A comprehensive but practical methodology for selecting biological indicators for long-term monitoring.

The selection of the many biological indicators described in scientific literature is rarely based on systematic or clear-cut processes, and often takes into account only a single or very few taxa, or even disregards the complex interactions that exist between the components of biodiversity. In certain cases, the particular context of a site-for example in the Mediterranean Basin-makes it difficult to apply the choice of indicators to other regions proposed in the literature. Therefore, the selection of appropriate methodologies for generating relevant indicators for a particular site is of crucial importance. Here, we present a simple quantitative methodology capable of incorporating multidisciplinary information for assessing and selecting appropriate methods and indicators for monitoring local biodiversity. The methodology combines several ecological levels (species, habitats, processes, and ecosystem disturbances), and embraces biological interactions and common functional guilds (detritivores, producers, herbivores, and carnivores). We followed an iterative selection procedure consisting of five phases: 1) collection focal area useful information; 2) classification of this information into interrelated datasets; 3) assessment and selection of the relevant components using a quantitative relevance index; 4) the adding of taxonomic, physiognomic and functional similarities to the relevant components; and 5) the quantitative selection of the priority indicators in the study area. To demonstrate the potential of this methodology, we took as a case study the biodiversity components and their ecological interactions present in a protected area. We show that our methodology can help select appropriate local and long-term indicators, reduce the number of components required for thorough biodiversity monitoring, and underline the importance of ecological processes.

1. Introduction

Ecosystems have many different biological components, a fact that hinders any attempt to attain knowledge of the entirety of the elements that constitute our natural biodiversity [1]. At the ecological level, the elements of biodiversity are organized into complex networks that interact with biotic and abiotic components in ecosystem processes and produce ecosystem services for use by human societies [2]. Therefore, understanding the causes and consequences of the current loss of biological components is fundamental [3, 4]. The availability of useful information regarding the state and trends operating in our biodiversity is a crucial step in the construction of biological indicators [5]. These indicators interact with the ecosystem and reflect the changes occurring in a habitat, community, or ecosystem; they provide information about complex ecological processes, act as early warning signals, help diagnose the cause of ecological problems, and are important tools for use in conservation planning and management [6-10].

Understanding the trends and drivers of biodiversity change is vital when attempting to decide on appropriate conservation measures [11, 12]; nevertheless, to do so requires robust and comprehensive information obtained from biodiversity monitoring programs [13]. Despite the enormous challenges facing global biodiversity conservation [14], it is essential to quantify and predict local and regional variations to be able to address and protect all aspects of biodiversity. The Group on Earth Observations Biodiversity Observation Network (GEO BON) has developed the concept of Essential Biodiversity Variables (EBVs) [15], which can be used to link local and global needs and aims [16]. Conceptually, EBVs are located between primary data observation and indicators [15, 17], and have been developed to help prioritize a minimum set of essential measures for the consistent study, reporting, and management of all the major elements of biodiversity change. GEO BON has proposed 22 candidate variables belonging to six EBV classes: genetic composition, species populations, species traits, community composition, ecosystem functioning, and ecosystem structure [16]. Some EBVs such as population abundance–a continuous variable for all taxa–may be difficult to obtain due to the large number of taxa found at a single site. Thus, it may be more advantageous in a particular region to select just a few local biodiversity components as candidates for monitoring and it follows that standardised local sampling at fine resolutions and the datasets it generates will become a necessary and useful part of the development of global biodiversity indicators [14]. These specific monitoring systems will provide accurate information for creating local indicators for decision-making that, at the same time, can be incorporated into global EBVs [16].

Despite the extensive scientific literature that exists on the selection of indicators, this process is often neither systematic nor methodical [1, 18–22]. The selection criteria for indicators may be related to the distribution, abundance, richness, functional importance, or sensitivity of taxa to environmental change [15, 20, 23–28]. However, the choice of indicators is usually based on previously cited research, the conservation status of taxa, and/or the ease with which data can be sampled, sorted, and identified [29]. Furthermore, choices may even be based on subjective criteria unrelated to ecological criteria [20, 30, 31] or be driven by the availability of data [32]. Consequently, given the huge number of taxa that meet these requirements [33], a multitude of indicators have been described in the literature [29]; thus, when selecting appropriate indicators their relation to the local context and ecosystems must be taken into account. Accordingly, the knowledge of experts or specialists in local taxa is essential since one aspect of biodiversity (species, habitats, ecological processes, and biotic, abiotic, and anthropic problems) may affect a focal region differently and so require its indicator [6]. It is also important to assess which indicators are valid and informative for a region and which are redundant, overvalued, or unnecessary.

The Mediterranean Basin is a biodiversity hotspot that, due to the unique ecological processes and heterogeneous climatic conditions that drive its ecosystems, harbours numerous endemic plant and animal species [34-37]. Moreover, this region is experiencing a multitude of environmental impacts related to the great anthropic presence (e.g. urbanisation, infrastructures, resource overexploitation, or frequent wildfires) that complicate the study, monitoring, and biodiversity predictions of its vast number of multidirectional ecological relationships. Hence, a suitable selection of indicators is as essential as is the correct design of monitoring protocols with specific objectives. In Mediterranean ecosystems, objective methods for selecting an appropriate set of biodiversity indicators are lacking, or are limited to just one or a few groups [38, 39], and assessments often cannot be contrasted or are difficult to put into practice [40]. In other regions protocols and selection methods already exist and these methodologies generate a large variety of context-specific indicators [20, 28, 41–46].

Our overall aim was thus to develop a simple quantitative methodology capable of incorporating multidisciplinary information for assessing and selecting appropriate methods and indicators for monitoring local biodiversity. This methodology combines several ecological levels (species, habitats, processes, and ecosystem disturbances [1]), and embraces biological interactions and common functional guilds (producers, herbivores, carnivores, and detritivores [47]). We applied our methodology to Sant Llorenç del Munt i l’Obac Natural Park (NE Spain), a protected area possessing an important and representative range of Mediterranean ecosystems. Despite focussing on Mediterranean areas, our approach is relevant to other regions, landscapes and ecosystems in which there are similar challenges to biodiversity conservation.

2. Materials and methods

2.1. Methodology approach

We developed a multi-criteria [48] methodology to select relevant biodiversity components (species, habitats, ecological processes, and ecosystem disturbances), and then used these components to create biodiversity indicators for monitoring biodiversity in particular sites. We followed a hierarchical selection procedure (1) so that the resulting indicators combining trends in diversity, reproductive success, and growth rates would be effective in detecting ecological changes and useful for assessing management impacts [29, 49, 50]. The process of selection of priority indicators to implement in long-term monitoring biodiversity following five steps: 1) the collection of published and unpublished information on the species, habitats, and biological communities present in the action scope; (2) the classification of the components of biodiversity in candidates of species, habitats, and ecosystem disturbances; (3) the establishment of the relevant components of species, habitats, ecological processes, and ecosystem disturbances employing the assessment with an Index of Relevance focusing on ecological networks; (4) the creation of a Monitoring Catalogue of relevant components grouped by similarity; and (5) the establishment of the Priority indicators by a quantitative Priority index.

Fig 1

The work schedule for select priority indicators.

The work schedule carried out for the selection of Priority indicators and its long-term monitoring program respects the following states: (1) search of published or unpublished information on the species, habitats, and biological communities present in the action scope; (2) classification of the components of biodiversity in candidates of species, habitats, and ecosystem disturbances; (3) establishment of and Relevant components catalogue of species, habitats, ecological processes and ecosystem disturbances employing the assessment with an Index of relevance, and make an ecological network to determine the Relevant components of the local ecological process; (4) creation of a Monitoring Catalogue of candidates; and (5) establishment of the Priority indicators by a quantitative Priority index.

2.1.1. State 1: Collection of available information

The primary components of the biodiversity (i.e. species and habitats) cited in the focal area were identified and collated. Subsequently, information regarding the stressors and drivers–i.e. the specific functional, compositional, and structural components or ecosystem disturbances such as natural and anthropogenic stressors [51]–of these biodiversity components was compiled (Fig 1).

2.1.2. State 2: Classification of the components of biodiversity in datasets of candidates of species, habitats, and ecosystem disturbances

Based on a literature review, the collected information was classified into different datasets (Fig 1). To ensure a precise and rigorous selection, we divided the initial components of biodiversity into three ecological levels: species, habitats, and ecosystem disturbances. These datasets incorporate different variables affecting the stressors and drivers of the elements used to assess and select the relevant components. An effective collection of prior information on the area to be applied, especially not published reports, is essential. If relevant prior information is not available, you need to back to State 1 to search for or generate relevant information.

2.1.3. State 3: The establishment of the relevant components of monitoring biodiversity

2.1.3.1. Quantitative method. To select the relevant components from the datasets we developed a quantitative Relevance Index focusing on different variables specified and related to species, habitats, and ecological processes (Table 1), that in turn each one was subdivided into different levels and criteria (Table 2).

Table 1

The quantitative scoring ranges and the relevance index formula used in our multi-criteria analysis for the relevant candidates of species, habitats, and ecosystem disturbances.

	Species	Habitats	Ecosystem disturbances
a) Degree of threat	0 to 3	0 to 2
b) Ecological interest	0 to 3	0 to 2
c) Representativeness		0 to 2	0 to 2
d) Habitats			0 to 2
e) Expert or specialist criterion	0 to 2	0 to 2	0 to 2
Relevance index	a + b + e	a + b + c + e	c + d + e
	= max. 8	= max. 8	= max. 6

Table 2

Summary of the assessment parameters used to select the relevant candidates, the ecological level (species, habitat, or ecosystem disturbances), the valuation for the calculation of the relevance index, the parameter’s description, and the specific criterion to assess each of the parameters.

Assessment parameters	Level	Value	Description	Criterion
Degree of threat	Species and habitats	0 to 1	Absence/presence on local plans or reports.	A value of 1 is assigned when the species or taxon appeared in one or more strategic reports or management plans at the local scale.
	Species	0 to 1	Absence/presence on current legislation.	When the species appears in the current legislation or annexe thereof, at the level of the autonomous community, country, or European community, it receives a value of 1.
	Species	0 to 1	Cataloguing on the IUCN Red List.	When the species is catalogued (vulnerable, endangered, or critically endangered) for the IUCN Red List at the country level, it is assigned a value of 1.
	Habitat	0 to 1	Cataloguing on European Directive	When the habitat is a priority or of interest for the European directive, it receives a value of 1.
Ecological interest	Species	0 to 2 (if fulfil a criterion = 1; if accomplish more than one = 2)	Habitat specialization	Species representative of a particular habitat, especially of rare habitats.
			Geographic distribution	Species with a disjoint geographical distribution (broad geographical separation between populations)
			Effects of climate	Species with their distribution boundary in the area of study and, therefore, the variations in the climate can affect it.
			Ecological process	Species relevant in some ecological processes (pollination, herbivorism, production of trophic resources, predator-prey relationship, seed dispersion, parasitism, etc.)
			Ecosystem disturbances	Species considered ecological indicators of environmental quality, water quality, unsustainable management, being affected by forest pests, being a hunting object, invasive/ allochthon species, abundant in undisturbed areas, affected by human frequentation, etc.
			Other aspects	Other relevant aspects of ecological interest such as the specialized diet, rarity or symbolism of the species, short-term population trends, etc.
		0 to 1	Absence/presence on monitoring programs	If it fulfils, the species obtain a value of 1 in this section as they correspond to common species (commonly detected species in monitoring plans).
	Habitat	0 to 2 (if fulfil a criterion = 1; if accomplish more than one = 2)	Abundance	Habitats with a considerable extension in the area of study.
			Singularity	Rare, regressive, or poorly represented habitat in the area of study.
			Ecological processes	Habitats in which important ecological processes occur for the global functioning of ecosystems, e.g. production of trophic resources, herbivorism, etc.
			Ecosystem disturbances	Habitats that are more susceptible to suffer relevant ecosystem disturbances, such as forest exploitation, afforestation, human frequentation, wildfires, etc.
			Other aspects	Other important or relevant characteristics of the habitats.
Representativeness	Habitat and Ecosystem disturbances	0 to 2	Number of affected species	Those habitats with a greater number of species or are affected by the ecosystem disturbances, have a higher value. The total number of species is relocated over a value of 2
Habitats	Ecosystem disturbances	0 to 2	Number of affected habitats	Ecosystem disturbances that affect a greater number of habitats present a higher value. The total number of habitats is relocated over a value of 2.
Expert or specialist criterion	Species, Habitat and Ecosystem disturbances	0 to 2	Local value awarded by scientific experts	The criterion of external expert or taxon specialist grants a value of 2 to the species, habitats, and/or ecosystem disturbances with high importance and relevance in the context of the study area, and 1 to species, habitats, and/or ecosystem disturbances with relative importance in the study area.

For species, the quantitative Relevance Index value lies in the range 0 to 8 (Table 1) and selects all values with a threshold of ≥ 4. The species’ assessment parameters are related to the degree of threat [52], their ecological interest [53, 54], and the expert criterion [6] for each taxonomic group (Table 2). The ecological interest parameter gives priority to species that occupy a wide-ranging habitat, have rich habitat requirements (patch size, structure, and configuration), and depend on particular ecological processes (Table 2), given that species restricted to fewer habitat types are more susceptible to local impacts [55].

At habitat level, the quantitative Relevance Index value is established from 0 to 8 (Table 1) and selects values with a threshold of ≥ 4. For each habitat catalogued, the parameters are the degree of threat, ecological interest, the representativeness of preselected species that depend on each one of the assessed habitats, and the expert criterion (Table 2).

In the case of ecosystem disturbances, the Relevance Index runs from 0 to 6 (Table 1), and, once assessed, values with a threshold of ≥ 3 are selected. In this case, the parameters used were related to the species and habitats potentially affected by each ecosystem disturbances, and to expert criteria (Table 2).

Once the species, habitats, and ecosystem disturbances have been assessed, those that reach the threshold value of the Relevance Index are selected. Because the threshold value needs to be sufficiently robust but at the same time to include potential under-detected components, we established as a criterion to maintain the components with a Relevance index equal to or above half of the maximum global potential value that included valuable ecosystem components, as we have verified in our previous tests (see Table 1).

For the component assessment for species, habitats, and ecosystem disturbances accurate knowledge of ecological requirements and responses to environmental changes in the candidates is required when choosing the elements that reflect our ultimate aim. Here, the bibliography and expert criteria are essential (Table 2), and will help highlight species or habitats that are not present or not evaluated as threatened. The ecological interest contribute to minimize the weight of these threatened and singular component, which do not necessarily reflect the spatial and temporal trends of biodiversity [23, 56] but, rather, respond to the local conservation status [57].

2.1.3.2. Construct an interrelated diagram to determine the ecological processes. Ecological processes were qualitatively assessed strictly on preselected relevant candidates (species, habitats, and ecosystem disturbances) and their diversity of interactions in the focal area, i.e. the diversity and structure of multi-trophic interactions between organisms and habitat types. These processes can be complex to define and elucidate and their monitoring is often difficult to perform. Thus, basic processes such as trophic relationships and mutualism interactions are required to select the processes [46]. Here, based on the previously relevant candidates, a diagram is established that depicts all possible direct and indirect trophic relationships, as well as the interactions between species, habitats, and external ecosystem disturbances (abiotic and anthropic). From here, the main ecological processes in the study area are defined as the water cycle, nutrient cycle, ecological succession, trophic networks, mutualisms, and other local relevant processes (Fig 2). This exercise forces to define the biological and geographical limits (monitor scale) and, then, make practical decisions about how much can be done. In addition, it emphasises that to monitor any ecosystem you must have a great deal of background data [46].

Fig 2

Interactions between the monitoring candidates to elucidate ecological processes.

Diagram of the possible interactions that are established between the monitoring candidates, where highlight five main groups of organisms in the trophic network, the detritivores (green box), the primary producers, the primary and secondary consumers, and the predators. However, within these groups, interactions occur, even within the same species communities. On the other hand, this entire complex trophic network is conditioned by abiotic and anthropogenic external ecosystem disturbances (ecosystem disturbances; red box), such as climatology, perturbations, pollution, forest management, etc. Continuous lines denote direct relationships, while discontinuous lines describe diffused relationships often characteristic of opportunistic species.

2.1.4. State 4: Monitoring catalogue

Once the different relevant components have been assessed and selected, a Monitoring Catalogue (Fig 1) can be established to act as a strategic guide, facilitate understanding, and identify which variables need to be sampled and measured to generate the biological indicators in an ecological sense and to avoid redundancies. To attain the list of relevant components, (1) we grouped components by taxonomic, physiognomic, and functional similarities (e.g. common birds, freshwater invertebrates, decomposers). Then, (2) classified the grouped relevant components in four ecological levels: species, habitat, ecological process, and ecosystem disturbances (see the case study below as a practical example). (3) For each group of relevant components we selected the adequate variables to be sampled, e.g., species presence and abundance, reproductive and survival taxes, habitat preferences, soil structure, and composition. Lastly, (4) incorporated into each level all the monitoring components whilst bearing in mind that each can be part of one or more levels depending on their role and interactions within the ecosystem (see State 3). For example, species abundance and richness of butterflies provide information about the species level and their community respectively, so in the latter case indicate the variation on ecosystem functional capabilities (e.g. as primary consumers), climate change or afforestation effects.

2.1.5. State 5: Priority indicators

Priority Indicators are useful for acquiring knowledge of biological and ecological changes in a given area and indispensable to optimize the efforts in monitoring. In our scheme, they were selected using a quantitative Priority Index, which awards importance from 0 to 5 based on the criteria of specificity (key species or habitats, etc.), generality (abundant species, representative and common processes, etc.), importance (endemic species, determining ecosystem disturbances, etc.), interactions between organisms, and usefulness for management and decision-making (Table 3). In this case, only those Monitoring Components that reach maximum importance (Priority Index ≥ 4) are chosen as Priority Indicators in the study area.

Table 3

Summary of the assessment parameters used to select the priority indicators by a quantitative priority index using five parameters with its specific criterion.

Parameter	Value	Criterion
Specificity	0 to 1	A value of 1 is assigned when the indicator allows capturing the tendencies and dynamics of the species, communities, or habitats and relevant ecological processes or ecosystem disturbances, in the study area.
Generality	0 to 1	A value of 1 is assigned when the indicator is represented by abundant species or communities, representative habitats, common ecological processes, or relevant and extensive ecosystem disturbances, in the study area.
Importance	0 to 1	A value of 1 is assigned when the indicator involves the presence of singular species, unique habitats, key ecological processes, determining ecosystem disturbances, etc.
Interactions between organisms	0 to 1	A value of 1 is assigned when the indicator involves different organisms, communities, key ecological processes, are altered by relevant ecosystem disturbances, etc.
Usefulness for management and decision making	0 to 1	A value of 1 is assigned when the indicator involves components that can provide useful information for the management or conservation.

Priority Indicators were divided into four interrelated ecological levels:

Species. The indicators of the state of species, in general, are designed to identify the population and conservation status of the most characteristic species in ecosystems, whose presence or abundance indicates population trends and the state of the habitats and ecosystems in which they live.

Habitats. The indicators of the state of habitats are based on knowledge of the condition of plant communities (floristically and functionally) and land use, and, in particular, of the evolution of their structural, spatial, and temporal evolution and distribution.

Ecological processes. The indicators highlight the relationships that link organisms or groups of organisms with each other and with the environment (habitat or ecosystem) that hosts them and the specific interactions between them.

Ecosystem disturbances. These indicators reveal the effects of the ecosystem disturbances that affect and alter the natural functioning of ecosystems and their constituent organisms. Disturbances in ecosystems may be the product of natural cycles or anthropogenic activity.

2.2. Pilot study area

To apply our quantitative and qualitative methodology for selecting biodiversity indicators at the local scale, Sant Llorenç del Munt i l’Obac Natural Park (henceforth PNSLL; Fig 3) was chosen as the focal region. This mountainous area forms part of the Catalan Prelitoral Mountain Range and has an altitudinal range of 280–1,100 m a.s.l. and a typical mid-altitude Mediterranean montane climate. Its orography and geographical situation afford it great climatic variability, with an annual mean rainfall of 500–800 mm and a mean annual temperature of 15 C°. Its lithology consists of permeable conglomerates with an argillaceous and calcareous matrix [58]. It is characterized by rocky outcrops and cliffs [59], whose forested crags and ridges are covered chiefly by holm oak (Quercus ilex L.) forests and, in the more humid valleys, patches of deciduous pubescent (Quercus pubescens Willd.) and sessile (Quercus petraea (Matt.) Liebl) oak forests. Lower areas are dominated by Aleppo pine (Pinus halepensis Mill.) woodland with an evergreen oak understory [60], while black (Pinus nigra subsp. salzmannii (Dunal) Franco) and Scots (Pinus sylvestris L.) pine forests cover smaller areas of terrain. Shrubland is also present mainly as a result of the wildfires that occurred during the 2000s. Currently, this 13,694-ha protected area is subject to intense human pressure (e.g. wildfires, human frequentation, and forest exploitation) since is located near the Barcelona conurbation (4 million inhabitants), one of the largest metropolitan areas in southern Europe.

Fig 3

Location of the pilot study area.

The geographic location of the Sant Lorenç del Munt i l’Obac Natural Park (white ring) in the nearby of the metropolitan area of Barcelona (black ring) in Catalonia (NE Iberian Peninsula) and its perimeter. Squares correspond to 100×100m Permanent Monitoring Plots, where COD correspond to Rocky areas habitat, MAT: Shrublands; PMD: Mediterranean pine forests, PHD: Wet pine forests; BMX: Mixed forests, AMY: Mountain holm oak forests, and BCF: Deciduous forests.

3. Results

3.1. Information collection and classification into datasets

In all, 387 literature sources were consulted (S1 Appendix), including 213 publications, 66 unpublished reports, 10 technical plans, 19 datasets, and 79 reports from monitoring programs, as well as unpublished data by experts and researchers. A total of 3,226 species were catalogued and assessed for the PNSLL as candidates as indicators, including 52 algae, 159 fungi, 159 lichens, 69 bryophytes, 27 pteridophytes, 1,081 vascular plants (100 allochthones) [61], 1,404 invertebrates (1,179 insects, 53 gastropods, 139 arachnids, 30 from other groups and 7 allochthones), and 270 vertebrates (3 freshwater fish, 13 amphibians, 21 reptiles, 174 birds, 27 raptors, 51 mammals and 12 allochthones).

3.2. Relevant components selection

The relevant components were obtained through a Relevance Index which was calculated for all 3,226 assessed species (Table 4) of which 406 (12.6%) were selected as Relevant Components (S1 Table): plants contributed with 69 species (17%—of which 10 species were threatened species, 13 pteridophytes, 41 vascular plants, and five alien species), 191 species (47%) were invertebrates (of which 26 species were Odonata, 12 Gastropoda, 43 decomposers, 22 Formicidae, 30 Orthoptera; 36 Lepidoptera, ten other invertebrates of special interest, seven plague species and five were alien species), while 146 species (36%) were vertebrates (of which tree species were freshwater fish, seven amphibians, seven reptiles, 13 raptors, 70 common birds, 21 bats, seven small-mammals, two common medium-sized preys, two ungulates, five carnivores and nine were alien species).

Table 4

Reduction in the number and percentage of candidates to relevant components through the use of the relevance index; and reduction of the number and percentage of monitoring components to priority indicators by a quantitative priority index.

	Species	Habitats	Ecological processes	Ecosystem disturbances
Candidates	3,226	96	8	14
Relevant components	408	11	6	9
Percentage of reduction	-87.4%	-89.6%	-25.0%	-35.7%
Monitoring components	17	2	5	8
Priority indicators	13	2	4	8
Percentage of reduction	-23.5%	0%	-20.0%	0%

We identified a total of 96 habitats in the PNSLL [62-65], which, to simplify the assessment and selection of habitats, were grouped according to floristic, structural, and ecological similarities, thereby generating 15 habitat categories to be assessed as candidates as indicators (S2 Table). Of the total 15 assessed habitats, eleven were selected (Table 4) as relevant components (using Relevance Index value; S3 Table): caves, rocky areas, dry meadows, shrublands, Mediterranean pine forests, humid pine forests, mixed forests, mountain holm oak forests, deciduous forests, riparian forests and freshwater habitats.

To identify the relevant ecosystem disturbances affecting Mediterranean ecosystem biodiversity and, in particular, our study area, an intense literature search was conducted. From the candidates’ list of 14 ecosystem disturbances, nine were selected as Relevant Components (Table 4), corresponding to those with a Relevance Index value ≥ 3 and including disturbances related to global change (climate change, wildfires, and alien species) and local ecosystem disturbances (afforestation, fragmentation, freshwater alteration, silvicultural activities, hunting, human frequentation, and fatalities on human infrastructures) (S4 Table).

At a global level, and using our quantitative method, from 3,344 candidates including species, habitats, and ecosystem disturbances a set of 434 Relevant Components were finally obtained. The main ecological processes operating in the study area were defined first drawing a diagram that illustrates trophic relationships (S1 Fig), as well as the interactions between species, habitats, and external ecosystem disturbances (abiotic and anthropic) resulting in decomposition, plant production, consumers and predators and mutualisms (pollination and zoochory). Other relevant ecological processes such as the water cycle or ecological succession were included in the ecosystem disturbances (e.g. climate change, alteration of aquatic ecosystems, wildfires, and afforestation; Table 5). Thus, the final number of relevant components was 434, an 87% decrease from the list of initial candidates (Table 4).

Table 5

Priority indicators (n = 27) that quantitatively meet selection criteria on Sant Llorenç el Munt i l’Obac Natural Park.

The indicators are divided into four categories and in different attributes or pressures [1], consequently, each require different variables from monitoring candidates.

Category	Attribute or pressure	Priority indicator	Variables from monitoring candidates
State of the species	Dispersal-limited and special interest species	Threatened flora	Species presence and abundance
			Flowering rate
			Occupied extension
		Freshwater fishes	Species presence and abundance
		Amphibian	Species presence and adult abundance
			Number of laying and larvae
		Chiroptera	Species presence and abundance
	Umbrella species	Raptors	Species presence and abundance
			Reproductive and survival taxes
	Link species	Decomposers	Species presence and abundance
			Diversity
		Orthoptera	Species presence and abundance
		Small-mammals	Species presence and abundance
		Common medium-size preys	Species presence and abundance
			Number of hunted individuals
	Indicator species	Butterflies	Species presence and abundance
			Habitat preferences
		Freshwater macroinvertebrates	Species presence and abundance
			Diversity
		Common birds	Species presence and abundance
	Ecological engineers	Ungulates	Species presence and abundance
			Number of hunted individuals
State of the habitats	Landscape	Structure	Habitat structure
			Regeneration
			Soil structure and composition
			Volume of necromass
		Plant composition	Community composition
			Plant distribution
State of the ecological processes	Ecosystem recourses	Primary production	Vegetal production
			Mushroom production
			Acorn production
			Pinecone production
	Trophic network	Decomposition	Abundance and diversity of detritivores
			Volume of necromass
		Consumers	Presence and abundance of orthopteran, small-mammals, common medium-size preys, common birds, and ungulates
		Predators	Presence and abundance of carnivores and raptors
			Vital taxes
			Diet
Ecosystem disturbances	Resistance and resilience	Climatic change	Rainfall
			Temperature
			Relative humidity
			Insolation
			Evapotranspiration
			Extreme episodes
		Wildfires	Presence and abundance of detritivores, ants, orthopteran, butterflies, reptiles, common birds, small-mammals and medium-size preys
			Plant composition
			Habitat structure
			Regeneration
			Soil erosion
			Trophic network
		Afforestation	Presence and abundance of ants, orthopteran, reptiles, butterflies, common birds, and medium-size preys)
			Plant composition
			Habitat structure
		Freshwater alterations	Flow level
			Contaminants and physical-chemical parameters
			Presence, abundance, and diversity of macroinvertebrates and freshwater fishes
			State of the riparian forest
	Biological invasions	Alien species	Species presence and abundance
			Distribution
	Anthropic pressure	Exploitation of natural resources (silvicultural and hunting)	Presence and abundance of detritivores, common birds, raptors, chiropters, medium-size preys, and ungulates
			Plant composition
			Habitat structure
			Soil erosion
			Trophic network
			Number of hunted individuals
		Human frequentation	Presence and abundance of threatened flora, xeric gastropods, ants, orthopteran, butterflies, and common birds.
			Reproductive taxa of rocky and cavern species
			Plant composition
			Habitat structure
			Soil erosion
			Trophic networks
			Number of visitors
		Human infrastructures	Roadkill taxa
			Risk and taxa of electrocution
			Risk and taxa of collision with power lines
			Fragmentation of river connectivity
			Alteration of fluvial flows

3.3. Monitoring catalogue and priority indicators

To constitute the Monitoring catalogue, we grouped the 434 Relevant components into independent levels of species, habitats, ecological processes, and ecosystem disturbances by similarities. In the case of 408 species were grouped by taxonomic, physiognomic, and functional similarities into 17 groups of species (threatened plant species, xeric gastropods, granivorous ants, decomposers, orthoptera, butterflies, freshwater macroinvertebrates, freshwater fishes, amphibians, reptiles, common birds, raptors, bats, small mammals, common medium-size preys, ungulates and carnivores). A total of 96 habitats were grouped in 11 habitat relevant categories (Table 4). Then, the habitat category resulted in two monitoring parameters (habitat structure and composition), which will be carried out in the 11 selected relevant habitats. Moreover, it is in these habitats where the specific transversal monitoring of species, processes and disturbances will be developed. Ecological processes were grouped by ecological functions in 6, decomposition, primary production, consumers, predators, zoocory, and pollination. Ecosystem disturbances were maintained in 8 (Table 4), climatic change, wildfires, afforestation, freshwater alterations, alien species, exploitation of natural resources, forests pests, human frequentation, human infrastructures). So resulted in a total of 32 specific Monitoring Components. Finally, to obtain the Priority Indicators we used the quantitative Priority Index, and 27 of these Monitoring Components were selected to monitor the biodiversity in the PNSLL (Table 5).

4. Discussion

Our methodology was established using a systematic and transparent conceptual framework. First, we divided our set of components into four interrelated ecological levels: species, habitats, ecological processes, and ecosystem disturbances. Species and habitat indicators provide information about specific states but other ecological levels are required if they are to be correctly interpreted since, for example, different bird species respond to specific habitat structure, the availability of resources, and environmental conditions [24, 38, 66]. Therefore, a single indicator cannot reveal ecosystem dynamics on its own, as all indicators are interlinked and their impacts are connected via a web of complex relationships (Fig 2).

In the components selection process quantitative criteria were given priority over more subjective qualitative criteria, and a multi-criteria analysis [48] was used to generate a set of candidates and assess these indicators in terms of their degrees of importance to the ecosystem. The evaluation of each species, habitat, and ecosystem disturbance based on a points score using specific multi-criteria analysis allowed us to reduce the initial candidates to the final selected Priority Indicators (see State 3 in Methods). Exceptionally, we used qualitative selection in the particular case of ecological processes (see Discussion below). The use of a quantitative selection of indices also allowed us to drastically reduce the number of species and habitats to be monitored, which will thus optimise both future efforts and the use of resources. Some possible biases could arise from the exclusive use of the weight values in the Relevance Index as some potential candidates (e.g. plants and invertebrate species) would be undervalued so expert criteria could be needed and this knowledge will reinforce the selection of candidates prioritized. It is a key of the scheme and process of selection to have a good information base beforehand (literature, reports and databases). The higher and better the quality of the information, the more rigorous the selection of components will be. Experts and specialists examined each biodiversity component and judged their importance in the focal region to minimise the risk of benefiting certain charismatic taxa or a predetermined species as an indicator. Expert criteria and ecological importance require a thorough knowledge of species, habitats, ecological processes, and ecosystem disturbances, which is vital when selecting the appropriate relevant candidates for each focal region in which this methodology is to be implemented. This also allowed us to minimise the tendency to select rare species [67, 68] and helped us obtain more site-specific candidates [69]. The selection of the relevant candidates (species and habitats) to be included in a monitoring catalogue and their subsequent inclusion amongst the different priority indicators led to a third assessment of indicators via a multi-criteria analysis based on their specificity, generality, relevance, interactions with other organisms, and usefulness in management and decision-making.

Nevertheless, we placed special emphasis on the indicator selection for trophic networks and used a qualitative method strictly based on preselected indicators (species, habitats, and ecosystem disturbances) and their interrelations in the study area. All these interactions and relationships are depicted in a diagram (see State 3 of Methods, Fig 1), which provides information about the number of directly and indirectly related indicators. The integration and assessment of trophic networks–from primary production to predators–is a key component in the elaboration, selection, and monitoring of indicators [70, 71]. At the same time, an understanding of trophic networks is key for predicting trends and anticipating the conservationist or adaptive measures required in ecosystems [72]. Our method enables us to combine local results with the exploration of questions at broader geographical scales [73, 74].

This practical methodology for selecting indicators can be applied simply by conservation managers and nature stakeholders in local areas everywhere and networks such as the Barcelona Provincial Council’s Network of Natural Parks (https://parcs.diba.cat/). Nevertheless, it is also useful as a management tool since it allows for better-informed and more cost-effective decision-making in a particular site [29, 49, 50]. Therefore, besides providing information about the current situation of ecosystems, the selected indicators can be used as decision criteria and as early warning signals of change in a specific region. This is especially relevant and necessary in a constantly changing world affected by natural and anthropic impacts [75]. It also allows us to formulate and implement biodiversity conservation strategies in changing landscapes through the use of a comprehensive and structured information system. Finally, these indicators also offer metrics for ecosystem status and provide interpretable information regarding changes [76].

4.1. Practical considerations before implementing indicators

After the selection of the definitive indicators, the following logical factors should be considered before implementing any monitoring program at the local scale: (1) an appropriate selection of monitoring points with which to monitor the relevant candidates; (2) the generation of user-friendly protocols; and (3) an assessment of the ability of the protocols to obtain the required information. Indicator monitoring must be planned in the long term since complex dynamics and relationships in ecosystems may potentially affect decision quality. Long-term data can tackle questions not easily addressed in the short term [74] and are an excellent means of understanding how ecosystem disturbances impact ecosystem functioning and species dynamics. For example, Failing and Gregory [77] defend prescribed burns and replicating natural disturbance regimes to achieve long-term sustainable levels of biodiversity. Krebs [46] also highlights the need to have monitoring programs that report continuous information and data, having hundreds of years as time frame, and remarks on the importance of maintaining extended discussion of the monitoring problem, what should be monitored, and what the costs will be.

Monitoring scales should also be defined according to the sampling needs of each of the biological indicators. If a parameter cannot be adequately sampled, its usefulness for monitoring is greatly reduced [50]. Here three different scales are proposed: point-scale, plot-scale, and macro-scale (or landscape). If it is to be statistically robust, the monitoring scale has to offer the possibility of replicas, which do not suppose huge efforts, which can be used in the subsequent comparison.

Point-scale monitoring is intended for site-specific indicators that require very accurate, specific, and concrete monitoring due to the scarcity or specificity of the habitat they occupy (e.g. threatened and rare flora), or to their linear distribution in space (e.g. riparian communities).

Plot-scale monitoring within a homogeneous habitat with continuity at a fine-scale reveals the variations and trends occurring in different indicators. The monitoring of the selected indicators in the same plot will thus establish interactions according to each habitat (e.g. small-mammals, birds, habitat structure, trophic networks, decomposition, etc.). To obtain robust results, a minimum of four 100×100m Permanent Monitoring Plots are required in each selected habitat.

Macro-scale (or landscape) monitoring corresponds to cases in which indicators require wide-ranging monitoring (a whole protected area, a mountain range, a slope, a cliff, etc.) given the high mobility, ubiquity, or impact of the indicators (e.g. afforestation, predators, etc.). This type of monitoring will follow protocols and dimensions dependent on the specific indicator to be followed; however, they have in common the fact that they will all cover considerable parts of the region.

User-friendly monitoring protocols should be standardised and adapted to the selected indicators in consensus with experts and bearing in mind the initiatives and monitoring processes that are already being carried out at local, national, and international levels. Indicators can form part of more than one level of monitoring, and some may require more than one protocol since certain species, habitat components, and ecological processes necessarily demand specific methodologies.

Data sources.

(DOCX)

Click here for additional data file.

List and selection values of species.

List of the Individual species indicators of the Natural Park of Sant Llorenç del Munt i l’Obac following the criterion of achieving a value of the Relevance index. The list includes the taxon community or aggrupation (bold font), the accepted name of the species, the Relevance index for each species, the average for the community or aggrupation (bold font), and the four subsections to extract this index: the degree of threat (scoring ranges from 0 to 3), the monitoring programs set (0 to 1), the ecological interest (0 to 2) and the expert or taxon specialist criterion (0 to 2).

(DOCX)

Click here for additional data file.

List of aggregated habitats.

List of the 15 aggregated habitats (by similarity and type of plant formation) that collect in them the 96 habitats present in the Natural Park of Sant Llorenç del Munt i l’Obac. This list shows, the name of the aggregated habitat, the CORINE codes that are grouped in each aggregated habitat, and the description of the aggregated habitat.

(DOCX)

Click here for additional data file.

Selection values of habitats.

List of the valued habitats (aggregation by similarity and type of plant formation) of the Natural Park of Sant Llorenç del Munt i l’Obac following the criterion of achieving a value of the Relevance index greater than 4 points on a total of 8. The Relevance index of each selected grouping habitat corresponds to the average of the Relevance indexes of the different habitats that make up the group according to the selection criteria (relevance ≥ 4), being the sum of the four subsections to extract this index: degree of threat (scoring ranges from 0 to 2), the ecological interest (0 to 2), representativeness (0 to 2) and the expert or specialist criterion (0 to 2). Representativeness corresponds to the number of selected species that appear in the grouping habitat relativized on 2, being 380 the maximum of species assessed.

(DOCX)

Click here for additional data file.

List and selection values of ecosystem disturbances.

List of the valued ecosystem disturbances on Natural Park of Sant Llorenç del Munt i l’Obac following the criterion of achieving a value of the Relevance index greater than 2.5 points on a total of 5 (in bold). The list includes the ecosystem disturbances, the relevance index, and the three subsections to extract this index: the representativeness (scoring ranges from 0 to 2), the affected habitat (0 to 2), and the expert or specialist criterion (0 to 1). Representativeness corresponds to the number of selected species affected by the ecosystem disturbances relativized on 2, being 380 the maximum of species assessed. Habitat grouping is affected by the ecosystem disturbances relative to 2, with 11 being the maximum number of habitats assessed.

(DOCX)

Click here for additional data file.

Selection of ecological processes.

The scheme used to select ecological processes in of the Natural Park of Sant Llorenç del Munt i l’Obac by a diagram of interactions between primary producers, food sources, primary consumers, secondary consumers, predators, decomposers, and abiotic and anthropogenic external ecosystem disturbances, such as climatology, perturbations, pollution, forest management, etc.

(DOCX)

Click here for additional data file.

19 Nov 2021

A rebuttal letter that responds to each point raised by the academic editor and reviewer(s). You should upload this letter as a separate file labeled 'Response to Reviewers'.

A marked-up copy of your manuscript that highlights changes made to the original version. You should upload this as a separate file labeled 'Revised Manuscript with Track Changes'.

An unmarked version of your revised paper without tracked changes. You should upload this as a separate file labeled 'Manuscript'.

If you would like to make changes to your financial disclosure, please include your updated statement in your cover letter. Guidelines for resubmitting your figure files are available below the reviewer comments at the end of this letter.

If applicable, we recommend that you deposit your laboratory protocols in protocols.io to enhance the reproducibility of your results. Protocols.io assigns your protocol its own identifier (DOI) so that it can be cited independently in the future. For instructions see: https://journals.plos.org/plosone/s/submission-guidelines#loc-laboratory-protocols. Additionally, PLOS ONE offers an option for publishing peer-reviewed Lab Protocol articles, which describe protocols hosted on protocols.io. Read more information on sharing protocols at https://plos.org/protocols?utm_medium=editorial-email&utm_source=authorletters&utm_campaign=protocols.

We look forward to receiving your revised manuscript.

Kind regards,

Ignasi Torre

Academic Editor

PLOS ONE

Journal Requirements:

When submitting your revision, we need you to address these additional requirements.

1. Please ensure that your manuscript meets PLOS ONE's style requirements, including those for file naming. The PLOS ONE style templates can be found at

https://journals.plos.org/plosone/s/file?id=wjVg/PLOSOne_formatting_sample_main_body.pdf and

https://journals.plos.org/plosone/s/file?id=ba62/PLOSOne_formatting_sample_title_authors_affiliations.pdf

2. We note that Figure 3 in your submission contain map images which may be copyrighted. All PLOS content is published under the Creative Commons Attribution License (CC BY 4.0), which means that the manuscript, images, and Supporting Information files will be freely available online, and any third party is permitted to access, download, copy, distribute, and use these materials in any way, even commercially, with proper attribution. For these reasons, we cannot publish previously copyrighted maps or satellite images created using proprietary data, such as Google software (Google Maps, Street View, and Earth). For more information, see our copyright guidelines: http://journals.plos.org/plosone/s/licenses-and-copyright.

We require you to either (1) present written permission from the copyright holder to publish these figures specifically under the CC BY 4.0 license, or (2) remove the figures from your submission:

1. You may seek permission from the original copyright holder of Figure 3 to publish the content specifically under the CC BY 4.0 license.

We recommend that you contact the original copyright holder with the Content Permission Form (http://journals.plos.org/plosone/s/file?id=7c09/content-permission-form.pdf) and the following text:

“I request permission for the open-access journal PLOS ONE to publish XXX under the Creative Commons Attribution License (CCAL) CC BY 4.0 (http://creativecommons.org/licenses/by/4.0/). Please be aware that this license allows unrestricted use and distribution, even commercially, by third parties. Please reply and provide explicit written permission to publish XXX under a CC BY license and complete the attached form.”

Please upload the completed Content Permission Form or other proof of granted permissions as an "Other" file with your submission.

In the figure caption of the copyrighted figure, please include the following text: “Reprinted from [ref] under a CC BY license, with permission from [name of publisher], original copyright [original copyright year].”

2. If you are unable to obtain permission from the original copyright holder to publish these figures under the CC BY 4.0 license or if the copyright holder’s requirements are incompatible with the CC BY 4.0 license, please either i) remove the figure or ii) supply a replacement figure that complies with the CC BY 4.0 license. Please check copyright information on all replacement figures and update the figure caption with source information. If applicable, please specify in the figure caption text when a figure is similar but not identical to the original image and is therefore for illustrative purposes only.

The following resources for replacing copyrighted map figures may be helpful:

USGS National Map Viewer (public domain): http://viewer.nationalmap.gov/viewer/

The Gateway to Astronaut Photography of Earth (public domain): http://eol.jsc.nasa.gov/sseop/clickmap/

Maps at the CIA (public domain): https://www.cia.gov/library/publications/the-world-factbook/index.html and https://www.cia.gov/library/publications/cia-maps-publications/index.html

NASA Earth Observatory (public domain): http://earthobservatory.nasa.gov/

Landsat: http://landsat.visibleearth.nasa.gov/

USGS EROS (Earth Resources Observatory and Science (EROS) Center) (public domain): http://eros.usgs.gov/#

Natural Earth (public domain): http://www.naturalearthdata.com/

[Note: HTML markup is below. Please do not edit.]

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: Yes

Reviewer #2: Yes

**********

2. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

Reviewer #2: N/A

**********

3. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

Reviewer #2: Yes

**********

4. Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #1: Yes

Reviewer #2: Yes

**********

5. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: Dear author (s),

Thanks a lot for your valuable info that you provided in your targeted study and really sounds great for all the stockholders in the fields of biological and environmental science. Only you need to rewrite the abstract to be more clear and insert your aims of study also.

Reviewer #2: In their manuscript „a comprehensive but practical methodology for selecting biological indicators for long-term monitoring” the authors present how they selected a set of indicators to monitor changes in biodiversity at different levels in a Mediterranean ecosystem. The selection of suitable indicators is an important aspect to set up an effective monitoring system. The comprehensive approach is of interest to researchers facing the same challenge. This makes the paper a timely and important contribution.

The paper is very readable. The tables and figures are informative and I found the supporting information good and well organized. In the legend of figure 3 (L290) BMX appears twice (BMX: mixed forests AND BMX deciduous forests).

The method presented in the manuscript is described by the authors as simple and practical. The steps in themselves are logical and easy to understand, however the exact execution is less clear. The authors build on a large amount of literature and prior research and their work flow includes the consultation of experts. Line 377 discusses the risk of undervaluing some components; line 392 the use of a qualitative method strictly based on preselected indicators. While the authors discussion of these points is relevant and comprehensible, they do contribute to the impression that the method is not as easy to implement as the authors initially imply. The pre-requirements and the time aspect could be discussed further, especially as the aim of the method is to make monitoring more manageable.

Some of the aspects of the methodology, specifically the step from biodiversity components to indicators, were not entirely clear to me. The methodology does not sufficiently distinguish between parameters (species, habitats, processes, factors) and the assessment methods (abundances, phenology etc.). For instance, line 233 gives butterflies as an example, but does not specify an actual parameter that is measured. On page 8 and 9, the authors set a threshold of >= 4 for their index. I was unsure how this threshold was set and of its implications for the selection of species. In Table 2, “ecological process” stood out to me as a criterion: the description could apply to all species. The methodology that allowed to group the 434 relevant components into 32 specific monitoring components requires more details.

Relevant components selection: I’m not sure about the grouping of species: while I understand that the % of alien species is of interest, they could also fit into the taxonomic groups. I think it would be more easily understandable to provide the numbers separately (classification into organism groups and a separate % for status). It might also be more easy to give species numbers rather than % - please check the numbers, as the % in the text do not always seem to match up with the number of species given in Annex 2.

**********

6. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files.

If you choose “no”, your identity will remain anonymous but your review may still be made public.

Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy.

Reviewer #1: Yes: Salwan Ali Abed

Reviewer #2: No

[NOTE: If reviewer comments were submitted as an attachment file, they will be attached to this email and accessible via the submission site. Please log into your account, locate the manuscript record, and check for the action link "View Attachments". If this link does not appear, there are no attachment files.]

While revising your submission, please upload your figure files to the Preflight Analysis and Conversion Engine (PACE) digital diagnostic tool, https://pacev2.apexcovantage.com/. PACE helps ensure that figures meet PLOS requirements. To use PACE, you must first register as a user. Registration is free. Then, login and navigate to the UPLOAD tab, where you will find detailed instructions on how to use the tool. If you encounter any issues or have any questions when using PACE, please email PLOS at figures@plos.org. Please note that Supporting Information files do not need this step.

31 Dec 2021

Reviewer #1:

Thanks a lot for your valuable info that you provided in your targeted study and really sounds great for all the stockholders in the fields of biological and environmental science. Only you need to rewrite the abstract to be more clear and insert your aims of study also.

Many thanks for the review. We have tried to clarify our abstract.

Reviewer #2:

In their manuscript „a comprehensive but practical methodology for selecting biological indicators for long-term monitoring” the authors present how they selected a set of indicators to monitor changes in biodiversity at different levels in a Mediterranean ecosystem. The selection of suitable indicators is an important aspect to set up an effective monitoring system. The comprehensive approach is of interest to researchers facing the same challenge. This makes the paper a timely and important contribution. The paper is very readable. The tables and figures are informative and I found the supporting information good and well organized.

Many thanks for your point of view about our contribution to the selection of suitable indicators.

In the legend of figure 3 (L290) BMX appears twice (BMX: mixed forests AND BMX deciduous forests).

Thanks. We changed the abbreviation error.

The method presented in the manuscript is described by the authors as simple and practical. The steps in themselves are logical and easy to understand, however the exact execution is less clear. The authors build on a large amount of literature and prior research and their work flow includes the consultation of experts. Line 377 discusses the risk of undervaluing some components; line 392 the use of a qualitative method strictly based on preselected indicators. While the authors discussion of these points is relevant and comprehensible, they do contribute to the impression that the method is not as easy to implement as the authors initially imply.

Thanks for the comments. To improve the comprehensibility of our process we have improved the explanation from the 125 to 133 lines (Previously lines 119 to 146) and we have modified figure 1. Furthermore, we have rewritten and reordered the methods, especially State 3, State 4 and the new redaction of Results (sections 3.2 and 3.3).

The pre-requirements and the time aspect could be discussed further, especially as the aim of the method is to make monitoring more manageable.

Our method presents certain key steps that can compromise the good selection of indicators. Especially by the own intricacies of each taxon. For this reason, our method presents three evident prerequisites:

• Make an exhaustive collection of information and especially the most difficult, unpublished reports.

• Having knowledgeable naturalists in the area.

• To be able to consult specialists.

Then, understanding your concern, we have made some adjustments into State 2 of M&M;

New lines from 158 to 161: “An effective collection of prior information on the area to be applied, especially not published reports, is essential. If relevant prior information is not available, you need to back to State 1 to search for or generate relevant information”.

We also incorporate into the discussion, some new arguments about prerequisites (new lines from 394 to 399):

“Some possible biases could arise from the exclusive use of the weight values in the Relevance Index as some potential candidates (e.g. plants and invertebrate species) would be undervalued so expert criteria could be needed and this knowledge will reinforce the selection of candidates prioritized. It is a key of the scheme and process of selection to have a good information base beforehand (literature, reports and databases). The higher and better the quality of the information, the more rigorous the selection of components will be.”

This complements other phrases that emphasize the expert importance in the introduction, e.g. new line from 86 to 90: “[...] the knowledge of experts or specialists in local taxa is essential since one aspect of biodiversity (species, habitats, ecological processes, and biotic, abiotic and anthropic problems) may affect a focal region differently and so require its indicator [6]. It is also important to assess which indicators are valid and informative for a region and which are redundant, overvalued or unnecessary”.

Moreover, in methods, e.g. new lines from 198 to 200: "[...] the bibliography and expert criteria are essential (Table 2), and will help highlight species or habitats that are not present or not evaluated as threatened [...]".

Some of the aspects of the methodology, specifically the step from biodiversity components to indicators, were not entirely clear to me. The methodology does not sufficiently distinguish between parameters (species, habitats, processes, factors) and the assessment methods (abundances, phenology etc.). For instance, line 233 gives butterflies as an example, but does not specify an actual parameter that is measured.

To facilitate the discrimination between parameters and method, we reformulate the new lines 228 to 244, of the State 4: Monitoring Catalogue section of the document:

“Once the different relevant components have been assessed and selected, a Monitoring Catalogue (Figure 1) can be established to act as a strategic guide, facilitate understanding, and identify which variables need to be sampled and measured to generate the biological indicators in an ecological sense and to avoid redundancies. To attain the list of relevant components, (1) we grouped components by taxonomic, physiognomic, and functional similarities (e.g. common birds, freshwater invertebrates, decomposers). Then, (2) classified the grouped relevant components in four ecological levels: species, habitat, ecological process, and ecosystem disturbances (see the case study below as a practical example). (3) For each group of relevant components we selected the adequate variables to be sampled, e.g., species presence and abundance, reproductive and survival taxes, habitat preferences, soil structure, and composition. Lastly, (4) incorporated into each level all the monitoring components whilst bearing in mind that each can be part of one or more levels depending on their role and interactions within the ecosystem (see State 3). For example, species abundance and richness of butterflies provide information about the species level and their community (indicators at species level), about the variation on ecosystem functional capabilities, e.g., as primary consumers (indicators at ecological process level), or climate change and afforestation effects (indicators at ecosystem disturbance level)”

On page 8 and 9, the authors set a threshold of >= 4 for their index. I was unsure how this threshold was set and of its implications for the selection of species.

We understand the question raised. Here we tried to establish a criterion that was rigorous and, at the same time, did not rule out underrepresented components. For this purpose, the criterion was half of the Relevance index. Thus, this criterion was chosen to greatly reduce the number of initial components but, at the same time, to ensure that those assessed by the experts were not lost.

We have incorporated the following explanation in new lines 198 to 193 of the State 3: The establishment of the relevant components of monitoring biodiversity selection for clarification:

“Because the threshold value needs to be sufficiently robust but at the same time to include potential under-detected components, we established as a criterion to maintain the components with a Relevance index equal to or above half of the maximum global potential value that included valuable ecosystem components, as we have verified in our previous tests (see Table 1).”

In Table 2, “ecological process” stood out to me as a criterion: the description could apply to all species.

Here, the selection criterion corresponding to the description was applied to all species. Unfortunately, this information is not known in-depth for all species, especially invertebrates.

The methodology that allowed to group the 434 relevant components into 32 specific monitoring components requires more details.

We have redefined the section State 4: Monitoring Catalogue section (new lines from 228 to 244) of the document for clarification, and wrote all grouped components:

“Once the different relevant components have been assessed and selected, a Monitoring Catalogue (Figure 1) can be established to act as a strategic guide, facilitate understanding, and identify which variables need to be sampled and measured to generate the biological indicators in an ecological sense and to avoid redundancies. To attain the list of relevant components, (1) we grouped components by taxonomic, physiognomic, and functional similarities (e.g. common birds, freshwater invertebrates, decomposers). Then, (2) classified the grouped relevant components in four ecological levels: species, habitat, ecological process, and ecosystem disturbances (see the case study below as a practical example). (3) For each group of relevant components we selected the adequate variables to be sampled, e.g., species presence and abundance, reproductive and survival taxes, habitat preferences, soil structure, and composition. Lastly, (4) incorporated into each level all the monitoring components whilst bearing in mind that each can be part of one or more levels depending on their role and interactions within the ecosystem (see State 3). For example, species abundance and richness of butterflies provide information about the species level and their community respectively, so in the latter case indicate the variation on ecosystem functional capabilities (e.g. as primary consumers), climate change or afforestation effects. ”

Ant the 3.3. Monitoring catalogue and Priority indicators selection (new lines from 355 to 373):

“To constitute the Monitoring catalogue, we grouped the 434 Relevant components independently by the level of species, habitats, ecological processes, and ecosystem disturbances by similarities. In the case of 408 species were grouped by taxonomic, physiognomic, and functional similarities into 17 groups of species (threatened plant species, xeric gastropods, granivorous ants, decomposers, orthoptera, butterflies, freshwater macroinvertebrates, freshwater fishes, amphibians, reptiles, common birds, raptors, bats, small mammals, common medium-size preys, ungulates and carnivores). A total of 96 habitats were grouped in 11 habitat relevant categories (Table 4). Then, the habitat category resulted in two monitoring parameters (habitat structure and composition), which will be carried out in the 11 selected relevant habitats. Moreover, it is in these habitats where the specific transversal monitoring of species, processes and disturbances will be developed. Ecological processes were grouped by ecological functions in 6, decomposition, primary production, consumers, predators, zoocory, and pollination. Ecosystem disturbances were maintained in 8 (Table 4), climatic change, wildfires, afforestation, freshwater alterations, alien species, exploitation of natural resources, forests pests, human frequentation, human infrastructures). So resulted in a total of 32 specific Monitoring Components. Finally, to obtain the Priority Indicators we used the quantitative Priority Index, and 27 of these Monitoring Components were selected to monitor the biodiversity in the PNSLL (Table 5).”

Relevant components selection: I’m not sure about the grouping of species: while I understand that the % of alien species is of interest, they could also fit into the taxonomic groups.

Although the reviewer is right (alien species could be grouped into the respective taxonomic groups), in previous discussions with managers and experts, we decided to give a category for relevant alien species, as an independent indicator. However, they are noted in surveys when autochthonous species (e.g. vegetation) are noted, i.e., they do not need a special survey. However, Then, in this case, as in others, the same species indicates on more than one level: as an indicator of species status, as well as an indicator of the landscape and ecosystem disturbances (as biological invasion).

I think it would be more easily understandable to provide the numbers separately (classification into organism groups and a separate % for status). It might also be more easy to give species numbers rather than % - please check the numbers, as the % in the text do not always seem to match up with the number of species given in Annex 2.

Thanks, we reviewed the numbers.

Reviewer suggestion was incorporated on 3.2. Relevant components selection section (new lines from 314 to 323):

“The relevant components were obtained through a Relevance Index which was calculated for all 3,226 assessed species (Table 4) of which 406 (12.6%) were selected as Relevant Components (S1 Table): plants contributed with 69 species (17% - of which 10 species were threatened species, 13 pteridophytes, 41 vascular plants, and five alien species), 191 species (47%) were invertebrates (of which 26 species were Odonata, 12 Gastropoda, 43 decomposers, 22 Formicidae, 30 Orthoptera; 36 Lepidoptera, ten other invertebrates of special interest, seven plague species and five were alien species), while 146 species (36%) were vertebrates (of which tree species were freshwater fish, seven amphibians, seven reptiles, 13 raptors, 70 common birds, 21 bats, seven small-mammals, two common medium-sized preys, two ungulates, five carnivores and nine were alien species)”

Thanks, we reviewed the numbers.

Submitted filename: 211229_Review_Comments.docx

Click here for additional data file.

11 Feb 2022

24 Feb 2022

Academic Editor:

I acknowledge the huge effort performed to summarise and synthetize all the information recorded, in such an easy way to be processed by the readers. However, there are still some points to be addressed prior to acceptance:

Line 174: include references for “monitoring programs”* and “expert criterion”.

*Torre, I., López-Baucells, A., Stefanescu, C., Freixas, L., Flaquer, C., Bartrina, C., Coronado, A., López-Bosch, D., Mas, M., Míguez, S., Muñoz, J., Páramo, F., Puig-Montserrat, X., Tuneu-Corral, C., Ubach, A., Arrizabalaga, A., 2021. Concurrent Butterfly, Bat and Small Mammal Monitoring Programmes Using Citizen Science in Catalonia (NE Spain): A Historical Review and Future Directions. Diversity 13, 454. doi:10.3390/D13090454.

Thanks for the reference, we have added the paper to the references. On the other hand, we have included “Duelli P, Obrist MK. Biodiversity indicators: the choice of values and measures. Agriculture, ecosystems & environment. 2003; 98(1):87-98. doi: 10.1016/S0167-8809(03)00072-0” for the expert criterion.

The scale values assigned to each criterion need to be further justified. Why “Threat” is in a scale 0-3, “Ecological Interest” in a scale 0-2, and “presence on monitoring programs” in scale 0-1? Why assigning more weight to Threat than to Ecological Interest? This represents a subjective starting point decided by authors.

Thanks for your comment. Degree of threat and Ecological Interest were to be given equal weight in the selection process. In the specific case of species, initially the "presence on monitoring programs" count as part of Ecological interest, as they correspond to common species (commonly detected species in monitoring plans). However, to avoid the misunderstandings and subjectivity that this implies, we have placed this category ("presence on monitoring programs") within Ecological interest. In this way, Ecological interest and Degree of threat have equal weight (from 0 to 3) in the final model (see Table 1, Table 2, and Table S1).

This is well explained in the new lines 76-90, 197-204 and 406-422 of the manuscript. However, we have also modified the following sentence in the new line 201-202 of the manuscript, to facilitate understanding:

“The ecological interest contributes to minimize the weight of these threatened and singular component, which do not necessarily reflect the spatial and temporal trends of biodiversity [23, 55] but, rather, respond to the local conservation status”.

In supporting Table, I see some data that not match my knowledge on the species selected. Some small mammals having monitoring programs, scored “0”. Also, expert criterion is low for species with reduced populations (M.glareolus, E.quercinus), so it’s disappointing. This needs to be corrected or justified. I don’t know if such differences are affecting to other taxonomic groups. This will affect final scores, and need to be recalculated, if necessary.

In the first case, the criterion was to assign a “1” if the species was detected previously in a specific monitoring program. Due to E.quercinus was captured by the monitoring program in 2018 and M.glareolus has the potential to be captured by SEMICE monitoring program; have updated the assessment of the two species. However, we have not updated the rest of the species since in the monitoring that has been carried out since 2018 (the database used here was last updated in 2017) in the natural park there have not been any new records, see Puig-Gironès R. 2020 Catàleg de Flora Vascular, Fauna Invertebrada i Fauna Vertebrada del Parc Natural de Sant Llorenç del Munt i l’Obac. Diputació de Barcelona, Barcelona.

The document presented is not intended to be an immovable document, and it is proposed to be re-evaluated in a couple of years (2024 or 2025). Therefore, we have decided not to modify the evaluation of the expert criterion since this revaluation will be made for the totality of the species. Experts and specialists were consulted to evaluate the ecological and conservation interest of each species (see acknowledgements for the list of the consulted experts), and they examined each biodiversity component and judged their importance in the focal region to minimise the risk of benefiting certain charismatic taxa or a predetermined species as an indicator.

Also raised by reviewers…Why a threshold is set in the mean, >=4 ? A rank criterion, ordering the species from 0-8, will be better since between 5-8 there are significant differences of suitability that could help selecting in particular situations (ex. limited resources). If one wants to select the most relevant indicators, I will choose those having 8 scores.

Thanks for the comment. We understand the reasoning of the editor and reviewers. However, the aim of this paper, as detailed in the introduction and discussion, is to be a balanced method (see again, the new lines 76 – 90, 197-204 and 406-422).

Given the dissimilarities in the knowledge and subsequent assessment of the species, to put a very high threshold as 8 entail biased indicators to more visible, detectable and studied species. For example, selecting those species with a Relevance index of 8, eight of the ten species of threatened and endemic flora would not be chosen, such as: Arenaria conimbricensis subsp. conimbricensis, Arenaria fontqueri or Erodium glandulosum, nor the endemic gastropods like Montserratina bofilliana, Xerocrassa montserratensis or Abida secale subsp. bofilli.

On the other hand, if the threshold were ≥ 7, then 57 species would be selected, of which: 8 would be plants (14%), 5 invertebrates (8.8%) and 44 vertebrates (77.2%) so at first instance represents a high biased frequency compared to listed plants (32%), invertebrates (61%) and vertebrates (7%) in the Sant Llorenç del Munt Natural Park (see Puig-Gironès R. 2020 Catàleg de Flora Vascular, Fauna Invertebrada i Fauna Vertebrada del Parc Natural de Sant Llorenç del Munt i l’Obac. Diputació de Barcelona, Barcelona).

These enormous differences are also maintained in the threshold ≥ 6 and 5. Therefore, we set the threshold at ≥ 4, since this limit allowed us to have reasonable percentages between plants, invertebrate and vertebrate species to follow as indicator.

Expert criteria seem a rather subjective selection criterion. Indeed, experts use similar criteria for selecting indicators regarding threat and ecological interest of the species, so it could be a redundant variable. Furthermore, a “0” indicates that experts consider the species irrelevant, or there is no information available?

Our main objective was a collaborative design process to enhance the acceptance of diverse values and prioritizations embedded in sustainability assessments. This emphasis on the process would make assessments more transparent, transformative and enduring. Even so, in certain species, habitats or processes there may be some redundancy, but we believe that the expert criterion based on the local scale minimized this effect. Furthermore, given the lack of monitoring systems or the lack of catalogues of endangered species (in the case of Catalonia we do not have a Catalogue of Endangered Fauna), expert judgement fills this gap.

On the other hand, expert criterion is necessary to assess which indicators are valid and informative for a region. As detailed in Table 2: “the criterion of external expert or taxon specialist grants a value of 2 to the species, habitats, and/or ecosystem disturbances with high importance and relevance in the context of the study area, and 1 to species, habitats, and/or ecosystem disturbances with relative importance in the study area”.

Table 1. why “z”? It should correspond to “f”. Please, justify.

Thanks to finding the error, we have changed 'z' to 'f'.

Table 2. Correct “Specie” to “Species”

Thanks, we have fixed the error.

Submitted filename: 220221_Editor_Comments.docx

Click here for additional data file.

28 Feb 2022

A comprehensive but practical methodology for selecting biological indicators for long-term monitoring

PONE-D-21-29903R2

Dear Dr. Puig-Gironès,

We’re pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it meets all outstanding technical requirements.

Within one week, you’ll receive an e-mail detailing the required amendments. When these have been addressed, you’ll receive a formal acceptance letter and your manuscript will be scheduled for publication.

An invoice for payment will follow shortly after the formal acceptance. To ensure an efficient process, please log into Editorial Manager at http://www.editorialmanager.com/pone/, click the 'Update My Information' link at the top of the page, and double check that your user information is up-to-date. If you have any billing related questions, please contact our Author Billing department directly at authorbilling@plos.org.

If your institution or institutions have a press office, please notify them about your upcoming paper to help maximize its impact. If they’ll be preparing press materials, please inform our press team as soon as possible -- no later than 48 hours after receiving the formal acceptance. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information, please contact onepress@plos.org.

Kind regards,

Ignasi Torre

Academic Editor

PLOS ONE

Additional Editor Comments (optional):

Reviewers' comments:

4 Mar 2022

PONE-D-21-29903R2

A comprehensive but practical methodology for selecting biological indicators for long-term monitoring

Dear Dr. Puig-Gironès:

I'm pleased to inform you that your manuscript has been deemed suitable for publication in PLOS ONE. Congratulations! Your manuscript is now with our production department.

If your institution or institutions have a press office, please let them know about your upcoming paper now to help maximize its impact. If they'll be preparing press materials, please inform our press team within the next 48 hours. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information please contact onepress@plos.org.

If we can help with anything else, please email us at plosone@plos.org.

Thank you for submitting your work to PLOS ONE and supporting open access.

Kind regards,

PLOS ONE Editorial Office Staff

on behalf of

Dr. Ignasi Torre

Academic Editor

PLOS ONE