Orobanche arpica Piwow., Ó. Sánchez & Moreno Mor., 2018

Piwowarczyk, Renata, Pedraja, Óscar Sánchez, Moral, Gonzalo Moreno, Góralski, Grzegorz, Kwolek, Dagmara, Denysenko-Bennett, Magdalena & Pawełek, Barbara, 2018, Orobanche arpica (Orobanchaceae): a new species from the Caucasus, and contributional notes on O. ser. Krylowianae, Phytotaxa 361 (1), pp. 65-76 : 66-74

Orobanche arpica Piwow., Ó. Sánchez & Moreno Mor. , sp. nov. ( Figs. 1 , 2 )

Type: — ARMENIA. Shirak prov.: Javakheti (Javakhq) range (Lesser Caucasus), E of villages Ghazanchi – Ashotsk, subalpine meadow, NW exposure, 2225 m, 22 July 2017, R. Piwowarczyk s.n. (holotype KTC!, isotype ERCB!).

Description: —Plant holoparasitic, achlorophyllous, pale yellow or dirty pink. Stem simple and ± slender, (20–)27– 30(–35) cm × (3.0–)4.0–5.0(–6.0) mm in diameter in the upper part, (5.0–)6.0–7.0 mm in the middle, and (6–) 7–8 mm towards the base, this slightly widening or bulbous; slightly striate (clearly striate when dry); glandular pubescent especially in the upper part, with whitish or pale orange glandular hairs 0.2–0.3(–0.4) mm; stem pale white-yellow or rarely pale yellow-brown-pink (brown when dry). Basal leaves (17–)23–24(–25) mm × (4–) 5–6 mm, lanceolate, glabrous abaxially, or shortly ciliate at the edge with hairs ca. 0.2–0.3 mm. Upper leaves (15–)20–21(–25) mm × 6–7(–8) mm, lanceolate, becoming sparse above, pale yellow-brown, pinkish-brown, changing early to brown when drying, especially at the top; sparsely glandular pubescent, especially at the edge, with hairs 0.2–0.4 mm. Inflorescence (10–)12–13(–15) cm × 3.5–4.0 cm, cylindrical, shorter than the remaining stem; (12–)20–23(–26)-flowered, usually dense or rarely lax. Flowers are ± erecto-patent and medium sized. Bracts (16–)20–23(–26) mm × 3.5–4.0 mm, shorter or rarely equal to the corolla, broad ovate-lanceolate, pale, yellow-pink or pinkish-brown, changing early to brown when drying, with dense white or pale yellow glandular hairs, 0.2–0.3 mm long. Bracteoles absent. Calyx (11–)13–14(–15) mm long, (2.5–)3.0–4.0 mm wide at the widest point, shorter than half of the corolla tube; two segments, free, clearly separate, simple, entire, or rarely unequally bidentate, with second small teeth, narrow, with teeth long, acuminate almost filiform at apex and with bases gently ovate, pale yellowish-pink, with dense glandularhairy, hairs ca. 0.2–0.3(–0.4) mm, white or pale yellow. Corolla (22–)24–26(–28) mm long, (8–)9–10(–11) mm in diameter in the central part; tubular to narrowly campanulate (slightly widening towards the mouth), strongly inflated above the insertion of the filaments, and strongly constricted below; the dorsal line slightly curved in the lower part, less curved or nearly straight in the middle part and ± flexed forwards or straight at the apex, externally glandular-pubescent with white or pale yellow (in dry form with orange glands), glandular hairs of 0.1–0.2(–0.3) mm, more or less abundant, hair cover slightly denser at upper lip; corolla pale ± pinkish-yellow, pinkish-white or yellowish-pink, with dirty pale pink veins, very sensitive to damage and browning; upper lip slightly emarginate, with two broad lobes, these usually facing downward, more rarely slightly bent backward laterally, sparsely glandular hairy also in the inner part, hairs ca. (0.1–) 0.2–0.3 mm; lower lip with three oval, or ± rectangular to spathulate lobes, central lobe slightly larger than lateral lobes, lobes slightly and irregularly dentate on margins and sparsely glandular hairy also in the inner part. Stamens obliquely inserted, adaxial at 5–6 mm above the corolla base, and abaxial at 4–5 mm, all them slightly widened at base. Filaments 16–18 mm long, 1.5 mm wide, white to pale yellow or rarely pale pink, clearly geniculate, entire densely villous; from base to ± middle part, hairs 0.8–10 mm long, not glandular, white or white-yellowish, upper part densely glandular-pubescent, usually ca. 0.1–0.2 mm long, short, white with orange glands. Anthers 2.5–3.0 mm long, ca. 1.5 mm wide, oblongoid, mucronate, pale brown; basally and along ¾ of suture distinctly and numerous hairy (ca. 0.2–0.4 mm). Ovary 8–9 mm × 3.5–4.0 mm, glabrous, pale, yellowish or pinkish; lateral opening by two longitudinal slots. Style 12–15 mm long, pale yellow to whitish, glandular-pubescent, with short and glandular hairs (these ca. 0.1–0.2 mm long and more abundant, conspicuous and dense on the upper part). Stigma bilobed, lobes elongated-spherical to ovate and ± flattened, with numerous warts (orange when dry) on the lobes, and yellow or bright yellow. Seeds usually oblongoid; seed coat reticulate with polygonal shallow cells, which range from more or less isodiametric to irregular; seed coat is pitted. Pollen inaperturate, spheroidal.

Distribution and habitat: ―So far, the species is known only from the type locality, in the northwestern part of Armenia, in the Shirak province, 30 km NNE of Gyumri, 4 km SEE of a place between the villages of Ghazanchi and Ashotsk. Orobanche arpica grows on the SWW slope of Mt Ampasar (3046 m), with NW exposure, 2225–2230 m a.s.l., in a subalpine meadow, in a neovolcanic area with chernozem soil ( Fig. 2 ).

Phenology: ―Flowering in the first half of July, fruiting in end of July–August.

Ecology: ―Parasitic on Psephellus pulcherrimus (Willd.) Wagenitz [ Centaurea pulcherrima Willd. ; Aetheopappus pulcherrimus (Willd.) Cass. ] ( Asteraceae ) ( Fig. 2 ), host attachments were verified. This species is native to the Caucasus and Turkey.

Etymology: ―The epithet ‘ arpica ’ derives from the name of Lake Arpi in Arpi Lich National Park, nearby which the new species was discovered.

Conservation: ―The new species grows outside of, but very close to the SW border of the protected areas of Arpi Lich National Park. The population is medium-sized (ca. 30 individuals), habitats are quite stable, however in some places there is overgrazing.

Phylogenetic studies: ―Both ITS and trnL-trnF ( Figs. 3 , 4 ) trees show that O. arpica is distinct from the rest of the species studied. The branches containing O. arpica sequences in most cases have strong bootstrap/posterior probability support. For trnL-trnF trees, the bootstrap on the Maximum Likelihood tree equals 99 and the posterior probability value on the Bayesian tree reaches 1.00. On ITS trees, these values equal 85 and 0.76 respectively. The last value seems to be rather weak, however in that case the separate common branch containing the rest of the Krylowianae series is supported by 1.00 Bayesian posterior probability. Generally, the closest neighbours of O. arpica on all trees are species belonging to the series Krylowianae , however there is one difference between ITS and trnL-trnF trees. On the ITS tree, there is a common branch of O. arpica and all the O. ser. Krylowianae species ( O. inulae , O. krylowii , O. lycoctoni and O. mlokosiewiczii ) with support by strong bootstrap/posterior probability values (100, 1.00). The trnL-trnF trees show a slightly different view: sequences of O. arpica and most of Krylowianae species ( O. inulae , O. krylowii and O. mlokosiewiczii ) form a common branch with strong support (98, 1.00) but O. lycoctoni is placed close to such species as O. lucorum , O. reticulata , O. flava and O. alba . ITS sequences of O. arpica differ from O. mlokosiewiczii and O. inulae by three substitutions, from O. krylowii in 8 sites (substitutions and indels) and much more from O. lycoctoni (46 indels and substitution sites). Differences in trnL-trnF sequences between O. arpica and other Krylowianae members comprise: three substitutions ( O. mlokosiewiczii ), five substitutions ( O. krylowii ), four substitutions and one indel ( O. lycoctoni ) and six substitutions ( O. inulae ). The number of substitutions between all Krylowianae species for both sequences (except trnL-trnF from O. lycoctoni ) do not exceed six. Such similarity suggests that all these species, including O. inulae , whose phylogenetical position was previously unclear ( Piwowarczyk et al. 2017a), are closely related and including them in one series is reasonable.

Contribution to the O. ser. Krylowianae : ―The three Turkish specimens, with inflorescence few-flowered, calyx segments shortly bidentate (wide-ovate at the base and triangular-lanceolate acute teeth shorter than the tube) and corolla with straight dorsal line largely and helmet-shaped at the apex, of the sheet P02968068 ( Sánchez Pedraja et al. 2016 +), correspond, from our point of view, to a species of the O. subsect. Orobanche (O. trib. Galeatae sensu Beck), probably small specimens of O. caryophyllacea , of course, assuming that Balansa mistook the host plant, because O. teucrii , an exclusive parasite of Teucrium species, has never been seen in Turkey. These specimens do not seem to be related to O. inulae , as it has been presented in the work of Rätzel & Uhlich (2017), which is usually characterized by its calyx teeth being entire and corolla not helmet-shaped at the apex ( Novopokrovskij & Tzvelev 1958; Sánchez Pedraja et al. 2016 +).

We also present supplements to the distribution of O. mlokosiewiczii , species described from Georgia ( Piwowarczyk et al. 2017a). A new locality was found in the herbarium collection from Russia – Caucasus, Stavropol Territory, Karachay-Cherkessia, Kabardino-Balkaria, Teberdinsky Reserve, Malaya Hatipara Mt, a high grassy glade, on Aconitum [roots attached to parasite], 11 August 2006, A.S. Zernov (MW0723406!) ( Seregin 2018, sub O. alsatica ).

Discussion: ―The newly-described species of the O. subsect. Curvatae (Beck) Piwow., Ó. Sánchez & Moreno Mor. ( Piwowarczyk et al. 2017a) is very characteristic and different from other known Orobanche , however clearly belongs morphologically and phylogenetically to species from the O. ser. Krylowianae Piwow., Ó. Sánchez & Moreno Mor. ( Piwowarczyk et al. 2017a). Morphological similarity is observable, e.g., in the habit and pale yellowish colour of the plant, and the strong narrowing of the corolla in the place of insertion of the filaments. However, O. arpica differs very clearly in morphology of flowers and host (see key and Table 2) from these species. The new species may be confused with O. inulae , but differs clearly, with such features as: longer inflorescence, scales, bracts, and corolla; shape of corolla – tubular to narrowly campanulate; shape of lobes; and in particular a much higher filaments insertion; as well as host species (see key and Table 2). As far as we know, this is the first report of Psephellus pulcherrimus [ Aetheopappus pulcherrimus ] as host of any Orobanche s.l. species. Of course, we already knew that O. kochii can parasitize other species of the genus Psephellus [e. g., Psephellus sibiricus , syn. Centaurea sibirica ] ( Plantarium, 2011 +; Sánchez Pedraja et al. 2016 +), but this species is completely different in all aspects (morphological and genetic).

and O. cicerbitae ; following Novopokrovsky & Tzvelev (1958), Carlón et al. (2002, 2005, 2008), Rätzel & Uhlich (2004),

Frajman et al. (2013), Uhlich (2015), Rätzel et al. (2016), Piwowarczyk et al. (2017a) and the authors’ own studies in herbaria and in the field.

Species

O. arpica O. inulae O. mlokosiewiczii O. lycoctoni O. krylowii O. cicerbitae Character Host Asteraceae : Asteraceae : Inula Ranunculaceae : Ranunculaceae : Ranunculaceae : Asteraceae : Psephellus aspera [ I. cordata ], Aconitum Aconitum Thalictrum minus Prenanthes pulcherrimus I. orientalis [I. cymbulatum lycoctonum petiolata [ Cicerbita glandulosa!, incl. petiolata ] (Rätzel I. grandiflora ], I. & Uhlich 2004). magnifica Senecio propinquus ( Rätzel et al. 2016) Distribution Lesser Caucasus: Greater Caucasus: Greater Caucasus: S W Europe to C S E Europe Greater Caucasus: Armenia Azerbaijan?, Georgia, Russia Europe: Cantabrian (Albania) Adygaea (Rätzel Georgia, Russia, S Mts. (Spain), and E Europe & Uhlich 2004) Ossetia, Endemic Alps (Austria, (Russia) to Asia: and Azerbaijan France, Germany, China?, Russia, ( Rätzel et al. 2016) Italy, Slovenia, Kazakhstan, Endemic Switzerland) Kyrgyzstan, Mongolia

Phylogenetic studies support the supposition that the studied specimens belong to a new species ( O. arpica ) and that it should be taxonomically ordered into the ser. Krylowianae . ITS and trnL-trnF trees indicate the close relation of O. arpica , as well as O. inulae to the rest of the Krylowianae species ( O. krylowii , O. mlokosiewiczii and O. lycoctoni except trnL-trnF tree) but O. arpica seems to be more distant to them than these species differ from each other. Comparisons of DNA sequences of all Krylowianae species show close relations between them (with the exception of trnL-trnF from O. lycoctoni ) and probably these species diverged relatively recently. This hypothesis seems to be supported by the fact that all of these species live in high mountains. Such environments typically contain a lot of small, isolated ecosystems, which promote endemism and speciation events ( Steinbauer et al. 2016). Consequently, species from the series Krylowianae can be an example of recent rapid radiation of mountain Orobanche species, however this assumption needs to be verified in future studies. These results also seem to resolve the problem of taxonomic relationships of O. inulae reported previously ( Piwowarczyk et al. 2017a). Phylogenetic analysis indicates that it belongs to the O. ser. Krylowianae .

Another interesting phenomenon that requires closer investigation is the placement of trnL-trnF sequence of O. lycoctoni on phylogenetic trees near O. lucorum , O. reticulata , O. flava and O. alba that are relatively distant to the other Krylowianae species. This may be a sign of some interesting genetic phenomena, such as gene flow caused by interspecies crosses, but it cannot be confirmed at the current state of the studies.

Identification key (modified from Frajman et al. 2013 and Piwowarczyk et al. 2017a)

1 Plant always yellowish or whitish; upper lip ± porrect (continuing the dorsal line of the corolla tube) or flexed forwards............. 2

- Plant usually strongly pigmented, rarely yellowish-white; upper lip erect or recurved..................................................................... 3

2 Corolla tubular to narrowly campanulate (slightly widening towards the mouth), dorsal line flexed forwards or straight at the apex; on Psephellus pulcherrimus ( Asteraceae ) .......................................................................................................................... O. arpica *

- Corolla broadly tubular-campanulate to campanulate (conspicuously widening towards the mouth); dorsal line ± porrect at the apex; on other species......................................................................................................................................................................... 4

4 Bracts usually longer than the corolla; calyx segments 10.4–16.8 × 3.5–7.8 mm, usually unequally bidentate; style glabrous or nearly; on Aconitum lycoctonum ( Ranunculaceae ) .......................................................................................................... O. lycoctoni

- Bracts usually shorter than the corolla; calyx segments 6–15 × 1.5–5 mm, usually entire or more unequally bidentate; style hairy; on other species .................................................................................................................................................................................. 5

5 Stem usually densely scaly at the base, but with few and scattered scales on the rest. Calyx segments 10–15 × 3–4 mm, c. 3/4 corolla length, lanceolate; on Inula sp. ( Asteraceae ) ......................................................................................................... O. inulae *

- Stem usually with few and scattered scales. Calyx segments 6–12 × 1.5–2.5 mm, c. 1/2–2/3 or less corolla length, ovate-subulate or ovate-lanceolate; on other species.................................................................................................................................................. 6

6 Corolla 17–21 mm, tubular; dorsal line gently and regularly curved (but straight near the apex); lobes with finely eroded-denticulate margin; on Thalictrum minus ( Ranunculaceae )................................................................................................................. O. krylowii

- Corolla (17–)20–23(–25) mm, tubular to campanulate; dorsal line slightly or strongly curved (but ± deflexed at the apex); lobes with irregularly dentate or finely eroded-denticulate and glandular margin; on other species .......................................................... 7

7 Corolla (17–)20–23(–25) × (7–)10–12(–14) mm, usually campanulate or more rarely tubular-campanulate, dorsal line slightly curved (but straight at the apex); lobes oval or rectangular to triangular, with irregularly dentate margin; on Aconitum cymbulatum ( Ranunculaceae ) ...................................................................................................................................................... O. mlokosiewiczii

- Corolla (21–)22–23(–25) × (6.8–)7.0–7.8(–8.3) mm, tubular, strongly curved (but ± deflexed at the apex); lobes spathulate rounded, with finely and irregularly denticulate and glandular margin; on Prenanthes petiolata ( Asteraceae )............................ O. cicerbitae

3 Corolla with distally straight dorsal line, with a distinctly flattened subapical area; lower lip cruciform, with concave, spathulate, very divergent lobes; on Centaurea sp. [s.l.], Echinops sp. and Ptilostemon sp. ( Asteraceae )............................................. O. kochii

- Corolla with regularly curved dorsal line; lower lip with less divergent, elliptic or rounded and flat to convex lobes..................... 8

8 Corolla variably (orange, reddish, yellow, yellowish-brown), but usually vividly pigmented; filaments hairy almost to the apex; on Petasites sp. ( Asteraceae ) ....................................................................................................................................................... O. flava

- Corolla brownish, purple or reddish to yellowish brown, rarely yellow, but usually feebly pigmented; filaments hairy at most in its lower two thirds.................................................................................................................................................................................. 9

9 Plants up to 85 cm, tall and stout; inflorescences long, cylindrical, often covering the upper two thirds of the stem; corolla with dorsal line regularly curved, sometimes more or less deflexed at the apex; upper corolla lip shallowly emarginate; exclusively on Centaurea sp. ( Asteraceae ) ......................................................................................................................................... O. elatior s. str.

- Plants up to 60 cm, usually shorter and more slender, with relatively narrower stems; oval inflorescences, dense, at most covering the upper half of the stem; corolla with regularly and strongly curved dorsal line, but somewhat straight at the apex; upper corolla lip more deeply and distinctly emarginate; on Apiaceae (mainly on Peucedanum sp. , Seseli sp. ).... O. alsatica (incl. O. bartlingii )

*Note to the Identification key:

Stem usually with few and scattered scales. Inflorescence (12–)20–23(–26) flowered. Bracts (16–)20–23(–26) mm. Corolla (22–)24– 26(–28) mm, tubular to narrowly campanulate (slightly widening towards the mouth), dorsal line flexed forwards or straight at the apex. Stamens inserted 4–6 mm above the corolla base. Parasitic on Psephellus pulcherrimus (Tribus Cardueae Cass., Familia Asteraceae ) ...... .......................................................................................................................................................................................................... O. arpica Stem usually densely scaly at the base, but with few and scattered scales on the rest. Inflorescence 7–13 flowered. Bracts 13–16(–18). Corolla (15–) 17–20 mm, broadly tubular-campanulate to campanulate (conspicuously widening towards the mouth); dorsal line ± porrect at the apex; on other species. Stamens inserted 1.5–3 mm above the corolla base. Parasitic on Inula sp. (Tribus Inuleae Cass., Familia Asteraceae ) ....................................................................................................................................................................................... O. inulae

Figures

FIGURE 1. Orobanche arpica. A. Plant.B–D. Flower, side view.C. Flower,front view.E. Bract. F.Leaf.G. Open corolla and androecium. H. Calyx. K. Anther. I. Gynoecium. J. Stigma. Illustration by Jolanta Urbanik.

FIGURE 2. Orobanche arpica. A. General habit. B. Flower. C. Inflorescence. D. Habitat with host species Psephellus pulcherrimus. E. Flower of host species. Photos by Renata Piwowarczyk.

FIGURE 3. Rooted a) Maximum Likelihood and b) Bayesian phylogenetic trees constructed for ITS sequences. As an outgroup, Phelipanche bohemica was used. Numbers near branches show bootstrap and Bayesian posterior probabilities respectively. The bars represent the amount of genetic change (nucleotide substitutions per site). Species belonging to series Krylowianae are marked by gray fields.

FIGURE 4. Rooted a) Maximum Likelihood and b) Bayesian phylogenetic trees constructed for trnL-trnF sequences. As an outgroup, Phelipanche bohemica was used. Numbers near branches show bootstrap and Bayesian posterior probabilities respectively. The bars represent the amount of genetic change (nucleotide substitutions per site). Species belonging to the series Krylowianae are marked by gray fields.