UNIVERSIDADE ESTADUAL PAULISTA “JÚLIO DE MESQUITA FILHO” INSTITUTO DE BIOCIÊNCIAS – RIO CLARO unesp PROGRAMA DE PÓS-GRADUAÇÃO EM CIÊNCIAS BIOLÓGICAS (BIOLOGIA VEGETAL) CARACTERES VEGETATIVOS E REPRODUTIVOS E DESENVOLVIMENTO PÓS-SEMINAL EM BROMELIACEAE KLEBER RESENDE SILVA Tese apresentada ao Instituto de Biociências do Câmpus de Rio Claro, Universidade Estadual Paulista, como parte dos requisitos para obtenção do título de Doutor em Ciências Biológicas (Biologia Vegetal. RIO CLARO/SP Novembro - 2018 KLEBER RESENDE SILVA CARACTERES VEGETATIVOS E REPRODUTIVOS E DESENVOLVIMENTO PÓS- SEMINAL EM BROMELIACEAE Orientadora: Profa. Dra. Aline Oriani Coorientador: Prof. Dr. Leonardo M. Versieux Tese apresentada ao Instituto de Biociências do Câmpus de Rio Claro, Universidade Estadual Paulista, como parte dos requisitos para obtenção do título de Doutor em Ciências Biológicas (Biologia Vegetal). Rio Claro – SP 2018 S586c Silva, Kleber Resende Caracteres vegetativos e reprodutivos e desenvolvimento pós-seminal em Bromeliaceae / Kleber Resende Silva. -- Rio Claro, 2018 102 f. : il., tabs., fotos Tese (doutorado) - Universidade Estadual Paulista (Unesp), Instituto de Biociências, Rio Claro Orientadora: Aline Oriani Coorientador: Leonardo Versieux 1. Anatomia. 2. Envoltório seminal. 3. Campos rupestres. 4. Germinação. 5. Plântula. I. Título. Sistema de geração automática de fichas catalográficas da Unesp. Biblioteca do Instituto de Biociências, Rio Claro. Dados fornecidos pelo autor(a). Essa ficha não pode ser modificada. Com carinho, à minha família e amigos e gratidão àqueles que me incentivaram e abriram portas AGRADECIMENTOS O presente trabalho foi realizado com apoio da Coordenação de Aperfeiçoamento de Pessoal de Nível Superior – Brasil (CAPES) – Código de Financiamento 001. Obteve-se também financiamento pelo DAAD (Co-financed Short-Term Research Grant Brazil, 2017) para realização de estágio no exterior (Ruhr Universität Bochum, Alemanha). Agradeço à Universidade Estadual Paulista (UNESP) - Câmpus de Rio Claro e ao Departamento de Botânica do Instituto de Biociências pela infraestrutura concedida. Ao Programa de Pós-graduação em Ciências Biológicas (Biologia Vegetal), professores e funcionários. À Profa. Dra. Alessandra Ike Coan que foi coordenadora do PPG da data do meu ingresso até meados do doutorado e ao Prof. Dr. Douglas Silva Domingues atual coordenador desse PPG. À Celinha Hebling, secretária do Departamento de Botânica e a todos os funcionários da Seção de Pós-Graduação. Agradeço a Ruhr Universität Bochum, Alemanha, pela infraestrutura concedida durante estágio no exterior e também aos funcionários do Welcome Centre desta universidade por todo o suporte com os procedimentos burocráticos e pelas atividades integrativas promovidas aos estudantes de outros países. À minha orientadora, Dra. Aline Oriani, o meu muito obrigado por aceitar me orientar, me incentivar sempre, sua amizade e confiança, compartilhando do seu tempo e conhecimento, contribuindo à minha formação profissional e pessoal. Ao meu coorientador, Dr. Leonardo Versieux pelo incentivo, por contribuir na elaboração do projeto de pesquisa, pelas dicas, sugestões e suporte sempre que solicitados, e auxílio em campo durante expedições de coleta na Chapada Diamantina, Bahia. Agradeço também à Profa. Dra. Daniela Guimarães Simão, minha professora na graduação e orientadora no mestrado, por toda sua dedicação, amizade, sabedoria e empenho durante minha formação. Ao Prof. Dr. Denis Coelho de Oliveira do Instituto de Biologia da Universidade Federal de Uberlândia por abrir as portas do seu laboratório e auxiliar com as análises imunohistoquímicas. Também agradeço ao Prof. Dr. Thomas Stützel por me supervisionar durante o estágio na Alemanha, providenciando todas as medidas para minha chegada e estadia, e a todos do seu departamento, em especial à Sabine Adler, Petra Lerch, Christoph Elpe e Julian Herting por todas as dicas e informações, paciência e auxílio. Vielen danke! Também agradeço ao Núcleo de Apoio à Pesquisa em Microscopia Eletrônica aplicada à Pesquisa Agropecuária (NAP/MEPA) da ESALQ/USP pelo uso do microscópio eletrônico de varredura durante a execução dos capítulos 1 e 2 dessa tese. Aos membros da banca pela disponibilidade e pelas contribuições. Ao Prof. Dr. Paulo Sano pelo auxílio em campo e pelos momentos divididos em Diamantina, Minas Gerais. Agradeço aos meus amigos e colegas, em especial à Kaire pela amizade desde a graduação, sempre ajudando e contribuindo; à Ana Angélica, Arthur, Blanca, Fernanda, Gisele, Letícia, Lucimara (Mara), Luís, Mariana, Naiara, Paula, Rafael e Thales, pela troca de vivências, convívio em laboratório e por estarem dispostos a ajudar. Ao Marcos pelo auxilio na coleta de material botânico e pela troca de informações. À Ester pelas aulas de inglês, contribuindo com meu aprendizado. Ao André pelo apoio sempre, dedicação e por estar sempre torcendo por mim. Ao Bernardo (que inclusive auxiliou na coleta de materiais na Serra do Cipó) e Danilo, que mesmo a distância, sempre me ajudaram e dedicaram do seu tempo para dar conselhos pessoais e profissionais. À Deicy, Laiza e Watson pela amizade e por estarem sempre torcendo por mim. Ao Helson e Missiani por toda a ajuda, amizade e troca de vivências em Bochum. À Carla que também me ajudou em Bochum, antes e durante a minha estadia. À minha família, especialmente aos meus pais e irmãos, pelo amor incondicional, por me apoiarem e se preocuparem comigo. Lembrarei com carinho de todos os momentos compartilhados. ʻʻO que vale na vida não é o ponto de partida e sim a caminhada. Caminhando e semeando, no fim terás o que colher (Cora Coralina)ʼʼ. Ainda tenho o que caminhar, semear e colher, e levo comigo um pouco de todos, que de forma direta ou indireta, contribuíram à minha formação e a quem sou grato por mais essa etapa da minha vida. “O sentido da vida é um sentimento.” (Rubem Alves) RESUMO Análises filogenéticas apontam Bromeliaceae como uma família monofilética e basal dentre as Poales. Contudo, a sistemática enfrenta dificuldades quanto ao estabelecimento de gêneros e espécies. Caracteres anatômicos, especialmente das folhas, têm sido utilizados na sistemática, taxonomia e para entender respostas ao ambiente. Já entre os órgãos reprodutivos, as sementes e os frutos são importantes para o reconhecimento de suas subfamílias. Bromelioideae é uma das subfamílias com divergência mais recente, apresentando frutos do tipo baga e sementes com envoltório mucilaginoso, o que difere da maioria das demais subfamílias, que possuem frutos do tipo cápsula e sementes aladas ou plumosas. Essa tese teve como objetivos identificar caracteres anatômicos de órgãos vegetativos e do eixo da inflorescência úteis para resolver dois complexos de espécies (Neoregelia bahiana – Bromelioideae, e Vriesea oligantha – Tillandsioideae), devido à variação morfológica de seus representantes; avaliar a germinação, anatomia de sementes e desenvolvimento pós-seminal de N. bahiana para entender adaptações e características do desenvolvimento relacionadas ao estabelecimento das plântulas; elucidar a ontogênese de frutos carnosos em espécies de Bromelioideae, relacionando sua estrutura a características do desenvolvimento das sementes para um melhor entendimento de aspectos funcionais. Dentre os caracteres vegetativos, a anatomia foliar e a arquitetura da planta são importantes para o reconhecimento de populações, indicando a presença de mais de um táxon em cada complexo. A germinação em N. bahiana inicia-se com o desenvolvimento da raiz primária e do hipocótilo. O cotilédone permanece preso ao endosperma no interior das sementes (hiperfilo haustorial) até o estabelecimento das plantas jovens. Cotilédone haustorial e o desenvolvimento da raiz primária por certo período de tempo, indicam maior necessidade nutricional para as plântulas de Bromelioideae. A anatomia dos frutos, formados por tecidos carpelares e não carpelares, que se tornam carnosos e envolvem diversas sementes, caracteriza-os como bagas. A disposição e número de óvulos no ovário estão relacionados com a área disponível nos lóculos para o desenvolvimento das sementes e à presença ou não e comprimento dos apêndices nas sementes. A suculência dos frutos, conferida pelas células do mesocarpo e pela secreção de substâncias no interior dos lóculos pelo obturador, assim como a testa mucilaginosa das sementes, aparecem em estágios posteriores do desenvolvimento e se destacam como estratégias à dispersão zoocórica. Palavras-chave: anatomia; bagas; envoltório seminal; campos rupestres; germinação; imunohistoquímica; Neoregelia; órgãos vegetativos; plântula; Vriesea. ABSTRACT Phylogenetic analyzes point to Bromeliaceae as a monophyletic and an earlyer-diverging family within Poales. However, the systematics faces difficulties in genera and species delimitation. Anatomical characters, especially for the leaves, have been used in systematics, taxonomy and to understand adaptive responses to the enviroment. Among the reproductive structures, seeds and fruits are important for the recognition of Bromeliaceae subfamilies. Bromelioideae is one of the latest-diverging subfamilies and exhibits berry-type fruits with mucilaginous seeds, different from the majority of the remaining subfamilies which has capsules with winged or plumose seeds. This thesis aimed to identify anatomical characters with taxonomic value in the vegetative organs and inflorescence axis of two species complexes (Neoregelia bahiana – Bromelioideae, and Vriesea oligantha – Tillandsioideae), due to the great morphological variation of their individuals; to analyse the germination, seed anatomy and the post-seminal development in N. Bahiana to understand adaptations and the traits related to seedling establishment; and to elucidate the ontogeny of fleshy fruits in species of Bromelioideae, relating their structure to the seed features and to functional aspects. Among the vegetative characters, the leaf anatomy together with the plant architecture are important to delimit populations, evidencing the presence of more than one taxon in each species complex. The germination in N. bahiana occurs by the development of the primary root and hypocotyl. The cotyledon remains attached to endosperm inside the seeds (haustorial hyperphyll) until the establishment of the young plants. This condition together with the development of the primary root for a certain period of time indicate a great nutritional need of the seedlings. The fruit anatomy, showing its formation by carpellary and non-carpellary fleshy tissues with several seeds characterizes it as a true berry. The arrangement and number of ovules in the ovary are related to the available area inside the locule for seed development and to the presence and length of seed appendages. The juiciness of the fruits, given by the mesocarp cells and by secretion of substances inside the locules by the obturator, and the mucilaginous testa of seeds, appear later in the development, and are strategies to dispersal by animals. Palavras-chave: anatomy; berries; campos rupestres; germination; immunohistochemistry; Neoregelia; seed coat; seedling; vegetative organs; Vriesea. SUMÁRIO INTRODUÇÃO GERAL..........................................................................................................11 Referências bibliográficas ........................................................................................................19 CAPÍTULO 1. Anatomy of the vegetative organs, inflorescence axis and pedicel in the Neoregelia bahiana complex (Bromeliaceae): taxonomic and ecological importance ...……25 Abstract....…………………………………………………………………………………….26 Introduction ...……………………………………………………………….……………......27 Material and methods………………………………………………………..………..............28 Results.......................................................................................................................................29 Discussion.................................................................................................................................33 References.................................................................................................................................42 Illustrations................................................................................................................................47 Tables........................................................................................................................................66 CAPÍTULO 2. Variações morfológicas e anatômicas de raiz, folha, eixo da inflorescência e bráctea no complexo Vriesea oligantha (Bromeliaceae): perspectivas para a taxonomia e aspectos adaptativos..................................................................................................................71 Resumo.....…………………………………………………………………………………….72 Introdução…...……………………………………………………………………………......73 Material e métodos.....…………………………………………………..…………….............74 Resultados.................................................................................................................................75 Discussão..................................................................................................................................80 Referências................................................................................................................................86 Ilustrações.................................................................................................................................92 Tabelas....................................................................................................................................106 CAPÍTULO 3. Aspectos estruturais da semente, germinação e desenvolvimento pós-seminal em Neoregelia bahiana (Bromeliaceae): entendendo o desenvolvimento das plântulas..................................................................................................................................109 Resumo.....…………………………………………………………………………………...110 Introdução…...……………………………………………………………………………....111 Material e métodos.………....……………………………………………………….............112 Resultados...............................................................................................................................114 Discussão................................................................................................................................120 Referências..............................................................................................................................131 Ilustrações...............................................................................................................................138 Tabelas....................................................................................................................................158 CAPÍTULO 4. Seed development and its relation to fruit structure in species of Bromelioideae (Bromeliaceae) with fleshy fruits…………………………………………...161 Abstract...……………………………………………………………………………………162 Introduction...……………………………………………………………….…………….....163 Material and methods………………………………………………………..………............165 Results.....................................................................................................................................165 Discussion...............................................................................................................................169 References...............................................................................................................................174 Illustrations..............................................................................................................................178 Table........................................................................................................................................198 CONSIDERAÇÕES FINAIS..................................................................................................199 11 INTRODUÇÃO GERAL Bromeliaceae: morfologia e relações filogenéticas em um panorama atual Bromeliaceae inclui cerca de 3400 espécies, distribuídas em 58 gêneros que ocorrem, principalmente, nas regiões tropicais e subtropicais da América (LUTHER 2014). Pitcairnia feliciana (A. Chev.) Harms & Mildbr. é a única exceção, ocorrendo na África (JACQUES- FELIX 2000). Os representantes de Bromeliaceae crescem em diferentes ambientes e habitats, desde florestas úmidas a campos de altitude, com solos arenosos e pedregosos (SMITH 1934; PITTENDRIGH 1948; RUNDEL; DILLON 1998), o que demonstra grande versatilidade morfológica e ecofisiológica (BENZING 2000). No Brasil, a família está bem representada, com 46 gêneros e 1340 espécies (com pelo menos 1177 espécies endêmicas) relatadas até o momento, ocorrendo em diferentes fitofisionomias dos domínios Amazônia, Caatinga, Cerrado, Mata Atlântica, Pampa e Pantanal (BFG 2015). O domínio Mata Atlântica se destaca como um dos seus principais centros de diversidade (MARTINELLI ET AL. 2008). Seus representantes, em geral, são plantas herbáceas, epífitas, rupícolas, ou terrícolas, com raízes absortivas, por vezes apenas para a fixação (comum em espécies epífitas). O caule pode ser aéreo (geralmente curto), rizomatoso ou estolonífero. As folhas, com margens inteiras ou espinescentes, estão dispostas em roseta, podendo ou não formar um tanque armazenador de água. O eixo da inflorescência, com comprimento variável entre as espécies, geralmente está inserido no interior da roseta, porta inflorescências simples ou compostas, com flores monoclinas, que podem ser sésseis ou pediceladas, protegidas por brácteas. Sépalas e pétalas podem ser livres ou conadas. O androceu é diplostêmone. O gineceu possui estigma trífido e ovário súpero, ínfero ou semi-ínfero, tricarpelar e trilocular, com placentação axilar e nectários septais. Os frutos são do tipo cápsula (na grande maioreia das espécies) ou baga e as sementes podem ser aladas ou plumosas nos frutos do tipo cápsula. (SMITH; DOWNS 1974; SMITH; TILL 1998; BENZING 2000). Devido à coloração das folhas e brácteas, e pelo arranjo da inflorescência, grande parte de seus representantes apresenta potencial ornamental (BENZING 2000; SOUZA; LORENZI 2012), como espécies de Aechmea Ruiz & Pav., Billbergia Thunb., Neoregelia L.B.Sm e Vriesea Lindl., estudadas nessa tese. Do ponto de vista filogenético, a família está incluída em Poales (APG IV 2016) e, em conjunto com Thyphaceae, são as primeiras linhagens divergentes na ordem (BOUCHENAK- KHELLADI ET AL. 2014). Seu status monofilético é sustentado por dados moleculares, 12 citogenéticos e morfológicos (TOMLINSON 1969; BROWN; GILMARTIN 1989a, b; GIVNISH ET AL. 2007; BENZING 2000). Bromeliaceae foi tradicionalmente dividida em três subfamílias: Bromelioideae, Tillandsioideae e Pitcairnioideae, pela combinação de certos caracteres, como a morfologia da margem da folha, inteira ou espinescente; posição do ovário; tipo de fruto; e morfologia das sementes (SMITH; DOWNS 1974, 1977, 1979; SMITH; TILL 1998). Atualmente, a família encontra-se dividida em oito subfamílias: Brocchinioideae, Bromelioideae, Hechtioideae, Lindmanioideae, Navioideae, Pitcairnioideae, Puyoideae e Tillandsioideae, a partir de dados moleculares (GIVNISH ET AL. 2007). Isso porque nas análises filogenéticas a subfamília Pitcairnioideae mostrou-se parafilética e, a partir dela, foram criadas novas subfamílias, a fim de tornar os grupos monofiléticos. Nessas análises, Brocchinioideae aparece como grupo irmão das demais subfamílias, enquanto Bromelioideae aparece como uma das linhagens com divergência mais recente. Essas duas subfamílias compartilham flores epíginas, apesar do tipo de fruto ser diferente, cápsula ou baga (SAJO ET AL. 2004; GIVNISH ET AL. 2007). Segundo GIVNISH et al. (2007) a origem de Bromeliaceae se deu a partir de representantes terrícolas, no Planalto das Guianas, e se expandiu para outras regiões da América do Sul e Central, há cerca de 15 milhões de anos. Durante esse processo ocorreram especializações paralelas nas espécies que ocupam nichos semelhantes, o que pode explicar a similaridade morfológica (como o tanque armazenador de água), fisiológica (como o mecanismo CAM) e de formas de vida (como o epifitismo) observadas em linhagens distintas da família. Os complexos de espécies Neoregelia bahiana e Vriesea oligantha e a importância de caracteres anatômicos para a taxonomia e ecologia Apesar de monofilética, Bromeliaceae apresenta problemas de delimitação de gêneros e espécies (SMITH; DOWNS 1974, 1979; BENZING 2000), como se observa para os complexos de espécies Neoregelia bahiana (Bromelioideae) e Vriesea oligantha (Tillandsioideae), que são endêmicos dos campos rupestres da Cadeia do Espinhaço, nos Estados de Minas Gerais e Bahia, no Brasil (VERSIEUX ET. AL. 2008; BFG 2015), inclusive com ocorrência simpátrica. Neoregelia bahiana (Ule) L.B.Sm. apresenta grande variação morfológica (principalmente com relação à forma, coloração e textura das folhas), sendo considerados como seus sinônimos: Nidularium bahianum Ule, Aregelia bahiana (Ule) Mez, Neoregelia 13 bahiana var. bahiana Spruce ex Engl., Neoregelia bahiana var. viridis L.B.Sm., Neoregelia bahiana f. viridis (L.B.Sm.) L.B.Sm., Neoregelia hatschbachii L.B.Sm., Neoregelia pabstiana E.Pereira, Neoregelia diamantinensis E.Pereira, Neoregelia intermedia E.Pereira e Neoregelia bahiana f. bahiana (SMITH; DOWNS 1979; LEME 1998; BFG 2015). Todos esses nomes têm intrigado os pesquisadores a respeito de sua identidade como uma única espécie. Neoregelia foi indicado como um gênero parafilético no estudo de Santos-Silva et al. (2017), que incluiu dados estruturais foliares de gêneros próximos filogeneticamente, a saber: Canistrum E.Morren, Canistropsis (Mez) Leme, Edmundoa Leme, Nidularium Lem. e Wittrockia Lindm.; todos esses gêneros constituem o complexo Nidularioide (SANTOS- SILVA ET AL. 2017). Desta forma, esforços buscando indicar caracteres diagnósticos para espécies desse gênero são necessários a fim de estabelecer relações filogenéticas mais robustas, ainda mais levando-se em conta as variações estruturais das mesmas em resposta ao ambiente. Já Vriesea oligantha (Baker) Mez em conjunto com outras duas espécies Vriesea lancifolia (Baker) L.B.Sm. e Vriesea pseudoligantha Philcox compartilham, além de sua distribuição geográfica, caracteres vegetativos, como folhas densamente recobertas por escamas e propagação vegetativa com novos indivíduos se desenvolvendo na base da planta e no seu eixo da inflorescência, e reprodutivos, como a inflorescência simples com flores secundas na antese (SMITH; DOWNS 1977). Dessa forma, a caracterização anatômica de folhas, raiz, eixo da inflorescência e brácteas pode contribuir na delimitação destas espécies, bem como determinar se existem variações intraespecíficas decorrentes do ambiente. Ressalta-se que caracteres anatômicos vegetativos, principalmente das folhas, já foram abordados em vários estudos com representantes de Bromeliaceae, os quais trouxeram resultados relevantes para a taxonomia, contribuindo com a caracterização da família, gêneros e espécies (p. ex. TOMLINSON 1969; ROBINSON 1969; SAJO ET AL. 1998; AOYAMA; SAJO 2003; SCATENA; SEGECIN 2005; HORRES ET AL. 2007; PROENÇA; SAJO 2004, 2007; ALMEIDA ET AL. 2009; VERSIEUX ET AL. 2010; MONTEIRO ET AL. 2011; FARIA ET AL. 2012; GOMES-DA-SILVA ET AL. 2012; SANTOS-SILVA ET AL. 2014). Estudos anatômicos como esses têm indicado caracteres com potencial diagnóstico, como variações de espessamento de parede, posição dos estômatos em relação à epiderme, número de camadas no mesofilo e forma das células braciformes que preenchem os canais de ar. Grande parte desses trabalhos ainda discute a relação entre caracteres anatômicos foliares e o ambiente, indicando respostas quanto ao estresse hídrico e proteção contra o excesso de 14 luminosidade (p. ex. BENZING ET AL. 1978; LOESCHEN ET AL. 1993; REINERT; MEIRELLES 1993; PIERCE ET AL. 2001; FRESCHI ET AL. 2010; VOLTOLINI; SANTOS 2011; PEREIRA ET AL. 2013; SANTOS-SILVA ET AL. 2014; VIEIRA ET AL. 2017). Dentre esses caracteres, destacam-se as escamas peltadas, características da família, cuja evolução está relacionada com a redução estrutural e funcional do sistema radicular e com o desenvolvimento dos tecidos parenquimáticos e de suporte, e da hipoderme aquífera que armazena água no mesofilo (TOMLINSON 1969; BRAGA 1977). Já as raízes e o caule apresentam estrutura mais constante na família, como a presença de velame, córtex heterogêneo e cilindro vascular poliarco com células esclerenquimáticas na medula das raízes, e feixes vasculares com distribuição aleatória no caule (TOMLINSON 1969). As escamas peltadas e o parênquima clorofiliano das folhas, a organização das folhas formando um tanque armazenador de água e a presença de velame nas raízes são as principais características que permitiram a evolução da forma de vida rupícola e epifítica na família. Desenvolvimento de frutos em Bromelioideae Em relação aos frutos, apesar da importância dos mesmos para o reconhecimento de subfamílias e da forma de dispersão (SMITH; DOWNS 1974, 1977, 1979; SMITH; TILL 1998; BENZING 2000), são poucos os estudos que descrevem sua ontogenia (FAGUNDES; MARIATH 2010; SANTOS-SILVA ET AL. 2015; THADEO ET AL. 2015). Dentre esses estudos, destaca-se aquele de Fagundes e Mariath (2010) que estudaram espécies de Aechmea, Billbergia (Bromelioideae), Dyckia Schult. & Schult.f., Pitcairnia L’Hér (Pitcairnioideae), Tillandsia L. e Vriesea (Tillandsioideae), relacionando a estrutura do fruto com a deiscência: cápsulas de Pitcairnioideae e Tillandsioideae exibem linhas de deiscência pré-determinadas enquanto que em Bromelioideae os frutos são indeiscentes pela fusão de tecidos carpelares e não carpelares. Em Bromelioideae parece ocorrer a redução de tecidos esclerenquimatosos, os quais se relacionam com a indeiscência de seus frutos (FAGUNDES; MARIATH 2010). Nas outras subfamílias estudadas células lignificadas ocorrem no exocarpo, mesocarpo e endocarpo em frutos de Tillandsioideae e no exocarpo ou no mesocarpo e endocarpo em frutos de Pitcairnioideae (FAGUNDES; MARIATH 2010; SANTOS-SILVA ET AL. 2015). No estudo recente de Thadeo et al. (2015), analisou-se a ontogenia de frutos carnosos de monocotiledôneas, incluindo uma espécie de Bromelioideae, Aechmea aquilega (Salisb.) Griseb. Segundo Thadeo et al. (2015), frutos carnosos tiveram várias origens independentes nas monocotiledôneas, inclusive em Poales. No entanto, pouco se sabe sobre a ontogenia e 15 anatomia dos frutos carnosos em Bromeliaceae e estudos são necessários para que se possa entender sua estrutura. Estrutura da semente e desenvolvimento das plântulas Tradicionalmente, as sementes de Bromeliaceae são apontadas como ótimas ferramentas para a circunscrição de subfamílias, além de suas características estarem relacionadas ao modo de dispersão. Tillandsioideae apresenta sementes filiformes e com apêndices plumosos, enquanto que em Pitcairnioideae s.l. as sementes podem ser discóides ou aladas, indicando dispersão pelo vento em ambas as subfamílias (MÜLLER 1895; SMITH; DOWNS 1974, 1977; VARADARAJAN; GILMARTIN 1988; SMITH; TILL 1998; BENZING 2000). As Bromelioideae, entretanto, possuem sementes com envoltório mucilaginoso, usualmente denominado de sarcotesta (Smith; Till 1998), associado à dispersão zoocórica (SMITH; DOWNS 1974, 1979; VARADARAJAN; GILMARTIN 1988; SMITH; TILL 1998; BENZING 2000). Em relação à ontogenia das sementes, essa foi estudada em espécies de Tillandsioideae cujos apêndices plumosos se formam pelo alongamento e posterior separação das células da testa (SZIDAT 1922; PALACÍ ET AL. 2004). Para Pitcairnioideae s.l. também foi discutido que o aspecto morfológico mais variável de suas sementes (nuas, aladas ou caudadas) se deve à estrutura da testa (VARADARAJAN; GILMARTIN 1988). Já para Bromelioideae foi hipotetizado que o envoltório mucilaginoso das sementes deve desintegrar no interior dos frutos, fazendo parte da suculência dos mesmos (SZIDAT 1922; SMITH; DOWNS 1974; BENZING 2000); desta forma, o envoltório na semente madura seria formado apenas pelo tégmen. O envoltório mucilaginoso também tem sido associado com a função de proteção das sementes, evitando o dessecamento das mesmas no ambiente (SILVA; SCATENA 2011). Nesse sentido, os contituintes das células desse tecido merecem ser investigados para uma posterior comparação com a dispersão das sementes ou com sua capacidade de retenção de água. Seguindo a segunda hipótese, já foi relatado por Wester e Zotz (2011) que um substrato mais estável e úmido permite maiores taxas de germinação e de desenvolvimento de plântulas de Catopsis sessiflora (Ruiz & Pav.) Mez (Tillandsioideae). Abaixo da testa, as sementes maduras de Bromeliaceae apresentam duas camadas no tégmen, constituindo o exotégmen com células de paredes espessadas e o endotégmen armazenando compostos fenólicos no interior de suas células (SZIDAT 1922; SMITH; DOWNS 1974; PALACÍ ET AL. 2004; MAGALHÃES; MARIATH 2012; PRADO ET AL. 16 2014). Apesar da estrutura uniforme desse tecido, em relação ao número de camadas, o grau de espessamento de parede de suas células pode ser importante para o reconhecimento de gêneros (BENZING 2000). As sementes de Bromeliaceae são, na sua maioria, albuminosas, com endosperma diferenciado na camada de aleurona, adjacente ao envoltório seminal, e no parênquima armazenador de amido (SZIDAT 1922; SMITH; DOWNS 1974; BENZING 2000; PALACÍ ET AL. 2004; MAGALHÃES; MARIATH 2012; PRADO ET AL. 2014). O embrião nas Bromeliaceae ocupa, geralmente, 1/3 da semente e é diferenciado em cotilédone e eixo hipocótilo-radicular (MÜLLER 1895; BILLINGS 1904; SZIDAT 1922; SMITH; DOWNS 1974; BENZING 2000; SCATENA ET AL. 2006; MENDES ET AL. 2010; MAGALHÃES; MARIATH 2012; PRADO ET AL. 2014). Durante a germinação o embrião se desenvolve e promove a ruptura do envoltório seminal (TILLICH 2007). A germinação dos representantes de Tillandsioideae parece ser marcada pelo desenvolvimento do cotilédone (MÜLLER 1895; SCATENA ET AL. 2006; PEREIRA ET AL. 2008, 2009; SILVA; SCATENA 2011), enquanto que em representantes de Bromelioideae e Pitcairnioideae s.l. ocorre primeiramente o desenvolvimento da raiz primária (MÜLLER 1895; PEREIRA 1988; BENZING 2000; MANTOVANI; IGLESIAS 2005; SCATENA ET AL. 2006; TILLICH 2007; PEREIRA ET AL. 2008, 2009, 2010; SILVA; SCATENA 2011). Dentre esses estudos, destaca-se o trabalho de Tillich (2007), que descreve não apenas para Bromeliaceae, mas para Poales em geral, a estrutura do cotilédone durante o desenvolvimento pós-seminal. Segundo esse autor, a porção superior do cotilédone (hiperfilo) de Bromeliaceae se mantém aderida ao endosperma, com função haustorial. Já a porção basal (hipofilo) é mais variável, podendo ser modificada em uma bainha pouco desenvolvida, ou com um lobo mediano (hipofilo compacto), ou como uma estrutura laminar (hipofilo laminar). Outro trabalho importante é o de Pereira (1988), que descreve o desenvolvimento pós-seminal de várias espécies de Bromelioideae, comparando a morfologia de plântulas entre os gêneros, como por exemplo, para Neoregelia, cujas espécies possuem diferentes formas de vida e crescem em diferentes fitofisionomias (BFG 2015), sendo um gênero-chave para se investigar as adaptações dos indivíduos em estágios juvenis. As modificações durante a transição entre os estágios de desenvolvimento da planta são importantes para o estabelecimento de Bromeliaceae em diferentes ecossistemas. Sabe-se, por exemplo, que as folhas jovens de Tillandsia deppeana Steud. são mais densamente cobertas por tricomas do que as dos indivíduos adultos, sendo tolerantes a longos períodos de seca (ADAMS; MARTIN 1986). Isso mostra que alguns caracteres estruturais estão 17 envolvidos com o desenvolvimento e estabelecimento das plântulas e podem indicar influências do habitat na seleção de caracteres morfológicos e anatômicos durante o desenvolvimento pós-seminal. Um ponto que ainda permanece com poucas informações para a família se refere à anatomia dos órgãos das plântulas, cujo desenvolvimento envolve processos de divisão e alongamento celular, diferenciação dos tecidos e órgãos, os quais são extremamente dependentes da dinâmica dos componentes da parede celular durante a ontogênese vegetal (ALBERSHEIM ET AL. 2011). Assim, elucidar esses processos mostra- se essencial para o entendimento da estrutura do corpo da planta e suas modificações/adaptações ao longo do desenvolvimento. Estrutura geral da tese e apresentação dos capítulos Pelo conjunto de fatores aqui apresentados quanto à dificuldade no reconhecimento de espécies, e daqueles que são úteis para o entendimento sobre a dispersão e o estabelecimento de suas espécies, essa tese foi organizada em quatro capítulos:  Capítulo 1. Anatomy of the vegetative organs, inflorescence axis and pedicel in the Neoregelia bahiana complex (Bromeliaceae): taxonomic and ecological importance Nesse capítulo foram estudas 12 populações ao longo da Cadeia do Espinhaço, buscando descrever as variações morfológicas de raiz, estolho, folhas, eixo da inflorescência e pedicelo de seus representantes em condições naturais e em cultivo. Identificou-se um conjunto de caracteres que delimitam a espécie (maior parte das populações estudadas), enquanto que caracteres exclusivos para apenas três populações mostraram-se úteis para a delimitação destas.  Capítulo 2. Variações morfológicas e anatômicas de raiz, folha, eixo da inflorescência e bráctea no complexo Vriesea oligantha (Bromeliaceae): perspectivas para a taxonomia e aspectos adaptativos Nesse capítulo foram estudadas 17 populações ao longo da Cadeia do Espinhaço, buscando descrever as variações morfológicas de raiz, folhas, eixo da inflorescência e brácteas de seus representantes em condições naturais e em cultivo. Os caracteres com valor diagnóstico foram 18 submetidos à análise de similaridade entre as populações e indicam a existência de mais de um táxon.  Capítulo 3. Aspectos estruturais da semente, germinação e desenvolvimento pós- seminal em Neoregelia bahiana (Bromeliaceae): entendendo o desenvolvimento das plântulas Já nesse capítulo, sabendo da ampla variação morfológica e distribuição geográfica de N. bahiana, sementes de duas de suas populações foram colocadas para germinar e as estapas do desenvolvimento pós-seminal foram descritas. Adicionalmente, a anatomia e o desenvolvimento dos órgãos das plântulas, incluindo análises imunohistoquímicas, foram realizadas a fim de contribuir com o entendimento sobre o estabelecimento das plântulas.  Capítulo 4. Seed development and its relation to fruit structure in species of Bromelioideae (Bromeliaceae) with fleshy fruits Por fim, nesse capítulo foi estudado o desenvolvimento de frutos e sementes de três espécies de Bromelioideae. Os resultados foram discutidos em um panorama evolutivo, taxonômico e ecológico. 19 Referências bibliográficas ADAMS, W. W; MARTIN, C; E. 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Oriani, Departamento de Botânica, Instituto de Biociências, Universidade Estadual Paulista (UNESP), C. Postal 199, Rio Claro, SP 13506-900, Brazil. – L. M. Versieux, Departamento de Botânica e Zoologia, Centro de Biociências, Universidade Federal do Rio Grande do Norte, Campus Universitário, Natal, RN 59078-970, Brazil. Corresponding author: K. R. Silva, Departamento de Botânica, Instituto de Biociências, Universidade Estadual Paulista (UNESP), C. Postal 199, Rio Claro, SP 13506-900, Brazil. E- mail: kleber_resende@hotmail.com 1 Artigo publicado no periódico Nordic Journal of Botany (2018:e01800) 26 Abstract Delimitation of Bromeliaceae into genera or species is often problematic, as observed for the Neoregelia bahiana complex, distributed throughout the rocky fields of Espinhaço Range, Brazil. Considering that the anatomical characterisation of different organs is potentially important for taxonomic and ecological interpretation of this complex, we analysed roots, stems (stolon), leaves, inflorescence axes (peduncle), and pedicels in individuals from different populations. In all the studied individuals, the roots are composed of velamen, a heterogeneous cortex, and a polyarch vascular cylinder with sclerenchymatous pith. The stolon features a parenchymatous cortex and collateral vascular bundles randomly distributed in the vascular cylinder. This organ may increase in diameter by the formation of new vascular bundles and a multi-layered cork. The leaf blade has epidermal cells with U-shaped thickened walls and peltate scales occur on the adaxial surface. The mesophyll consists of mechanical and water-storage hypodermis and a heterogeneous chlorenchyma. The inflorescence axis and the pedicel have a parenchymatous cortex and vascular bundles randomly distributed in an aerenchyma. Some variable leaf characters, such as presence of air lacunae in the mesophyll, are related to the size of the individuals and were interpreted as phenotypic variations related mainly to sunlight incidence. Leaf characters such as lamina shape, distribution of the peltate scales, and number of cell layers forming the water-storage hypodermis distinguish the populations of the Serra do Cabral and one population of the Diamantina (Minas Gerais) from the remaining studied populations, suggesting the existence of more than one taxon. Keywords: leaf; Neoregelia; Poales; root; rocky fields; stolon 27 Introduction Bromeliaceae are an early-diverging lineage of the order Poales (Linder and Rudall 2005, Givnish et al. 2010). Although monophyletic, the family is problematic in terms of generic and specific delimitation (Smith and Downs 1974, 1979, Benzing 2000, Santos-Silva et al. 2017), as observed in some complexes of species such as the Neoregelia bahiana (Ule) L.B.Sm. (Bromelioideae). Neoregelia bahiana is a rupicolous species endemic to Brazil, restricted to the rocky fields of the Espinhaço Range, a mountain chain located in the states of Minas Gerais and Bahia (Versieux et. al. 2008, BFG 2015). The species exhibits a wide morphological variation (particularly in the shape, size and colouration of the leaves) and a broad synonymy exists in the literature, including both homotypic (Nidularium bahianum Ule, Aregelia bahiana (Ule) Mez) and heterotypic synonyms and taxa of different hierarchical levels: Neoregelia bahiana var. viridis L.B.Sm., Neoregelia bahiana forma viridis (L.B.Sm.) L.B.Sm., Neoregelia hatschbachii L.B.Sm., Neoregelia pabstiana E.Pereira, Neoregelia diamantinensis E.Pereira, Neoregelia intermedia E.Pereira, Neoregelia bahiana forma bahiana (Smith and Downs 1979, BFG 2015). In this context, the anatomical characterisation of individuals from different populations of Neoregelia bahiana is a potential source of diagnostic characters for the species, and may help ascertain the existence of intraspecific or interpopulational variations related to the environment. The importance of anatomical data for the taxonomic and ecological understanding of Bromeliaceae has already been demonstrated in many studies involving different organs such as root, stem, leaf and the inflorescence axis (e.g. Tomlinson 1969, Sajo et al. 1998, Pita and Menezes 2002, Aoyama and Sajo 2003, Segecin and Scatena 2004a, 2004b, Scatena and Segecin 2005, Horres et al. 2007, Proença and Sajo 2004, 2007, 2008a, 2008b, Versieux et al. 2010, Monteiro et al. 2011, Silva and Scatena 2011a, 2011b, Faria et al. 2012, Reinert et al. 2013, Santos-Silva et al. 2014). For the Nidularioid complex, which comprises the genera Nidularium Lem., Canistrum E. Morren, Canistropsis (Mez) Leme, Edmundoa Leme, Wittrockia Lindm., and Neoregelia L. B. Sm., a recent phylogenetic study of representatives of these genera, including N. bahiana, showed that the leaf anatomical characters are useful for the phylogeny of the group (Santos-Silva et al. 2017). This study indicated that Neoregelia is paraphyletic, highlighting the importance of obtaining new anatomical data that can be used in future phylogenetic analyses of the genus. 28 Considering the wide variation in leaf morphology and plant architecture within the Neoregelia bahiana complex, the objective of this study was to describe the morphology and anatomy of the vegetative organs, inflorescence axis and pedicel of individuals collected from different locations along the Espinhaço mountain range, seeking to answer the following questions: Which characters have taxonomic value and contribute to the delimitation of the species or other taxa? How do the presented results increase the anatomical data available for the family? Which characters may be interpreted as adaptations to the environment and to the rupicolous life form? Materials and methods Individuals of Neoregelia bahiana were collected in rocky fields in the Brazilian states of Minas Gerais- MG (Grão Mogol, Santo Antônio do Itambé, Serra do Cabral, Serra do Cipó, and Diamantina municipalities) and Bahia- BA (Abaíra, Mucugê and Rio de Contas municipalities), as listed in Table 1. The general characteristics of luminosity for each collection point were recorded and on the basis of these characteristics the plants were classified as sciophyte (growing in total or partial shade) or heliophyte (growing under direct sunlight) (Table 1). Figure 1 shows the map of the populations’ distribution, which was generated based on the geographic coordinates of the collection points using the public available Quantum Geographic Information Systems software (QGIS ver. 2.12.3). We kept some individuals in a greenhouse to observe possible variations in leaf morphology and plant architecture under cultivation conditions. Voucher specimens for each population are deposited in the Herbarium Rioclarense (HRCB) and duplicates in the herbarium of Universidade Federal do Rio Grande do Norte (UFRN). For anatomical studies, roots, stolons, leaves (margin and central region of the lamina), inflorescence axis (peduncles) and pedicels of at least three individuals (rosettes) from each population were fixed in FAA 50 (37% formaldehyde, glacial acetic acid, 50% ethanol, 1:1:18 v/v) (Johansen 1940) and stored in 70% ethanol. Subsequently, the samples were dehydrated in a n-butyl alcohol series and embedded in (2-hydroxyethyl)-methacrylate (Leica Historesin Embedding Kit) (Gerrits and Smid 1983). Paradermal, cross and longitudinal sections were made in the middle portion of the studied organs using a rotary microtome (Leica RM 2245). For the stolon, cross sections were also made in its distal portion. The sections were stained with periodic acid–Schiff’s reagent (PAS) and toluidine blue (O’Brien et al. 1964, Feder and O’Brien 1968) and mounted on permanent slides using 29 Entellan (Merck). We also performed cross and longitudinal handmade sections, which were bleached in a 20% sodium hypochlorite solution, double stained with safranin and Astra Blue (Bukatsch 1972) and mounted on slides with water. The images were obtained with a microscope (Leica DM 4000B) equipped with a camera (Leica DFC 450), using the software LAS (Leica Application Suite Version 4.0.0). Histochemical tests were performed with Sudan III for lipid (Sass 1951), phenol for silica compounds (Johansen 1940), and hydrochloric acid and glacial acetic acid solutions for calcium oxalate detection (Chaberlain 1932). For the scanning electron microscopy (SEM) study, the samples were dehydrated in an ethanol series (80, 90 e 100%), critical point dried (Balzers CPD 050), and coated with gold (Bal-Tec SCD 050). The results were documented by images obtained with a camera coupled to microscope (Zeiss, LEO 435 VP). Quantitative data on the average size of the leaf rosette (average ± standard deviation) relative to the height and diameter (measured from the top view of the rosette) were obtained using the measurements from three adult individuals per population. We also measured the entire leaf length (including the leaf sheath) and the lamina width of three leaves on each of three individuals per population (totalizing nine leaves per population). Results Morphology – Neoregelia bahiana individuals grow on rocky outcrops (Fig. 2a) and occasionally on trees (Fig. 2b, Table 1). Its leaves, green to reddish in colour, are spirally arranged and overlapping on a short aerial stem, forming a small tank capable of storing water (Fig. 2c-d). In some populations from Minas Gerais (Populations 8, 10 and 12), the leaf apex is pinkish (Fig. 2d, Table 2). In the remaining populations, the leaf apex is green or reddish green (Figs. 2c, 3d, Table 2). In most of the studied individuals (Populations 1-5, 8-10 and 12), the leaf sheath is narrow (Fig. 3a) and the lamina is linear, varying in length (Fig. 3d - leaves 3 and 4). In the Populations 6 and 7 (Serra do Cabral, MG), the lamina is lanceolate with a wide sheath (Fig. 3b, d - leaf 1), whereas in the Population 11 (Diamantina, MG) the lamina is rounded to oblong with also a wide sheath (Fig. 3c, d - leaf 2, Table 2). Regarding the position of the leaf lamina relative to the plant’s axis, the erect ascending position is predominant (Fig. 3a-b), except for the individuals of Population 11, which exhibit suberect- patent laminas (Fig. 3c). 30 The inflorescence axis, which bears the flowers and fruits, is achlorophyllous and nested in the leaf rosette (Fig. 2e). The vegetative propagation of the species occurs naturally through the formation of stolons, from which new ramets are formed (Figs. 2e, 3a-b, arrows). Adventitious roots develop in the distal portion of the stolon and from the short aerial stem (Fig. 3a-b, arrowheads). In general, the individuals of the populations from Minas Gerais (Populations 5-12) (Figs. 2d-e, 3b-c) and of one population from Mucugê, BA (Population 3) are larger than those of the remaining populations from Bahia, exhibiting rosettes of greater height and diameter as well as longer leaves (Figs. 2d, 3b-d, Table 2). Only one population from Minas Gerais (Population 8) had smaller individuals, with an average size similar to that recorded for Populations 1, 2 and 4 from Bahia (Table 2). All of the smaller individuals are heliophilic, growing under high luminosity (Table 1). The individuals that grew in shaded areas (Table 1) generally exhibited rosettes of larger diameter and longer leaves with narrower laminas (Table 2). Root anatomy – The roots consist of the velamen, heterogeneous cortex and vascular cylinder (Fig. 4a-b). The outermost layer of the velamen is the epivelamen, composed of cells differentiated into root hairs. The remaining layers of the velamen are formed by cells with slightly thickened and lignified walls (Fig. 4b). The outer cortex has 1–3 layers of cells with thickened and lignified walls (Fig. 4b). The middle cortex is parenchymatous and is formed by about 10 layers of cells in compact arrangement (Fig. 4b). Idioblasts occur in the velamen, outer cortex and middle cortex (Fig. 4b, asterisks). These idioblasts are elongated longitudinally and contain raphides as well as mucilage, which reacted positively to the PAS, indicating the presence of polysaccharides (Fig. 4d, arrow). The inner cortex has 6–7 layers of rounded, thin-walled cells that delimit conspicuous intercellular spaces and are arranged radially around the endodermis (Fig. 4a-c). Sclereids occur between the middle cortex and inner cortex (Fig. 4b, detail). The endodermis is one-layered and composed of cells with thickened anticlinal walls (Fig. 4c). The pericycle is one-layered and formed by thin-walled cells (Fig. 4b-c). The vascular cylinder is polyarch, with 20–30 protoxylem poles and a sclerenchymatous pith (Fig. 4c). Stolon anatomy – The stolon has an atactostele, with a defined cortex and vascular bundles distributed randomly in the pith (Fig. 5a). During its development, the stolon becomes more rigid due to thickening and lignification of the cell walls, mainly of the epidermis, hypodermis 31 and pericycle, which can be observed when comparing cross sections of the younger, distal portion (Fig. 5b, f) to the middle portion (Fig. 5d, g). In the distal portion of the stolon, the epidermis is single-layered and its cells have U- shaped thickened walls and contain silica bodies (Fig. 5b, arrow). The cortex is composed of rounded, thin-walled cells (Fig. 5e) and exhibits vascular traces (Fig. 5f), fiber bundles, and adventitious roots (Fig. 5k). The endodermal cells are undifferentiated from the other cortical cells (Fig. 5f). The vascular cylinder is delimited by a multi-layered pericycle, which is parenchymatous (Fig. 5f). The vascular bundles are collateral, simple or compound, and surrounded by a fibrous sheath (Fig. 5f). In the pith, as in the cortex, the cells are rounded, thin-walled, and delimit conspicuous intercellular spaces (Fig. 5e). Idioblasts containing raphides and polysaccharides occur in both the cortex and the pith (Fig. 5f, asterisks). In the middle portion of the stolon, the thickening of the epidermal cell walls is more pronounced, thus their lumen is greatly reduced (Fig. 5c-d). In this portion of the stolon a new multi-layered protective tissue is produced in the outer layers of the cortex (Fig. 5c-d) and later its cells become suberized. Vascular traces and fiber bundles (Fig. 5i) are also observed in the cortex (Fig. 5g). The pericycle cells have thickened, lignified walls and there are more vascular bundles in the vascular cylinder, which are surrounded by fibrous sheaths (Fig. 5g- h). The cells of the pith may contain starch grains (Fig. 5j). Leaf anatomy – The leaf lamina is composed of epidermis, hypodermis, chlorenchyma, and the vascular system (Fig. 6a-e). The epidermis is single-layered, with U-shaped thick-walled cells (Fig. 7a, d). The epidermal cells may contain silica bodies (Fig. 7b, f, arrow) and are covered by a relatively thick cuticle (Fig. 7a, f). From the frontal view, the silica-containing cells have a quadrangular shape and sinuous walls (Fig. 7b). The stomata are tetracytic (Fig. 7e), with wide substomatal chambers (Figs. 6g-h, 7c-d, asterisks). They are arranged in bands on the abaxial surface of the lamina (Fig. 8b-c, f, arrowheads), slightly below the remaining epidermal cells (Figs. 6g-h, 7c), at the same level as the mechanical hypodermis (Fig. 7e). In the individuals of Populations 6, 7 (Serra do Cabral, MG), 10 (Diamantina, MG) and 12 (Serra do Cipó, MG), the stomata lie in intercostal grooves (Fig. 7d). These grooves are more pronounced in Populations 6 and 7, giving the abaxial surface of the lamina a more sinuous contour (Fig. 6e). In the remaining populations, the grooves occur between the bands of stomata, in the costal regions (Figs. 7c, 8b). Peltate scales are present on the adaxial surface of the lamina in all the studied individuals (Fig. 8a-d). However, they are present on the abaxial surface only in individuals of 32 Populations 6, 7 (Serra do Cabral, MG), 10 (Diamantina, MG) and 12 (Serra do Cipó, MG), occurring in the intercostal grooves, together with the stomata (Fig. 8c-d, Table 3). In Populations 6 and 7, the scales are abundant and arranged in bands, with their shields overlapping (Fig. 8d, Table 3). In Populations 10 and 12, the scales are less numerous and arranged in rows (Fig. 8c). Each scale consists of 1–2 basal cells, a stalk with 3–4 cells, and a multicellular shield (Fig. 7f). The number of shield cells varies between individuals, or even between scales on the same leaf (Fig. 8e-f). The shield has polygonal cells, and the peripheral cells are elongated (Fig. 8g). All shield cells exhibit thickened inner periclinal and anticlinal walls (Figs. 7f, 8e-g). A hypodermis differentiated into supporting tissue (mechanical hypodermis) and water-storage tissue is present on both surfaces of the lamina (Figs. 6a-e, g-h, 7a, d, f). The mechanical hypodermis is composed of lignified, thick-walled cells (Figs. 6b, d, 7e), with two or more layers of cells facing the adaxial surface (Fig. 7a) and one or two layers of cells facing the abaxial surface (Fig. 7d, f). The water-storage tissue, which is more developed on the adaxial surface, especially in the centre of the lamina, is composed of 4–6 layers of thin- walled cells with large lumen (Fig. 6a). Only in the individuals of Populations 6, 7 (Serra do Cabral, MG) and 11 (Diamantina, MG), the water-storage tissue is composed of more than 10 layers of cells (Fig. 6c, Table 2). In the centre of the lamina, the cells of the innermost layers of the water-storage tissue have elongated and sinuous anticlinal walls (Fig. 6c). Toward the margin of the lamina, the water-storage cells are rounded (Fig. 6b, d), except in the leaves of individuals of Populations 6 and 7, for which all the cells of the innermost layers of the water- storage tissue are anticlinally elongated (Fig. 6e). On the abaxial surface, the water-storage tissue is formed by 2–3 layers of rounded cells (Figs. 6g-h, 7d, f). These layers are interrupted by the substomatal chambers, which are supported by arm cells (Figs. 6g-h, arrowhead, 7c-d). Idioblasts containing raphides and polysaccharides occur in the water-storage tissue facing the adaxial surface and in the chlorenchyma near the leaf margin (Fig. 6j). The leaf margin in cross section is obtuse to acute in the individuals of Populations 1–5, 8–10 and 12 (Fig. 6b) and always acute in the individuals of Populations 6, 7 and 11 (Fig. 6d–e). The margin also exhibits spinescent emergences formed by the elongation of the thickened, lignified cells of the mechanical hypodermis (Fig. 6f). The chlorenchyma is heterogeneous, with palisade cells facing the adaxial surface and rounded cells facing the abaxial surface of the lamina (Fig. 6a-c, i). Small druses (calcium oxalate crystals) occur in the chlorenchyma cells (Fig. 6k, arrow). In individuals of Minas Gerais (Populations 5–7, 9–12) and one population of Mucugê, BA (Population 3), air lacunae 33 are interspersed with the vascular bundles in the centre of the lamina; they are connected to the substomatal chambers and may be large (Fig. 6a, c, g) or small (Fig. 6i, Table 2). In the remaining populations, the air lacunae are restricted to the substomatal chambers (Fig. 6h, Table 2). These lacunae are supported by arm cells (Fig. 6g, i). The vascular bundles are distributed in a single row in the cross section of the lamina and are not buttressed to surface layers (Fig. 6a-e). They are collateral and surrounded by a double sheath, the outer parenchymatous and the inner sclerenchymatous (Fig. 6g-i). The fibrous caps on the xylem and the phloem are more developed in the smaller vascular bundles, which alternate with the larger ones (Fig. 6g, i). In the individuals of Population 11 (Diamantina), the fibrous cap on the phloem are prominent and may form fiber strands isolated in the mesophyll (Fig. 6i). Commissural vascular bundles interconnect the vascular bundles that are distributed longitudinally through the leaf (Fig. 6l). Anatomy of the inflorescence axis and pedicel – The inflorescence axis and the pedicel are cylindrical, with sinuous contours (Fig. 9a-b). They have a single-layered epidermis, composed of papillose thin-walled cells (Fig. 9d). The cortex and the vascular cylinder are easily distinguishable (Fig. 9a-c). The cortex is narrow relative to the vascular cylinder (Fig. 9a-b) and consists of rounded, thin-walled cells (Fig. 9c). In the inflorescence axis, vascular traces occur in the cortex (Fig. 9e). The vascular bundles are simple (Fig. 9h) or compound (Fig. 9i), and arranged randomly within the vascular cylinder (atactostele) (Fig. 9a-b). Most of the vascular bundles are collateral (Fig. 9h-i); however, some bundles formed by only phloem were also observed (Fig. 9j, arrowhead). The vascular bundles are surrounded by cellulosic thin-walled cells, except in the pedicels of the individuals of Population 1, in which the vascular bundles are surrounded by cells with lignified walls (Fig. 9j). The pith is composed of both rounded cells and arm cells (Fig. 9c, g), the later delimiting air spaces, which are larger in the pedicel (Fig. 9b). Idioblasts containing raphides and polysaccharides occur in both cortex and pith (Fig. 9f). Starch grains were found in the pith cells of the inflorescence axis, near the vascular bundles, confirmed by the positive reaction to PAS. Discussion Contributions to the taxonomy of Neoregelia bahiana – The Populations 1–5, 8–10 and 12 show the same anatomical pattern, without qualitative variations, in the roots, stolons, leaves, 34 inflorescence axes and pedicels, indicating that they belong to a single species: Neoregelia bahiana. Therefore, we can point to a set of characters that are useful in delimiting the species: root with velamen, heterogeneous cortex and polyarch vascular cylinder; stolon with atactostele, presenting growth in thickness; linear-shaped lamina without peltate scales on the abaxial surface or, when present, not overlapping; heterogeneous mesophyll, with mechanical and water-storage tissues on both surfaces of the lamina and palisade chlorenchyma facing the adaxial surface; water-storage tissue consisting of up to 10 layers of cells; and the presence of idioblasts containing raphides and polysaccharides in all organs. The individuals of Populations 6 and 7 (Serra do Cabral, MG) are differentiated by the presence of lanceolate lamina, always with acute margins and with water-storage tissue composed of more than 10 layers of anticlinally elongated cells, as well as the presence of overlapping peltate scales on the lamina abaxial surface. The individuals of Population 11 (Diamantina, MG) also exhibit laminas with acute margins and with water-storage tissue consisting of more than 10 layers of cells; however they are differentiated by their rounded to oblong, suberect-patent laminas and by the presence of a prominent fibrous cap on the phloem, which may form fiber strands dispersed in the chlorenchyma. Such characters together delimit these populations and suggest the presence of more than one taxon in the complex. It must be emphasized that the Populations 6 and 7 occur in the Serra do Cabral, Minas Gerais, in a more xeric environment than the other rocky fields in the central or southern part of the Espinhaço montain range. Santos-Silva et al. (2017) recently studied the leaf anatomy of Neoregelia bahiana from a phylogenetic perspective and most of the character states identified by the authors agree with those described here, namely adaxial and abaxial epidermis with cells with thickened walls and a small lumen; scale stalk having more than two cells; adaxial mechanical hypodermis composed of more than one layer of thick-walled cells, and abaxial hypodermis composed of one layer of thick-walled cells and one or more transition layers; abrupt transition between water-storage tissue and chlorenchyma; and smaller vascular bundles with fibrous cap (taller than wide). The extension of the water-storage tissue described by Santos-Silva et al. (2017) (up to 1/3 of the thickness of leaf blade) agrees with that observed in most of the populations studied here, except for the Populations 6, 7 (Serra do Cabral) and 11 (Diamantina), indicating the diagnostic potential of this character. The presence of extravascular fiber strands in the leaves of Population 11 (indicated as absent by Santos-Silva (2017) can also be used to delimit this population. Our results also demonstrate that the following character states described by 35 Santos-Silva et al. (2017) as diagnostic for N. bahiana: presence of air lacunae composed of cells with short arms in the mesophyll, and anticlinally elongated cells forming the water- storage tissue, may vary among the populations, and should be used with caution in future phylogenetic analyses. In addition to these interpopulational structural variations, we also observed the presence of palisade parenchyma in the leaves of all studied populations, a character described as absent in N. bahiana by Santos-Silva et al. (2017). These authors also indicated the occurrence of stomata at the same level as the remaining epidermal cells; however, our results show that the stomata are located slightly below the other epidermal cells, and they can occur in intercostal grooves, as in the individuals of Populations 6, 7 (Serra do Cabral), 10 (Diamantina) and 12 (Serra do Cipó). These anatomical differences emphasize the need for the taxonomic delimitation of N. bahiana, and the need to understand how representatives of the Nidularioid complex respond to the environmental factors for a more robust phylogeny. The air lacunae alternated with the vascular bundles in the mesophyll is a character with interpopulational variation. They are absent in most of the populations from Bahia (Populations 1, 2 and 4) and in one population from Minas Gerais (Population 8), whose individuals are smaller (reduced diameter and height of the rosette and shorter leaf length) and grow in areas of high light intensity. Therefore, the presence of air lacunae may be related to the size of the lamina (longer leaves have air lacunae), which is in turn related to environmental factors, particularly sunlight incidence. Dai et al. (2009) demonstrated that in Tetrastigma hemsleyanum Diels et Gilg (Vitaceae), the plants growing in a 67% shade condition showed a higher photosynthetic activity and growth. When the light intensity exceeded that of 50% shade, photosynthetic activity was depressed likely due to photoinhibition (Dai et al. 2009). In the same way, the presence of air lacunae in the longer leaves of N. bahiana may be related to a higher photosynthetic activity and growth, since the air lacunae may facilitate the gas flow within the lamina, improving the photosynthetic performance and the carbon gain. A recent study that associated the leaf anatomy with the photosynthetic pathways in Bromeliaceae showed that CAM plants, in general, show a higher chlorenchyma thickness and reduced internal air spaces (Males 2018). In this sense, the absence of air lacunae in the heliophyte individuals of N. bahiana may indicate an ability to express a degree of CAM in response to stresses such as high sunlight, as demonstrated for other bromeliad species (Pierce et al. 2002). In addition to the length, sunlight incidence may also influence the width of the lamina, since the individuals occurring in shady areas have narrower laminas compared to those occurring in sunny areas of the same location 36 (Populations 2 and 3, 6 and 7, 8 and 9). However, it should be noted that light is not the only factor influencing leaf ontogeny and regulating the morphological aspects of the organ. In the Populations 10 (Diamantina, MG) and 12 (Serra do Cipó, MG), for example, whose individuals grow exposed to high luminosity, the leaves are long and have wide air lacunae. This longer leaf length may be related to the fact that, in these individuals, the stomata lie in the intercostal grooves, protected by the scales, conferring an adaptive advantage. Another variable character between populations of N. bahiana is the pink colouration of the leaf apex, present in the individuals of Population 8, 9 (Santo Antônio do Itambé, MG), 10 (Diamantina, MG) and 12 (Serra do Cipó, MG). The most uniform morphological aspect among these populations may be due to their relative geographic proximity (Fig. 1), sharing more genetic similarities among themselves. Despite the genetic factor, we observed that the light incidence can also influence this pattern of colouration. For example, individuals of the Population 9 that grow in shaded areas of the gallery forests have a green leaf apex, while those individuals growing in areas between the gallery forest and the rocky outcrops (more exposed to the sun) have a pinkish leaf apex. Furthermore, in individuals of the four populations transplanted and maintained under cultivation in a greenhouse, the leaf apices ended up presenting a less intense pinkish colour. Pereira et al. (2013), comparatively analysing sun and shade leaves of Billbergia elegans Mart. ex Schult. & Schult. f. and N. mucugensis Leme, also observed that individuals that grew in sunny environments exhibited leaves that were reddish and smaller than those of the individuals in shaded areas, which were green and larger. It must be noted, however, that other environmental factors may be influencing the pinkish colouration of the leaves, since low temperatures and nutritional and water stresses can increase anthocyanin production (Close and Beadle 2003). All N. bahiana populations with pinkish leaves occur in altitudinal rocky fields with low average temperatures; hence, climate is another environmental factor to be considered. The diversity in leaf morphology illustrates the phenotypic plasticity of the N. bahiana populations and points to this species as one of the most variable in leaf texture, colour and size in the genus. Contributions to the knowledge on Bromeliaceae – Many of the characters observed in Neoregelia bahiana are common in the family, including the root with multi-layered velamen, heterogeneous cortex, one-layered endodermis and pericycle, polyarch vascular cylinder, and pith with thick-walled cells (Tomlinson 1969, Pita and Menezes 2002, Segecin and Scatena 2004a, Proença and Sajo 2008a, Silva and Scatena 2011a). 37 An endodermis with cells presenting only thickened anticlinal walls in the roots characterises Bromelioideae and Pitcairnioideae, as it is also observed in the species of Acanthostachys Klotzshc, Aechmea Ruiz & Pav., Ananas Mill., Billbergia Thunb. (Bromelioideae), Dyckia Schult. & Schult. f. and Encholirium Mart. ex Schult. & Schult. f. (Pitcairnioideae) (Pita and Menezes 2002, Proença and Sajo 2008a, Silva and Scatena 2011a). On the other hand, endodermal cells with thin walls or with O-shaped or U-shaped thickened walls are common among the Tillandsioideae (Segecin and Scatena 2004a, Proença and Sajo 2008a). The radial arrangement of the cells of the inner cortex in N. bahiana may be due to meristematic activity of the endodermis, as demonstrated by Menezes et al. (2005) for other monocotyledons that grow in thickness. Regarding the stem, Tomlinson (1969) presents a general pattern, with distinct cortex and vascular cylinder, common for most Bromeliaceae. The short aerial stem has been studied anatomically by Segecin and Scatena (2004a) and Proença and Sajo (2008a), who termed it rhizome. However, rhizomes are generally characterised as subterranean stems with storage tissue, originating from the plumule, and constituting a monopolar system (Appezzato da Glória 2015), differing from what is observed in Bromeliaceae. Therefore, we use here the term short aerial stem to refer to the stem axis that bears the leaf rosette, and stolon to refer to that which develops from an axillary bud of the short aerial stem and grows horizontally, producing a new ramet. The stolon has defined nodes and internodes and cataphylls that protect the organ. It has no storage parenchyma and develops during vegetative propagation, possibly functioning to place the new ramet away from the mother-plant. The anatomy of the stolon in Bromeliaceae is described here for the first time: it is very similar to the short aerial stem, with a well-defined cortex and bundles distributed randomly in a parenchymatous pith, and may increase in diameter, by the formation of new vascular bundles and a multi-layered protective tissue with suberized cells. The presence of such suberized tissue on the stem has been reported for the species of Acanthostachys, Aechmea, Ananas, Billbergia, Bromelia L. (Bromelioideae), Dyckia (Pitcairnioideae) and Tillandsia L. (Tillandsioideae) (Segecin and Scatena 2004a, Proença and Sajo 2008a), indicating that the short aerial stem probably grows in thickness as well. This growth is probably promoted by the pericycle and/or endodermis (primary thickening meristem), as occurs in other monocotyledons (Diggle and DeMason 1983, Menezes et al. 2005), including Poales such as Cyperaceae (Rodrigues and Estelita 2009), Eriocaulaceae (Scatena and Menezes 1995, Oriani et al. 2008, Oliveira and Oriani 2016) and Rapateaceae (Menezes et al. 2005). Ontogenetic studies are necessary to elucidate 38 the stem structure in Bromeliaceae and its relationship to the different life forms found in the family. Regarding the leaves, characters common to the family include the presence of tetracytic stomata restricted to the abaxial surface, as well as epidermal cells with silica bodies and a small lumen (Tomlinson 1969, Sajo et al. 1998, Aoyama and Sajo 2003, Proença and Sajo 2004, 2008a, Scatena and Segecin 2005, Sousa et al. 2005). The silica bodies are frequent in monocotyledons, especially in the commelinid clade, which includes the Poales (Prychid et al. 2003). For this order, the presence of silica bodies is considered a plesiomorphic condition (Prychid et al. 2003) and also occurs in the leaf and inflorescence axis epidermis of Rapateaceae (Ferrari et al. 2014, Daltin et al. 2015), a family closely related to Bromeliaceae (Linder and Rudall 2005, Givnish et al. 2010). The structure of the scales observed in N. bahiana follows the pattern reported for Bromelioideae, with respect to the number of cells in the stalk and the arrangement of cells in the shield (Tomlinson 1969, Smith and Downs 1974, Smith and Till 1998, Benzing 2000). Furthermore, the presence of elongated peripheral cells in the shield corroborates the pattern described for Neoregelia (Smith and Till 1998). Another characteristic of Neoregelia is the spinescent leaf margin, though it is not exclusive to this genus, but a characteristic of Bromelioideae and Pitcarnioideae (Tomlinson 1969, Leme 1998). In the literature, these projections on the leaf margin are described as spines, including for Neoregelia bahiana (Tomlinson 1969, Leme 1998, Benzing 2000); however, we refer to them here as spinescent emergences, since they are not vascularized, but formed from both epidermal and hypodermal cells, conforming to the definition of emergences (Theobald et al. 1979, Evert 2006). Also, the use of aculeate margins as descriptive terminology is confuse, since the emergences observed are not a simple epidermal projection (i.e. not an aculeus or prickle). Regarding the inflorescence axis, it has been studied anatomically in the species of Acanthostachys, Aechmea, Ananas, Billbergia, Dyckia, Tillandsia and Vriesea Lindl. (Segecin and Scatena 2004b, Proença and Sajo 2008b, Silva and Scatena 2011b) and has been termed as floral scape. However, due to the presence of nodes and internodes, as well as the presence of bracts in the nodes, we use here the term inflorescence axis. The anatomical characterisation of the inflorescence axis in the Neoregelia bahiana complex is similar to that described for other genera (Segecin and Scatena 2004b, Proença and Sajo 2008b, Silva and Scatena 2011b). The anatomy of the pedicel is described here for the first time for Bromeliaceae and shows to be very similar to the anatomy of the inflorescence axis, 39 differentiated only by the absence of vascular traces in the cortex and by the presence of wider intercellular spaces in the pith. The presence of raphide-containing idioblasts, in both the vegetative and the reproductive organs, is another common feature of Bromeliaceae (Tomlinson 1969), being the raphides a typical calcium oxalate crystal among monocotyledons (Prychid and Rudall 1999, Prychid et al. 2008). The presence of mucilage is related to the formation of this type of crystal during the differentiation of the idioblasts (Kausch and Horner 1983, Webb et al. 1995, Prychid and Rudall 1999, Franceschi and Nakata 2005). In the Neoregelia bahiana complex, raphides occur in all the studied organs, in addition to small druses in the mesophyll of the leaf lamina. The type and shape of different calcium oxalate crystals are genetically regulated during cellular expansion and differentiation processes (Prychid and Rudall 1999, Franceschi and Nakata 2005) and therefore have taxonomic value. Morphological and anatomical characters understood as adaptations to the rupicolous life form and to the environment – Considering that Neoregelia bahiana has a wide geographic distribution, growing on rocky outcrops, characters such as the water storage tank, roots with velamen, stolons with a suberized protective tissue, and leaves with water-storage tissue favour the rupicolous life form. Although individuals sometimes grow on trees, as observed for Populations 5 (Grão Mogol) and 7 (Serra do Cabral), the representatives of the complex cannot be classified as epiphytes, because these populations occur in areas where outcrops and large trees intersperse, and during vegetative propagation individuals that grow on the rocky outcrops emit stolons toward the tree trunks, where new ramets develop. Studies on seed dispersion, germination, and post-seminal development in different species of Neoregelia are necessary in order to understand whether there are distinctions and adaptations related to the establishment of rupicolous and/or epiphytic species. The water storage tank formed by the leaves is a common feature of Bromelioideae (Benzing 2000, Schulte et al. 2009). The formation of this tank has great adaptive value for this subfamily, as it facilitates the accumulation and absorption of water and nutrients (Benzing 1970, Martin 1994). This is especially important for rupicolous species such Neoregelia bahiana, which grow in exposed soil with little available organic material and are subject to the climatic oscillations observed in the rocky fields. An epivelamen with root hairs has been reported for other species of Bromeliaceae (Pita and Menezes 2002, Segecin and Scatena 2004a, Proença and Sajo 2008a, Silva and Scatena 2011a). It contributes to the fixation of the plants on the rocks and increases the 40 capacity to absorb water and nutrients (Peterson and Farquhar 1996). However, an epivelamen with cells undifferentiated into root hairs also occurs in Tillandsia (Segecin and Scatena 2004a, Proença and Sajo 2008a, Silva and Scatena 2011a) and is probably associated with the epiphytic life form of most of its species. In N. bahiana, the velamen as well as the outer cortex with lignified cells may favour the rupicolous life form by providing mechanical support and minimising water loss, as demonstrated for epiphytic orchids (Dycus and Knudson 1957, Zotz and Winkler 2013). Variation in leaf size among the different populations of Neoregelia bahiana may be related to the sunlight incidence, as previously discussed. For other Bromelioideae species, such as Billbergia elegans and Neoregelia mucugensis from rocky fields (Pereira et al. 2013) and Neoregelia cruenta (R. Graham) L. B. Sm. from the Atlantic Forest (Reinert et al. 2013), a statistical difference in leaf size was registered, with longer and narrower leaves in low-light conditions, as was seen in shade leaves of the N. bahiana complex. Furthermore, Pereira et al. (2013) observed a larger area and thickness of the lamina in shade leaves, as well as a higher stomatal density and fewer scales density. Reinert et al. (2013) also found larger leaf area and fewer spinescent emergences on the margins of shade leaves. Differences in the plant architecture have also been observed: shade plants have a larger diameter and fewer leaves in the rosette (Reinert et al. 2013). All of these results show the ability of Bromelioideae species to respond to the environment with morphological adjustments, maximising their light- harvesting capacity and growth in low-light conditions. The scales, typical of the family (Tomlinson 1969), protect the leaf against direct sunlight and have been linked to the absorption of water and nutrients (Benzing 1970, 2000, Benzing et al. 1976, Brighigna et al. 1984). The grooves that form on the abaxial surface of the lamina are modulated by the presence or absence of scales, and they function as drainage channels for water that is then stored in the tank. In Populations 6, 7 (Serra do Cabral), 10 (Diamantina) and 12 (Serra do Cipó), the grooves in the stomatal bands reduce the stomata’s exposure to the environment, and it is probable that some of the water that flows through these grooves is assimilated along the lamina due to the absorptive properties of the scales. The epidermis and the mechanical hypodermis both with lignified thick-walled cells protect the leaf against excessive water loss and maintain the integrity of the lamina if the water-storage tissue wilt (Scatena and Segecin 2005). The well developed water-storage tissue on the adaxial surface of the lamina contribute to the survival of the plant under water stress, and simultaneously protect the chlorenchyma against significant deterioration of the photosynthetic system due to high light intensity (Smith and Downs 1974, Brighigna et al. 41 1984), which is common in environments such as the rocky fields. The variations in shape and size of the water-storage cells and the sinuosity of the cell walls indicate metabolic activity in the process of water storage and loss, as demonstrated for Pitcairnia burchellii Mez, a desiccation tolerant bromeliad species (Vieira et al. 2017), and is a common xeromorphic feature in many Cactaceae (Calvente et al. 2008). The characteristics of the inflorescence axis and the pedicel, such as the achlorophyllous aspect, sinuous contour, thin-walled cell, and pith with aerenchyma, were interpreted as adaptations to the inclusion of these structures in the water storage tank formed by the leaves. Nogueira et al. (2015), studying the anatomy of the ovary and ovule in the Nidularioid complex, also related the formation of aerenchyma in the ovary wall to hypoxic conditions because the inflorescence is nested in the leaf rosette. Regarding the crystals present in both the vegetative and the reproductive organs, different functions are attributed to them and related to environmental factors or to the developmental stage of the organ or tissues in which they are formed (Franceschi and Nakata 2005). Among other functions, the occurrence of these metabolites in specialised cells (idioblasts) may be related to a defence against herbivores, regulation of calcium in the cells (Franceschi and Nakata 2005), and an increase in resistance and mass of the tissues (Evert 2006). The druses present in the cells of the chlorenchyma (crystal cells) may optimise the distribution of light in this tissue, especially among the palisade cells beneath the well- developed water-storage tissue facing the adaxial surface. Assuming that cellular metabolism during the photosynthesis and transpiration processes, as well as calcium availability in the environment, stimulate crystal formation (Franceschi and Nakata 2005), the presence or absence, size and abundance of these crystals may be related to the age of the leaf and to chemical characteristics of the substrate. In conclusion, our results demonstrate the morphological and anatomical versatility of the Neoregelia bahiana complex, pointing to adaptive characters related to the rupicolous life form, different light conditions, water and nutrients availability, as well as characters with taxonomic value. From the evolutionary perspective, it is difficult to establish whether the populations of this complex can be considered as isolated entities, though the results presented here suggest the occurrence of speciation in some populations. Acknowledgments – The authors thank CAPES - Coordenação de Aperfeiçoamento de Pessoal de Nível Superior and CNPq - Conselho Nacional de Desenvolvimento Científico e Tecnológico for the financial support (#455510/2014-8). We also thank NAP/MEPA - 42 ESALQ/USP for the use of the scanning electron microscope, Marcos Vinicius Dantas de Queiroz and Kaire de Oliveira Nardi for assistance in the fieldwork, and the two anonymous referees for their valuable comments and suggestions on previous version of the manuscript. Conflict of interest – The authors declare that there is no conflict of interest. References Aoyama, E. M. and Sajo, M. G. 2003. Estrutura foliar de Aechmea Ruiz & Pav. subgênero Lamprococcus (Beer) Baker e espécies relacionadas (Bromeliaceae). – Braz. J. Bot. 26: 461-473. Appezzato da Glória, B. 2015. Morfologia de sistemas subterrâneos de plantas. – 3i Editora. Benzing, D. H. 1970. 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