I. Phylogenetic Systematics 

1. The evolution of apomixis in ferns

The evolution of apomixis in ferns

Apomixis can be defined as, simply, asexual reproduction  [“apo” = without, “mixis” = mixing/sex]. In plants, this definition encompasses vegetative asexual reproduction and asexual reproduction through the alternation of generations (including apospory, apogamy, and parthenogenesis). In the Grusz lab, we study the evolution (and evolutionary implications) of apogamous reproduction, in particular. In ferns, this involves two major modifications to the normal, sexual life cycle: the production of unreduced diplospores via meiosis (diplospory), and the apogamous development of a new sporophyte from somatic gametophyte tissue, without the fusion of sperm and egg (apogamy). Sporogenesis takes place within the fern sporangium, where spores develop via meiosis. In sexual lineages, these meiotic products (spores) are normally haploid, with half the parental number of chromosomes. Instead, apomictic ferns use one of two alternative spore-generating pathways to yield chromosomally unreduced diplospores: Döpp-Manton (premeiotic endomitosis) or Braithwaite (meiotic first division restitution) sporogenesis.  

> Grusz. 2016. Journal of Systematics and Evolution.

 apogamous fern life cycle (left). Döpp-Manton and braithwaite Sporogenesis in ferns (right). 

apogamous fern life cycle (left). Döpp-Manton and braithwaite Sporogenesis in ferns (right). 

Reticulate evolution and origins of apomixis

 Evolutionary relationships in the  Cheilanhtes  (=  Myriopteris )  yavapensis  species complex.

Evolutionary relationships in the Cheilanhtes (= Myriopteris) yavapensis species complex.

We use a variety of experimental approaches—spanning cytogenetics (chromosomes!) to phylogenetics (character evolution!)—to explore the evolutionary origins (and consequences) of asexual (apomictic) reproduction in polyploid ferns. Our research focuses primarily on the (mostly) New World fern genus, Myriopteris Fée (previously Cheilanthes Sw.). We use spore studies, flow cytometry, and chromosome counts to distinguish the number of genome copies per cell (ploidy level: x = haploid, 2x = diploid, etc.) for species in the myriopterid clade to determine, How many full genome copies interact in each nucleus? Meanwhile, morphological data illuminate (and often confirm) the origin(s) of apomictic polyploid hybrids. And enzyme electrophoresis—used to observe polymorphism in the size/shape of particular proteins—along with plastid and nuclear DNA/RNA sequencing of hybrid taxa, together, can reveal, Which divergent genomes comprise each nucleus, in what dosage?  And What (if any) are the short- and long-term evolutionary effects? 

Grusz et al. 2009. American Journal of Botany.     

Phylogenetic systematics of apomictic groups    

 chromosome number, ploidy level, and reproductive mode across  Myriopteris  (Pteridaceae). C =  covillei clade , L = lanosa clade, A =   alabamensis  clade.

chromosome number, ploidy level, and reproductive mode across Myriopteris (Pteridaceae). C = covillei clade, L = lanosa clade, A =  alabamensis clade.

Complex diversification in lineages rampant with morphological convergence, paired with polyploidy, hybridization, and apomixis, makes determining phylogenetic relationships and stabilizing the taxonomy especially difficult for some groups. Cheilanthoid ferns (Pteridaceae) are no exception. The poster-child genus of the clade, Cheilanthes Sw., is a perfect example of taxonomic instability that can result from confounding factors like these (Windham et al. 2009). Phylogenetic analyses of cheilanthoid ferns demonstrated Cheilanthes to be grossly polyphyletic, with species assigned to the name dispersed over five of the six major cheilanthoid clades (Windham et al. 2009). But recent molecular studies confirmed the existence of a strongly supported, monophyletic clade comprising ca. 45 primarily North and Central American taxa historically included in Cheilanthes. Using information from morphology, cytology, life history, and molecular (DNA) sequencing, we evaluated relationships among these taxa and explored character evolution in the group. Our recently updated circumscription of these taxa (in the resurrected genus, Myriopteris Fée; Grusz and Windham 2013) guides toward a more stable taxonomy for the genus—one that better reflects evolutionary history. Our phylogenetic analyses indicate strong statistical support for intrageneric species relationships, and shed light on morphological, cytological, and reproductive character evolution (Grusz et al. 2014).

Grusz et al. 2014. Systematic Botany.

Grusz and Windham. 2013. Phytokeys

Windham et al. 2009. American Fern Journal


2. Molecular marker development

Bulk identification of single copy nuclear gene regions (RNA-Seq)

  PIPELINE FOR THE identifICATION OF single copy orthologous regions.

 PIPELINE FOR THE identifICATION OF single copy orthologous regions.

The sequencing of transcribed RNA can aid greatly in the simplification of complex and/or repetitive genomes. Leveraging bioinformatic tools, these data facilitate a variety of molecular studies, including whole genome assembly and low-copy molecular marker development. Using data gathered for the 1KP Project (onekp.com), this project utilized high-throughput bioinformatic pipelines to identify and extract >2000 orthologous gene regions from Illumina PE RNA-Seq data from species spanning the fern family Pteridaceae. 


> Grusz et al. 2016. BMC Genomics.

Repetitive DNA:  simple sequence repeats (SSRs)  

 polymorphic microsatellite (SSR) marker: (GCC)^3 and (GCC)^6.

polymorphic microsatellite (SSR) marker: (GCC)^3 and (GCC)^6.

Ferns are notorious for their large, repetitive genomes. We exploit this tendency to develop genetic markers that are polymorphic for DNA repeat length. With these molecular tools, we examine genetic and genotypic variation in nature to shed light on micro- and macroevolutionary processes.

> Grusz and Pryer. 2014. Applications in Plant Sciences.

Low-copy nuclear DNA markers

 MAP OF NUCLEAR gapCp in ferns.

MAP OF NUCLEAR gapCp in ferns.

In organisms with large genomes and frequent polyploidy (genome duplication), sequence data for low copy genes can sometimes be in short supply. We use traditional and next generation sequencing approaches to develop DNA markers that are used to inform on genotypes / phenotypes, and provide insight into the evolutionary origins of polyploids. 

> Schuettpelz et al. 2008. Systematic Botany.


II. Evolutionary Genomics

Extreme evolutionary rate variation in ferns

 Rapid molecular evolutionary rates in the morphologically extreme vittarioid ferns. 

Rapid molecular evolutionary rates in the morphologically extreme vittarioid ferns. 

In the fern tree of life, some lineages evolve at mostly consistent rates (relative to one another), but others have vastly different evolutionary rates compared to closely related groups (most notably, the filmy ferns, tree ferns, and vittarioid ferns). Using transcriptome / gene expression (RNA) data, we study evolutionary rate discrepancies among the genomes of taxa spanning the Pteridaceae — a cosmopolitan fern family with ca. 50 genera and > 1000 species (ca. 10% of documented fern diversity).  In collaboration with E. Schuettpelz (Smithsonian Institution, NMNH) and C. Rothfels (UC Berkeley), with support from the 1KP Project (onekp.com), our goal is to uncover genomic clues driving disparate morphologies, ecologies, life histories, and rates of molecular evolution across the family. This is part of an ongoing project with K. M. Pryer (Duke University) aimed toward identifying drivers of rapid molecular and morphological divergence between Adiantum and the vittarioid ferns, and sheds light on the mechanisms influencing evolutionary rates across the family. 

Grusz et al. 2016. BMC Genomics.