined.Egg, larva and pupa of X. brunneopicta are described and illustrated.

Materials and methods
The larval material originates from a female collected during an expedition to the province of Chita in East Siberia, N56.87133, E118.18302, at an elevation of 750 m.a.s.l. on 7.vii.2013by Hannu Saarenmaa.Seventy-two eggs were laid on the needles of Larix gmelini Rupr.(Pinaceae) in a plastic jar at Chara Sands 7 -10 July.The rearing of eggs was carried out in Finland by HS and 60 larvae hatched after two weeks on 20-21 July.All larvae were reared under different lamps from 18 to 24 hours of light every day, because the northern larvae grow faster in continuous daylight.Twenty larvae grew to maturity during autumn and went into diapause instead of pupating.We follow Beck (2000) in descriptions of cuticular ornaments.Two larvae were prepared by dry inflating.Larval chaetotaxy nomenclature follows Hinton (1946) while pupal follows Patočka and Turčáni (2005).The hypopharyngeal complex, mandibles and labrum were dissected and preserved on a slide to study the morphology.

Descriptions of immature stages
Egg (Fig. 6): The eggs were laid on the needles of Larix gmelini.Shortly after laying they were whitish grey, but fertile eggs darkened in two days.The micropyle area became dark reddish brown, and the narrow zone around the micropyle was reddish brown Morphology of full-grown larva (Figs 1-5): Spinneret flat, 2.5× as long as wide, ventral lip straight, dorsal lip short-fringed, longitudinal grooves present on dorsal surface.Base segment of labial palp (Lps1) about 2× as long as wide, second segment (Lps2) 1/5 as long as Lps1, labial palp seta Lp1 slightly longer than Lps2, seta Lp2 shorter than Lps1 (3/5) and 2× as long as Lp1.Hypopharynx with long spines on anterior surface above spinneret, distolateral spines slightly longer and stouter than distomedian spines, spines on lateroposterior part forming row of 8-10 differentiated, triangular teeth, lateral surface above this row densely covered with tiny spinules, posterior area of medial part bare.Stipular setae below spinneret shorter than seta Lp2 of labial palp, situated in front of prementum.Mandible with two setae on outer surface, six teeth on cutting margin, three ridges on inner surface terminating in low protuberances before cutting margin and triangular tooth on first ridge.Maxillary palp three-segmented, second segment longer than galeal lobe, sensillum styloconicum of galeal lobe as long as end segment of maxillary palp, three sensilla trichodea present.Labrum with low, rounded notch.Epicranial suture slightly shorter than height of frons.Six stemmata present, distance between second (Oc2) and third (Oc3) stemma greater than those between Oc1-Oc2 or Oc3-Oc4, distance Oc1-Oc2 greater than Oc3-Oc4.Abdominal prolegs on abdominal segments 3-6 (Ab3-6) equal in size, crochets uniordinal, 17-20 on Ab3, 22-24 on Ab6 and 26-28 in Ab10.Body without warts or other protuberances.
Chaetotaxy resembling that of other members of Pachnobia: Setae of head and body rather long when compared to the height of the spiracle on Ab8: P1 on head 3.1-3.3×,D2 on Ab2    1.
Third instar larva: Length about 7-9 mm.Head brown, stripes visible but weak.Shields pale brown, dorsal lines yellowish, not sharp-edged on shields.Dorsal zone green with white, narrow, middorsal and subdorsal lines without darker margins, setal bases small, pale green.Subdorsal zone dark olive green, spiracular line yellowish white, broad.Pleural and ventral zones pale green (Fig. 9).
Fourth instar larva (Fig. 10): Length 14-18 mm: Head brown, netfields visible but translucent, frons and anterior zone green, brown setal points on brown bases.Thoracic and anal shields pale brown, dorsal and subdorsal lines present as yellowish white flecks on prothoracic shield but absent on anal shield.Body green on dorsal region with pale green or with whitish elements, yellowish green between abdominal segments, dark green on ventral subdorsal zone.Middorsal line nearly white, broad, continuous, subdorsal line slightly narrower, broken into spots.Spiracular line broad, yellowish white, sharply bordered above dark green ventral subdorsal zone.Ventral region pale green.Setal points black on dorsal region with dark green dorsal/whitish ventral bases.
Penultimate and last instar : Length of last instar larva 35-40 mm.Head brown, stripes brown, reticulate structure with brown bands and pale brown, weakly visible netfields.Frons and anterior zone greenish, adfrons brown, ocellar zone pale yellowish brown, setal points dark brown.Prothoracic shield darker green than body, with narrow pale grey middorsal line, subdorsal line not visible, shield caudally bordered with narrow, blackish grey colour.Anal shield greenish brown, lines not visible, sutures brown, setal points blackish brown.Dorsal and ventral regions of body green, middorsal and subdorsal Table 1.Relevant distances between setae of larva of Xestia brunneopicta.Ab = abdominal segment.

Notes on natural environments
The collecting site of the female in Chara Sands is a peculiar dune habitat with occasional springs, bogs and coniferous tree patches.It does not represent the typical habitat of the species.).X. brunneopicta was common across all sites, but it seemed to be more frequent in the lowlands than in the mountain valleys.Typical habitat for the species is forested bog with Larix gmelini (Fig. 15).However, it was most numerous in low Salix and Alnus vegetation on the banks of the Chara River.Other species typical for these localities include several other Xestia (Pachnobia) species such as X. atrata (Morrison, 1874), and also Polia altaica (Lederer, 1853), P. conspicua (A.Bang-Haas, 1912), P. vespertilio (Draudt, 1934), P. vesperugo Eversmann, 1856, and the arctiine Borearctia menetriesii Eversmann, 1846.

Key to the larvae of subgenus Pachnobia
Larvae of the subgenus Pachnobia differ from subgenera Megasema and Xestia in chaetotaxy and morphology.Dorsal setae are long in Pachnobia, seta D2 on Ab8 is more than twice as long as height of spiracle of same segment, whereas it is about as long as height of spiracle in subgenera Megasema and Xestia.Also, setal distances P1-P2 (head) and V1-V1 (Ab7) differ on average (Table 2).Spinneret of larvae in subgenus Pachnobia is long, about 1.5-5.0×as long as wide, flat (except X. liquidaria), dorsal margin with short fringes (without fringes in X. liquidaria) and ventral margin straight.Megasema and Xestia larvae have a short spinneret, 1.0-1.5×as long as wide (except 2-3× in X. collina, castanea and agathina), dorsal margin with longer fringes and ventral margin more or less bilobed.Body colour and pattern vary a lot.Larva of X. liquidaria is peculiar with a tubular spinneret but long dorsal setae as in subgenus Pachnobia.

Discussion
The appearance of the larval stages of X. brunneopicta differs greatly from those of X. gelida and X. fabulosa (Ferguson, 1965).This is a surprise considering the adult of X. brunneopicta has been considered to be closely related to them (Lafontaine et al. 1998).The larva of X. fabulosa is similar to that of X. gelida and therefore we supposed that the larva of X. brunneopicta could resemble this species as well.Ahola and Silvonen (2011) described larvae close to X. gelida as possible X.brunneopicta from Kuusamo and Kuhmo in Finland.These three larvae differ from X. gelida in having a paler dorsal zone, shorter visible part of the middorsal line, the subdorsal line is broken into flecks, has a narrower black dorsal margin of subdorsal line, is not enlarged on Ab7-8 and has a wider white dorsal part of the spiracular line.Also, setal positions are slightly different, and SD1 is more distant from the spiracles.However, we now rather see these differences as variation in X. gelida.Many Noctuidae have green larvae.For example, in Europe there are more than 140 species with such larvae.Larvae with green bodies and narrow white dorsal and subdorsal lines are not so common, but still about 40 species have such larvae.However, a quarter of them can occur in the same northern areas with X. brunneopicta.In Finland larvae of Orthosia gothica (Linnaeus, 1758) and O. incerta (Hufnagel, 1766) resemble those of X. brunneopicta.The brown head and position of the spiracles above the spiracular line on Ab7 separates X. brunneopicta readily from Orthosia species.Xestia includes also two European species with green larvae, namely X. ochreago (Hübner, 1790) and some variations of X. castanea (Esper, They have, however, short dorsal setae, and the head is green. DNA barcodes of X. brunneopicta differ from those of X. gelida and X. fabulosa by a minimum of 6.47% and 6.73% genetic distance, respectively (Marko Mutanen, pers.comm.), also suggesting that X. brunneopicta is perhaps not a very close relative of these species.Based on the COI sequences from one Finnish and one Russian specimen of X. brunneopicta, the closest relatives of X. brunneopicta are X.lorezi (4.44%), X. sincera (4.60%) and X. ursae (McDunnough, 1940) (4.61%), but many other Xestia species show less than 6% divergence as well.Based on DNA barcodes, no other Xestia species is a very close relative of X. brunneopicta, and based on both larval morphology and DNA barcodes, its sister species remains unclear.

Table 2 .Figure 15 .
Figure 15.Typical habitat of X. brunneopicta and the other species mentioned in the article.Chara River with richer vegetation on its banks is 100 meters to the left (Photo: H. Saarenmaa).