Disentangling Cuscuta identification in Brazil: a first taxonomic contribution to the northeast region species

Cuscuta is a genus of Convolvulaceae distributed worldwide and comprises about 200 species, of which 26 were reported to Brazil. They are characterized by being holoparasites, leafless, gamopetalous, bisexual and usually pentamerous flowers. Studies available for South American Cuscuta are mostly from late XIX and early XX centuries. The restriction of taxonomic bibliographies, together with the fact that taxonomic informative characters are restricted to their tiny flowers, make the species identification a challenge. In this context, this work aimed to study the diversity of Cuscuta for the northeast region of Brazil and allow species identification. Field works were carried out in all northeast region states and about 150 specimens from 17 herbarium collections were analyzed. A lectotypification is proposed for C. racemosa. Cuscuta orbiculata, cited to Mexico, Guatemala and Brazil (Goiás and northeast region), is synonymized under C. tinctoria. Eight species were recognized, occurring mainly in areas of Caatinga, that predominates in the region. Descriptions, taxonomic comments, illustrations and an identification key are presented.


Introduction
Cuscuta L. (1753a: 124) is represented by leafless plants, with haustoria, aclorophyllate, nonphotosynthetic and therefore, these individuals suck the elaborate sap of a host plant (Andrade et al. 2007). It comprises about 200 species, and according with BFG (2018) 26 species were recognized, of which 11 were recorded in the northeast region. About 15 to 20 species of Cuscuta are pests that can cause up to 80% yield losses in plants cultivated, ornamental and wild plants (Pereira 1998) preventing healthy growth of the host species.
See supplementary material at <https: //doi.org/10.6084/m9.figshare.14669889.v1> In addition of being an economic concern, as parasitic weeds, some species also stand out by its medicinal and ecological importance. Popular recommendations for treatment of urinary disorders, icterus, muscular pain, cough, biliary disorders, as purgative also for itchy skin are found in literature (Wright et al. 2011;Khan et al. 2010). Ecologically, because of their parasitic condition, they can compromise the development of the host plant, possibly generating changes in the competitive interactions and influencing the community structure, observed diversity, nutrient cycling and vegetation zoning (Pennings & Callaway 2002).
The taxonomic position of Cuscuta in a historical perspective has been largely discussed, with disagreement about its belonging either to Convolvulaceae (Jussieu 1789;Choisy 1841;Engelmann 1842Engelmann , 1859Engler & Gilg 1924) or to a monogeneric family, Cuscutaceae (Cronquist 1981;Pfeiffer 1845;Progel 1869;Austin 1998). This discordance is because Cuscuta is quite distinct from other Convolvulaceae, for being holoparasites, with leaves reduced to scales and calyx usually gamosepalous (vs. photosynthetic plants, foliolated and calyx dialisepalous), although remnant flower morphology is mostly congruent. The most recent phylogenetic hypothesis indicated the monophyly of Convolvulaceae only with the inclusion of Cuscuta, despite its position within the family has not been completely elucidated (Stefanović et al. 2003;Stefanović & Olmstead 2004).
The last comprehensive taxonomic treatment of the genus was carried out in 1932 by Yuncker, for the American species. Despite its importance, this treatment is sometimes hard to use, since the identification key starts with fruit charactersfrequently missing in herbarium material -and also species descriptions are not comparable to each other since they have different formats. Besides that, the infrageneric classification proposed by Yuncker was considered unnatural by recent phylogenetic studies (Costea et al. 2015), indicating that more in depth studies are needed.
The lack of recent taxonomic literature together with informative characters restricted to its small flowers, led to severe difficulty to identify taxa. Most of the available literature that that includes Cuscuta species in Brazil deals with the consequences of the parasitic relationship to the host plants, or are references in local floras (Simão-Bianchini & Pirani 1997;Andrade et al. 2007; Alves & Kolbek 2009;Simão-Bianchini et al. 2016), and more rarely new species descriptions, as Cuscuta taimensis P.P.A. Ferreira & Dettke (Ferreira et al. 2014). The present study aimed to evaluate the diversity of Cuscuta in northeast region of Brazil, providing tools to the correct identification of species.

Material and Methods
This study was carried out in the northeast region of Brazil ( Fig. 1), which is composed by nine states: Alagoas (AL), Bahia (BA), Ceará (CE), Maranhão (MA), Paraíba (PB), Pernambuco (PE), Piauí (PI), Rio Grande do Norte (RN) and Sergipe (SE). It comprises an area of 1.561.177,8 km 2 (18,3% of Brazilian territory) and is limited by the Atlantic ocean to the east and north, by the states of Espírito Santo and Minas Gerais to the south and by the states of Pará, Tocantins and Goiás to the west (SUDENE 2015). Caatinga is the predominant phytogeographic domain, occupying more than 50% of all states. In this region, other phytogeographic domains such Amazonia (in a small portion of Maranhão state), Cerrado and the Atlantic Forest are also found (IBGE 2017).
Initially, a literature survey together with consultation of collections available online were carried out to determine species occurring in the study area, in addition to identifying priority areas for fieldwork. Collections of 17 herbaria were consulted (ALCB, CEPEC, EAC, HRB, HST, HUEFS, HUFRN, HVASF, IPA, JPB, MAC, MBM, MOSS, PEUFR, RB, SPF and UFP) (acronyms according to Thiers, continuously updated), resulting in about 150 specimens analyzed; whenever was possible identifications were updated. Specimens identification were made by comparison with species previously identified by experts, type material available online, Digital Atlas of Cuscuta (Costea 2007-onwards) and by using specialized literature (Progel 1869;Yuncker 1921Yuncker , 1922Yuncker , 1923Yuncker , 1932. Field work have been carried out by the Laboratory of Integrative Systematics of Angiosperms team (LASIA-UFRPE) focusing on Convolvulaceae since 2015. Expeditions were carried out in all northeast region states of Brazil. Specimens collected were processed and deposited in PEUFR herbaria according to the usual botanical material management techniques (Mori et al. 1985). Duplicates were sent to JPB, ALCB and K. A database was compiled containing information available about all vouchers studied (Tab. S1, available on supplementary material <https://doi. org/10.6084/m9.figshare.14669889.v1).
The terminology used followed Harris & Harris (2001) and Yuncker (1932) and measurements were made using the ImageJ© software (Schneider et al. 2012;Rasband 1997Rasband -2018 calibrated in centimeters. An identification key was elaborated accompanied by detailed morphological descriptions, taxonomical comments and geographical distribution. Diagnostic characters were illustrated, and distribution maps were constructed using DIVA-GIS© 7.5 software (Hijmans et al. 2012).

Results and Discussion
Despite all effort employed in field trips, only 15 specimens of five different species were found ( Fig. 2) Choisy (1841: 284) andC. tinctoria Mart. ex Engelm. (1859: 480-481)), which indicate the rarity of this group in the NE Brazil. Some species showed preferences for humid habitat such as C. americana, while others for shaded environments such as C. globosa.
A total of eight species of Cuscuta are listed occurring in the NE Brazil, in discordance with the 11 species indicated in BFG (2018); no records for Cuscuta indecora Choisy (1841: 278-279) and Cuscuta insquamata Yunck. (1923: 12) were found. A specimen identified in herbaria as C. indecora (Mendes et al. 548,UB) is actually C. partita. However, considering that fruit may present a late dehiscence in C. partita, to observe this character is sometimes challenging. Other morphological difference between these two species are the corolla papillate and calyx smooth in C. partita, while C. indecora has both the calyx and corolla papillate.
Cuscuta insquamata was indicated for Brazil only by one record from Abaíra, Bahia state. This specimen (Stannard et al. 51588, NY) could not be seen. Cuscuta insquamata presents flowers with less than 2 mm long, with calyx and corolla lobes acute and 4-divided. Two specimens from Bahia (ALCB 3456 e ALCB 9900) that present similar size and shape of the lobes, but are 5-divided and with ovary and infrastaminal scales still in early stages of development were observed. Comparing with floral buds of C. partita specimens, since it also presents calyx and corolla lobes acute and 5-divided, we noted that these buds are quite similar to the ALCB samples. Thus, since only one record of C. insquamata is cited and it could not be checked, this species is not included in this treatment. Cuscuta insquamata is considered here as occurring only in Bolivia (Yuncker 1932), therefore.
Phylogenetic studies in which C. globulosa and C. americana samples were included, shows C. globulosa together with Cuscuta cozumeliensis Yunck. (1922: 108), and Cuscuta macrocephala Yunck. (1921: 126), appearing as sister of C. americana with high support (Stefanović et al. 2007). Thus, considering that characters used by Yuncker (1932) does not seem consistent enough to differentiate these species, and phylogenetic studies show them as closely related, further morphological or population level investigations are recommended to improve specific delimitation.
Despite the overlapping characters, specimens are here identified as C. americana due to predominant characteristics and wider distribution, considering that C. globulosa seems to be more common throughout West India and Mexico and is not cited for South America (Yuncker 1932). In NE Brazil, is similar to Cuscuta corymbosa var. grandiflora Engelm. (1859: 483) because it presents similar calyx but can be distinguished by infrastaminal scales with bridge measuring ca. 1/2 or more of the total scale size in C. americana, and bridge measuring ca. 1/3 or less of the total scale size in C. corymbosa var. grandiflora.
Flowering from May to December. Occurs in Brazil, Venezuela and Colombia (Yuncker 1932). Registered to Brazil in Bahia, Goiás, Rio de Janeiro, Santa Catarina and Rio Grande do Sul (BFG 2018), with this study it was verified its occurrence also in Pernambuco state. Even though it was not found in the field, herbarium labels indicated the occurrence of the species in irrigated areas.

Cuscuta corniculata
It resembles C. tinctoria by its spherical flowers but can be distinguish by the flowers ca. 6 mm long, bracteoles present, calyx lobes completely overlapped and corolla lobes rounded in C. tinctoria vs. flowers ca. 2 mm long, bracteoles absent, calyx lobes slightly overlapping and corolla lobes acute in C. corniculata.

Cuscuta corymbosa
Flowering in August. It is endemic to northeast region of Brazil, where it occurs mainly in the states of Bahia and Pernanbuco (Yuncker 1932) in both Caatinga and Atlantic Forest areas. According to the BFG (2018) it also occurs in Alagoas, Ceará and Paraíba states. We also record it to Rio Grande do Norte and Sergipe states. Cuscuta globosa was usually found in shaded areas parasitizing the stem of herbs.

Cuscuta globosa
It is easily identified by its glomeruliform inflorescence, with flowers of ca. 2.6 mm long, with acute apex in calyx and corolla lobes. Considering species occurring in the northeast region of Brazil is the only one that can also present individuals with tetramerous flowers. Yuncker (1932) still indicates its similarities with C. acuta Engelm. (1859: 497) and C. micrantha Choisy (1841: 271) for its flowers with about 1.5 mm in length and calyx with acute lobes, and the C. globosa differs from these by its long pedicels, minute flowers and long stamens. In addition, C. acuta is endemic to Ecuador and C. micrantha to Chile.
Flowering from July to December.    In the study area can be confused with C. umbellata Kunth (1818: 121-122) for presenting corolla with acute lobes, but can be distinguished by its umbelliform cymes, calyx with lobes overlapping at base and erect, smooth corolla and opaque fruit, while C. partita has a dichasial inflorescence, calyx with lobes not overlapped, often reflexed, papillate corolla and translucent fruit. Specimens branches are usually orange when dried. Its fruit may present    Cuscuta racemosa is endemic to Brazil, being common in the Southern region. According to the BFG (2018), this species should be widely distributed in northeast region, reported to all states but Maranhão. However, in this study only records for the states of Bahia and Pernambuco were found. In Bahia it was found in waterlogged regions, one on the roadside near areas of anthropic influence and another on the trail of the Fumaça waterfall (Chapada Diamantina National Park). In Pernambuco, the species was found parasitizing ornamental plants in a small urban park.

Cuscuta partita
From Yuncker's (1932) identification key, these specimens could be identified either as C. decipiens Yunck. (1921: 55) (a rare species from Mexico) or C. gronovii Willd. (1820: 205) (widespread in North America). But when comparing these specimens with illustrations and morphological descriptions (Yuncker 1932), the determination can easily reach C. racemosa. Futhermore, all morphological descriptions of C. decipiens, C. gronovii and C. racemosa are very similar and could be accepted to these specimens analyzed here. Thus, our decision is taken from the geographic distribution, since C. racemosa was described as endemic to Brazil. However, type material besides specimens from these other localities should be studied before a taxonomic decision as a synonym or not is undertaken.
The name "Cuscuta racemosa" was used arbitrarily in Brazilian collections for any species, sometimes completely different from each other (C. americana and C. partita, for example). This probably occurs due to the lack of elucidative bibliography for support correct identifications.
In the protologue, Martius does not report any examined material. However, exsiccates with the author's handwrites were found in M. According to Art 9.4 of the International Code of Nomenclature for Plants, algae and fungi (ICN) adopted in Shenzhen (Turland et al. 2018) an original material comprises those specimens that the author associated with the taxon, and that were available to the author prior to, or at the time of, preparation of the description and/or diagnosis validating the name. Therefore, the material deposited in Munich is considered as original material because it was analyzed by Martius. We found two exsiccates with his handwriting (M0184374, M0184376) and we choose the M0184374 as lectotype because it is a complete collection with flowers and fruits and matches closely with the description provided by Martius.
Flowering from March to May. Cuscuta tinctoria is cited for Mexico, Guatemala and Brazil. In the latter, is recorded for Goiás (Yuncker 1932) and has a wider distribution in the northeast region, being here presented new records for Alagoas, Paraíba and Rio Grande do Norte states.

Cuscuta tinctoria
The species can be readily distinguished by its spherical flowers of ca. 5 mm and its calyx deeply divided and lobes totally overlapped, often coming to be confused with sepals free. In the northeast region of Brazil it resembles Cuscuta corniculata because both present spherical flowers, but C. tinctoria present flowers with ca. 5 mm, with bracteoles, calyx lobes completely overlapped and corolla lobes rounded, while C. corniculata has flowers with ca. 2 mm, without bracteoles, calyx lobes slightly overlapped and corolla lobes acute.
After analyzing the protologues, comparing descriptions and type materials we concluded that C. tinctoria and C. orbiculata do not present significant differences to be treated as different species. Moreover, the proportions calyx/corolla (calyx as long as the corolla tube in C. tinctoria vs. calyx shorter than corolla tube in C. orbiculata) indicated by Yuncker (1932) as the only character to distinguished them, can be observed in the same individual (see Miranda et al. 857 -ALCB 25577, HST 9727). Cuscuta tinctoria has priority over the C. orbicularia. Cuscuta orbiculata was described as endemic to Brazil, referred only for Goiás, Pernambuco (Fernando de Noronha) and Bahia being known only by the holotype and the one paratype.

Cuscuta umbellata
It resembles C. partita due to corolla with acute to acuminate lobes but can be distinguished by a set of characteristics pointed out in the comments of this species.
Flowering from May to June.