Trebouxia lynnae sp. nov. (Former Trebouxia sp. TR9): Biology and Biogeography of an Epitome Lichen Symbiotic Microalga

[EN] Simple Summary In this work, we present the formal description of a new species of lichen photobiont (i.e., Trebouxia lynnae) isolated from the lichen Ramalina farinacea. The findings reported here provide an exhaustive characterization of the cellular ultrastructure, physiological traits and genetic and genomic diversity of the new species. Our results contribute to the knowledge of lichen-forming symbiotic green microalgae with their diversity and distribution. Two microalgal species, Trebouxia jamesii and Trebouxia sp. TR9, were detected as the main photobionts coexisting in the thalli of the lichen Ramalina farinacea. Trebouxia sp. TR9 emerged as a new taxon in lichen symbioses and was successfully isolated and propagated in in vitro culture and thoroughly investigated. Several years of research have confirmed the taxon Trebouxia sp. TR9 to be a model/reference organism for studying mycobiont-photobiont association patterns in lichen symbioses. Trebouxia sp. TR9 is the first symbiotic, lichen-forming microalga for which an exhaustive characterization of cellular ultrastructure, physiological traits, genetic and genomic diversity is available. The cellular ultrastructure was studied by light, electron and confocal microscopy; physiological traits were studied as responses to different abiotic stresses. The genetic diversity was previously analyzed at both the nuclear and organelle levels by using chloroplast, mitochondrial, and nuclear genome data, and a multiplicity of phylogenetic analyses were carried out to study its intraspecific diversity at a biogeographical level and its specificity association patterns with the mycobiont. Here, Trebouxia sp. TR9 is formally described by applying an integrative taxonomic approach and is presented to science as Trebouxia lynnae, in honor of Lynn Margulis, who was the primary modern proponent for the significance of symbiosis in evolution. The complete set of analyses that were carried out for its characterization is provided.PROMETEO 2021/005 (Excellence in research, Generalitat Valenciana) and the Grants New Generation EU (Ministry of Universities) to Salvador Chiva (MS21-058) and Cesar Bordenave (ZA21-046)Barreno, E.; Muggia, L.; Chiva, S.; Molins, A.; Bordenave, C.; García-Breijo, F.; Moya, P. (2022). Trebouxia lynnae sp. nov. (Former Trebouxia sp. TR9): Biology and Biogeography of an Epitome Lichen Symbiotic Microalga. Biology. 11(8):1-19. https://doi.org/10.3390/biology1108119611911

Multidisciplinary approach to describe Trebouxia diversity within lichenized fungi Buellia zo-haryi from the Canary Islands.

[EN] The Canary Islands are famous for their extraordinary biodiversity; however, lichenized algae have only been studied partially. Buellia zoharyi is a circum-Mediterranean/Macaronesian species that usually occurs in semi-arid areas of the Mediterranean, but occasionally some interesting communities of this species grow on basaltic lava flows in Lanzarote, Fuerteventura and Tenerife. Those three locations showed similar ecological conditions, but different mean annual temperatures. Here we applied a multidisciplinary approach to describe microalgae diversity from B. zoharyi covering the entire described range of distribution in the Canary Islands. Photobionts were characterized in symbiosis using molecular and microscopic techniques. Different Trebouxia spp. were detected as primary photobiont in each island (Trebouxia cretacea-Fuerteventura, T. asymmetrica-Lanzarote and Trebouxia sp. `arnoldoi '-Tenerife). Coexistence of various Trebouxia spp. within a thallus were detected by using specific primers-PCR. Those three photobionts were isolated and cultured under laboratory conditions. Different phytohormone profiles were obtained in the isolated strains which suggest different internal signalling needs. In addition, we characterized the response of the isolated strains to different temperatures using chlorophyll fluorescence. T. asymmetrica did not modify their F-v/fm values with respect to temperature acclimation. In contrast, Trebouxia sp. `arnoldoi'and T. cretacea were more sensitive to changes in growing temperature decreasing Fv/fm at 17 degrees C. Our results indicate that B. zoharyi is flexible regarding the photobiont choice depending on the region, and suggest that bioclimatic factors could influence the myco/photobiont association patterns.Funding for field and laboratory work for this study was provided by the Ministerio de Economia y Competitividad (MINECO and FEDER, Spain) (CGL2016-79158-P) and Prometeo Excellence in Research Program (Generalitat Valenciana, Spain) (PROMETEOII/2013/021; PROMETEO/2017/039). Daniel Sheerin revised the English manuscriptMolins, A.; Chiva, S.; Calatayud, A.; Marco, F.; García-Breijo, F.; Reig-Arminana, J.; Carrasco, P.... (2020). Multidisciplinary approach to describe Trebouxia diversity within lichenized fungi Buellia zo-haryi from the Canary Islands. 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Symbiotic microalgal diversity within lichenicolous lichens and crustose hosts on Iberian Peninsula gypsum biocrusts

Abstract This study analyses the interactions among crustose and lichenicolous lichens growing on gypsum biocrusts. The selected community was composed of Acarospora nodulosa, Acarospora placodiiformis, Diploschistes diacapsis, Rhizocarpon malenconianum and Diplotomma rivas-martinezii. These species represent an optimal system for investigating the strategies used to share phycobionts because Acarospora spp. are parasites of D. diacapsis during their first growth stages, while in mature stages, they can develop independently. R. malenconianum is an obligate lichenicolous lichen on D. diacapsis, and D. rivas-martinezii occurs physically close to D. diacapsis. Microalgal diversity was studied by Sanger sequencing and 454-pyrosequencing of the nrITS region, and the microalgae were characterized ultrastructurally. Mycobionts were studied by performing phylogenetic analyses. Mineralogical and macro- and micro-element patterns were analysed to evaluate their influence on the microalgal pool available in the substrate. The intrathalline coexistence of various microalgal lineages was confirmed in all mycobionts. D. diacapsis was confirmed as an algal donor, and the associated lichenicolous lichens acquired their phycobionts in two ways: maintenance of the hosts’ microalgae and algal switching. Fe and Sr were the most abundant microelements in the substrates but no significant relationship was found with the microalgal diversity. The range of associated phycobionts are influenced by thallus morphology

How did terricolous fungi originate in the Mediterranean region? A case study with a gypsicolous lichenized species

Aim: The historical causes responsible for the wide distribution of terricolous, crustose lichenized fungi across the Mediterranean Basin and the Canary Islands have never been explored. Here, we used the terricolous, circum-Mediterranean/Macaronesian species Buellia zoharyi (Caliciaceae, Ascomycota) to infer the time frame, and the climatic, geological and ecological factors influencing the origin and current spatial distribution of this species. Location: Mediterranean Basin and Canary Islands. Methods: Data from two nuclear markers (nrITS and tef1) obtained from 226 specimens of 23 populations covering the entire distribution range of B. zoharyi were used to calculate genetic diversity indices and haplotype networks and to investigate population size changes and structure. Three secondary calibrations were used to estimate the timing of the divergence of B. zoharyi from its hypothesized sister species, B. elegans, and the diversification of B. zoharyi. Results: We found low nucleotide diversity and two geographically differentiated haplogroups, with a contact zone in the Iberian Peninsula. The three dating approaches established wide temporal windows for the divergence of B. zoharyi from B. elegans (Eocene-Pliocene) and its diversification (Miocene-Pleistocene). These intervals overlap with the origin and diversification ages found in other lichen-forming fungi and vascular plants inhabiting the Mediterranean region. Main conclusions: In the context of lichen biogeography, our results support ecological specialization as well as geological and climatic events as drivers of the evolutionary history of B. zoharyi in the Mediterranean. In particular, the combined effects of the Messinian salinity crisis and the subsequent Zanclean Flood on the availability of gypsum soils in the Mediterranean Basin, as well as the Quaternary climatic oscillations, seem to have collectively shaped the amount and distribution of B. zoharyi population genetic diversity

Trebouxia maresiae Garrido-Benavent, Chiva & Barreno 2022, sp. nov.

<i>Trebouxia maresiae</i> Garrido-Benavent, Chiva & Barreno, sp. nov. (Figs 1-3) <p> <i>Trebouxia maresiae</i> Garrido-Benavent, Chiva & Barreno, sp. nov. shows crenulate chloroplasts that bear a crenulata-type pyrenoid when cells are in culture, whereas in the lichenized state it acquires a hybrid structure (maresiae-type), characterized by the periphery of the pyrenoid being rather gigantea-type, with thylakoid membranes forming short, branched tubules. These traits distinguish the new species from <i>T. decolorans</i>, which has deeply lobed chloroplasts with a decolorans-type pyrenoid. The two species also differ by the relatively high number of nucleotide differences in their nrITS DNA sequences. Vegetative cells of <i>T. maresiae</i> sp. nov. tend to be smaller in diameter (<15 µm) than those of other species in <i>Trebouxia</i> clade A like <i>T. decolorans</i> and <i>T. solaris</i>.</p> <p> HOLOTYPE. — <b>Spain</b>. Balearic Islands, Mallorca, Es Trenc, phycobiont of <i>Seirophora villosa</i> (Ach.) Frödén collected on branches of <i>Juniperus phoenicea</i> subsp. <i>turbinata</i> (Guss.) Nyman in coastal dunes, 39°20’40.71”N, 2°59’8.24”E, 3 m a.s.l., 07.VIII.2008, leg. G. Salvà. The lichen specimen was deposited in the herbarium of the University of Valencia (VAL-Lich) with the following code: VAL-Lich 31798.</p> <p>TYPE STRAIN. — Deposited at the Symbiotic Algal collection from the University of Valencia (ASUV), as item type ASUV 142.</p> <p>REFERENCE STRAINS. — ASUV 142 and ACOI 3420, deposited at the Coimbra Collection of Algae (ACOI), University of Coimbra.</p> <p> GENBANK ACCESSION NUMBERS. — MZ724411 (nrITS), MZ687825 (<i>rbc</i> L) and MZ687826 (COX2).</p> <p>ETYMOLOGY. — According to the Canarian Academy of Language, the term “maresía”, from which the specific epithet “maresiae” derives, means “air laden with marine humidity in areas close to the seashore”. Therefore, this term refers to the particular habitat where the lichen phycobiont lives, in coastal sea-shore environments.</p> <p> ECOLOGY. — In association with lichen-forming fungi of the genera <i>Seirophora</i> Poelt, <i>Ramalina</i> Ach. (this work), and <i>Xanthoria parietina</i> (L.) Th. Fr. (isolate no. P-280-II-a-Sc; Nyati <i>et al</i>. 2014).</p> <p> DISTRIBUTION. — Currently, the known geographic distribution of the new alga includes the Mediterranean basin and the Cape Verdean Island of Santiago (this study; Nyati <i>et al</i>. 2014; Garrido- Benavent <i>et al</i>. 2022).</p> DESCRIPTION <p>Cells are mostly unicellular,rarely in tetrads and octads.Vegetative mature cells usually spherical, rarely oval, 7-11 (15) µm diam. Cell wall is usually thin and arranged in a bilayer. Chloroplast central, dissected into crenulated lobes with several tree-like lobes elongated at their ends. Some cells containing two chloroplasts may be also observed. The chloroplast usually contains a single pyrenoid of the crenulata-type with wide, branched tubules of curved profile, often crossing the entire pyrenoid. In symbiosis, however, the pyrenoid shows a hybrid structure (maresiae-type), characterized by displaying two discrete regions that can be each clearly assigned to different pyrenoid types; here, the pyrenoid periphery is more similar to the gigantea-type. Secondary pyrenoids may be present.Pyrenoglobuli present in the closest parts of the chloroplast stroma.The cytoplasm of old cells is sometimes yellowish, probably due to the presence carotenoid pigments, and vacuolated. Asexual reproduction involves the formation of autospores. Autosporangia (10-11 µm diam.) usually contain 4-16 (32) autospores. Sexual reproduction was not observed.</p>Published as part of <i>Garrido-Benavent, Isaac, Chiva, Salvador, Bordenave, César D., Molins, Arantzazu & Barreno, Eva, 2022, Trebouxia maresiae sp. nov. (Trebouxiophyceae, Chlorophyta), a new lichenized species of microalga found in coastal environments, pp. 135-145 in Cryptogamie, Algologie 20 (9)</i> on pages 139-143, DOI: 10.5252/cryptogamie-algologie2022v43a9, <a href="http://zenodo.org/record/7829260">http://zenodo.org/record/7829260</a&gt