Seasonal diversity and distribution of decomposing macrofungi in three forest communities: why do they differ?

The diversity of macrofungal species, especially those that are not edible, receives less attention because people do not realize the importance of their ecological role. This study aimed to reveal the inter-seasonal diversity of macrofungal species in three forest communities in South Sulawesi: Karst forest, Lowland forest, and Pine forest. Field surveys were carried out twice (i.e. in the wet and dry seasons) in 25 units plots measuring 10 m x 10 m which were systematically spread in an area of approximately 1 ha in each forest community. The macrofungi found in each plot were recorded by the species name, photo, and the area of cover that measured using Crown Diameter Method to analyse the area of cover. In total there were found 130 species of macrofungi from 38 families across forest communities both in wet and dry seasons. The highest number of macrofungi species was found in the Lowland Forest (67 species), followed by Karst Forest (65 species), and the lowest was in Pine forest (42 species). Some species are specialists in specific forest communities. Given that most species of macrofungi are monophagous that specialist to an only litter of one tree species, the difference in the diversity of macrofungi among forest communities is likely related to the difference in tree species diversity among forest communities. In addition, microhabitat conditions were found in line with the diversity of macrofungal species inter-seasons and inter-forests.


Introduction
Fungi are organisms that digest food externally and absorb nutrients directly through their cell walls [1].Most fungi produce fleshy fruiting bodies with sexual reproductive structures, however, some also have asexual reproductive structures [2].As heterotrophic organisms, fungi can be distinguished based on their size: microfungi and macrofungi.In general, people can easily recognize macrofungi because they have a macroscopic life phase in the form of a fruiting body which people call it mushroom.[3,4].
Ecologically, macrofungi can be classified into three major groups: saprophytes, parasites, and symbiotic species [2,5].Almost all terrestrial macrofungi are saprophytes and mycorrhizal symbionts, but some of them are plant pathogens.Saprophytic fungi obtain nutrients through dead plants or animals for growing [1].While symbiotic mycorrhizal fungi and parasites obtain their nutrition from a living host (plant or animal) [2,6].Through symbiotic associations, mycorrhizal fungi obtain carbohydrates (plant-produced sugars) from plants, and in return the plant obtains minerals from the fungi [6].Parasitic fungi, on the other hand, obtain their nutrition by killing the living cells of their host [7].
Macrofungi have now attracted people's interest because of their broadly important role they play in an ecosystem: i.e. as agents for decomposing organic matter [2], foodstuffs, pharmaceuticals, and even biocontrol agents used in many industries [8][9][10][11].However, behind these benefits, an understanding of macrofungi is still insufficient due to difficulties in studying them, especially those that grow naturally in the field.This is due to its sporadic and short-lived nature, as well as its growth and distribution pattern which depend on several environmental factors.Hence, they have been mainly neglected and overlooked in nature conservation both at the national and international levels.
Macrofungi are cosmopolitan species, but they require certain environmental conditions to grow.Each macrofungal species requires a different habitat, different substrates, and different climatic conditions to grow [12].Naturally, most macrofungal species grow during the wet season, but there are also several species that grow during the dry season [13,14].Mahmud et al. [15] revealed that environmental conditions that mainly affect the growth of fungi are temperature, humidity, and CO2.Hu et al. [16] added that the emergence of macrofungi is strongly influenced by the type of vegetation.Humidity, rainfall, and wind speed are other environmental factors reported by Hu et al. [16] which was also found to have a significant impact on the occurrence of macrofungi.
The number of macrofungal species in the world is estimated at about 1.5 million, of which only approximately 120,000 species are known to date [17,18].Among the 120,000 known species, 16,000 are found in the Indonesian archipelago, particularly on island of Java [19].Meanwhile, data on the presence and diversity of macrofungi from outside Jawa is lacking.
The lack of data on the macrofungi diversity is also a particular problem for Sulawesi, the largest island in the Wallacea region, which is known biogeographically for its unique biodiversity [20].This study aims to compare the differences in the diversity of macrofungi in three forest communities with different habitats and species composition seasonally: a 47-year-old secondary broadleaf karst forest, a 56-year-old secondary broadleaf lowland forest, and a 60-year-old Pinus merkusii plantation forest.Given that each species of macrofungi is usually associated with a particular tree species as its host [21], we predict that most macrofungi species found in each forest community vary according to differences in tree species composition.The results of this study will enrich the lack of information about macrofungi in Sulawesi and become a database to support conservation efforts and sustainable use of macrophytes in the future.

Study site
This study was carried out in 2021 in three secondary forest communities on the southern peninsula of Sulawesi Island, Indonesia.The three secondary forest communities are 47-old broadleaf karst forest (119 o 44'14.9"E, 05 o 01'46.8"S, hereafter referred to as Karst forest), 56-old broadleaf lowland forest (119 o 46'35.0"E, 04 o 58'06.9"S, hereafter referred to as Lowland forest), and 60-old Pinus merkusii plantation forest (119 o 45'56.7"E, 05 o 00'17.3"S, hereafter referred to as Pine forest).The Karst forest is located within the Karaenta forest complex of the Bantimurung Bulusaraung National Park, while Lowland and Pine forests are located in the Education Forest of Hasanuddin University.To reveal the habitat characteristics of each forest community, vegetation analyses were carried out in each forest community by Putra et al. [22], as shown in Table

Climate and environmental factors
Rainfall data for the study site for 2021 were obtained from the Indonesian Agency for Meteorology, Climatology, and Geophysics that located in Maros City (Figure 1).According to Andrew et al. [14], microhabitat conditions affect the growth of macrofungi.For this reason, we measured several factors of microhabitat conditions, including temperature, soil moisture, and soil pH every 1 st to 2 nd of each month during the study period.Temperature data was measured by installing a maximum-minimum thermometer on a tree trunk at the height of about 1 m from the ground.Soil moisture and pH measurements were carried out using the Takemura Dm-5 Soil pH and Moisture Tester in 10 systematically selected plots.Meanwhile, the measurement of light intensity was only carried out once in the entire plots using a Lux meter type LX-100.Measurements were made during the day between 11.00 -13.00 in cloudless conditions.

Macrofungal diversity and distribution
Observations of macrofungi were carried out in 25 plots measuring 10 m x 10 m which were systematically distributed over an area of approximately 1 ha in each forest community.Observations on macrofungi included the identification of species, area of cover, and type of substrate.Identification of species in the field was carried out using a macrofungal field manual.All macrofungal species found were photographed.When identification could not be carried out directly in the field using a field guidebook, species identification was carried out in the laboratory through several online identification manuals.For each macrofungal colony that we found in each plot, we measured the colony diameter using the Crown Diameter Method [23].According to this method, the diameter of an imperfect circle is measured four times from different directions, and the diameter used is the average of the four measurements.In addition, the type of substrate on which macrofungi grew was also recorded, such as litter (leaves, twigs, fruits), decayed wood, living tree trunks and roots, cow dung, soil, and rooks.Observations of macrofungi were made twice: first in the wet season (March 2021) and second in the dry season (September 2021).

Data analysis
The area of cover of each colony of macrofungal species in the three forest communities, we analyzed using the following formula: [24] ‫ݎ݁ݒܥ‬ Where: π = Constants (3.14) We used the Analysis of Variance (ANOVA) with Tukey Honest Significant Difference (HSD) method to detect the differences in (a) the soil moisture among the forest communities in the wet and dry seasons; (b) the soil pH among the forest communities in the wet and dry seasons; (c) the Light Intensity among the forest communities.All statistical analyses were performed using the R version 4.2.1 application [25].

Environmental conditions during the wet season and dry season
Different macrofungal species grow in different microclimatic environmental tolerance ranges [26,27].Among the three forest communities studied, the highest temperature maximum during the wet season was recorded in the Pine forest, while the lowest was recorded in the Karst forest (Table 2).However, during the dry season, the maximum temperature did not show a difference among the three forest communities.The same result was also recorded for the minimum temperature during the wet season, the highest minimum temperature was recorded in the Pine forest, while the lowest minimum temperature was recorded in the Karst forest.Minimum temperature during the dry season did not differ between the Karst forest and the Pine forest but was lower in the Lowland forest.
For the soil moisture during the wet season, the highest value was noted in the Karst forest, followed by the Lowland forest, and lowest in the Pine forest (F = 70.8473,df = 2, P <0.001).In the dry season, the mean soil moisture was significantly highest in the Lowland forest, followed by the Karst forest, and lowest in the Pine forest (F = 114.7268,df = 2, P <0.001).The mean pH during wet season was not significantly different between the Karst and Lowland forests but was much lower in the Pine forest (F = 14.3889, df = 2, P = 0.0016).Meanwhile, the mean pH during the dry season was significantly highest in the Lowland forest, followed by the Karst forest, and lowest in the Pine forest (F = 17.4577, df = 2, P <0.001).For the mean light intensity, the highest value was noted significant in the Pine forest, followed by the Karst forest, and lowest in the Lowland forest (F = 7.3939, df = 2, P = 0.0012).
Boddy et al. [28] stated that among the microhabitat factors, temperature, soil moisture, and light intensity are ecologically the most important environmental factors for fungal growth.Macrofungi generally grow in a range of temperature intervals between 25°C -35°C [29] and soil moisture at an interval range of 70% -90% [30].Relative high soil moisture is strongly associated with high rainfall [31].On the other hand, the higher the intensity of sunlight in a habitat, the lower the diversity of macrofungi species because in such habitat conditions, the temperature increases which then triggers evaporation so that soil moisture decreases.Yusef and Allam [32] revealed that high irradiation intensity would inhibit the growth of macrofungi populations because it would inhibit the formation of reproductive organs and fungal spores.

Seasonal diversity of macrofungi in the three forest communities
The total species of macrofungi found both in the wet and dry seasons were 130 species from 38 families (Figure 2).Three families with the largest number of species were Polyporaceae (19 species), Marasmiaceae (17 species), and Ganodermataceae (12 species).Prasetyaningsih and Rahardjo [33] reported that, families from the Order of Polyporales, especially Polyporaceae and Ganodermataceae, were less affected by changes in physical environmental factors.Their large and hard fruit body allows them to adapt well in various habitats, both in wet and dry habitat conditions [34].Meanwhile, although their fruiting bodies are relatively more minor and have a smoother appearance, species from Marasmiaceae have the ability to survive the dry season by wrinkling their fruiting body, then quickly returning to their normal state in the following wet season [35].
Of the 130 species of macrofungi found, 81 species were recorded in the wet season, 22 species were recorded in the dry season, and 27 other species were recorded in both the wet and dry seasons (Table 3).The highest number of species was recorded in the Lowland forest (67 species), the second highest in the Karst forest (65 species), and the lowest in the Pine forest (42 species).During the wet season, the largest number of macrofungal species was recorded in the Karst forest and the lowest number was in the Pine forest.During the dry season, the largest number was recorded in the Lowland forest, and the lowest number was in the Karst forest.For macrofungal species that were found in both seasons, the largest number of species was found in the Lowland forest while the lowest was in the Pine forest.
In all forest communities, the diversity of macrofungal species was distinctly higher in the wet season than in the dry season.Ideal climate conditions during the wet season, where the relative humidity is higher, the temperature is lower, and light intensity is also lower, support fungal fruiting [28,36,37].This finding was also supported by Tibuhwa [38], who studied the phenology of the macrofungal community at the University of Dar es Salaam main campus in Tanzania, and by Sutjaritvorakul et al. [39] who studied the seasonal (wet and dry season) and climatic changes on macrofungal diversity in Sai Yok District, Kanchanaburi Province, China.This study found that the fruiting bodies of several macrofungal species from the families of Auriculariaceae, Boletaceae, Hygrophoraceae, Hymenogastraceae, Marasmiaceae, Pleurotaceae, Psathyrellaceae, Pyronemataceae, and Tricholomataceae grew in the dry season.Meanwhile, a previous study reported that in general, macrofungal species from these families are difficult to grow in the dry season because they require relatively high humidity [30].These contradictory findings may be caused by climate change which makes the dry season not completely dry, but rainfall still occurs at the study site even though with low intensity (Figure 1).The monthly rainfall in September 2021 was 72 mm.According to the climate classification by Schmidt and Ferguson [40], the rainfall of 72 mm per month falls into the category of humid enough to support the growth of the saprophyte macrofungi group with soft and easily crushed fruiting bodies.Sutjaritvorakul, et al. [39] revealed that macrofungal species that grow during the dry season or throughout the year generally come from the family of Ganodermataceae and Polyporaceae.The characteristics of hard or rigid fruiting bodies and their large size are the reasons why species of these families can grow across seasons [41].The hard fruiting bodies have denser hyphal systems and thicker cell walls so as to prevent excessive evaporation [42,43].
The variation in the diversity of macrofungal species among forest communities can be related to differences in soil moisture, light intensity, and temperature.Hu et al. [16] and Santos-Silva et al. [44] revealed that the combination of canopy cover and soil moisture is the main factor in supporting the growth of macrofungi in forests.Thick and large canopy cover will keep the soil moist due to the lack of intensity of sunlight penetrating the forest floor [45].In addition, the type of bedrock and soil may be related to water holding capacity and this in turn determines soil moisture.
Data from this study indicate that in the wet season, soil moisture was significantly the highest in the Karst forest, the second highest was in the Lowland forest, and the lowest was in the Pine forest.Meanwhile, in the dry season the highest soil moisture was observed in the Lowland forest, followed by the Karst forest and the lowest was also in the Pine forest.Rendzina soil with a clay texture on porous limestone bedrock in Karst forest is likely to be able to hold more water during the wet season compared to Cambisol soil with dusty clay mixed with small, medium and large stones in the Lowland forest.However, during the dry season, denser canopy cover in the Lowland forest, characterized by lower light intensity, can better retain moisture under the forest stand, causing soil moisture in the Lowland forest to be higher than Karst forest.Low soil moisture in Pine forests compared to Karst forests and Lowland forests can be caused by the needle cover of Pine forests which allows a lot of sunlight to reach the forest soil surface [46,47], as well as caused by the higher transpiration rate of Pine forest stands compared to broadleaf forest stand [48].In general, data on variations in inter-forest microhabitat conditions were consistent with macrofungal species' inter-forest diversity.More species of macrofungi were found in forests with higher soil moisture, lower light intensity, and higher soil pH.
Macrofungi cover area in the wet season was largest in the Pine forest, followed by the Karst forest, and lowest in the Lowland forest Meanwhile in the dry season, the cover area of macrofungi showed the largest value in the Karst forest, followed by Lowland forest and the lowest was in Pine forest.Trametes ochracea, Elmerina sp. and Coltricia cinnamomea were macrofungal species with the largest cover area respectively in Karst, Lowland, and Pine forests during the wet season.Meanwhile, in the dry season, Inocybe rimosa, Microporus xanthopus, and Russula Fragilis were the species with the largest cover area respectively in Karst, Lowland, and Pine forests.
The distinct difference in the cover area of macrofungi between forest communities, either in the wet or the dry season, is apparently related to the cover area (colony size) of species unique to certain forest communities in a certain season.For example, two macrofungal species (Coltricia cinnamomea and Ganoderma sp.5) that were only found in Pine forest in the wet season have large colony size covering up to 85878.50 cm 2 or 94% of the total cover of all macrofungal species in this forest community.While in the dry season, the existence of Polyporus sp, Favolus sp., and Inocybe rimosa with large colony size (82332.25 cm 2 or 88% of the total cover area of macrofungi in this forest) only in Karst forest made the total cover area of macrofungi in this forest became the largest.Thus, the diversity of macrofungal species must not always be in line with the cover area since each species performs a different size of fruiting body and colony size.Most of the macrofungal species found in this study were saprophytic and growing on litter or soil substrate, while some species were ectomycorrhiza fungi (Table 3).Litter was the substrat most supported the growth of macrofungal species (81 species), followed by soil (46 species), weathered wood (25 species), and living trees (17 species).One species was found to grow on cow dung.As to the substrate on which they grow, saprotrophic fungi are generally monophagous, using only the litter of a single tree species as a substrate [38].Only a few species can grow on different substrate types.This is related to the differences in carbon and nutrient requirements for each macrofungal species, which are determined by the chemical properties of the substrate, such as the ratio of lignin, hemicellulose, cellulose, and pH [49].Therefore, differences in the diversity of macrofungal species among forest communities are inseparable from the type of forest community.This is because different forest communities are composed of and dominated by different tree species.Shay [50] further stated that the diversity of macrofungi has a strong correlation with the diversity of tree species that compose forest stands, especially in tropical habitats.
Several species of macrofungi were found to be specialists only in a certain forest community (Table 3).Differences in the successional stage and environmental conditions may cause structure and species composition differences among forest communities.This enables one tree species to grow in a certain forest community but not in other forest communities.Given that most fungi are monophagous, it is not surprising that there are some macrofungal species that are only found in certain forest communities, along with their host plants that are only found in that forest community.
When compared between broadleaf forests and pine forests, the data from this study show that the diversity of macrofungal species in the broadleaf forests (Karst and Lowland forests) is distinctly higher compared to Pine forest.This appears to be due to the higher diversity of three species in broadleaf (Table 1).In addition, microclimatic conditions such as high soil moisture, lower light intensity, and higher soil pH in the broadleaf forest are more favorable for the growth of many macrofungal species than those in Pine forests.The acidic pH will affect the quality of carbon needed by macrofungi to grow [51,52].Rousk et al. [53] revealed that the soil pH gradient often correlates with the vegetation structure, which will indirectly form the colonization of different fungal species.

Conclusion
In total, we found 130 species of macrofungi in the three forest communities identified in 38 families.Seasonally, more species were found during the wet season in all forest communities, and some species were found to be adaptable to both seasons.Among the forest communities, more species were found in broadleaf forests than in Pine forests.This study reveals that species composition and structure of forest influence the microhabitat (soil moisture, soil pH, and light intensity) conditions in each forest community, which in turn cause the difference in the diversity of macrofungal species across the tree forest communities.

Figure 1 .
Figure 1.Monthly total rainfall during the study period (Source: Indonesian Agency for Meteorology, Climatology, and Geophysics in Maros City).

Figure 2 .
Figure 2. The number of species of each macrofungal family found in the three forest communities.

Table 3 .
Number of species and cover area of macrofungi in the wet and dry seasons (Cont.)

Table 1 .
. Pinus merkusii dominates 88% of the total basal area of the Pine forest.1230 (2023) 012059 Description of the research location.

Table 2 .
Microhabitat conditions in the three forest communities.
jDifferent lowercase letters after the mean values of soil moisture, soil pH, and light intensity in a row indicate significant differences between forest communities during the wet season (a, b, c for soil moisture, e, f for soil pH, and i, j for light intensity), while different uppercase letters after the mean values of soil moisture and soil pH in a row indicate significant differences between forest communities during the dry season (A, B, C for soil moisture and E, F, G for soil pH)

Table 3 .
Number of species and cover area of macrofungi in the wet and dry seasons

Table 3 .
Number of species and cover area of macrofungi in the wet and dry seasons (Cont.)

Table 3 .
Number of species and cover area of macrofungi in the wet and dry seasons (Cont.)

Table 3 .
Number of species and cover area of macrofungi in the wet and dry seasons (Cont.)

Table 3 .
Number of species and cover area of macrofungi in the wet and dry seasons (Cont.)