Unexpected microbial diversity in new Caledonia’s ultramafic ecosystems with conservation implications in a biodiversity hotspot

A higher microbial diversity on iron crust soils

Surprisingly, fungal species richness was higher in iron crust soils (Goro and Tiébaghi sites). These soils are composed of ironstones and gravels and are, subsequently, empirically thought to harbour more stressful conditions than lateritic soils¹⁵. However, nickel content is known to gradually decrease from the bottom to the top along the weathering profiles of soils from ultramafic rocks¹⁶. Considering their lower nickel content and, subsequently, the potential lower nickel bioavailability, iron crust soils may thus represent less restrictive environments regarding this parameter than previously thought. One hypothesis would thus be that limitation in nickel constraints may promote fungal diversity. However, to the best of our knowledge, no studies have been performed concerning the constraints that iron crust soils have on living organisms, nor has a comparison been made with other soils originating from ultramafic rocks (e.g., in terms of nickel and the bioavailability of other heavy metals, water drainage and temperature). Another hypothesis would be an adaptation to iron crust soils, leading to fungal speciation and diversification. Further, we cannot also exclude a plant composition effect, since maquis on iron crust soils are distinct vegetation types compared to others¹⁷.

For bacteria, greater species richness was only observed in iron crust soils at the Goro site. A previous comparison¹⁸ of the molecular diversity of the symbiotic nitrogen-fixing bacteria genus Frankia from the root system of two Gymnostoma species (Casuarinaceae), endemic to New Caledonia, revealed that G. deplancheanum hosted a greater diversity of Frankia than G. chamaecyparis. G. deplancheanum occurs on iron crust soils, whereas G. chamaecyparis is found on hypermagnesian soils that develop at the base of ultramafic massifs. A greater selective effect of hypermagnesian soils on bacteria was given as a probable explanation¹⁸. This is consistent with our hypothesis of iron crust soils being less constraining in nickel content, but a vegetation composition effect must also be considered. Indeed, G. deplancheanum is a major component of the maquis in the South of the main island and could thus drive bacterial community diversity in this area. The highest values of fungal richness being observed in the G. deplancheanum-dominated vegetation suggest that the presence of this Casuarinaceae species may also locally contribute to increased soil fungal diversity, through direct (e.g., beneficial mutualistic interactions) or indirect (e.g., litter composition) effects.

Soil microbial phyla composition as markers of ecological succession, land degradation and geographic origin

Beyond variations in soil microbial diversity, differences in relative abundances of phyla among vegetation types were observed. Ascomycota and Basidiomycota, the two most abundant fungal phyla in the large-scale dataset, varied in their proportions, with Ascomycota being relatively well-represented in the first stages of the plant successions, especially at the Goro site (open low maquis), as also found by Fernandez Nuñez et al.¹¹. At the Rivière Blanche and Kopéto sites, Ascomycota was also well present in the open vegetation types (i.e., sedge and Tristaniopsis formations); nevertheless, the pattern was not as strong as that detected in Gourmelon et al.¹⁰. Interestingly, we found that at these two sites, Mucoromycota were more prevalent in the first stages of the succession and decreased along the succession. Among Mucoromycota taxa, ASVs assigned to Bifiguratus adelaidae and GS23 clade were detected as the most common in terms of relative abundance and richness. B. adelaidae has been recovered mostly from soil and to a lesser extent as a plant endophyte¹⁹. Despite the broad geographic distribution and abundant site occurrence of this novel taxon, its functional roles are still unknown. The GS23 clade, to which most of Mucoromycota reads and ASVs were assigned, forms a new monophyletic lineage from tropical and subtropical acidic forest soils (2.5 to 4.0)²⁰. Recently, fungal isolation and identification from plant roots in acidic and oligotrophic soil of northeast America led to the description of two new species, belonging to a new genus and a new family, Pygmaeomycetaceae, proposed to correspond to the Clade GS23²¹. The functions of Pygmaeomycetaceae members are not yet fully understood, however, Walsh et al.²¹ suggest that these fungi may be capable of degrading diverse substrates, allowing mobilization of nitrogen and subsequently contributing to the success of their associated host plants in acidic and nutrient-poor environments. Such a hypothesis suggests that ASVs belonging to the GS23 Clade are root symbionts that strongly influence ecosystem functioning at the two distant sites that are Rivière Blanche and Kopéto, especially in vegetation with poor plant coverage and soil nutrition. This reinforces the necessity of determining to which functional groups (in terms of guilds and trophic modes) these Mucoromycota members belong to. Their unknown functionality may explain why no clear trend in fungal functional groups was detected across the successions, or conversely, along the land degradation gradients, at least at Rivière Blanche and Kopéto sites. In addition to this, akin to Ascomycota, high relative abundances of Mucoromycota may be an indicator of land degradation¹⁰ as well as a sign of early stages of an ecological succession¹¹. However, we do not know whether the prevalence of this GS23 Clade in ultramafic substrates is mainly restricted to lateritic soils in general or linked to particular vegetation types.

Similarly, the relative abundances of soil bacteria varied between vegetation types. These discrepancies were overall due to the higher relative abundances of Chloroflexi in the open vegetation and Proteobacteria in the closed vegetation. Despite their low proportions, Cyanobacteria were also mostly encountered in closed maquis and rainforests. These results allow us to generalize the findings made at a local scale by Fernandez Nuñez et al.¹¹, regardless of the type of ultramafic soil (i.e., iron crust soils versus lateritic soils). Indeed, the higher proportion of Chloroflexi that characterized the open low maquis at Goro¹¹ can be extended to the sedge-dominated maquis, two ecosystems characterized by a more or less herbaceous layer with sparse shrubs, which results in lower plant coverage compared to other vegetation types (approximately 10% for Goro’s open low maquis and 30–80% for the sedge maquis). The large relative occurrence of Proteobacteria, as well as the increased presence of Cyanobacteria, previously highlighted from Goro’s closed vegetations¹¹, were also found in most of the closed maquis and rainforests. The prevalence of Chloroflexi and Proteobacteria, in three out of the four successions investigated in our analysis (i.e., at Goro, Rivière Blanche and Kopéto sites, but not at Tiébaghi), may be explained by the availability of nutrients in soil along ecological successions: Chloroflexi suspected as oligotrophs^11,22, survive in nutrient-poor environments, while Proteobacteria, usually copiotrophs²³, prefer nutrient richer substrates.

Another striking finding was the high relative abundance of bacteria belonging to the Actinobacteria phyla at Tiébaghi site independent of the vegetation type. Acidothermus, Conexibacter and Mycobacterium were by far the most dominant taxa in terms of relative abundance and ASVs richness. The Acidothermus genus contains to date only one described species, A. cellulolyticus, that has been isolated and characterized from acidic hot springs²⁴, and amplicons assigned to this genus have since been recovered from acidic soils from China^25,26. The complete genome sequencing of A. cellulolyticus revealed the presence of several genes encoding for enzymes involved in the breakdown of plant and fungal cell wall components²⁷. We, therefore, hypothesise that Acidothermus species possess the ability to use a range of carbon sources and could play a significant role in the degradation of biomass and carbon cycling. For the second most abundant actinobacterial genus, Ma et al.²⁸. recently found that Conexibacter may be involved in carbon and nitrogen biogeochemical cycles in natural forests of eastern China. Both genera may therefore play a central role in nutrient cycles at the Tiébaghi site. In addition, the slow-growing and high genomic GC content observed in Conexibacter strains are suggested as potential traits that could favour the survival of this group in stressful environments^29,30. It is not worthy that besides the Tiébaghi site, members of this taxonomic group were mostly recovered from Goro’s iron crust soils. The third most abundant genus was Mycobacterium which is usually positively related to the prevalence of certain iron minerals in soil^31,32. Overall, the over-representation of these three genera in iron crust environments supports the interest of further characterizing and comparing environmental constraints of distinct ultramafic soils, as well as identifying the key microbial taxa involved in soil biogeochemical cycle regulation.

Focus on ectomycorrhizal fungi

A remarkable finding was the considerable proportion of ectomycorrhizal fungal ASVs that did not find matches at the species level to the existing databases. Out of the 317 delineated ASVs only 41 were assigned to known species (more exactly to 23 described species; several ASVs can be assigned to one species). Based on this dataset, this would indicate a hypothetical endemism rate of 87% in New Caledonia. This result, obtained from a soil eDNA and high-throughput sequencing approach, is consistent with the 95% calculation made by Carriconde et al.⁵ on ectomycorrhizas and fruit bodies typed by Sanger sequencing. New Caledonia clearly hosts a high and unique ectomycorrhizal fungal diversity. This raises questions about the underlying evolutionary and ecological mechanisms that have led to this exceptional and exclusive diversity in this remote territory. The New Caledonian biodiversity is also facing various severe threats (e.g., open cast mining, pollution, frequent and extended wildfires and invasive species introduction) and fungi, as all other living forms, are prone to extinction risks. This high endemism rate points out the effort that should be made to identify fungal species under threat and establish on-the-ground species-oriented conservation plans, an aspect that has never been considered and addressed in this biodiversity hotspot.

Concerning the fungal ectomycorrhizal taxonomic composition, the Cortinarius genus was over-represented in numerous ultramafic vegetation types, corroborating the observations made by Carriconde et al.⁵. on ectomycorrhizas and fruit bodies collected in N. aequilateralis, A. gummiferum and mixed rainforest stands in the south of New Caledonia. With respect to the generalization of this pattern on a larger geographical scale and to other vegetation types than rainforests, it is likely that ultramafic rainforests and shrublands dominated by ectomycorrhizal plant species are characterized by the prevalence of Cortinarius. Nevertheless, despite the limited number of samples, Cortinarius was not the main group in ectomycorrhizas collected in ultramafic substrates from Acacia spirorbis roots³³, a shrub legume that can dominate the ecosystems where it occurs. Future in-depth examinations must be undertaken to confirm or not the pre-eminence of Cortinarius in ecosystems dominated by a single plant species in New Caledonia. Such dominance by this fungal ectomycorrhizal group has direct implications in terms of ecosystems functioning: it appears that Cortinarius has retained from their saprophyte ancestors the capacity to degrade organic matter for potentially mobilizing nitrogen^34,35, a limited element in ultramafic soils⁵.

Finally, it is noteworthy that, in addition to the high relative abundances of ectomycorrhizal fungi in ultramafic vegetation dominated by ectomycorrhizal trees (N. aequilateralis or A. gummiferum) in forests and shrubs (Tristaniopsis spp.) in maquis, this functional group was also prevalent in Goro’s closed vegetation, where only sparse individuals of known ectomycorrhizal plants have been recorded. Isolated ectomycorrhizal plants are unlikely to account for such high ectomycorrhizal fungal abundance and richness (G. deplancheanum-dominated vegetation harbouring the highest ectomycorrhizal fungal richness). As stated by Fernandez Nuñez et al.¹¹, these observations clearly highlight the need to determine the mycorrhizal status of the New Caledonian flora, with only 14 plant species inventoried as ectomycorrhizal to date. This will improve our understanding of the extent to which this mutualistic association influences the ecosystems functioning in this extraordinary hotspot of biodiversity.

A geographical structure of soil microbial communities

Finally, in our analysis, structure investigations (through PERMANOVA analyses, bipartite networks and communities’ partitioning) revealed a geographical clustering of soil microbial communities, a result in accordance with the initial work of Gourmelon et al.¹⁰ undertaken at two distant massifs (Rivière Blanche and Kopéto). For fungi, the two studied iron crust sites, Tiébaghi and Goro, were particularly apart from the other locations and distinct from each other as well. For bacteria, only Goro appeared clearly different. Overall, such geographical structure seems to indicate that each site exhibits its own microbial community. Ultramafic outcrops are patchily distributed and represent edaphic islands. This spatial discontinuity could have led to geographical (allopatric) speciation, contributing to this species diversity³⁶. Soil microorganisms, like plants³⁷, may thus display microendemic patterns.

In terms of soil microbial conservation, this geographical structure has a direct implication: it indicates that each ultramafic massif might be considered as a conservation unit by itself, especially for the isolated massifs such as Tiébaghi and Kopéto. For the “Massif du Grand Sud”, with a surface area of 3015 km², making it the largest continuous ultramafic outcrop in the world³⁸, different hotspots of fungal and bacterial diversity likely exist and may result in many priority areas for soil conservation. Distinct and unique microbial communities may also be hosted by the other types of soils encountered in New Caledonia. Samples from Maré island collected on aluminic-rich soils (gibbsic Ferralsols) were, indeed, completely dissimilar from ultramafic soils. The interest in focusing on soil biota for establishing priority areas has been very recently spotlighted by Guerra et al.³⁹ at the Earth scale. However, despite the input of this work, the approach remains imprecise, with many regions being not considered, including well-known biodiversity hotspots like New Caledonia. A high-resolution investigation of microorganisms, and other soil-living organisms (e.g., protists, nematodes, earthworms, and arthropods), using high-throughput sequencing technologies, on a large array of soil types could constitute a next future research challenge. At the scale of this territory and more widely, it will undeniably help to put on the map this still largely neglected hidden biodiversity and will contribute to a more holistic perception of nature conservation.