Taxonomy and palaeoecology of the fossil anamorphic fungus Mycoenterolobium eccentricum (R.K. Kar) G. Worobiec, n. comb.

Microremains of fungi from Neogene deposits from the Gray Fossil Site (Tennessee, United States) and the Bełchatów Lignite Mine (Poland), similar to the enigmatic fossil-species Kutchiathyrites eccentricus R.K. Kar, 1979, were reconsidered as representatives of the modern mitosporic genus Mycoenterolobium Goos, 1970. A new combination, Mycoenterolobium eccentricum (R.K. Kar) G. Worobiec, n. comb., is proposed. The geographical and stratigraphical


INTRODUCTION
As one of the kingdoms of eukaryotic organisms, fungi most likely appeared in the late Proterozoic (Loron et al. 2019;Berbee et al. 2020;Bonneville et al. 2020). Their taxonomic diversity is estimated to be between 2 and 4 million species (Hawksworth & Lücking 2017), but the list of fungal taxa, both modern and fossil, is expanding continually. Many fossil fungal taxa have the potential as a non-pollen palynomorph (NPP) palaeoecological proxy (Lange 1976;Conran et al. 2016;Worobiec & Worobiec 2017;Worobiec et al. 2018Worobiec et al. , 2020. The particular group of fossil fungi that play an essential role as palaeoenvironmental proxies is wetland fungi saprophytic on decaying plant remains (Widera et al. 2021;Worobiec et al. 2021Worobiec et al. , 2022. On wetlands (swamps, bogs, fens, marshes, bottomland hardwoods) can be found various fungal taxa, both terrestrial and aquatic (Stephenson et al. 2013). Saprophytic fungi play an important ecological role in wetlands as decomposers of organic matter, mainly dead plant material (Dix & Webster 1995;Gessner et al. 2007;Gulis et al. 2019). Microremains of fungi, including those of wetland origin, were found during palynological investigations of Neogene deposits from the Gray Fossil Site (United States) and from the Bełchatów Lignite Mine in Poland (plant remains assemblages KRAM-P 218 and KRAM-P 226) as chitinous non-pollen palynomorphs (Zobaa et al. 2011;Ochoa et al. 2012;Worobiec et al. 2013Worobiec et al. , 2018Worobiec et al. , 2020Worobiec & Worobiec 2017). Pollen slides from these localities were reinvestigated then for new fungal forms. Among them, conidia of dictyosporous anamorphic fungus apparently similar to the enigmatic fossil-species Kutchiathyrites eccentricus R.K. Kar, 1979 were found. Kutchiathyrites eccentricus previously has been considered to be similar to "Microthyriaceous ascostromata" (Kar 1979), Mycoenterolobium platysporum Goos, 1970, Arbuscula eugeniae Bat. &Peres, 1965, andTretopileus sphaerophorus (Berk. &A. Curtis) S. Hughes & Deighton, 1960(Jain & Kar 1979 or as similar to Mycoenterolobium platysporum and classified as Fungi Imperfecti, Dictyosporae (Kalgutkar & Jansonius 2000;Berbee et al. 2015). Thus, the relationship of the fossil Kutchiathyrites eccentricus with modern fungal genera has been disputable so far. However, establishing the affinities of fossil fungi with their modern counterparts is essential for palaeoenvironmental and palaeoclimatic reconstructions (Saxena & Wijayawardene 2022). The aim of our study is to describe and re-interpret the Kutchiathyrites-like remains from Neogene deposits of the United States, and Poland and to consider their importance as a proxy in term of palaecology.

MATERIAL AND METHODS
The investigated fossils were found in palynological samples from two localities. Most fossil specimens came from the Gray Fossil Site, Washington County, Tennessee, United States. Two specimens were found in two sites from the Bełchatów Lignite Mine, Poland.
Lacustrine deposits of the Gray Fossil Site in Washington County, Tennessee, United States (36°23'9.6"N, 82°29'52.8"W) were suggested to be a multiple sinkholes/sub-basins fills that could represent asynchronous events (Zobaa et al. 2011;Worobiec et al. 2013). Laminated sediments of the Gray Fossil Site, previously dated to the latest Miocene-the earliest Pliocene (Wallace & Wang 2004;Shunk et al. 2006) and an updated analysis indicating an Early Pliocene age of latest Hemphillian to early Blancan (4.5-4.9 Ma) (Samuels et al. 2018), preserved various animal and plant remains (see Worobiec et al. 2018). Fungal remains from the Gray Fossil Site were earlier reported four times. Zobaa et al. (2011) found remains of Callimothallus sp. (now Neomycoleptodiscus pertusus (Dilcher) G. Worobiec, 2020), Ochoa et al. (2012 mentioned the presence of fungal remains, Worobiec et al. (2013) noted the occurrence of epiphyllous fungi (Microthyriales) and, finally, Worobiec et al. (2018)  channel fill in the floor part of clayey-sandy unit (I-P) of the Neogene deposits of the Bełchatów Lignite Mine (Worobiec & Szynkiewicz 2016;Worobiec & Worobiec 2016, considered to be Late Miocene in age (Szynkiewicz 2000). Investigated fungal remains were found in one palynological slide from the specimen (rock sample) with plant macroremains numbered KRAM-P 218/85. The assemblage KRAM-P 226 with plant macroremains came from the borehole core No. 1326/B from the mine outcrop. The stratigraphical position of the fossil leaf litter sample found in the core corresponds to the clayey-coal unit (I-W). The complicated tectonics of this part of the mine outcrop makes it difficult to establish the exact geological position and age of the plant macroremain assemblage KRAM-P 226. However, the age of the discussed plant assemblage is probably the Late Miocene (Worobiec 2014). In one palynological slide from this locality, one specimen of the fossil Mycoenterolobium eccentricum (R.K. Kar) G. Worobiec, n. comb. was found. Fungal remains from the fossil assemblage KRAM-P 218 of the Bełchatów Lignite Mine were previously reported by Worobiec & Worobiec (2017) and Worobiec et al. (2020 Worobiec (2014) described "bodies of epiphyllous, microthyriaceous fungus" on the abaxial epidermis of the fossil leaf of ?Magnolia sp. Detailed descriptions of the geology of the localities KRAM-P 218 and KRAM-P 226 were provided by Worobiec & Szynkiewicz (2016) and Worobiec (2014), respectively.
Palynological samples from the Gray Fossil Site and the Bełchatów Lignite Mine were processed in the laboratory of the W. Szafer Institute of Botany, Polish Academy of Sciences (Kraków), using hydrochloric acid, potassium hydroxide, and sulfuric acid (Moore et al. 1991). Additionally, hydrofluoric acid was used to remove mineral matter, and the residuum was sieved at 5 µm on a nylon mesh. The microscope slides were made using glycerine jelly as a mounting medium (Worobiec et al. 2013).
The fossil specimens are housed in the W. Szafer Institute of Botany, Polish Academy of Sciences, under catalogue numbers KRAM-P 218 and KRAM-P 226 as well as GFS/Bear Pit and GFS/Elephant Pit. Microphotographs were taken with a Nikon Eclipse E400 microscope equipped with a Canon A640 digital camera. Terminology for the morphology of fungal remains follows Goos (1970) and Calabon et al. (2020).  (4), KRAM-P 226B. Ten specimens.

Institutional abbreviations
revised diAgnosis. -Dictyosporous conidia, strongly flattened in one plane, variable in shape, but mature specimens mostly fan-shaped, sometimes lobed. Conidia composed of two layers of smooth-walled muriform cells radiating from a single cell at the point of attachment of conidiophore. Cells of conidium usually isodiametric, sometimes rectangular, with cell walls straight or rounded.

DISCUSSION
reconsiderAtion of fossil Kutchiathyrites eccentricus Dictyosporous fungal conidia above described have rather unique morphology, and they are the same as fungal remains described first by Kar (1979) from the Oligocene deposits of Kutch, western India, as new fossil-genus Kutchiathyrites R.K. Kar, 1979, represented by the fossil-species Kutchiathyrites eccentricus R.K. Kar, 1979. These remains were considered by Kar (op. cit.) as "Microthyriaceous ascostromata" and compared with the microthyriaceous fossil fungal genera Notothyrites Cookson, 1947 (synonym of Trichothyrites Rosend., 1943), Paramicrothallites K.P. Jain & R.C. Gupta, 1979, Parmathyrites K.P. Jain & R.C. Gupta, 1970, andPhragmothyrites W.N. Edwards, 1922. However, these genera in the opinion of Kar (1979) Worobiec, n. comb., by analogy with its modern counterpart Mycoenterolobium platysporum, are rather similar to the conidia of modern representatives of the anamorphic genus Cancellidium Tubaki, 1975(Calabon et al. 2020. Conidia of Mycoenterolobium differ from those of Cancellidium, having rows of cells radiating in a fan-shaped pattern from a basal cell attached to the conidiophore (Fig. 1A-D). The Cancellidium cells are arranged in parallel, adherent rows, and conidia of Cancellidium contain chains of monilioid cells developing from the base (Seifert et al. 2011;Zhao et al. 2013). Mycoenterolobium eccentricum (R.K. Kar) G. Worobiec, n. comb. also shows similarities to conidia of some modern representatives of helicosporous anamorphic fungi from the genus Xenosporium Penz. & Sacc., 1902, and especially to Xenosporium mirabile Penz. & Sacc., 1902(Zhao et al. 2007. Mycoenterolobium differs, however, from Xenosporium mirabile in lacking welldefined conidiophores and secondary conidia (Zhao et al. 2007), and also in cell arrangement of conidia. Conidia of other modern dictyosporous genera differ considerably from both the fossil Mycoenterolobium eccentricum (R.K. Kar) G. Worobiec, n. comb. and modern Mycoenterolobium platysporum.  (2000), however, did not point to any morphological features of this fossil that could indicate that Dictyosporites eccentricus should be included in the fossil-genus Kutchiathyrites. The classification of Dictyosporites eccentricus as a member of the fossil-genus Kutchiathyrites is questionable in our opinion. Dictyosporites eccentricus differs having some rows of cells that are longer than other contrary to muriform rows of cells of conidium of Mycoenterolobium which are proportionally of more or less equal length. Dictyosporites eccentricus can also represent other than Kutchiathyrites fungal genera having dictyosporous conidia. Thus, the taxonomic position of Kutchiathyrites canadensis seems still enigmatic, and we could not reconsider it as a fossil representative of the modern Mycoenterolobium genus.
Kutchiathyrites mehrotrae, originally described as Kutchiathyrites sp. (Singh et al. 1986), was then considered a new fossil-species of this fossil-genus by Saxena & Tripathi (2011). Original illustrations of Kutchiathyrites sp. "ascomata" (Singh et al. 1986: pl. 1, figs 11, 12) suggest that the specimen on pl. 1, fig. 11 could be related to the fossil Mycoenterolobium eccentricum (R.K. Kar) G. Worobiec, n. comb. (obovate conidium with similar row of cells) and the specimen on pl. 1, fig. 12, that differs considerably from the specimen on fig. 11, is somewhat similar to above discussed Kutchiathyrites canadensis (some rows of muriform cells are longer than those at the sides). Considering this, in our opinion Kutchiathyrites mehrotrae seems doubtful as a new fossil-species probably comprising two different species, and only specimen on pl. 1, fig. 11 fig. 14), Pliocene of Slovakia (as remains of Musci, Planderová 1972: fig. 1: 8) (1998) found fungal remains interpreted as Microthyriacites sp. as well. The morphology of these fossils, however, is different from the fossil-genus Microthyriacites Cookson, 1947. One specimen (Song 1998: pl. 5, fig. 18) presumably could be assigned to Mycoenterolobium and the second specimen (Song 1998: pl. 5, fig. 18 (Goos 1970;Kilbertus et al. 1980;Chamuris et al. 1985;Nakagiri 1993;Heredia-Abarca & Mercado-Sierra 1998;Arias Mota et al. 2008;Wang et al. 2008;Tianyu 2009;Leão-Ferreira et al. 2009 Modern Mycoenterolobium found both in aquatic and terrestrial localities were classified as aero-aquatic hyphomycetes (Goh & Hyde 1996). These mitosporic fungi are usually found in small and shallow water reservoirs with stagnant slow-running water like ponds or ditches as saprophytes on submerged leaves or woody substrates. They sporulate only when the substrate is exposed to air in a moist interface between air and water. It could happen when previously submerged substrate, like leaves or twigs, become exposed to air at a reservoir margin when the water dries up, for example, during the hot summer months (Dix & Webster 1995;Goh & Hyde 1996;Webster & Weber 2007;Markovskaja 2012 and Alnus composed swamp communities (swamp forests and bush swamps) along with representatives of Ericaceae, Cyrillaceae, Clethraceae as well as Myrica Linnaeus, 1753, and presumably Ilex Linnaeus, 1753. The floristic composition of the palaeovegetation of the plant assemblage KRAM-P 218 points to a warm and moderately wet temperate climate with mild winters, comparable to the Cfa climate type in the Köppen-Geiger climate classification with a presumable mean annual temperature of 13.5-16.5°C (Worobiec & Szynkiewicz 2016;Worobiec & Worobiec 2016). These climatic inferences are consistent with the results of the analysis of fungal remains that accompanied plant remains in the locality KRAM-P 218 (Worobiec & Worobiec 2017). Plant macroremains of the assemblage KRAM-P 226 from the Bełchatów Lignite Mine suggest the dominance of mesophytic vegetation; only a few taxa are typical for azonal, riparian, and swamp forests. Remains of aquatic vegetation were completely absent there, suggesting that the freshwater sedimentary reservoir formed the plant assemblage KRAM-P 226 was shallow and/or only periodically existed (e.g. after flooding). Plant taxa of this assemblage, along with remains of epiphyllous, microthyriaceous fungi indicate warm palaeoclimate with mild winters (Worobiec 2014). Worobiec G. et al. From the above data, we may conclude that palaeoenvironmental factors inferred from palaeobotanical investigations of both the Gray Fossil Site, United States, and the Bełchatów Lignite Mine, Poland, suggest a warm temperate to subtropical and humid climate associated with Mycoenterolobium eccentricum (R.K. Kar) G. Worobiec, n. comb. in Late Miocene to Early Pliocene of North America and Central Europe. It is in accordance with other fossil records of Mycoenterolobium eccentricum (R.K. Kar) G. Worobiec, n. comb. (see above), where this taxon was associated with subtropical to tropical palaeovegetation. The same concerns the distribution of the modern species of Mycoenterolobium which prefers tropical, subtropical to Mediterranean, usually humid climate.
Similarly to the living species of Mycoenterolobium classified as aero-aquatic hyphomycetes (Goh & Hyde 1996), Mycoenterolobium eccentricum (R.K. Kar) G. Worobiec, n. comb. found in the Gray Fossil Site and the Bełchatów Lignite Mine suggests the presence of small, shallow water bodies with accumulated plant debris (decaying leaves, wood, and bark) and with fluctuating water level or even presence of periodic reservoirs existing only in the wet season or after floods. It is especially true in the case of the locality KRAM-P 226 from the Bełchatów mine.

CONCLUSION
Considering all the above discussion, we may conclude that modern and fossil species of Mycoenterolobium prefer warm (tropical to warm temperate), usually humid climates, and lignicolous habitats associated with freshwater, aquatic or swampy environments. Thus, these investigations may be informative for mycologists, palaeobotanists, and geologists as Mycoenterolobium eccentricum (R.K. Kar) G. Worobiec, n. comb. could be considered as a non-pollen palynomorph proxy for the reconstruction of palaeoenvironment, and palaeoclimate, and could be used for the calibration of the divergence time estimations in the phylogenetic trees of fungi.