Taxonomic diversity and community structure of arbuscular mycorrhizal fungi (Phylum Glomeromycota) in three maritime sand dunes in Santa Catarina state,south Brazil

Community structure and species richness of arbuscular mycorrhizal fungi (Phylum Glomeromycota) were studied in sand dune sites at Itapiruba (southern), Joaquina (intermediate) and Praia Grande (northern) beaches along the coast of the state of Santa Catarina, Brazil. In each site, a 20 × 20 m plot was established and 20 soil samples collected in a regular grid pattern. Fungal spores were extracted from each sample, counted and identified to species level. A total of 25 species were recovered belonging to seven genera and five families in the Glomeromycota. Gigaspora albida and Acaulospora scrobiculata occurred in >50 % of samples at all three sites. Other common species whose sample frequency was >50 % in one or two sites were Scutellospora weresubiae, Scutellospora cerradensis and Racocetra gregaria, while the remaining majority of species were detected in <25 % of samples within a given site. Dune sites could be differentiated based on the higher frequency of occurrence of S. cerradensis and Acaulospora morrowiae in Itapiruba, S. weresubiae in Joaquina, and Scutellospora hawaiiensis in Praia Grande. No differences across sites were observed for species richness and total spore numbers, the latter averaging from 28.8 to 31.8 spores per 100 ml soil. Shannon diversity was significantly higher in Praia Grande compared to the other two sites. Differences in the relative spore abundance of genera among dunes were detected only for Scutellospora, which was significantly more abundant in the Joaquina beach. Community structure, as depicted by species rank/log abundance graphs, was not significantly different between areas according to the Kolmogorov–Smirnov two-sample test. Species accumulation curves demonstrated that 13 samples were enough to detect 90 % of all species. Overall, sand dune systems share similar arbuscular mycorrhizal fungal communities despite being geographically distant (150 km) from each other.     

Fungal endophyte–grass symbioses are rare in the California floristic province and other regions with Mediterranean-influenced climates

Research on agronomic grasses has shown that Class 1 fungal endophytes (Neotyphodium/Epichloë; Clavicipitaceae) can have profound effects on host plant fitness. However, in natural systems, even basic ecological knowledge of most endophyte symbioses is lacking. Here, I describe the distribution and abundance of endophytes across 36 native (or naturalized) grasses in a previously unsurveyed region, the California Floristic Province. Symbiosis was generally low: 8.33 % of species and 18.75 % of genera hosted endophytes. I then compared the proportions of symbiotic species and genera found in California and other Mediterranean regions to the proportions found in non-Mediterranean regions. Surveys of Mediterranean-influenced regions showed significantly lower proportions of species (～66 % lower) and genera (～65 % lower) hosting endophyte than surveys of non-Mediterranean regions. This pattern suggests that selection in Mediterranean climates may not favor endophyte symbioses.     

Rumen ciliates of domestic cattle (Bos taurus taurus) in Kastamonu,Turkey, with the description of a new species

Species composition and distribution of ciliates were investigated in the rumen contents of 25 domestic cattle (Bos taurus taurus L.) living in Kastamonu, Turkey. Forty-seven species and 37 morphotypes representing 15 genera were identified. Of them, a new species of Ostracodinium was recognized and described as Ostracodinium anatolicum n. sp. This new species has two caudal lobes. The dorsal lobe is small and rounded and the ventral lobe is triangular shaped and bent toward the dorsal side like a thick hook. Furthermore, the anterior end of the macronucleus (1/5 of the length) is bent toward the left like a hook. The density of rumen ciliates in cattle was 96.8 ± 43.3 × 10⁴ cells mL⁻¹ and the mean number of ciliate species per host was 14.2 ± 4.4. Entodinium longinucleatum, E. nanellum, E. simulans and Isotricha prostoma were the most abundant species, each with a prevalence of 88%. Entodinium chatterjeei, E. bifidum m. monospinosum, Hsiungia triciliata, Oligoisotricha bubali, Ostracodinium dogieli, O. mammosum and O. munham are new host records for cattle from Turkey.     

Testing the effectiveness of surrogates for assessing biological diversity of arthropods in cereal agricultural landscapes

Agricultural intensification is altering biodiversity patterns worldwide. Rapid and effective methods are needed to monitor these changes in farmland biodiversity, but it becomes both a cost- and time-prohibitive task, particularly for hyper-diverse groups such as arthropods. We evaluated the effectiveness of surrogates in irrigated and rainfed wheat fields in a Mediterranean farmland in NW Spain in order to get a rapid tool to assess arthropod biodiversity. We studied six groups with different ecological needs (i.e. Aphididae, Aphidiinae, Coccinellidae, Formicidae, Heteroptera and Syrphidae) at species level (147 species), genus (105), family (10, only Heteroptera) and order (19) level. Higher taxa, cross-taxa and subset-taxa or total richness approaches were tested as well as the correlation in composition between levels for the selected groups, and the influence of farming regime. Genus richness was a good surrogate of species richness in all six groups studied (R² = 0.38–0.60), like family and order were for Heteroptera (R² = 0.37 and 0.29, respectively). Cross-taxa analyses showed that Aphididae and Aphidiinae genera (R² = 0.19 and 0.30, respectively) and species (R² = 0.20 and 0.28, respectively) were good surrogates for Aphidiinae and Aphididae species respectively. Coccinellidae genera (R² = 0.26) and species (R² = 0.25) were good surrogates for Heteroptera species. Finally, Aphididae and Coccinellidae both at genera (R² = 0.14 and 0.20, respectively) and at species levels (R² = 0.12–0.22, respectively) were good surrogates for total species richness of all groups. Genera composition was the best surrogate for the species composition within each group. Farming regime had no influence on the relationships between surrogates and species patterns in most cases. Our results suggest that genera level is a useful surrogate for all the studied groups and family is appropriate for Heteroptera. Genus level provided a saving of 15% of identification time in Aphididae and 80% for Coccinellidae. This proves its usefulness to asses and monitor biodiversity in wheat croplands and the possibility to reduce costs.     

Shape matters in sampling plant diversity: Evidence from the field

The identification of shape and size of sampling units that maximises the number of plant species recorded in multiscale sampling designs has major implications in conservation planning and monitoring actions. In this paper we tested the effect of three sampling shapes (rectangles, squared, and randomly shaped sampling units) on the number of recorded species. We used a large dataset derived from the network of protected areas in the Siena Province, Italy. This dataset is composed of plant species occurrence data recorded from 604 plots (10 m × 10 m), each divided in a grid of 16 contiguous subplot units (2.5 m × 2.5 m). Moreover, we evaluated the effect of plot orientation along the main environmental gradient, to examine how the selection of plot orientation (when elongated plots are used) influences the number of species collected. In total, 1041 plant species were recorded from the study plots. A significantly higher species richness was recorded by the random arrangement of 4 subplots within each plot in comparison to the ‘rectangle’ and ‘square’ shapes. Although the rectangular shape captured a significant larger number of species than squared ones, plot orientation along the main environmental gradient did not show a systematic effect on the number of recorded species. We concluded that the choice of whether or not using elongated (rectangular) versus squared plots should dependent upon the objectives of the specific survey with squared plots being more suitable for assessing species composition of more homogeneous vegetation units and rectangular plots being more suited for recording more species in the pooled sample of a large area.     

Foraminiferal assemblage indices: A comparison of sediment and reef rubble samples from Conch Reef,Florida, USA

Benthic foraminiferal assemblages are increasingly utilized as indicators of water and sediment quality in coastal-marine environments. Most reef-dwelling foraminifers live on firm substrata such as reef or phytal surfaces, while most assessments have examined assemblages from sediments. This case study compared relative abundances of total foraminiferal-shell assemblages between sediment and phytal/rubble samples collected from one reef within one week. A total of 117 species within 72 genera were identified, with the same taxa in both sample sets in different proportions. Larger benthic foraminifers and some agglutinated taxa were concentrated about 1.5–3 fold in sediment samples, while nearly two-thirds of small, fragile shells were lost. Several common indices were compared, including Taxonomic Richness (number of genera), Shannon (H), Simpson's (D) and Fisher (α) diversity indices, Evenness (E), and the FORAM Index (FI). Highly significant differences (p < 0.001) between shell assemblages from 13 sets of phytal/rubble substrata and sediments were found in mean number (± standard deviation) of genera (49 ± 4 vs. 34 ± 10) and mean FI (5.6 ± 0.8 vs. 3.6 ± 0.4); both reflecting greater relative abundances of smaller foraminifers in the rubble samples. Fisher diversity was marginally significant (p = 0.05); other indices showed no significant differences between sample types. Although assessment of total assemblages is substantially less costly than distinguishing between specimens that were live or dead when collected, many researchers report those distinctions. The results of our study provide insight that can assist interpretations of studies that use live assemblages to calculate the FI, rather than total assemblages for which it was originally developed.     

Impacts of forest gaps on butterfly diversity in a Bornean peat-swamp forest

Forest degradation is leading to widespread negative impacts on biodiversity in South-east Asia. Tropical peat-swamp forests are one South-east Asian habitat in which insect communities, and the impacts of forest degradation on them, are poorly understood. To address this information deficit, we investigated the impacts of forest gaps on fruit-feeding butterflies in the Sabangau peat-swamp forest, Central Kalimantan, Indonesia. Fruit-baited traps were used to monitor butterflies for 3 months during the 2009 dry season. A network of 34 traps (n_gap = 17, n_shade = 17) was assembled in a grid covering a 35 ha area. A total of 445 capture events were recorded, comprising 384 individuals from 8 species and 2 additional species complexes classified to genera. On an inter-site scale, canopy traps captured higher species richness than understory traps; however, understory traps captured higher diversity within each site. Species richness was positively correlated with percent canopy cover and comparisons of diversity indices support these findings. Coupled with results demonstrating morphological differences in thorax volume and forewing length between species caught in closed-canopy traps vs. those in gaps, this indicates that forest degradation has a profound effect on butterfly communities in this habitat, with more generalist species being favored in disturbed conditions. Further studies are necessary to better understand the influences of macro-habitat quality and seasonal variations on butterfly diversity and community composition in South-east Asian peat-swamp forests.     

A novel flexible method to split feeding behaviour into bouts

Before meal patterns can be analysed properly, a biologically relevant meal criterion must be determined in order to group short-term feeding behaviour into meals. Existing methodologies are based on modelling of the frequency distributions of intervals between feeding events but these methods cannot be used if the proper distributions cannot be clearly identified. For such cases we developed two new methods – (1) based on the analysis of the distribution of between-meal interval lengths only and (2) based on the analysis of changes in the probability of animals starting to feed with time since the last feeding event. Both methods were developed using a data set of over 700,000 records of visits to feeders obtained with broilers (Gallus gallus) aged between 2 and 5 weeks. The two methods resulted in meal criteria estimates of 20.1 and 17.5 min, respectively, which, when applied to the data set, gave statistically significant but very small differences in meal characteristics. The new methods were tested against an independent cow (Bos taurus) data set and the resultant meal criteria compared with those predicted by an existing method. The two novel methods estimated meal criteria for cows at 27.9 and 35.5 min, compared with 28.9 min for the existing method. Again, these differences in meal criteria resulted in only very small differences in meal characteristics. Even though meal criteria were relatively similar for birds and cows, characteristics such as average daily number of meals (10.9 and 5.9), meal size (12.5 g and 7.4 kg) and meal duration (7.7 and 31.4 min) were very different. The analyses show that, if the appropriate distributions of intervals cannot be identified, meal criteria can still be estimated for species as diverse as mature cows and young birds by the novel methodologies developed here.     

Phylomineralogy of the Coralline red algae: Correlation of skeletal mineralogy with molecular phylogeny

The coralline algae in the orders Corallinales and Sporolithales (subclass Corallinophycidae), with their high degree of mineralogical variability, pose a challenge to projections regarding mineralogy and response to ocean acidification. Here we relate skeletal carbonate mineralogy to a well-established phylogenetic framework and draw inferences about the effects of future changes in sea-water chemistry on these calcified red algae. A collection of 191 coralline algal specimens from New Zealand, representing 13 genera and 28 species, included members of three families: Corallinaceae, Hapalidiaceae, and Sporolithaceae. While most skeletal specimens were entirely calcitic (range: 73–100 wt.% calcite, mean 97 wt.% calcite, std dev = 5, n = 172), a considerable number contained at least some aragonite. Mg in calcite ranged from 10.5 to 16.4 wt.% MgCO₃, with a mean of 13.1 wt.% MgCO₃ (std dev = 1.1, n = 172). The genera Mesophyllum and Lithophyllum were especially variable. Growth habit, too, was related to mineralogy: geniculate coralline algae do not generally contain any aragonite. Mg content varied among coralline families: the Corallinaceae had the highest Mg content, followed by the Sporolithaceae and the Hapalidiaceae. Despite the significant differences among families, variation and overlap prevent the use of carbonate mineralogy as a taxonomic character in the coralline algae. Latitude (as a proxy for water temperature) had only a slight relationship to Mg content in coralline algae, contrary to trends observed in other biomineralising taxa. Temperate magnesium calcites, like those produced by coralline algae, are particularly vulnerable to ocean acidification. Changes in biomineralisation or species distribution may occur over the next few decades, particularly to species producing high-Mg calcite, as pH and CO₂ dynamics change in coastal temperate oceans.     

Intestinal ciliate fauna of the Asian elephant from Gaziantep,Turkey and the description of Brevitentaculum antebum n. g., n. sp.

The aims of this investigation were to identify and quantify ciliates inhabiting the large intestine of Asian elephants living in Gaziantep Zoo, Turkey, and to describe a new suctorian genus and species. Species composition and distribution of intestinal ciliates voided in the feces of two Asian elephants were examined. Fifteen species of intestinal ciliates, representing 7 genera, were identified. One new suctorian genus and species, Brevitentaculum antebum n. g., n. sp., was described. This new species has two short truncated terminal projections, with two longitudinally lined bands located between the two projections, near the convex surface of the cell. Short clavate-like tentacles are in two fascicles near each base of terminal projections on the flattened surface of the body. Ciliate densities in the two fecal samples were 4.5 × 10⁴ mL⁻¹ and 10 × 10⁴ mL⁻¹.     

Sperm cryopreservation in guppies and black mollies—A generalized freezing protocol for livebearers in Poeciliidae

In this study, we evaluated various parameters of sperm cryopreservation in two livebearers, guppies (Poecilia reticulata) and black mollies (P. latipinna). Our results suggested a common freezing protocol for the guppies and mollies: suspend sperm in Hanks’ balanced salt solution (HBSS) at 300 mOsm/kg, use 14% glycerol as cryoprotectant, cool at 25 °C/min, and thaw at 40 °C in a water bath for 7 s. Live young were produced from females inseminated with frozen-thawed sperm in both species. In guppies, percent fertilization (F) and the number of embryos (N) produced with cryopreserved sperm (F = 50%, N = 74, from 26 females) were similar to those of fresh controls (F = 54%, N = 61, from 22 females). Interestingly, this same freezing protocol has been used successfully for sperm cryopreservation in green swordtails Xiphophorus helleri, and platyfish of X. couchianus with post-thaw motility as high as 80%. All these species belong to the family of Poeciliidae, and their sperm are similar in morphology exhibiting the absence of acrosome, elongate sperm head, and the long mitochondrial sheaths. Besides their internal fertilization reproduction mode, these fish are also small in size (2–4 cm) and live in a freshwater environment. Sperm cryopreservation in fish has been generally recognized as species specific, and new protocols are required for new species. However, results presented in this study suggested otherwise. Thus, sperm cryopreservation methods optimized for one species may be applicable to others if they are taxonomical closely related species with similar sperm morphology and reproduction mode. Considering the enormous number of fish species on the planet, development of generalized sperm freezing protocols for species in groups could have additional advantages for genetic conservation.     

Achilles tendon stress is more sensitive to subject-specific geometry than subject-specific material properties: A finite element analysis

This study used subject-specific measures of three-dimensional (3D) free Achilles tendon geometry in conjunction with a finite element method to investigate the effect of variation in subject-specific geometry and subject-specific material properties on tendon stress during submaximal isometric loading. Achilles tendons of eight participants (Aged 25–35 years) were scanned with freehand 3D ultrasound at rest and during a 70% maximum voluntary isometric contraction. Ultrasound images were segmented, volume rendered and transformed into subject-specific 3D finite element meshes. The mean (±SD) lengths, volumes and cross-sectional areas of the tendons at rest were 62 ± 13 mm, 3617 ± 984 mm³ and 58 ± 11 mm² respectively. The measured tendon strain at 70% MVIC was 5.9 ± 1.3%. Subject-specific material properties were obtained using an optimisation approach that minimised the difference between measured and modelled longitudinal free tendon strain. Generic geometry was represented by the average mesh and generic material properties were taken from the literature. Local stresses were subsequently computed for combinations of subject-specific and generic geometry and material properties. For a given geometry, changing from generic to subject-specific material properties had little effect on the stress distribution in the tendon. In contrast, changing from generic to subject-specific geometry had a 26-fold greater effect on tendon stress distribution. Overall, these findings indicate that the stress distribution experienced by the living free Achilles tendon of a young and healthy population during voluntary loading are more sensitive to variation in tendon geometry than variation in tendon material properties.     

Global phylogeny and new classification of the Rapaninae (Gastropoda: Muricidae), dominant molluscan predators on tropical rocky seashores

The monophyly of the muricid subfamily Rapaninae has recently been confirmed with molecular techniques, but its composition and the relationships among its constituent genera remain unclear. We use four genes (28S rRNA, 12S rRNA, 16S rRNA and cytochrome c oxidase subunit I, COI) to construct a Bayesian phylogeny of 80 rapanine species (73% of the approximately 109 currently accepted), representing 27 of the 31 nominal genera. This is the most complete phylogeny of this taxonomically confusing subfamily yet produced. We propose a revised phylogenetic classification of the Rapaninae, assigning the recognized species to 28 genera. Most of the morphologically-defined rapanine genera are considered valid, including Purpura, Drupa, Thais and Nassa, but many of them are here restricted or redefined so that they are monophyletic. In particular the familiar genus Thais is narrowly restricted to a single species. Many groups previously accepted as subgenera, including Mancinella, Vasula, Thalessa and Thaisella, are here accorded full generic rank. We describe one new genus, Indothais. While we do not formally alter species-level taxonomy, we show molecular evidence for two cryptic species and several instances of probable species synonymy. We estimate the age of diversification of the Rapaninae as Late Cretaceous (75.9 Ma) and of many of its genera as Miocene.     

Fruit nutritional composition and non-nutritive traits of indigenous South African tree species

Frugivorous animals play a major role in dispersing tropical, and to a lesser extent, temperate tree species. In order to attract potential seed dispersers, plants generally offer a reward of fleshy fruit pulp. Criteria for fruit choice by avian frugivores are influenced by a number of non-nutritive (e.g. fruit size and colour) factors; and nutritional composition of the fruit. There is a paucity of nutritional composition and other fruit trait data of indigenous South African fruit. This information is necessary in order to determine which frugivores are likely to ingest which fruits and consequently act as potential seed dispersal agents. This information would provide us with an understanding of the inter-relationships between indigenous fruit and frugivores in South Africa. Consequently nutritional composition was investigated in various indigenous fruit species that avian frugivores feed on. Fruits were collected from 38 indigenous tree species found in KwaZulu-Natal Afromontane and coastal forests. Pulp was freeze-dried to constant mass and then analysed for sugar, lipid and protein content; and for water content determination. Fruit width in this study ranged from 4 mm (Searsia rehmanniana and Trema orientalis) to 40 mm (Annona senegalensis, Ficus sur and Xylotheca kraussiana). Of the fruits examined in this study 29% were black and 43% were red when ripe. Most (84%) fruit species analysed for sugar content were hexose dominant with 50% being fructose and 34% being glucose dominant. Only 16% of the fruit species analysed were sucrose dominant. Fruits in this study were generally observed to be high (mean: 68.1 ± 3.3%; n = 30) in water content; and low in protein and lipid content respectively (mean: 8.2 ± 0.5%; 9.3 ± 2.2%; n = 30) indicating that these fruit species could be considered as nutrient-dilute. Future studies need to determine the nutritional composition of the remaining indigenous South African fruit in order to develop a comprehensive database as well as examining non-nutritive factors.     

Estimating field metabolic rates for Australian marsupials using phylogeny

Field metabolic rate (FMR) is a useful measure for the energy expenditure in free-ranging animals. Field metabolic rates for species that have not been measured are usually predicted by allometric equations on the basis of their body mass (BM). Phylogenetically informed methods improve estimates of both allometric relationships and species-specific FMR values by considering the evolutionary history of species. Further improvement is possible by incorporating isolated measurements on BM and FMR, but most existing methods force the user to discard such incomplete data. In the present study the FMR of most Australian marsupial species was predicted for the first time using a phylogenetic method that was explicitly designed to handle incomplete data. This allows full use of the dataset containing 35 samples of FMR and 130 samples of BM. Cross-validation demonstrated that FMRs were estimated with high accuracy. The resulting prediction equation was FMR (kJ day^? 1) = 5.27 BM (g)^0.69. Field metabolic rate and BM were highly phylogenetically correlated (r = 0.96), i.e. FMR and BM co-evolved. Differences between species-specific and generic marsupial estimates of FMR revealed that herbivores have lower energy expenditure than carnivores. Specifically, herbivorous macropods have on average lower relative FMR (kJ/d) (3.75 ± 0.53 BM^0.69; mean ± SD) than carnivorous dasyurids (7.64 ± 0.84 BM^0.69). Phylogenetically informed estimates for most extant Australian marsupial species are now available.     

Barcoding forest insect pests in South Korea: Constructing a basic endemic species dataset

A total of 103 barcode (mitochondrial COI) sequences were newly provided for 77 forest insect pests from 66 genera belonging to Coleoptera, Hemiptera, and Lepidoptera. All 77 species had distinct COI sequences, revealing low intraspecific genetic divergence (< 1.20%) and high interspecific genetic divergence (> 7.30%). Among the 66 genera, 32 COI sequences of 25 species belonging to 16 genera were compared with 280 COI sequences of 117 species belonging to the same 16 genera archived in GenBank, showing that most species were clearly distinguished by barcode sequences. Based on these results, we conclude that a DNA barcode is effective for identifying forest insect pest species.     

Cow traffic in relation to social rank and motivation of cows in an automatic milking system with control gates and an open waiting area

Cow traffic in automatic milking (AM) systems can be controlled with gates at the entrances to feeding areas, where cows are either let through (gate opens) or redirected (gate remains closed) to an open waiting area in front of the milking unit (MU). The motivation of a cow to enter the MU after a redirection is assumed to be reflected in redirection time, which is the elapsed time from a redirection in a control gate (CG) until the next registration in the MU (i.e. CG → milking in MU/passage through MU). In order to study the possibility of guiding cows in this kind of system and to increase the understanding for the emergence of redirection time, a behavior study was performed at the Swedish University of Agricultural Sciences, Uppsala, Sweden. Two groups of 12 cows each (high and low social rank, respectively) in an AM-system with control gates and an open waiting area were used as focal cows. After a redirection in a CG, the location of each focal cow was continuously observed until entrance in the MU or during a maximum observation time of 1 h. This was repeated five times for each focal cow, generating 120 observations in total. The social rank of the cows was established from registered visits to roughage stations. Cows of high social rank spent, on an average, 13 min in the waiting area during the observation period, compared to 20 min for cows of low social rank. In 22% of all observations, the cows entered cubicles in a resting area during the 1 h observation period and when they did, they, on an average, spent 41 min of the 1 h long observation time in the cubicles. Both time since last feeding and time since last milking were in negative relationship to redirection time. The number of cows in the waiting area significantly prolonged the redirection time. Redirection time was influenced by individual characteristics, such as social rank and by situation-specific factors, such as time since last feeding, time since last milking and the number of cows in the waiting area.     

Is there a significant relationship between the benthic status of an area,determined by two broadly-used indices,and best professional judgment?

Two benthic indices to assess the quality status (the AZTI's Marine Biotic Index (AMBI) and multivariate-AMBI (M-AMBI)) are being used extensively in different habitats worldwide. We try to interpret what is behind these indices making them suitable for different habitats. To demonstrate that, we used best professional judgment (BPJ), applying it to a dataset from southern Chile, to determine the criteria proposed by 12 experts in assessing the status. The experts were provided with raw species abundance data, from 12 stations within a gradient of disturbance, from unaffected to severely affected. There was a very good agreement among experts (kappa values 0.72–0.77), with highly significant (p < 0.001) correlation between BPJ and AMBI and M-AMBI classifications, and an agreement of 76.4% and 81.9%, respectively. When comparing BPJ in Chile with other results in Europe, USA and northern Africa, similar patterns can be identified: (i) the number of criteria identified for classification is very high (range 7–12); (ii) the experts use several criteria together in the BPJ assessment; and (iii) the rank of the most important criteria is indicator species, richness, and diversity/dominance. These criteria are included in indices such as AMBI and M-AMBI. Hence, although experts are classifying samples subjectively when applying BPJ, they are corroborated in their opinions when using such indices. This fact can explain why these indices are so widely used.