Lecture 2: Common red algal families in the vicinity of Bocas del Toro: Gracilariaceae Photo: O.Camacho Hadad & Anna Fricke

Halymeniaceae Rhodymeniaceae Gracilariaceae

vegetative structure: cellular medulla family, ordinal level Gracilaria Hydropuntia flat sp. terete sp. mixed-sized cells

Gracilariaceae Solieriaceae

Solieriaceae (Solieria) loose filiform, or mixed cellular-filiform medulla

Halymeniaceae (Cryptonemia)

-cortical unit of 3 surface cells

Conjunctor cells fuse with neighboring cells, resulting in secondary pit connection formation

-inherently uniaxial

-uniaxial growth quickly disguised -axial cells: same shape as medullary cells

Generic distinction: -comparative post-fertilization events of female reproductive system in establishment of cystocarp: nature of fusions - complexity of male reproductive system

Gracilaria intermedia cystocarps

pericarp carposporangia medulla cystocarp floor inner gonimo- blast

Gracilaria: multinucleate tubular nutritive cells to roof, to base Gracilariopsis: uninucleate conjunctor cells at base Gracilariophila (parasite) Hydropuntia: multinucleate tubular nutritive cells at base of cystocarp “TAPPING BACK”

-origin: 3-cortical unit again -lower cell of unit = cell 1 = supporting cell cutting off carpogonial branch initial -cells 2 & 3 = nutritive cells of carpogonial branch unit unfertilized female structure

unfertilized 2-celled carpogonial branches

-2-celled carpogonial branch on supporting cell -straight trichogyne reaching thallus surface -partly fusion of cortical nutritive cells to carpogonium

Partly fusion of neighboring cortical cells with carpogonium; vegetative nuclei remain in unfused portion

-diploid zygote nucleus in uninucleate carpogonium; -fused cortical cells = nutritive cells

-young carpospore-bearing filaments (gonimoblasts) cut off from carpogonial fusion cell

How lower cells of young carpospore-bearing filaments fuse back to multinucleate carpogonial fusion cell Generic character:

young cystocarp: Gracilariopsis -very regular, progressive growth of carpospore- bearing filaments

Fusion of lower carposporophyte cells to vegetative gametophytic cells Gracilariopsis

Gracilaria: multinucleate tubular nutritive cells to roof, to base Gracilariopsis: uninucleate conjunctor cells at base Gracilariophila (parasite) Hydropuntia: multinucleate tubular nutritive cells at base of cystocarp “TAPPING BACK”

Gracilaria:Gracilariopsis: Gracilariophila: Hydropuntia: male structures in shallow or deep pits super- ficial in deep pits, con- necting to vege- tative cells

Gurgel & Fredericq 2004 = rbcL phylogeny of the Gracilariaceae, Bayesian Tree.

Global Phylogeny of the Gracilariaceae Bayesian Inferred Phylogeny Gurgel & Fredericq 2004

Bayesian Tree A meaningful global phylogeny of the Gracilariaceae and the evolutionary patterns found in it could only be obtained after reaching a dataset composed of 35+ taxa Before that, taxa sampling played a major issue in obtaining stable and meaningful phylogenies. So, nowadays, when new taxa (i.e. distinct rbcL DNA sequences) are added to the global phylogeny they do not change the overall topology. Ready for final taxonomic conclusions.

What has been established so far? G. tenuistipitata G. chilensis G. vermiculophylla Gp. hommersandii (outgroup) H. urvillei H. aff. changii H. changii H. crassissima H. cornea H. caudata H. secundata H. perplexa H. preissiana H. edulis H. rangiferina H. millardetii H. eucheumatoides G. beckeri 83 G. acuelata G. textorii G. ornata G. bursa-pastoris G. cervicornis G. tikvahiae G. mammillaris G. canaliculata G. salicornia G. gracilis G. pacifica Hydropuntia Gracilaria ‘New Genus’ Genera: Chilensis-type spermatangia ‘Verrucosa’-type spermatangia Rounded carposporophyte Lobed carposporophyte Textorii-type spermatangia Verrucosa-type spermatangia Sub-Genera:

Is high quality-agar (gel strength) correlated to phylogeny? Is there character congruence between agar gel strength (phenotype) and molecular-based phylogenies (haplotype)? Can sound phylogenies identify lineages characterized by high-quality agar? Gurgel & Fredericq 2004

G. vermiculophylla G. tenuistipitata G. chilensis (90% agar worldwide, Zemke-White & Ohno 1999) Proposed as a distinct genus by Gurgel & Fredericq 2004

Agar Properties:Yield( %, dw/dw ) Gel Strength( g. cm -2 ) 3,6 Anhydro-galactose( %, dw/dw ) Gelling Point( °C ) Boiling Point( °C ) 1 st Source of Variation: (in the Agar Properties) Strain / population ‘Tissue’ / Thallus Age Light Regime Temperature Life-cycle Reproductive status ≠ Methodologies (Whyte et al. 1984):2 nd Source of Variation: * Extraction: Standard vs. Alkali * Strength: gel shape, Ø and crosshead speed (plunger) * 3,6 AG: Resorcinol vs. IR Spectrophotometer Data in the Literature: Fred Gurgel, pers. comm

Searching for Phylogenetic Patterns of Native Gel Strengths Sampling for Published Data: Obtain agar parameters for species for which we have phylogenetic data (= rbcL DNA sequences). Select methodologies that reflect the algae’s most natural conditions. –Extraction: Standard (shredding  boiling  gelling  freezing  thawing  washing) –Strength method: 1.5% gels, 1.0 cm Ø plungers Minimize Method Variation Total number of Papers Surveyed= 26 Total number of Papers Selected= 09 Fred Gurgel, pers. comm

G. chilensis Clade Gel Strength (g. cm -2 ) Number of species with comparable agar data Gracilaria s. s. Clade Gracilariopsis Clade Fred Gurgel, pers. comm

Conclusions RbcL-based phylogenies reevaluated character evolution in the Gracilariaceae and inferred a new classification for Gracilaria sensu lato. The G. chilensis-clade and the genus Hydropuntia are reinforced as distinct taxa in the Gracilaria complex. Based on a literature review, data suggest that there is a strong correlation between phylogeny and agar gel strength (agar quality) between genera, and between average genetic divergence and gel strength values within each genus.

ordinal & family-level characters: -based on how unfertilized female system develops in relation to vegetative growth cf. Kylin system

Gracilaria: multinucleate tubular nutritive cells to roof Gracilariopsis: uninucleate conjunctor cells Gracilariophila (parasite) Hydropuntia: multinucleate tubular nutritive cells at base of cystocarp

Problem: *differentiate among stages that are are functional, versus abortive?? *no auxiliary cells!

Gracilaria: male structure development

Gracilariopsis: male structure development

-tetrasporangial initials are surface cortical cells -cruciate divisision of tetrasporangia all genera:

Gracilariaceae from the vicinity of Bocas del Toro

Gracilaria curtissiae -thick thallus; thinner lateral blade-like proliferations Photo: A. Fricke & O. Camacho-Hadad

Gracilaria intermedia

Photo: A. Planas & C. Fernández)

Gracilaria cervicornis

Photo: M.C. Díaz Ruiz

Photo: R. Fikes

Gracilaria domingensis

Photo: A. Fricke & O. Camacho-Hadad

Hydropuntia caudata

Gracilaria galetensis

Gracilaria hayi

Global Phylogeny of the Gracilariaceae Bayesian Inferred Phylogeny GTR+inv+gama 100,00 generations, 4 chains Burn in = 20,000 Gurgel & Fredericq 2004