Karst Habitats Research Articles

A new species of Bent-toed Gecko (Squamata, Gekkonidae, Cyrtodactylus)from the Shan Plateau in eastern Myanmar (Burma).

An integrative taxonomic analysis of the Cyrtodactylus linnwayensis group of the Shan Plateau recovered two new populations from isolated karst habitats near Pinlaung Town, Shan State as a new species, C. pinlaungensis sp. nov. Cyrtodactylus pinlaungensis sp. nov. is most closely related to a clade comprising C. linnwayensis and C. ywanganensis from the western edge of the Shan Plateau approximately 90 km to the northwest. Cyrtodactylus pinlaungensis sp. nov. can be distinguished from all members of the C. linnwayensis group by a number of statistically different morphological characters, discrete color pattern differences, and its heavy tuberculation. It also bears an uncorrected pairwise sequence divergence of 5.0-7.6% from all other species combined based on the mitochondrial gene ND2 and its flanking tRNAs. The discovery of this new species on the Shan Plateau continues to underscore the fact that this region is rapidly emerging as a herpetological diversity hot-spot for Myanmar.

Uncovering karst endemism within Borneo: two new Cyrtodactylus species from Sarawak, Malaysia.

The island of Borneo lies within one of the most biodiverse regions in the world. Despite this, its documented gekkonid diversity is not commensurate with other areas of Southeast Asia. The megadiverse genus Cyrtodactylus is especially underrepresented. Limestone-karst ecosystems, in particular, harbor many endemic Cyrtodactylus species, but only one karst-dwelling species is currently recognized from Borneo. This paper adds two additional karst-dwelling Cyrtodactylus species-C. muluensis sp. nov. and C. limajalur sp. nov.-from Sarawak, Malaysia. Cyrtodactylus muluensis sp. nov. is endemic to Gunung Mulu and is distinguished from its congeners by having a precloacal groove, 31-38 ventral scales, a maximum SVL of at least 88 mm, enlarged subcaudals, 19-20 subdigital lamellae, and a banded dorsal body pattern. Cyrtodactylus limajalur sp. nov. is endemic to the Serian region and is distinguished from its congeners by having 33-42 ventral scales, enlarged subcaudals, a precloacal pit, a maximum SVL of at least 94 mm, 5-6 enlarged femoral scales, 19-22 subdigital lamellae, and five distinct bands on the dorsum. Both species are phylogenetically distinct and deeply divergent from all other congeners. The description of two new karst-dwelling species highlights the need to conserve karst habitats and the endemic species they harbor.

A new species of Achnanthes (Bacillariophyceae) from a freshwater habitat in a karst landform from south‐central China

SUMMARY Achnanthes maolanensis nov. sp. is a large diatom collected from subaerial, freshwater habitats in karst landforms in central‐south China. Living cells have two chloroplasts. Valves of this new species are panduriform in outline, slightly constricted in the middle part of the margins, with uniseriate areolae; the raphe is filiform, and there is a linear, thickened stauros in the central area of the RV. On the ARV, there is no central area, and as well there are no terminal orbiculi, marginal ridge, or terminal spine externally. The axial area (rapheless sternum) is located down the center, not offset from the center as in many other species of the genus and is narrow and almost straight. This suite of characters makes the new species easy to distinguish from other species in the genus. Its presence adds further documentation of the unique and diverse freshwater diatom flora from karst habitats in this region of China.

Expression analysis of calcium-dependent protein kinases (CDPKs) superfamily genes in Medicago lupulina in response to high calcium, carbonate and drought

The aim of this study is to identify calcium dependent protein kinases (CDPKs) superfamily genes (including calcium dependent protein kinase (CDPK) genes, CDPK-related kinase (CRK) genes, and phosphoenolpyruvate carboxylase kinase-related kinase (PEPRK) genes) and to analyze their expression in response to high calcium, bicarbonate, and water deficit in Medicago lupulina. Transcriptome sequencing databases and bioinformatics were used to identify CDPK superfamily genes. qRT-PCR analyses were used to study the expression of CDPK superfamily genes in response to high calcium, bicarbonate, water deficit, and their combined treatments. Ten MlCDPKs, four MlCRKs, and two MlPEPRKs genes were identified from transcriptome sequencing databases. The identified MlCDPK superfamily genes could be divided into six subgroups including CDPK I, CDPK II, CDPK III, CDPK IV, CRK, and PEPRK according to evolutionary relationships. Analysis of protein structure indicated that MlCDPK proteins contained a protein kinase domain and EF-hand domain. However, PEPRKs contained only a protein kinase domain while CRKs contained also degenerative EF-hands. Most of the MlCDPK superfamily genes were differentially expressed in different organs. According to their expression levels, MlCDPK superfamily genes such as MlCDPK1, MlCDPK9, and MlCDPK10 were more sensitive in response to individual treatments of high calcium, carbonate or PEG rather than their combination of treatments. The results indicated that MlCDPK superfamily genes may play an important role in response to those stresses. These results provide important insight into the role of CDPKs in M. lupulina, especially in the tolerance to Karst habitats, where high calcium and carbonate and drought conditions are the dominant ecological limiting factors.

Mitrephora chulabhorniana (Annonaceae), an extraordinary new species from southern Thailand

Mitrephora chulabhorniana, a remarkable new species with the smallest flowers in the genus, is described and illustrated. The new species, from a karst habitat of southern Thailand, morphologically most resembles M. andamanica, which is endemic to the Middle and North Andaman Islands, but differs primarily by having narrower leaves, shorter inner petals, and lower number of stamens and carpels per flower. The conservation status of M. chulabhorniana is assessed and a key to the species of Mitrephora in Thailand is given.

Two new rock-inhabiting species of Cyphellophora from karst habitats in China

During survey on rock-inhabiting fungi from karst habitats in Guizhou, China, two new species in Cyphellophora were discovered and identified. Phylogenetic analysis based on combined sequences of the nuclear large subunit (nucLSU) and internal transcribed spacer (ITS) region of ribosomal DNA revealed that the tested isolates, clustered into two clades that well affiliated in the genus of Cyphellophora. Morphological characteristics were also supported the estabolishment of the new species. Herewith Cyphellophora botryose sp. nov. and Cyphellophora guizhouensis sp. nov. were described and their differences from closely related species were discussed.

Nitrogen transfer from one plant to another depends on plant biomass production between conspecific and heterospecific species via a common arbuscular mycorrhizal network.

The formation of a common mycorrhizal network (CMN) between roots of different plant species enables nutrient transfers from one plant to another and their coexistence. However, almost all studies on nutrient transfers between CMN-connected plants have separately, but not simultaneously, been demonstrated under the same experimentation. Both conspecific and heterospecific seedlings of Cinnamomum camphora, Bidens pilosa, and Broussonetia papyrifera native to a karst habitat in southwest China were concurrently grown in a growth microcosm that had seven hollowed compartments (six around one in the center) being covered by 35.0-μm and/or 0.45-μm nylon mesh. The Ci. camphora in the central compartment was supplied with or without Glomus etunicatum and 15N to track N transfers between CMN-connected conspecific and heterospecific seedlings. The results showed as follows: significant greater nitrogen accumulations, biomass productions, 15N content, % Ntransfer, and the Ntransfer amount between receiver plant species ranked as Br. papyrifera≈Bi. pilosa > Ci. camphora under both M+ and M-, and as under M+ than under M- for Ci. camphora but not for both Bi. Pilosa and Br. papyrifera; the CMN transferred more nitrogen (15N content, % Ntransfer, and Ntransfer amount) from the donor Ci. camphora to the heterospecific Br. papyrifera and Bi. pilosa, with a lower percentage of nitrogen derived from transfer (%NDFT). These findings suggest that the CMN may potentially regulate the nitrogen transfer from a donor plant to individual heterospecific receiver plants, where the ratio of nitrogen derived from transfer depends on the biomass strength of the individual plants.

The composition and diversity of arbuscular mycorrhizal fungi in karst soils and roots collected from mulberry of different ages

ABSTRACT: Arbuscular mycorrhizal fungi (AMF) have been attracted more scientific attentions due to its critical role in enhancement of drought tolerance of plants for growth and vegetation restoration in karst fragile ecosystem. However, scientists know little about the AMF composition and diversity occurring in root systems of mulberry (Morus sp.), and in karst habitats which return land use from mulberry forestry, as well as the effects of soil environment change on the diversity of the AMF communities. To understand: (1) the AMF community composition and diversity at different stage of returning cropland to forest; and (2) the effects of soil environment change on the diversity of the AMF communities, soil and mulberry root samples were collected from Bijie and Libo sites, China, which experienced one and ten years, respectively, after returning croplands to forest. With the high throughput 454-sequencing technology, 8 known genera including 83 virtual species were distinguished and the genera Glomus, Paraglomus, Archaeospora and Diversispora were found to be dominant in soil and root sample. Compared to the samples in Libo, the genera Glomus, Paraglomus, Acaulospora and Claroideoglomus in root samples at Bijie site had a relatively abundance of species indicating that the returning cropland to forest is benefit to the AMF diversity and abundance, which was attribute to the variation of soil physiochemical properties. This conclusion is of great significance for guiding the return of farmland to forests.

Behavioral response of the endemic Martino’s vole Dinaromys bogdanovi (Martino 1922) to environmental complexity

The Martino’s vole (Dinaromys bogdanovi) is a rare species that lives in differently structured karst habitats, varying from open rocky plains to deep fissures, mostly residing under boulders and in crevices. Populations of the species are declining, probably due to its strict habitat preferences and competition with the European snow vole. Since the species is difficult to study in the wild, we tested its behavioral response to a differently complex environment and novel object presence in captivity. We exposed 14 individuals to differentially complex setups, from open and unsheltered to rocky with covered tunnels. We measured the effect of the presented setup and season on vole behavior. Analysis showed seasonal differences with higher movement frequency and longer time spent still during the breeding season, and a higher number of attempts due to the lack of vole entry into the experimental terrarium in the non-breeding season. Additionally, movement significantly differentiated between an open and simple setup in comparison to more complex ones, with higher frequencies of moving and peeping and the duration of peeping in the simple setup, indicating restlessness. We also found significant differences between the novel object and control setup. The results of this study could be useful for future assessment of the influence of habitat complexity on mobility in the wild. Moreover, this is the first study on the behavior of this endemic and rare rodent.

Beauty underground: cavefishes in China

Cavefishes are a distinctive group of animals restricted to subterranean environments for at least for part of their lives. Their level of troglomorphism differentiates them into groups: stygobionts (troglomorphic) and stygophiles/stygoxenes (non-troglomorphic). China is home to the greatest diversity of cavefishes in the world, with more than 76 endemic stygobiotic species and an additional 70+ non-stygobiotic taxa. All but one species (Onychostoma macrolepis in northern China), are found in the southwest, primarily in the karst of the Yunnan-Guizhou Plateau. Sinocyclocheilus, the largest cavefish genus with 35 stygobiotic species, has undergone rapid evolution and diversification. Many species in this genus possess horns and humpbacks (i.e., horn-like structures and hyperdevelopment of a dorsal protuberance similar to a humpback), probably resulting from parallel evolution. Syntopy (inhabitation of the same cave system by multiple cavefishes) is common in China, and sometimes several stygobiotic species are found in the same cave pool or subterranean river. Caves and karst habitats are fragile environments, and cavefishes are extremely sensitive to environmental change. Threats from rapid economic growth in recent decades have increased the need for conservation efforts for cave-dwelling communities, and the conservation of Chinese cavefishes is an urgent issue, in particular. To this end, the Chinese Cavefish Working Group (CCWG) was created form a collaborative effort by researchers from China and the United States. The primary goal of the CCWG is to document the distribution and diversity of Chinese cavefishes, to gather information on their status and threats to wild populations and describe unknown aspects of their organismal biology. These data can be used to make sound conservation and management decisions. Research efforts are not limited to surveys and collection but also taxonomy, systematics, morphology, ecology, behavior, sensory neuroethology, evolution, population biology, and conservation science.

A new Cyrtodactylus Gray, 1827 (Squamata, Gekkonidae) from the Shan Hills and the biogeography of Bent-toed Geckos from eastern Myanmar.

A phylogenetic taxonomic analysis indicates that a newly discovered population of Cyrtodactylus from the vicinity of Ywangan Town in the Shan Hills, Shan State, Myanmar is a new species (C. ywanganensis sp. nov.) and the earliest diverging member of the linnwayensis group within the previously defined Indochinese clade. The DIVALIKE+J model of a BioGeoBEARS biogeographic analysis indicates that the Indochinese clade evolved in the Shan Hills and Salween Basin of eastern Myanmar and dispersed into Indochina on at least three separate occasions from 18.6-13.4 mya. Once there, uplift of the Tenasserim Mountains and Thai Highlands created the intermedius group, the oldhami group, and C. tigroides of western and southern Thailand which form sister lineages to the linnwayensis group, yathepyanensis group, and the sinyineensis group, respectively, of eastern Myanmar. Diverging lineages within the Indochinese clade highlight the importance of the Thai Highlands and Tenasserim Mountains in that group's evolution and speciation. The discovery of C. ywanganensis sp. nov. in karstic habitats in the Shan Hills continues to underscore the unrealized karst-associated herpetological diversity of this vast, relatively unexplored, upland region and the need for additional field studies.

Environmental drivers influencing stonefly assemblages along a longitudinal gradient in karst lotic habitats

Stoneflies are among the most sensitive aquatic insect taxa and therefore arguably the best indicator of the excellent, i.e. pristine, ecological status of surface streams. Karst habitats are one of the most exciting freshwater habitats in terms of biological-geological interplay. They, in turn, support a biodiversity far superior to surrounding freshwater habitats and hence these habitats are designated as biodiversity hotspots. Our study deals with both of these crucial ecological players. We studied stonefly assemblages, their microhabitat preferences and emergence patterns along a karst oligotrophic hydrosystem. The sampling was conducted monthly from March 2007 to December 2008 using pyramid-type emergence traps set in various habitats and associated microhabitats (e.g. springs, rivers, streams, tufa barriers × moss, angiosperm, cobble, sand, silt substrates). Favorable environmental conditions, such as a wide range of karst habitat types with low water temperature and high oxygen concentration, resulted in high stonefly species richness (31 recorded species). Water temperature and pH had the highest influence on stonefly assemblages. Species richness and diversity decreased in a downstream direction. We recorded a longitudinal shift from crenal-epirhithral to epirhithral-metarhithral assemblages with some hyporhithral and potamal elements. Upstream sites were dominated by shredders, while downstream sites had a higher proportion of gatherers-collectors. Several species showed a significant preference for a specific microhabitat type in accordance with their feeding strategies and food availability. The majority of recorded species exhibited univoltine life cycles slow or fast.

Ecological traits of water beetles in a karstic river from the Eastern Mediterranean region

This study presents new insights into ecological traits of water beetles in lotic karst habitats of the Eastern Mediterranean. The aims of the study were to investigate population aspects and ecological traits of water beetles. Data were collected monthly during one year from a large perennial river showing current anthropogenic disturbance. Altogether 11 families, 23 genera and 31 species were identified. Elmidae were the most diverse and abundant family, and the genus Elmis predominated at all sampling sites. The distribution of water beetles along the river was influenced primarily by river bed morphology and beetle substrate preference. Water beetles were more abundant in bryophytes but more diverse in angiosperms. Ecological traits of water beetles along the watercourse were defined primarily by alkalinity and current velocity. Larval stages correlated with water depth and current velocity, while the adult stage correlated only with water depth. Our results support the potential of water beetles as indicators of water quality. This report provides the most detailed investigation so far of water beetles in the Eastern Mediterranean region, providing a basis for further detailed studies of water beetles and their ecology in freshwater habitats in southeastern Europe.

Three new species of nocturnal geckos of the Paroedura oviceps clade from xeric environments of Madagascar (Squamata: Gekkonidae).

We describe three new gecko species of the Paroedura oviceps clade, diagnosed by deep divergences in mitochondrial DNA, absence of haplotype sharing in two nuclear genes (sacs and kiaa1239), and morphological differences. Paroedura spelaea sp. nov. is an extremely slender species from karst habitats in the limestone massif of the Tsingy de Bemaraha National Park in western Madagascar, morphologically reminiscent of P. homalorhina but distinguished by the absence of distinct dorsal rows of spiny tubercles. Paroedura fasciata sp. nov. from the small karstic island Nosy Hara in northern Madagascar is phylogenetically placed sister to P. spelaea despite radical morphological differences. It is morphologically most similar to P. hordiesi from the geographically close Montagne des Français massif, but is smaller, has slightly more spiny dorsal scales, and a distinctive colour pattern with light grey dorsal crossbands. The third species, Paroedura kloki sp. nov., is known from Ankarafantsika National Park and the western slopes of Makira, two sites of dry to transitional forest in western Madagascar. It is at least partly arboreal and morphologically very similar to P. oviceps, from which it differs by spiny scales extending over its entire tail. Although incomplete, the available phylogenetic evidence suggests that the karst specialists in the P. oviceps clade (P. fasciata, P. homalorhina, P. hordiesi, P. spelaea) form a monophyletic group which might have diversified by vicariance after becoming isolated, respectively, in their limestone habitats in Tsingy de Bemaraha, Ankarana, Nosy Hara and Montagne des Français.

Comparison of woody species composition between rocky outcrops and nearby matrix vegetation on degraded karst hillslopes of Southwest China

Habitats with different features such as soil depth and soil/rock conditions can provide favorable environments for species with different requirements, while anthropogenic disturbances normally exert additional effects on species composition. However, specific studies have rarely been conducted in the degraded karst regions of Southwest China despite the high heterogeneity of karst habitats and past human disturbances. In this study, woody species richness and composition on rocky outcrops on a typical karst hillslope were investigated and compared with those of nearby matrices on shallow and rocky soil. Our results showed that matrix vegetation was more diverse in genera and species than vegetation on rocky outcrops. This might relate to the contrasting substrate features and different disturbance histories of these two habitats. Unlike the significant effect of slope on species richness of the matrix vegetation, rocky outcrops exhibited no significant differences between upper and lower slope positions, largely because their microhabitats were similar in different slope positions. Although the study area has been reforested naturally for about 30 years, woody species of the matrix vegetation were still dominated by pioneer shrub species. Rocky outcrops were dominated by late-successional tree species, which was primarily related to their isolated features and resistance to certain disturbances. Most of these late-successional species were not habitat endemics, indicating the possibility for their encroachment into surrounding the matrix. From this aspect, further studies will be necessary to identify and address the limiting factors for the encroachment of these late-successional species into the surrounding environment.

Three more new species of Cyrtodactylus (Squamata: Gekkonidae) from the Salween Basin of eastern Myanmar underscore the urgent need for the conservation of karst habitats

ABSTRACTAn integrative phylogenetic taxonomic analysis recovers three additional new species of karst-associated Cyrtodactylus Gray (Squamata: Gekkonidae) – C. bayinnyiensis sp. nov., C. chaunghanakwaensis sp. nov. and C. naungkayaingensis sp. nov. – from a narrow zone in the Salween Basin of Kayin and Mon states in eastern Myanmar from which nine new species were recently described. This degree of unprecedented diversity and site-specific endemism will no doubt continue to rise when at least 44 unsurveyed karstic habitat-islands in this same area are also explored. These data indicate that karst habitats not only serve as foci for speciation, but their rugged terrain spares them from agricultural development and, as such, they are the only habitats in the Salween Basin wherein much of the pre-agricultural herpetofauna can survive. This continues to underscore the fact that karst habitats in Myanmar harbour a significant portion of that country’s herpetofauna, some of which remains undescribed. Despite eastern Myanmar constituting some of the most extensive karstic regions in South-east Asia, they are the least legally protected, with only 1% of their terrain recognised as vulnerable. Until karst habitats in Myanmar are thoroughly investigated, a significant portion of this country’s herpetological diversity will remain underestimated and unprotected. Therefore, issues associated with karst conservation and management in Myanmar should be elevated to a new level of urgency.

First Records ofMyotis altarium(Chiroptera: Vespertilionidae) from India and Vietnam

The Szechwan myotis, Myotis altarium, was first described in 1911 from southeastern China, and for the next 100 years was thought to be restricted to that country with just one additional individual known from Aung Kang region in northern Thailand. We captured eight individuals of this species during surveys in Copia Nature Reserve in northwestern Vietnam and two individuals from East Khasi Hills and East Jaintia Hills districts of Meghalaya state, northeastern India. They are described here as the first records for Vietnam and India. We also provide comparative mensural data including the holotype and other specimens from Southeast Asia. The record of M. altarium from Meghalaya significantly extends the westward distribution range of this species beyond Thailand. Our study affirms previous observations of the cave dwelling habits of M. altarium and suggests that this species temporarily hibernates during cold winter periods in Vietnam. It is also suggested that its distribution range may encompass other karstic habitats in the mainland Indomalayan region.

Microhabitat selection and distribution of functional feeding groups of mayfly larvae (Ephemeroptera) in lotic karst habitats

Research examining mayfly ecology in karst streams and rivers has increased in recent years, though microhabitat preferences remain poorly characterized. We examined mayfly assemblage taxonomy, functional feeding groups and microhabitat preferences in two contrasting lotic Dinaric karst catchments, one pristine and one anthropogenically impacted. At monthly intervals over a one-year period, all major microhabitats (i.e.dominated by boulders, cobbles, sand, silt, mosses, or angiosperms) were sampled at sites spanning springs, upper, middle and lower river reaches, and tufa barriers. In both catchments, mayfly species richness was comparable among microhabitats, while mayfly abundance was highest on mosses and lowest on silt. NMDS ordination did not group assemblages according to microhabitat type, which may reflect the greater influence of physical and chemical water properties. In both catchments and all microhabitats, mayfly assemblages were dominated by grazers/scrapers at upstream sites and by detritivores at downstream sites. Active filter feeders were more abundant in microhabitats with silt substrates and lower current velocities. This study demonstrated that certain mayfly species strongly preferred a specific microhabitat type, reflecting their water current preferences and feeding strategies, while other species shifted between microhabitats, likely in search of food resources and shelter. The results emphasize the importance of habitat heterogeneity in supporting diverse communities in karst rivers.

Analysis of Genetic Diversity and Structure Pattern of Indigofera Pseudotinctoria in Karst Habitats of the Wushan Mountains Using AFLP Markers

Indigofera pseudotinctoria Mats is an agronomically and economically important perennial legume shrub with a high forage yield, protein content and strong adaptability, which is subject to natural habitat fragmentation and serious human disturbance. Until now, our knowledge of the genetic relationships and intraspecific genetic diversity for its wild collections is still poor, especially at small spatial scales. Here amplified fragment length polymorphism (AFLP) technology was employed for analysis of genetic diversity, differentiation, and structure of 364 genotypes of I. pseudotinctoria from 15 natural locations in Wushan Montain, a highly structured mountain with typical karst landforms in Southwest China. We also tested whether eco-climate factors has affected genetic structure by correlating genetic diversity with habitat features. A total of 515 distinctly scoreable bands were generated, and 324 of them were polymorphic. The polymorphic information content (PIC) ranged from 0.694 to 0.890 with an average of 0.789 per primer pair. On species level, Nei’s gene diversity (Hj), the Bayesian genetic diversity index (HB) and the Shannon information index (I) were 0.2465, 0.2363 and 0.3772, respectively. The high differentiation among all sampling sites was detected (FST = 0.2217, GST = 0.1746, G’ST = 0.2060, θB = 0.1844), and instead, gene flow among accessions (Nm = 1.1819) was restricted. The population genetic structure resolved by the UPGMA tree, principal coordinate analysis, and Bayesian-based cluster analyses irrefutably grouped all accessions into two distinct clusters, i.e., lowland and highland groups. The population genetic structure resolved by the UPGMA tree, principal coordinate analysis, and Bayesian-based cluster analyses irrefutably grouped all accessions into two distinct clusters, i.e., lowland and highland groups. This structure pattern may indicate joint effects by the neutral evolution and natural selection. Restricted Nm was observed across all accessions, and genetic barriers were detected between adjacent accessions due to specifically geographical landform.

Biomass Production of Three Biofuel Energy Plants’ Use of a New Carbon Resource by Carbonic Anhydrase in Simulated Karst Soils: Mechanism and Capacity

To determine whether the bicarbonate in karst limestone soil could be used as a new carbon resource for biomass production by the catalysis of carbonic anhydrase (CA), a simulative karst drought stress experiment was designed and performed. Three plants used for biofuel energy, Orychophragmus violaceus L. (Ov), Brassica juncea L. (Bj), and Euphorbia lathyris L. (El), were grown under simulated karst drought stress. In response to drought stress, the photosynthesis of the three energy plants was inhibited, but their CA activity increased. The hypothesis was confirmed by plant physiological and stable isotope techniques. The obtained results showed that plant biomass was produced with atmospheric CO2 as well as bicarbonate under drought stress. Bicarbonate use was proportional to the CA activity of the plants. With high CA activity over a long period, El had the highest proportional bicarbonate use compared to Ov and Bj, reaching 26.95%. Additionally, a new method is proposed for the screening of plants grown for energy in karst habitats.