Capsicum benoistii Hunz. ex Barboza, 2019

Barboza, Gloria E., García, Carolina Carrizo, González, Segundo Leiva, Scaldaferro, Marisel & Reyes, Ximena, 2019, Four new species of Capsicum (Solanaceae) from the tropical Andes and an update on the phylogeny of the genus, PLoS ONE (e 0209792) 14 (1), pp. 1-26 : 4-23

Capsicum benoistii Hunz. ex Barboza , sp. nov. [ urn:lsid:ipni.org:names: 77192556–1]. Type: Ecuador. Tungurahua: Baños , 3 Apr 1931 (fl), M. R. Benoist 4204 (holotype, P) .

Fig 1

Diagnosis. Similar to Capsicum geminifolium (Dammer) Hunz. but differing in the length of the flowering pedicels, the shape of the corolla, and the presence of heterostylous flowers.

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Description. Shrubs few branched. Young stems light brown, glabrous or pubescent, striate; bark of older stems dark brown, glabrous, striate; lenticels absent. Sympodial units difoliate, geminate, leaf pair markedly anisophyllous in size and shape. Leaves simple, membranaceous, discolorous, adaxial surface dark green, abaxial surface light green, glabrous or with simple antrorse trichomes 0.3–1.1 mm long adaxially and abaxially, trichomes more abundant on main veins; the larger leaves with blades 8.5–12 cm long, 2.8–6 cm wide, ovate or elliptic, major veins 4–5 (6) on each side of midvein, base asymmetric and attenuate, margin entire, apex long-acuminate; petioles 0.5–1.0 cm long, glabrous or glabrescent with trichomes like those of the leaves; the minor leaves 2.4–6 cm long, 1.7–4 cm wide, ovate or elliptic, major veins 3–4 on each side of midvein, base rounded, asymmetric, margin entire, apex acute or rounded; petioles 0.1–1 cm long, glabrescent or pubescent. Flowers in fascicles of 3–6; flowering pedicels filiform, striate, pendent, not geniculate at anthesis, 1.3–2 cm long, moderately to densely pubescent, the trichomes simple, non-glandular, multicellular, antrorse, 0.30–0.75 mm long. Flower buds ovoid. Calyx 2–2.5 mm long, ca. 5 mm wide, cup-shaped, thick, the margin truncate, pubescent with the same trichomes as pedicels, with 5 appendages 2.5–3.5 mm long, ca. 0.5 mm wide, thick, erect, subulate, inserted close to the margin, pubescent with the same trichomes as calyx tube. Corolla ca. 12–13 mm long, deeply stellate, thick, without interpetalar tissue; tube ca. 3 mm long, glabrous inside and outside; lobes ca. 9 mm long, ca. 2 mm wide, narrowly triangular, erect, glabrous adaxially and abaxially, the tips and margins pubescent. Stamens 5, equal, filaments equal, 3–3.2 mm long, glabrous, inserted on the corolla 1.5 mm from the base, with inconspicuous auricles at point of insertion; anthers ca. 3 mm long, not connivent, elliptic. Ovary 1.3–1.7 mm long, 1.2–1.5 mm diam, subglobose, glabrous; nectary ca. 0.3 mm long, inconspicuous; long style ca. 6.5 mm long, short style ca. 3.6 mm long, widening distally, glabrous; stigma 0.3 mm long, 0.5 mm wide, globose. Berry unknown.

Distribution and ecology. Endemic to a restricted area in central-southern Ecuador (Tungurahua, Loja, Fig 2 ) growing in thickets in montane forests, between 1500–2600 m elevation.

Phenology. Flowering from March to May. Fruiting time unknown.

Etymology —The new species is named in honor to Raymond Benoist (1881–1970), a French botanist, who collected in French Guyana, Morocco and Ecuador; the holotype is a nice specimen collected by her in 1931.

Species Conservation Assessment. Following the IUCN Red List Criteria (IUCN 2017), this species is proposed as Endangered ( EN). The extent of occurrence is calculated to be 2050 km 2 (Criterion B1 <5000 km 2, Endangered), the area of occupancy, 12 km 2 (Criterion

B2 <500 km 2, Endangered) and the species is known from only three localities (Criterion

B1a � 5, Endangered). It is possible that its geographic range has declined (EOO and AOO, Criterion B2b i & ii) because the species has not been collected since 1978 despite recent intensive searches in the same locations.

Additional specimens examined. ECUADOR. Loja: Pueblo Nuevo , 04˚05’51’’S, 79˚ 11’55’’W, 2580 m, 21 May 1978 (fl), F. Vivar & Estudiantes 1066 (LOJA) ; Tungurahua: Río Verde Grande , 1500 m, 30 Mar 1956 (fl), E. Asplund 20070 (S) .

Capsicum benoistii was identified as a new species by the late Solanaceae specialist Armando T. Hunziker ( CORD) who annotated the epithet name benoistii on the specimen housed at P (Benoist 4204), but this name was never published. It is a poorly known species collected only three times in Ecuador; none of these collections have fruits. Extensive recent field explorations in Tungurahua were unsuccessful in finding this species. It is distinctive in its deeply lobed stellate corolla (lobes three times longer than the tube, Fig 1B) and in the presence of heterostylous flowers ( Fig 1G and 1H). These features plus the short flowering pedicels (1.3–2 cm long) distinguish C. benoistii from C. geminifolium , which has funnel-shaped corollas lobed about halfway, homostylous flowers, and longer pedicels (5 cm long).

The presence of heterostylous flowers as in C. benoistii is unusual among Capsicum species. It has been reported in C. baccatum L. varieties: var. baccatum [ 29] and var. umbilicatum (Vell.) Hunz. & Barboza [ 30, 31], and observed in other species ( C. tovarii Eshbaugh, P.G.Sm. & Nickrent and C. pubescens Ruiz & Pav., Barboza pers. obs.). This character deserves careful field observations to ascertain if short-styled flowers produce fruits.

As some data are still unknown (e.g. corolla color, fruit and seed characters, and chromosome number) for this species and freshly collected leaf material is not available for DNA extraction, we cannot suggest in which of the different clades of the current phylogeny of Capsicum [ 1] it could be placed.

Capsicum longifolium Barboza & S. Leiva , sp. nov. [ urn:lsid:ipni.org:names: 77192558–1]. Type: Ecuador. Zamora-Chinchipe: Area of Estación Científica San Francisco, road Loja- Zamora , ca. 35 km from Loja, transect Q2, 03˚58’S, 79˚04’W, 1900 m, 12 Jun 2005 (fl, fr), F. A. Werner 1548 (holotype, QCA [QCA-160608]; isotypes, LOJA, NY [NY-01130066]) .

Figs 3 and 4

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Diagnosis. Similar to Capsicum dimorphum (Miers) Kuntze but differing in the long and narrow coriaceous leaves, the number of flowers (3–9), the unequal thick calyx appendages, and the glabrous vegetative organs and calyx.

Description. Scandent shrubs (0.60) 1.40–3 m tall, laxly branched, slightly plagiotropic. Young stems green, fragile, glabrous, striate with abundant ovoid and light dark lenticels; bark of older stems dark green, glabrous, striate with sparse white lenticels. Sympodial units difoliate, geminate, the leaf pair markedly anisophyllous in size and shape. Leaves simple, coriaceous, slightly discolorous, adaxial surface dark green and shiny, abaxial surface light green and opaque, glabrous on both surfaces and margins; the larger leaves with blades (7) 8.5–17 (18) cm long, (0.8) 1–2.5 cm wide, narrowly elliptic (ratio 6–10.8), major veins (11)13-17 on each side of midvein, base asymmetric and attenuate, margin entire, apex acuminate; petioles 0.2 (0.5–1.4) cm long, glabrous; the minor leaves 2.5–5.7 cm long, 1–2 cm wide (ratio 1.78–4), ovate or broadly elliptic, major veins 4–5 on each side of midvein, base short attenuate, sometimes asymmetric, margin entire, apex obtuse; petioles 0.1–0.5 cm long, glabrous. Flowers in fascicles of 3–7 (9) on a short shoot leaving evident scars when fallen, rarely solitary; flowering pedicels green, filiform, terete, pendent, slightly curved, not geniculate at anthesis, widening to the apex, 0.3–0.8 cm long, glabrous. Flower buds ovoid, yellow or purplish yellow. Calyx 2.5–3 mm long, 2.8–3 mm wide, cup-shaped, very thin, transparent, light green or greenish purple, the margin truncate, glabrous, with 2–3 thick appendages like triangular-compressed wings, 2–2.5 mm long, 1.8–2.2 mm wide, green or purple, glabrous. Corolla 6–8.5 mm long, 8–11 mm diam, stellate-campanulate, thick, entirely yellow or yellow with red-brown coloration at margin lobes or inside, without interpetalar tissue; tube (3) 4–5 mm long, glabrous inside and outside; lobes 3–3.5 (4) mm long, ca. 3 mm wide, broadly ovate, erect or patent, glabrous adaxially and abaxially, the tips papillose and cucullate. Stamens 5, equal, filaments equal, 2–2.6 mm long, white or red-brown, glabrous, inserted on the corolla ca. 2 mm from the base, with inconspicuous auricles at point of insertion; anthers 2–2.75 mm long, not connivent, elliptic, purplish white or brown. Ovary 1.6–1.8 mm long, 1.2 mm diam, subglobose, white or light green, glabrous; nectary 0.3–0.5 mm tall, white; style 5–5.8 mm long, white and lilac at the apex, widening distally, glabrous; stigma 0.3 mm long, 0.2–0.4 mm wide, light green, somewhat bilobed. Berry 0.8–1.3 cm diam, globose, slightly flattened at the apex, green when immature, orange at maturity, glabrous, not pungent, the pericarp lacking giant cells (endocarp smooth) and stone cells; fruiting pedicels 1–1.6 cm long, pendent, terete, widened distally; the fruiting calyx persistent, non-accrescent, 4–5.5 mm diam, discoid, green-purple or green, the appendages spreading or reflexed, short and wide (2–2.8 mm long, 2.4–2.6 mm wide at base) or long and more slender (4.5–5.5 mm long, ca. 1.5 mm wide at base), fleshy and subulate. Seeds ca. 24 per fruit, 1.7–2.3 mm long, 1.7–2.2 mm wide, not compressed, obconic, black, the surface reticulate, cells rectangular or polygonal in shape, lateral walls straight or slightly sinuate.

Distribution and ecology. Endemic to northern Peru (Amazonas, Cajamarca and Piura) and southern Ecuador (Zamora-Chinchipe) ( Fig 2 ), growing in montane wet forests at mid elevations (1800–2200 m), associated with other Solanaceae shrubs ( Capsicum geminifolium (Dammer) Hunz. , Solanum spp. , and Deprea spp. ), Cyathea Sm. ( Cyatheaceae ), Miconia Ruiz & Pav. ( Melastomataceae ), Piper L. ( Piperaceae ), Ocotea Aubl. ( Lauraceae ), Anthurium Schott ( Araceae ), amongst other shrubs and trees. It grows in the interior of primary forest in shady areas.

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Phenology. Flowering and fruiting from December to August, and probably all year.

Etymology —The species epithet refers to the shape of the leaves, which are the longest and narrowest elliptic leaves known thus far in the genus.

Species Conservation Assessment. Following the IUCN Criteria (IUCN 2017), we suggest C. longifolium deserves a status of Endangered. The extent of occurrence is calculated to be 19207 km 2 (Criterion B1 <20000 km 2, Vulnerable), the area of occupancy, 24 km 2 (Criterion B2 <500 km 2, Endangered). Although the species has been collected many times in the last 12 years, in San Francisco Biological Reserve (SFBR, Zamora-Chinchipe, Ecuador), it is known from only other 5 locations (Criterion B1a � 10, Vulnerable), in areas not included in a National System of Protected Areas which would indicate a risk to the quality of its habitat (Criterion B2b).

Karyology. This taxon possesses a 2n = 2x = 26 karyotype with 9 m pairs (1–9) of decreasing but rather similar size, 3 sm pairs (10–12), and one st pair (13) ( Fig 5 , S 1 Table). Pair 10 (sm) is satellited. Two types of constitutive heterochromatin are found in this taxon, GC-rich heterochromatin ( CMA +/DAPI-) and moderately GC-rich heterochromatin ( CMA +/DAPIo). The fluorescent banding pattern is quite simple, with most of the chromosomes having similar-sized small terminal bands, except for pairs 8, 12 and 13, which are not banded, an intercalary band on the long arm of pair 3, all of them carrying moderately GC-rich heterochromatin, and the large heterochromatic nucleolar organizer region ( NOR)-associated block on the short arm of satellite pair 10, bearing GC-rich heterochromatin ( Fig 5 , S 4 Table).

Affinities. Capsicum longifolium is strongly resolved within the Andean clade, as the first diverging branch within the clade, sister to the rest of the species included in it ( Fig 6 ).

Additional specimens examined. PERU. Amazonas: Rodríguez de Mendoza, Omia, entre la Cordillera y Quebrada de Agua Sal , 06˚26’18’’-06˚25’4.4’’S, 77˚10’30.8’’-77˚10’2.3’’W, 2457– 2118 m, 22 Sept 2008 (fl, fr), V. Quipuscoa S. et al. 4374 ( HUSA, HUT, F, USM) . Cajamarca: San Ignacio, Huarango , Quebrada El Mirador, 05˚16’12"S, 78˚40’01"W, 2200 m, 13 Mar 2000, J. Campos et al. 6607 ( MO) . Piura: Huancabamba, distr. Carmen de la Frontera, Río Samaniego, margen derecha, zona de amortiguamiento del Santuario Nacional Tabaconas-Namballe , 2150–2200 m, 25 Apr 2003 (fl, fr), S. M. Baldeón et al. 5316 ( USM) . ECUADOR. Zamora-Chinchipe: Estación Biológica San Francisco ( EBSF) , camino hacia la antena, pasando el río San Francisco , 3˚58’22.5”S, 79˚04’40.9”W, 1830 m, 2 May 2017 (fl), G. E. Barboza & S. Leiva González 4821 ( CORD) ; at the same place, S. Leiva González 6531 ( HAO) ; EBSF, a unos 300 m después del cruzar el Río San Francisco, por el camino del Atajo , 3˚ 58’21.6”S, 79˚04’41.4”W, 1839 m, 17 Aug 2017 (fl, fr), G. E. Barboza & S. Leiva González 4846 & 4851 (LOJA, duplicates to be sent to CORD & HAO) ; EBSF, después de cruzar el Río San Francisco , 3˚58’22.5”S, 79˚04’39.3”W, 1888 m, 17 Aug 2017 (fl, fr), G. E. Barboza & S. Leiva González 4849 & 4850 (LOJA, duplicates to be sent to CORD) ; road Loxa-Zamora , 5 km W of Tambo, 2100 m, 14–19 Jul 1959 (fl), G. Harling 5867 (S) ; above Valladolid on road to Yanganá , 2700 m, 2 Feb 1985 (fl), G. Harling & L. Andersson 21464 ( GB) .

Capsicum longifolium is unique in the genus in having the longest and narrowest leaves and the striking calyx appendages that arise from the calyx tube as lateral compressed thick expansions or wings ( Fig 4C–4E and 4L–4N). Apart from that, it is morphologically most similar to C. dimorphum with which it shares the shape and color of the corolla, fruit and seeds. Capsicum longifolium can be distinguished by having completely glabrous vegetative organs and calyces, long and narrow (ratio 6–10.8) coriaceous major leaves, flowers in fascicles of 3–7 (9) on a short shoot and calyces with 2–3 thick appendages like triangular-compressed wings compared to the pubescent vegetative organs and calyces, the shorter and wider (ratio 4–5.25) membranaceous major leaves, the solitary or up to 5 axillary flowers, and the toothless calyx or with 3 tiny appendages of C. dimorphum . Another species of Capsicum sympatric with C. longifolium (especially in SFBR, Ecuador) is C. geminifolium that has a dense indumentum, long apiculate leaves, longer pedicels (5 cm long), thin calyx appendages, and funnel-shaped yellow corollas with many purple or maroon spots inside.

Variation in corolla color and length of the fruiting calyx appendages can be observed in the field in individuals growing under the same environmental conditions. The corolla is mainly pure yellow ( Fig 4E, 4H and 4I), but occasional specimens have corolla lobes red-to brown-edged ( Fig 4J and 4K), or with a red-brown ring inside the corolla limb ( Fig 4F and 4G); in this latter case, the filaments and the style are also red-brown. In general, the fruiting calyx appendages do not enlarge considerably ( Fig 4L and 4N) but some specimens have long appendages ( Fig 4M).

The chromosome number 2n = 26 found in C. longifolium is the same as that of C. rhomboideum (Dunal) Kuntze [ 32], C. lanceolatum (Greenm.) C.V. Morton & Standl. [ 33] and C. lycianthoides Bitter [ 34], all belonging to the Andean clade. Their karyotype formulas are quite similar, but that of C. longifolium is closest to C. lycianthoides (9 m + 3 sm + 1 st) than to C. rhomboideum (10 m + 1 sm + 2 st). The species of this clade share small amounts of heterochromatin, a single pair of NOR, short karyotype lengths, and small chromosomes in comparison with other species of the genus [ 32]. The karyotype of C. longifolium is almost half the length of C. rhomboideum , the latter with the shortest karyotype length known until now for the entire genus.

Capsicum piuranum Barboza & S. Leiva , sp. nov. [urn:lsid:ipni.org:names: 77192559–1]

Type : Peru. Piura: Prov. Huancabamba, borde de carretera y riachuelo, 5˚22’46”S, 79˚ 33’47”W, 2311–2459 m, 22 Mar 2011 (fl, fr), T. Mione 812 (holotype, CORD [CORD-00006936]; isotype, NY [NY-03231447]) .

Figs 7 and 8

Diagnosis. Like Capsicum caballeroi M. Nee but differing in the purple calyx, the 5 equal calyx appendages, the longer tubular-campanulate corolla, the globose orange non-pungent mature fruit, and the black seeds.

Description. Scandent shrubs 2–2.20 (3) m tall, densely branched. Young stems green, shiny, fragile, flexuous, glabrous, striate; bark of older stems green to dark brown, glabrous, striate; lenticels absent. Sympodial units difoliate, geminate, leaf pair markedly anisophyllous in size and shape. Leaves simple, membranaceous, discolorous, adaxial surface dark green and shiny, abaxial surface light green and opaque, glabrous or with sparse simple antrorse trichomes 0.5–1.2 mm long adaxially and abaxially, occasionally trichomes more abundant on main veins and margins; the larger leaves with blades (8) 12–17.7 cm long, (2) 2.5–4.5 cm wide, elliptic, major veins 7–9 on each side of midvein, base asymmetric and attenuate, margin entire, apex long-acuminate; petioles 0.7–1.4 (1.7) cm long, slightly winged from the decurrent leaf bases, glabrous or glabrescent with trichomes like those of the leaves; the minor leaves 2.5– 4.5 cm long, 1.5–2.6 cm wide, ovate or elliptic, major veins 3–4 on each side of midvein, base short attenuate or rounded, asymmetric, margin entire, apex acute or slightly rounded; petioles 0.2–0.5 cm long, glabrescent or pubescent. Flowers solitary or in fascicles of 3; flowering pedicels green, filiform, terete, pendent, slightly curved, not geniculate at anthesis, 1.9–2.6 cm long, glabrous or glabrescent, the trichomes simple, non-glandular, multicellular, antrorse, 0.30– 0.45 mm long. Flower buds ovoid, yellow or pale yellow. Calyx 1.5–2.6 (3) mm long, 3–4 mm wide, cup-shaped, thick, purple or greenish purple, the margin truncate, glabrescent to pubescent, with 5 appendages (0.9) 2.5–3 mm long, 0.5–0.8 mm wide, thick, erect, subulate, inserted close to the margin, glabrous or glabrescent with the same trichomes as pedicels and calyx tube. Corolla 14.5–17 mm long, 12–17 mm diam, tubular-campanulate, thick, entirely yellow; tube 11–12 mm long, glabrous inside and outside; lobes 3.5–5 mm long, 4.5–5 mm wide, broadly ovate, erect, glabrous adaxially and abaxially, the tips papillose. Stamens 5, equal, filaments subequal, 3–5 mm long, greenish white, glabrous, inserted on the corolla 3–4 mm from the base, with inconspicuous auricles at point of insertion; anthers 2–2.5 (2.8) mm long, slightly connivent before anthesis, elliptic, yellowish white. Ovary 1.25–1.5 mm long, 1.5 mm diam, subglobose, white, glabrous; nectary ca. 0.5 mm tall, inconspicuous, yellowish white; style 7.5–8 mm long, white, widening distally, glabrous; stigma 0.5 mm long, 0.8–1 mm wide, green, somewhat bilobed. Berry 0.9–1.2 cm in diameter, globose, slightly flattened at the apex, green or white when immature, orange to red at maturity, glabrous, not pungent, the pericarp lacking giant cells (endocarp smooth), sclerotic granules 2, polyhedral, yellowish white; fruiting pedicels 2.8–3.6 cm long, pendent, slightly striate and widened distally; the fruiting calyx persistent, non-accrescent, ca. 4 mm diam, discoid, green-purple or green, the reflexed appendages 5–6.1 mm long, 0.8–1 mm wide at base, fleshy and subulate. Seeds ca. 50–80 per fruit, 2–2.2 mm long, ca. 2.5 mm wide, somewhat compressed, subreniform or obconic, dark brown, the surface reticulate, cells polygonal in shape, lateral walls straight or slightly sinuate.

Distribution and ecology. Endemic to a restricted area in northern Peru (Piura, Fig 2 ) growing in montane misty rain forests, associated with other Solanaceae shrubs ( Solanum spp.

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and Streptosolen jamesonii (Benth.) Miers ), Begonia L. ( Begoniaceae ), Otholobium C.H.Stirt. ( Fabaceae ), Aphelandra R. Br. ( Acanthaceae ), Juglans L. ( Juglandaceae ) amongst other herbs and shrubs. It grows in margins of forest and near streams, between 2300–2860 m elevation, in areas of low temperature and rich soils.

Phenology. Flowering from November to May, with a peak of fruiting in March–May.

Etymology. The new species is named in allusion to its very restricted habitat in Department Piura ( Peru).

Species Conservation Assessment. According to IUCN criteria (IUCN, 2017), C. piuranum is proposed as Critically Endangered ( CR) species. The extent of occurrence is calculated to be 10.195 km 2 (Criterion B1 <100 km 2, Critically Endangered), the area of occupancy, 8 km 2 (Criterion B2 <10 km 2, Critically Endangered). The species is known from only three locations (Criterion B2a � 5, Endangered) and the number of mature individuals observed in each subpopulation is � 50 (Criterion C2a, Critically Endangered).

Karyology. A somatic chromosome number of 2n = 2x = 26 was found in this species. The karyotype comprises 9 m pairs of rather similar length (1–9), 3 sm pairs (10–12), and one st pair (13) ( Fig 9 , S 1 Table). One pair is satellited (10 sm). As in C. longifolium , this species bears two types of constitutive heterochromatin, GC-rich heterochromatin ( CMA +/DAPI-) located in the large heterochromatic band associated to the NOR in pair 10, and moderately GC-rich heterochromatin ( CMA +/DAPIo), located in the small terminal bands and in the intercalary band on the long arm of pair 3. The fluorescent banding pattern is quite simple and very similar to C. longifolium , except for the presence of 3 small bands that are not seen in that species ( Fig 9 , S 4 Table).

Affinities. Capsicum piuranum is resolved within the Andean clade, strongly supported as sister to the C. lycianthoides - C. geminifolium assemblage ( Fig 6 ).

Additional specimens examined. PERU. Piura: Prov. Huancabamba, carretera Canchaque-Huancabamba, km 98–99 (ca. 20 km de Canchaque rumbo a Huancabamba), subiendo a Cuello del Indio , 05˚22’43”-05˚22’44”S, 79˚33’34”-79˚33’37”W, 2341–2346 m, 8 May 2017 (fl, fr), G. E. Barboza & S. Leiva González 4841 & 4842 ( CORD); carretera Huancabamba-Canchaque, a una hora del Abra Cruz Blanca, 05˚22’25"S, 79˚34’02"W, 2860 m, 13 Apr 2017 (fl, fr), M. Cueva 2912 ( USM); distrito Canchaque, km 98–99 (ruta Canchaque-Huancabamba), 5˚ 22’44.1”-5˚ 22’42.7”S, 79˚ 33’37.2”-79˚ 33’34.1” W, 2341–2346 m, 8 May 2017 (fl, fr), S. Leiva González & G. E. Barboza 6561 & 6562 ( HAO); Prov. Morropon: Chalaco, Bosque Mijal, 05˚ 03’51.1"S, 79˚43’25.9"W, 2800 m, 1 Nov 2015 (fl, fr), M. Cueva et al. 2655 ( USM); same locality and date, M. Cueva et al. 2656, 2657 & 2658 ( USM, duplicates to be sent to CORD, HUSA) .

Capsicum piuranum is morphologically most similar to C. caballeroi M. Nee of the Bolivian yungas (Santa Cruz and Cochabamba) based on their campanulate yellow corollas. However, these species can be distinguished in the calyx color, the calyx appendages (number, size, and shape), the position of the corolla lobes at anthesis, the fruit size, shape, color and pungency, the presence of stone cells, and the seed color. Capsicum piuranum has a purple or greenish purple calyx with 5 equal subulate appendages ( Fig 8D and 8E ), while C. caballeroi has green calyx with 10 unequal linear appendages. Corolla lobes are erect compared to those of C. caballeroi which are recurved. Mature fruits are smaller (up to 1.2 mm diam), globose, orange and not pungent in C. piuranum but are larger (up to 1.6 mm diam), globose-depressed to globose, bright red and pungent in C. caballeroi . Capsicum piuranum has two stone cells ( Fig 8H ) and dark brown smaller seeds (2–2.2 mm long, ca. 2.5 mm wide) while C. caballeroi lacks of stone cells and the seeds are pale yellow or light brown and larger (3.2–4 mm long, 3.8–5 mm wide).

Capsicum piuranum is sympatric with other two Andean species, C. geminifolium (Dam-

mer) Hunz. and C. rhomboideum , both of which have also yellow corollas and non-pungent

fruits, but a moderate to dense pubescence on stems and leaves. Capsicum geminifolium differs

in having longer calyx appendages (3–6.5 mm long) compared to C. piuranum (2.5–3 mm

long) and funnel-shaped generally purple spotted yellow corollas (tubular-campanulate and

pure yellow in C. piuranum , Fig 8E ). Capsicum rhomboideum has ovate or rhomboid-ovate

leaves, up to 12 axillary flowers, campanulate-rotate smaller corollas (0.6–0.95 cm long), and

smaller (up to 0.9 cm diam) bright red to blackish red fruits in contrast to C. piuranum where

leaves are elliptic or narrowly elliptic (sometimes the minor leaves are ovate, Fig 8B and 8C ),

the flowers are solitary or in fascicles of 3 ( Fig 8E ), the corolla is tubular-campanulate and lon-

ger (14.5–17 mm long), and the fruits are larger (0.9–1.2 cm diam) and orange colored.

This species exhibits the same number of chromosomes as C. longifolium and the species that belong to the Andean clade [ 32 – 34]. In addition, C. piuranum and C. longifolium share the same karyotype formula, little heterochromatin, one only pair of NOR, and the smallest chromosomes in the genus [ 32].

The markedly anisophyllous leaves, the deflexed non-geniculate pedicels, the yellow corollas, the globose orange to red non-pungent fruits, the absence of giant cells and the presence of stone cells in the pericarp, the black seeds, and the chromosome number 2n = 26 place C. piuranum in the Andean clade proposed by Carrizo García et al. [ 1], as it has been determined in this work based on DNA data.

Capsicum neei Barboza & X. Reyes , sp. nov. [ urn:lsid:ipni.org:names: 77192560–1]. Type: Bolivia. Chuquisaca: Prov. Hernando Siles, a 4.1 km del puente nuevo de Monteagudo viniendo desde Monteagudo , sobre mano derecha, - 19.804617 S, - 64, 019923 W, 16 Dec 2017 (fl), G. E. Barboza 4927 (holotype, LPB; isotypes, CORD [CORD-00006935, CORD-00006956], NY) .

Figs 10 and 11

Diagnosis. Like Capsicum minutiflorum Rusby (Hunz.) but differing in the non-geniculate pendent flowering pedicels and the strongly nerved calyx with 10 unequal appendages.

Description. Small shrubs 0.70–2 (3) m tall, thin, erect, laxly branched above. Young stems green, slim, fragile, glabrescent, and slightly striate, without lenticels; bark of older stems light brown, glabrous, with a few oblong lenticels. Sympodial units difoliate, geminate, leaf pair not markedly anisophyllous in size and shape. Leaves simple, membranaceous, glabrescent on both surfaces and margins with 4-7-celled non glandular trichomes 0.2–0.5 mm long; the larger leaves with blades (5.5) 6.7–11 cm long, 2.1–4 (4.5) cm wide, elliptic or ovate, major veins 3–4 on each side of midvein, base attenuate, margin entire, apex acute; petioles 0.3–0.8 (1.5) cm long; the minor leaves 2.7–4.6 (6) cm long, 1.2–1.8 (2.3) cm wide, elliptic or ovate, major veins 2–3 on each side of midvein, base attenuate, margin entire, apex obtuse or acute; petioles 0.2–0.5 (0.8) cm long, with similar pubescence as in larger leaves. Flowers 2–4 per axil, rarely solitary; flowering pedicels green, filiform, striate, pendent, slightly curved, not geniculate at anthesis, (0.65) 0.8–1.5 cm long, with sparse 5-6-celled non-glandular trichomes and tiny dark glandular trichomes (stalk unicellular, head multicellular). Flower buds ovoid, greenish pale yellow. Calyx 1.7–2.5 mm long, 2–3 mm wide, cup-shaped, green, with 10 nerves clearly evident, the margin truncate, pubescent, with non-glandular trichomes 0.3–0.6 mm long outside and dense glandular pubescence inside (head multicellular, stalk unicellular), 10 unequal linear appendages, green, the five longer appendages (0.7) 0.9–1.75 (2) mm long, emerging almost from the margin, the five shorter 0.2–0.8 (1.2) mm long, emerging 0.8–1 mm below the margin, with the same non glandular trichomes of the calyx tube. Corolla (6) 8–10 mm long, 5–6 mm diam, stellate, delicate, entirely yellow or with small brownish green spots in the base of the lobes and tube inside, with a thin interpetalar tissue; tube 3–4.5 mm long, with tiny glandular trichomes (head and stalk one celled each) inside and glabrescent outside; lobes 3.5–5.5 mm long, ca. 2 mm wide, ovate, erect, glabrous adaxially and with sparse non-glandular trichomes abaxially, the tips papillose and cucullate. Stamens 5, subequal, filaments 1.4–1.75 mm long, cream, glabrous, inserted on the corolla ca. 1.2 mm from the base, with inconspicuous auricles at point of insertion; anthers (1.5) 1.8–2 mm long, not connivent, elliptic, light yellow. Ovary ca. 1.2 mm long, 1.3 mm diam, ovoid or subglobose, light green, glabrous; nectary ca. 0.3 mm tall, style 3.75 mm long, cream, widening distally, glabrous; stigma ca. 0.2 mm long, 0.3 mm wide, light green, somewhat bilobed. Berry 0.4–0.75 cm diam, globose, green when immature, red at maturity, glabrous; fruiting pedicels (1.3) 1.8–2.3 cm long, pendent, striate and widened distally; the fruiting calyx persistent, non-accrescent, ca. 4 mm diam, discoid, the appendages spreading 1–2 mm long, subulate. Seeds unknown.

Glandular trichome of the inside corolla. (L) Non-glandular trichome of the outside calyx. (M) Non-glandular trichome of the outside corolla lobes. Drawn by S. Montecchiesi.

https://doi.org/10.1371/journal.pone.0209792.g010

Distribution and ecology. Endemic to southeastern Bolivia ( Fig 12 ), mainly in the Serra-

nías Iñao, Yahuañanca and Khaskha Orkho (Dpt. Chuquisaca). A few collections have been

recorded from the Yungas (Dpt. Santa Cruz). Capsicum neei is most commonly collected in

the Boliviano-Tucumano Forest in both Departments [ 35] from understories at the foot of cloud forest hillsides and deciduous forests, between 1100–1750 m elevation. It grows associated with members of Juglandaceae , Lauraceae , Myrtaceae, Leguminosae , ferns and bryophytes.

Phenology. Flowering and fruiting from October to May.

Etymology. The epithet is in honor to Dr. Mike Nee ( NY), a solanaceous specialist who carried out extensive explorations in the Bolivian territory and separated specimens of this species as a rare or probable new species in various herbaria.

Species Conservation Assessment. According to IUCN criteria [ 15], C. neei is proposed as Near Threatened species. The species meets the area requirements under criterion B for threatened (EOO: 16912 km 2, B1 <20000 km 2, Vulnerable; AOO: 44 km 2, B2 <500 km 2, Endangered) and is declining, but the population is not severely fragmented and occurs in more than 10 locations. Capsicum neei has been collected many times in the last 23 years in a recently Protected Area: National Park and Integrated Management Natural Area “Serranía Iñao” [ 36], and in nearby areas which suggests that both the decline in its geographic range (EOO and AOO) and the population size will not be significantly affected in the forthcoming years.

Affinities. Capsicum neei is nested within the Bolivian clade, strongly resolved as sister to C. caballeroi ( Fig 6 ).

Additional specimens examined. BOLIVIA. Chuquisaca: Hernando Siles, ca. 7 km de Monteagudo, inicio del cañón Heredia , 19˚47’17’’S, 64˚02’08’’W, 1127 m, 13 Dec 2006 (fl, fr), H. Huaylla et al. 2178 ( HSB, MO); Parque Nacional y área natural de manejo integrado de la Serranía del Iñao , cuenca del río Limón , 19˚44’01"S, 63˚54’52"W, 1247 m, 15 Dec 2006 (fr), E. Portal et al. 108 ( HSB, MO); foot of Cerro Urkhal path before 2nd river crossing, 19˚48’S 63˚ 57’W, 1300 m, 4 Oct 2000 (fl), K. Wendelberger 170 ( HSB, MO); Luis Calvo, Ticucha, serranía del Iñao, 12 km al NO de la comunidad de Ticucha, 19˚35’0.4”S, 63˚53’12.7”W, 1431 m, 11 Apr 2003 (fl), A. Carretero et al. 824 ( HSB, MO, NY); Entierrillos, aprox. a 5 km de la escuela de Entierrillos, serranía del Iñao, 19˚31’S, 63˚52’W, 1700 m, 18 Dec 2003 (fl), A. Carretero

et al. 939 ( HSB, MO); Serranía del Iñao, pasando la Laguna, 19˚31’S, 63˚52’W, 18 Dec 2003 (fl), A. Carretero et al. 998 ( HSB, MO, NY); Las Frías, ca. a la la cima de la serranía de Ñahuañanca, 19˚09’30.6"S, 63˚50’40.6"W, 1930 m, 22 Dec 2003 (fl, fr), A. Carretero et al. 1067 ( HSB, MO, NY); Las Frías, ca. 1/ 2 km de la vivienda de Sr. Severino Daza, hacia la cima de la serranía de Yahuañanca, 19˚09’31"S, 63˚50’23"W, 1600 m, 23 Dec 2004 (fl), A. Carretero et al. 1085 ( HSB, MO, NY); Sud Cinti, ca. 3 horas en caballo al NW de la comunidad de Orocote entre los ríos Limonal y Cochayo, 20˚47’S, 64˚21’W, 1650 m, 29 Apr 2005 (fr), R. Lozano 1207 ( HSB, MO); Tomina, aprox. 800 m. antes de llegar a Llantoj, de La Florida subiendo hacia el E de la Serranía de Kaska Orcko, 19˚09’46"S, 64˚03’42"W, 1750 m, 11 Oct 2004 (fl, fr), J. Gutiérrez R. 1004 ( HSB, MO); Llantoj, aprox. 800 m antes de llegar a Llantoj, de la Florida subiendo hacia el E de la Serranía de Kaska Orcko, 19˚09’46"S, 64˚03’42"W, 1750 m, 15 Dec 2004 (fr), J. Gutiérrez R. 1072 ( HSB, MO); Rio Limón Valley between Padilla and Monteagudo, 1500 m, 1 Jan 1995 (fl), J. R. I. Wood 9104 ( NY). Santa Cruz. Prov. Florida, Mairana, La Yunga de Mairana , 18˚04’13”S, 63˚55’08”W, 2190 m, 15 Nov 2004 (fl), M. Serrano et al. 5482 ( NY).

Capsicum neei is morphologically most similar to the Bolivian C. minutiflorum in having stellate yellow corolla and red fruit at maturity. It can be distinguished by the non-geniculate pendent flowering pedicels and the strongly nerved calyx with 10 unequal appendages ( Figs 10C, 10E and 11B ) versus the geniculate and erect flowering pedicels and the calyx weakly nerved and with 5 equal short appendages in C. minutiflorum (Rusby) Hunz. The flowers in C. neei often appear to be solitary but the remains of 2–3 early deciduous bud or flower scars can be seen in the axils. Fruit features as pungency, presence of giant cells and sclerotic granules in the pericarp and mature seeds are unknown at present but it is probable that the fruits are pungent and have giant cells in the innermost layer of the pericarp as occur in the remaining species of the Bolivian clade where C. neei is positioned.

This new species is sympatric with C. baccatum L. var. baccatum , a taxon with a much wider distribution in South America, that has geniculate pedicels, calyx with 5 equal appendages, white corollas with greenish yellow spots inside and ovoid or globose red fruits.

Capsicum neei has been resolved as a new member of the Bolivian clade, which is coherent with its geographic range and the main common feature recognized for the clade, the yellow corollas [ 1]. However, the Bolivian clade has a weak support, most likely due to the apparent divergence of C. coccineum from the rest of the species; indeed, C. coccineum would deserve more attention considering some morphological variability observed in the species (GEB, pers. obs.).

Figures

Fig 2. Distribution of Capsicum species. https://doi.org/10.1371/journal.pone.0209792.g002

Fig 5. Somatic metaphase chromosomes and ideogram of Capsicum longifolium. (A) Methaphase chromosomes. (B) Ideogram. Solid black blocks or dots denote CMA+/DAPI- (NOR) or CMA+/DAPIo (terminal and intercalary) heterochromatic bands. The NOR is indicated as a separate block. https://doi.org/10.1371/journal.pone.0209792.g005

Fig 6. Bayesian majority-rule consensus tree of Capsicum. Posterior probabilities values indicated by each branch. New species are highlighted in bold-colored letters and the clade to which they belong is indicated. https://doi.org/10.1371/journal.pone.0209792.g006

Fig 8. Capsicum piuranum Barboza & S. Leiva. (A) Plant. (B) Leaves, abaxial surface. (C) Fruiting branch. (D) Flower bud. (E) Flower and immature fruit. (F) Mature fruit. (G) Fruit, transverse section, showing placenta and seeds. (H) Fruit transverse section, showing a stone cell at the apex (arrow). Photos by S. Leiva González and G. E. Barboza. https://doi.org/10.1371/journal.pone.0209792.g008

Fig 9. Somatic metaphase chromosomes and ideogram of Capsicum piuranum. (A) Methaphase chromosomes. (B) Ideogram. Solid black blocks or dots denote CMA+/DAPI- (NOR) or CMA+/DAPIo (terminal and intercalary) heterochromatic bands. The NOR is indicated as a separate block. https://doi.org/10.1371/journal.pone.0209792.g009

Fig 10. Capsicum neei Barboza & X. Reyes. (A) Flowering branch. (B) Inflorescence. (C) Flower bud. (D) Flower. (E) Calyx. (F) Opened corolla.(G) Gynoecium (H) Fruit. (I) Glandular trichome of the inside calyx. (J) Glandular trichome of the pedicels.(K)

Fig 11. Capsicum neei Barboza & X. Reyes. (A) Plant. (B) Flower bud. (C) Flowering branch. Photos by G. E. Barboza. https://doi.org/10.1371/journal.pone.0209792.g011

Fig 12. Distribution of Capsicum neei Barboza & X. Reyes. https://doi.org/10.1371/journal.pone.0209792.g012