Philippa Kaur

The Convention on International Trade in Endangered Species of Wild Fauna and Flora (CITES) last month released its annual report on elephant poaching. It reveals a downward trend across African range states, based on data from its Monitoring the Illegal Killing of Elephants programme. The decline correlates with reduced ivory trading over the period, particularly in the Chinese market.
Competing Interests
The author declares no competing interests. More

Description of the study areaThe study covers urban, peri-urban and rural wetlands in the Bamenda Municipality of the North West Region of Cameroon that have evolved concomitantly with different stages of urbanization (Fig. 1). In this study, urbanisation is considered a loose term that is aimed at giving a geographical expression to the variation in the economic, social and cultural practices in the milieu. The central town with many economic activities is termed the urban, the fringe area with sprawls is termed peri-urban while the rural has typical peasant activities and make-shift structures. From the variation of human activities in the three sub-zones, a variety of chemical substances are discharged into drains, playing a substantial role in soil quality, and therefore plant macrophyte diversity. The Plants studied were ubiquitous in the area and verification of their IUCN conservation status in the red data book of plants of Cameroon confirmed their abundance14. Information on protected sites in Cameroon does not place the study area under conservation status. In line with that, permits are not required to undertake academic and research studies as well as do a responsible collection of plants in the study area. The urbanization rate of Bamenda is 42%, and the population grew from 48,111 inhabitants in 1976 to 488,883 inhabitants in 201015, with 150–200 inhabitants/km2.Figure 1Relief Map of Bamenda showing the Bamenda escarpment, topography and the location for quadrat sites.Full size imageThe study area is part of the Bamenda escarpment that is located between latitudes 5° 55″ N and 6° 30″ N and longitudes 10° 25″ E and 10° 67″ E. The town shows an altitudinal range of 1200–1700 m and is divided into two parts by escarpments—a low-lying and gently undulating part with altitudes ranging from 1200 to 1400 m, with many flat areas that are usually inundated for most parts of the year, and an elevated part that range from 1400 to 1700 m altitude. Most of the streams take their rise from this elevated part (Fig. 1).This area experiences two seasons—a rainy season (mid-March to mid-October) and a short dry season (mid-October to mid-March). The thermic and hyperthermic temperature regimes dominate in the area. The mean annual temperature stands at 19.9 °C. January and February are the hottest months with mean monthly temperatures of 29.1 and 29.7 °C, respectively. This area is dominated by the Ustic and Udic moisture regimes with the Udic extending to the south9. Annual rainfall ranges from 1300 to 3000 mm16. The area has a rich hydrographical network with intense human activities and a dense population along different water courses in the watershed. The area is bounded on the West, North and East by the Cameroon Volcanic Line (made up of basalts, trachytes, rhyolites and numerous salt springs). The geologic history of this area originates from the Precambrian era when there was a vast formation of geosynclinal complexes, which became filled with clay-calcareous, and sandstone sediments9. These materials, crossed by intrusions of crystalline rocks, were folded in a generally NE-SW direction and underwent variable metamorphism9. The Rocks in the area are thus of igneous (granitic and volcanic) and metamorphic (migmatites) origin17, which gives rise to ferralitic soils18.Agriculture is the principal human activity in and around this region18. The area equally harbours the commercial center that has factories ranging from soap production, and mechanic workshops to metallurgy, which may be potential sources of pollutants that can influence wetland Geochemistry. Raffia farinifera bush, which is largely limited to the wetlands, is an important vegetation type in this area. R. farinifera provides raffia wine, a vital economic resource to the inhabitants who are fighting against the cultivation of these wetlands by vegetable farmers.Methods of the studyMacrophyte diversity studyThe plant diversity of the wetlands was evaluated using quadrats in the dry season for accessibility reasons. For each of the three wetlands (the urban, peri-urban and rural areas), three transects were established on which representative quadrats, each measuring 10 m × 10 m, were mapped out in uncultivated areas for the determination of plant species cover-abundance and diversity. It is perceived that the different zones receive different mixtures of chemical substances and thus influence macrophyte diversity differently.According to a publication by14 on the vascular plants of Cameroon and a taxonomic checklist with IUCN assessment, the plants of the area are placed under the Least Concern Category(LC), and therefore not in the risky category. Diversity studies involved the identification of a specific area called “relevé” by progressively increasing test quadrat size and sampling for specific diversity until the smallest area with adequate species representation was reached. The relevé size determined here was 1 m2, making a total of 300 sub-quadrats (relevé) in the entire study ie. 100 in each main quadrat). For each site (main quadrat), 10 representative relevés were sampled and all plant species were enumerated. Most plant species in each of them were identified in the field by the Botanist, Dr Ndam Lawrence Monah using visual observation of the morphology of the leaves and flowers, a self-made field guide, the Flora of West Africa and the Flora of Cameroon. 10 unidentified plants were appropriately collected where there were in abundance, placed onto a conventional plant press and dried in the field. Voucher specimens were tagged and transported to the Limbe Botanic Gardens (SCA: Southern Cameroon, the code of the Limbe Botanic Gardens Herbarium) for identification. Mr Elias Ndive, the Taxonomist of the Limbe Botanic Gardens compared unidentified specimens with authentic herbarium specimens and other taxonomic guides and finally identified them. Voucher specimens of the 10 plants were given identification numbers and deposited in the Herbarium of the Limbe Botanic Gardens.The Braun–Banquet method was used19 for the assessment of species cover abundance. Relative abundance and abundance ratings were determined using the Braun–Banquet rating scheme (Table 1).Table 1 Braun-Blanquet rating scheme for vegetation cover-abundance, Source19.Full size tableSimpson’s diversity indexSpecies richness was evaluated using Simpson’s diversity index (D), which takes into account both species richness and the Braun-Blanquet rating scheme for vegetation cover abundance and evenness of abundance among the species present. In essence, D measures the probability that two individuals that are randomly selected from an area will belong to the same species. The formula for calculating D is presented as:$${text{D}} = frac{{sum {{text{n}}_{i} left( {{text{n}}_{i} – 1} right)} }}{{{text{N}}({text{N}} – 1)}}$$where ni = the total number of each species; N = the total number of individuals of all species.The value of D ranges from 0 to 1. With this index, 0 represents infinite diversity and 1 represents no diversity. That is, the larger the value the lower the diversity.Alternatively, Simpson’s Diversity Index, = 1–D,1-D was used as a measure of diversity because it is more logical and less likely to cause confusion. The scale then gives a score from 0 to 1 with higher scores showing higher diversity (instead of being associated with low scores).The Simpson index is a dominance index because it gives more weight to common or dominant species. In this case, a few rare species with only a few representatives will not affect the diversity.
Soil sampling and analysisSoil sampling was done in and around the three quadrats laid in the urban, peri-urban and rural wetlands for macrophytes sampling. Twenty-one (21) composite samples (0–25 cm) were randomly collected (Fig. 2) and taken to the laboratory in black plastic bags. Each composite sample was a collection of 5 dried core soil samples. Due to the observed greater heterogeneity in the urban sector, the sampling density was intensified. The soil samples were air-dried and screened through a 2-mm sieve. They were analyzed in duplicate for their physicochemical properties in the Environmental and Analytical Chemistry Laboratory of the University of Dschang, Cameroon. Particle size distribution, cation exchange capacity (CEC), exchangeable bases, electrical conductivity (EC) and pH were determined by standard procedures20. Soil pH was measured both in water and KCl (1:2.5 soil/water mixture) using a glass electrode pH meter. Part of the soil was ball-milled for organic carbon (Walkley–Black method) and total nitrogen (Macro-Kjeldahl method) as largely described by20. Available phosphorus (P) was determined by Bray I method. Exchangeable cations were extracted using 1 N ammonium acetate at pH 7. Potassium (K) and sodium (Na) in the extract were determined using a flame photometer and magnesium (Mg) and calcium (Ca) were determined by complexiometric titration. Exchange acidity was extracted with 1 M KCl followed by quantification of Al and H by titration20. Effective cation exchange capacity (ECEC) was determined as the sum of bases and exchanged acidity.Figure 2Adapted from the 1980 land use map of the Bamenda City Area showing soil sampling points: Source Bamenda City Council.Map of the study area in freshwater wetlands of Bamenda Municipality.Full size imageApparent CEC (CEC at pH 7) was determined directly as outlined by20. Based on critical values of nutrients established for vegetables, nutrients were declared sufficient or deficient.
Statistical analysisThe data were subjected to statistical analysis using Microsoft Excel 2007 and SPSS statistical package 20.0. Soil properties were assessed for their variability using the coefficient of variation (CV) and compared with variability classes (Table 2).$$CV=frac{Sd}{X}X 100$$where: Sd = standard deviation; = X arithmetic mean of soil properties.Table 2 Grouping coefficient of variation into variability classes.Full size tableThe hierarchical cluster analysis (HCA) was used to group the area under managing units. The main goal of the hierarchical agglomerative cluster analysis is to spontaneously classify the data into groups of similarity (clusters). This is done by searching objects in the n-dimensional space that is located in the closest neighborhood and separating a stable cluster from other clusters. The sampling sites were considered objects for classification. Each object was determined by a set of variables (chemical concentrations of the soils in this case). More

These authors contributed equally: Laura H. Antão, Benjamin Weigel.These authors jointly supervised this work: Tomas Roslin, Anna-Liisa Laine.Research Centre for Ecological Change, Organismal and Evolutionary Biology Research Programme, Faculty of Biological and Environmental Sciences, University of Helsinki, Helsinki, FinlandLaura H. Antão, Benjamin Weigel, Giovanni Strona, Maria Hällfors, Elina Kaarlejärvi, Otso Ovaskainen, Marjo Saastamoinen, Jarno Vanhatalo, Tomas Roslin & Anna-Liisa LaineDepartment of Biological Sciences, University of South Carolina, Columbia, SC, USATad DallasDepartment of Biology, Lund University, Lund, SwedenØystein H. OpedalFinnish Environment Institute (SYKE), Helsinki, FinlandJanne Heliölä, Mikko Kuussaari, Juha Pöyry & Kristiina VuorioNatural Resources Institute Finland (Luke), Helsinki, FinlandHeikki Henttonen, Otso Huitu, Andreas Lindén, Päivi Merilä, Maija Salemaa & Tiina TonteriSection of Ecology, Department of Biology, University of Turku, Turku, FinlandErkki KorpimäkiFinnish Museum of Natural History, University of Helsinki, Helsinki, FinlandAleksi LehikoinenKainuu Centre for Economic Development, Transport and the Environment, Kajaani, FinlandReima LeinonenUniversity of Helsinki, Helsinki, FinlandHannu PietiäinenDepartment of Biological and Environmental Science, University of Jyväskylä, Jyväskylä, FinlandOtso OvaskainenCentre for Biodiversity Dynamics, Department of Biology, Norwegian University of Science and Technology, Trondheim, NorwayOtso OvaskainenHelsinki Institute of Life Science, University of Helsinki, Helsinki, FinlandMarjo SaastamoinenDepartment of Mathematics and Statistics, Faculty of Science, University of Helsinki, Helsinki, FinlandJarno VanhataloSpatial Foodweb Ecology Group, Department of Agricultural Sciences, University of Helsinki, Helsinki, FinlandTomas RoslinSpatial Foodweb Ecology Group, Department of Ecology, Swedish University of Agricultural Sciences, Uppsala, SwedenTomas RoslinDepartment of Evolutionary Biology and Environmental Studies, University of Zürich, Zürich, SwitzerlandAnna-Liisa Laine More

Phung, T. X., Nascimento, J. C. S., Novarro, A. J. & Wiens, J. J. Correlated and decoupled evolution of adult and larval body size in frogs. Proc. R. Soc. Lond. B 287, 20201474–10 (2020).
Google Scholar 
Hall, B. K. & Wake, M. H. The Origin and Evolution of Larval Forms (Gulf Professional Publishing, 1999).Hime, P. M. et al. Phylogenomics reveals ancient gene tree discordance in the amphibian tree of life. Syst. Biol. 70, 49–66 (2021).CAS 
PubMed 

Google Scholar 
Duellman, W. E. & Trueb, L. Biology of Amphibians (Johns Hopkins University Press, 1994).Nunes-de-Almeida, C. H., Batista Haddad, C. F. & Toledo, L. F. A revised classification of the amphibian reproductive modes. Salamandra 57, 413–427 (2021).
Google Scholar 
Vences, M. & Köhler, J. Global diversity of amphibians (Amphibia) in freshwater. Hydrobiologia 595, 569–580 (2007).
Google Scholar 
Blackburn, D. G. Evolution of vertebrate viviparity and specializations for fetal nutrition: a quantitative and qualitative analysis. J. Morphol. 276, 961–990 (2015).PubMed 

Google Scholar 
AmphibiaWeb. Electronic Database (University of California, Berkeley, CA, USA, 2019). https://amphibiaweb.org.Frost, D. R. Amphibian Species of the World: an Online Reference. Version 6.0 (date of access: 01.08.2019). Electronic Database (American Museum of Natural History, New York, USA, 2019). https://amphibiansoftheworld.amnh.org/.Bonett, R. M., Ledbetter, N. M., Hess, A. J., Herrboldt, M. A. & Denoël, M. Repeated ecological and life cycle transitions make salamanders an ideal model for evolution and development. Developmental Dynamics 251, 957–972 (2022).Salthe, S. N. Reproductive modes and the number and sizes of ova in the urodeles. Am. Midl. Naturalist 81, 467490 (1969).
Google Scholar 
Salthe, S. N. & Duellman, W. E. in Evolutionary Biology of the Anurans (ed. Vial, J. L.) 229–249 (University of Missouri Press Columbia, 1973).Haddad, C. & Prado, C. P. A. Reproductive modes in frogs and their unexpected diversity in the Atlantic Forest of Brazil. BioScience 55, 207–217 (2005).
Google Scholar 
Lutz, B. Ontogenetic evolution in frogs. Evolution 2, 29–39 (1948).CAS 
PubMed 

Google Scholar 
Crump, M. L. Anuran reproductive modes: evolving perspectives. J. Herpetol. 49, 1–16 (2015).
Google Scholar 
Schoch, R. Evolution of life cycles in early amphibians. Annu. Rev. Earth Planet. Sci. 37, 135–162 (2009).ADS 
CAS 

Google Scholar 
Meegaskumbura, M. et al. Patterns of reproductive-mode evolution in Old World tree frogs (Anura, Rhacophoridae). Zool. Scr. 44, 509–522 (2015).
Google Scholar 
Portik, D. M. & Blackburn, D. C. The evolution of reproductive diversity in Afrobatrachia: A phylogenetic comparative analysis of an extensive radiation of African frogs. Evolution 70, 2017–2032 (2016).PubMed 
PubMed Central 

Google Scholar 
San Mauro, D. et al. Life-history evolution and mitogenomic phylogeny of caecilian amphibians. Mol. Phylogenet. Evol. 73, 177–189 (2014).PubMed 

Google Scholar 
Pereira, E. B., Collevatti, R. G., de Carvalho Kokubum, M. N., de Oliveira Miranda, N. E. & Maciel, N. M. Ancestral reconstruction of reproductive traits shows no tendency toward terrestriality in leptodactyline frogs. BMC Evol. Biol. 15, 91 (2015).Gomez-Mestre, I., Pyron, R. A. & Wiens, J. J. Phylogenetic analyses reveal unexpected patterns in the evolution of reproductive modes in frogs. Evolution 66, 3687–3700 (2012).PubMed 

Google Scholar 
Wake, D. B. & Hanken, J. Direct development in the lungless salamanders: what are the consequences for developmental biology, evolution and phylogenesis? Int. J. Dev. Biol. 40, 859–869 (1996).CAS 
PubMed 

Google Scholar 
Dubois, A. Developmental pathway, speciation and supraspecific taxonomy in amphibians: 1. Why are there so many frog species in Sri Lanka? Alytes 22, 19–37 (2004).
Google Scholar 
Hedges, S. B., Duellman, W. E. & Heinicke, M. P. New World direct-developing frogs (Anura: Terrarana): molecular phylogeny, classification, biogeography, and conservation. Zootaxa 1737, 1–182 (2008).
Google Scholar 
Dugo-Cota, Á., Vilà, C., Rodríguez, A. & Gonzalez-Voyer, A. Ecomorphological convergence in Eleutherodactylus frogs: a case of replicate radiations in the Caribbean. Ecol. Lett. 22, 884–893 (2019).PubMed 

Google Scholar 
Simpson, G. G. The Major Features of Evolution (Columbia University Press, 1953).Vági, B., Végvári, Z., Liker, A., Freckleton, R. P. & Szekely, T. Parental care and the evolution of terrestriality in frogs. Proc. R. Soc. Lond. B 286, 20182737–10 (2019).
Google Scholar 
Furness, A. I. & Capellini, I. The evolution of parental care diversity in amphibians. Nat. Commun. 10, 1–12 (2019).CAS 

Google Scholar 
Furness, A. I., Venditti, C. & Capellini, I. Terrestrial reproduction and parental care drive rapid evolution in the trade-off between offspring size and number across amphibians. PLoS Biol. 20, e3001495 (2022).CAS 
PubMed 
PubMed Central 

Google Scholar 
Wollenberg, K. C., Vieites, D. R., Glaw, F. & Vences, M. Speciation in little: the role of range and body size in the diversification of Malagasy mantellid frogs. BMC Evol. Biol. 11, 217 (2011).PubMed 
PubMed Central 

Google Scholar 
Cayuela, H. et al. Determinants and consequences of dispersal in vertebrates with complex life cycles: a review of pond-breeding amphibians. Q. Rev. Biol. 95, 1–36 (2020).
Google Scholar 
Phillimore, A. B., Freckleton, R. P., Orme, C. D. L. & Owens, I. P. F. Ecology predicts large‐scale patterns of phylogenetic diversification in birds. Am. Nat. 168, 220–229 (2006).PubMed 

Google Scholar 
Chen, J.-M. et al. An integrative phylogenomic approach illuminates the evolutionary history of Old World tree frogs (Anura: Rhacophoridae). Mol. Phylogenet. Evol. 145, 106724 (2020).PubMed 

Google Scholar 
Zimkus, B. M., Lawson, L., Loader, S. P. & Hanken, J. Terrestrialization, miniaturization and rates of diversification in African Puddle Frogs (Anura: Phrynobatrachidae). PLoS ONE 7, e35118 (2012).ADS 
CAS 
PubMed 
PubMed Central 

Google Scholar 
Moen, D. S. Improving inference and avoiding over-interpretation of hidden-state diversification models: specialized plant breeding has no effect on diversification in frogs. Evolution 76, 373–384 (2022).PubMed 

Google Scholar 
Eastman, J. M. & Storfer, A. Correlations of life-history and distributional-range variation with salamander diversification rates: evidence for species selection. Syst. Biol. 60, 503–518 (2011).PubMed 

Google Scholar 
Bonett, R. M., Steffen, M. A., Lambert, S. M., Wiens, J. J. & Chippindale, P. T. Evolution of paedomorphosis in plethodontid salamanders: ecological correlates and re-evolution of metamorphosis. Evolution 68, 466–482 (2014).PubMed 

Google Scholar 
Akaike, H. A new look at the statistical-model identification. IEEE Trans. Autom. Control 19, 716–723 (1974).ADS 
MathSciNet 
MATH 

Google Scholar 
Wagenmakers, E.-J. & Farrell, S. AIC model selection using Akaike weights. Psychon. Bull. Rev. 11, 192–196 (2004).PubMed 

Google Scholar 
Beaulieu, J. M., Oliver, J. C., O’Meara, B. & Boyko, J. R package ‘corHMM’: hidden markov models of character evolution. (2020).Pagel, M. & Meade, A. BayesTraits, version 4. University of Reading, Berkshire, UK. http://www.evolution.rdg.ac.uk (2022).Pagel, M. & Meade, A. Bayesian analysis of correlated evolution of discrete characters by reversible-jump Markov chain Monte Carlo. Am. Nat. 167, 808–825 (2006).PubMed 

Google Scholar 
Maddison, W., Midford, P. & Otto, S. Estimating a binary character’s effect on speciation and extinction. Syst. Biol. 56, 701–710 (2007).PubMed 

Google Scholar 
FitzJohn, R. G., Maddison, W. P. & Otto, S. P. Estimating trait-dependent speciation and extinction rates from incompletely resolved phylogenies. Syst. Biol. 58, 595–611 (2009).PubMed 

Google Scholar 
Beaulieu, J. M. & O’Meara, B. C. Detecting hidden diversification shifts in models of trait-dependent speciation and extinction. Syst. Biol. 65, 583–601 (2016).PubMed 

Google Scholar 
Herrera-Alsina, L., van Els, P. & Etienne, R. S. Detecting the dependence of diversification on multiple traits from phylogenetic trees and trait data. Syst. Biol. 68, 317–328 (2018).
Google Scholar 
Strathmann, R. R. Hypotheses on the origins of marine larvae. Annu. Rev. Ecol. Syst. 24, 89–117 (1993).
Google Scholar 
Wray, G. A. Evolution of larvae and developmental modes. In Ecology of Marine Invertebrate Larvae. (ed. McEdward, L.) 413–447 (CRC Press, 1995).Raff, R. A. Origins of the other metazoan body plans: the evolution of larval forms. Philos. T R. Soc. B 363, 1473–1479 (2008).
Google Scholar 
Collin, R. & Moran, A. in Evolutionary Ecology of Marine Invertebrate Larvae 50–66 (Oxford University Press, 2018).Collin, R. & Miglietta, M. P. Reversing opinions on Dollo’s Law. Trends Ecol. Evol. 23, 602–609 (2008).PubMed 

Google Scholar 
Wiens, J. J. Re-evolution of lost mandibular teeth in frogs after more than 200 million years, and re-evaluating Dollo’s law. Evolution 65, 1283–1296 (2011).PubMed 

Google Scholar 
Wiens, J. J., Kuczynski, C. A., Duellman, W. E. & Reeder, T. W. Loss and re-evolution of complex life cycles in marsupial frogs: does ancestral trait reconstruction mislead? Evolution 61, 1886–1899 (2007).CAS 
PubMed 

Google Scholar 
Chippindale, P. T., Bonett, R. M., Baldwin, A. S. & Wiens, J. J. Phylogenetic evidence for a major reversal of life-history evolution in plethodontid salamanders. Evolution 58, 2809–2815 (2004).CAS 
PubMed 

Google Scholar 
Castroviejo-Fisher, S. et al. Phylogenetic systematics of egg-brooding frogs (Anura: Hemiphractidae) and the evolution of direct development. Zootaxa 4004, 1–75 (2015).PubMed 

Google Scholar 
Naumann, B., Schweiger, S., Hammel, J. U. & Müller, H. Parallel evolution of direct development in frogs—skin and thyroid gland development in African Squeaker Frogs (Anura: Arthroleptidae: Arthroleptis). Dev. Dyn. 250, 584–600 (2021).PubMed 

Google Scholar 
Goldberg, J., Taucce, P. P. G., Quinzio, S. I., Haddad, C. F. B. & Candioti, F. V. Increasing our knowledge on direct-developing frogs: the ontogeny of Ischnocnema henselii (Anura: Brachycephalidae). Zool. Anz. 284, 78–87 (2020).
Google Scholar 
Wassersug, R. J. & Duellman, W. E. Oral structures and their development in egg-brooding hylid frog embryos and larvae: evolutionary and ecological implications. J. Morphol. 182, 1–37 (1984).PubMed 

Google Scholar 
Kerney, R. R., Blackburn, D. C., Müller, H. & Hanken, J. Do larval traits re-evolve? Evidence from the embryogenesis of a direct-developing salamander, Plethodon cinereus. Evolution 66, 252–262 (2011).PubMed 

Google Scholar 
Theska, T. Musculoskeletal development of the Central African caecilian Idiocranium russeli (Amphibia: Gymnophiona: Indotyphlidae) and its bearing on the re-evolution of larvae in caecilian amphibians. Zoomorphology 138, 137–158 (2019).
Google Scholar 
Laslo, M., Denver, R. J. & Hanken, J. Evolutionary conservation of thyroid hormone receptor and deiodinase expression dynamics in ovo in a direct-developing frog, Eleutherodactylus coqui. Front. Endocrinol. 10, 307 (2019).
Google Scholar 
Gao, W. et al. Genomic and transcriptomic investigations of the evolutionary transition from oviparity to viviparity. Proc. Natl Acad. Sci. USA 116, 3646–3655 (2019).ADS 
CAS 
PubMed 
PubMed Central 

Google Scholar 
Altig, R. & Crother, B. I. The evolution of three deviations from the biphasic anuran life cycle: alternatives to selection. Herpetol. Rev. 37, 321–356 (2006).
Google Scholar 
Venturelli, D. P., da Silva, W. R. & Giaretta, A. A. Tadpoles’ resistance to desiccation in species of Leptodactylus (Anura, Leptodactylidae). J. Herpetol. 55, 265–270 (2021).
Google Scholar 
Seymour, R. S. Respiration of aquatic and terrestrial amphibian embryos. American Zoologist 39, 261–270 (1999).Blackburn, D. G. Convergent evolution of viviparity, matrotrophy, and specializations for fetal nutrition in reptiles and other vertebrates. Am. Zool. 32, 313–321 (1992).
Google Scholar 
Buckley, D., Alcobendas, M., García-París, M. & Wake, M. H. Heterochrony, cannibalism, and the evolution of viviparity in Salamandra salamandra. Evol. Dev. 9, 105–115 (2007).PubMed 

Google Scholar 
Kusrini, M. D., Rowley, J. J. L., Khairunnisa, L. R., Shea, G. M. & Altig, R. The reproductive biology and larvae of the first tadpole-bearing frog, Limnonectes larvaepartus. PLoS ONE 10, e116154–e116159 (2015).ADS 
PubMed 
PubMed Central 

Google Scholar 
Lanza, B. & Leo, P. Sul primo caso sicuro di riproduzione vivipara nel genere Speleomantes. 1–54 (2000).Lunghi, E. et al. Comparative reproductive biology of European cave salamanders (Genus Hydromantes): nesting selection and multiple annual breeding. Salamandra 54, 101–108 (2018).
Google Scholar 
Liedtke, H. C. et al. Terrestrial reproduction as an adaptation to steep terrain in African toads. Proc. R. Soc. Lond. B 284, 20162598–20162599 (2017).
Google Scholar 
Wake, M. H. The reproductive biology of Eleutherodactylus jasperi (Amphibia, Anura, Leptodactylidae), with comments on the evolution of live-bearing systems. J. Herpetol. 12, 121–133 (1978).
Google Scholar 
Jennings, D. H. & Hanken, J. Mechanistic basis of life history evolution in anuran amphibians: thyroid gland development in the direct-developing frog, Eleutherodactylus coqui. Gen. Comp. Endocr. 111, 225–232 (1998).CAS 
PubMed 

Google Scholar 
Callery, E. M. & Elinson, R. P. Thyroid hormone-dependent metamorphosis in a direct developing frog. Proc. Natl Acad. Sci. USA 97, 2615–2620 (2000).ADS 
CAS 
PubMed 
PubMed Central 

Google Scholar 
Callery, E. M., Hung, F. & Elinson, R. P. Frogs without polliwogs: evolution of anuran direct development. BioEssays 23, 233–241 (2001).CAS 
PubMed 

Google Scholar 
Buckley, L. B. & Jetz, W. Environmental and historical constraints on global patterns of amphibian richness. Proc. R. Soc. Lond. B 274, 1167–1173 (2007).
Google Scholar 
Pyron, R. A. & Wiens, J. J. Large-scale phylogenetic analyses reveal the causes of high tropical amphibian diversity. Proc. R. Soc. Lond. B 280, 20131622 (2013).
Google Scholar 
Gómez-Rodríguez, C., Baselga, A. & Wiens, J. J. Is diversification rate related to climatic niche width? Glob. Ecol. Biogeogr. 24, 383–395 (2015).
Google Scholar 
Moen, D. S. & Wiens, J. J. Microhabitat and climatic niche change explain patterns of diversification among frog families. Am. Nat. 190, 29–44 (2017).PubMed 

Google Scholar 
Kozak, K. H. & Wiens, J. J. Accelerated rates of climatic-niche evolution underlie rapid species diversification. Ecol. Lett. 13, 1378–1389 (2010).PubMed 

Google Scholar 
Jaramillo, A. F. et al. Vastly underestimated species richness of Amazonian salamanders (Plethodontidae: Bolitoglossa) and implications about plethodontid diversification. Mol. Phylogenet. Evol. 149, 106841 (2020).PubMed 

Google Scholar 
Jetz, W. & Pyron, R. A. The interplay of past diversification and evolutionary isolation with present imperilment across the amphibian tree of life. Nat. Ecol. Evol. 2, 850–858 (2018).PubMed 

Google Scholar 
Bars-Closel, M., Kohlsdorf, T., Moen, D. S. & Wiens, J. J. Diversification rates are more strongly related to microhabitat than climate in squamate reptiles (lizards and snakes). Evolution 71, 2243–2261 (2017).PubMed 

Google Scholar 
Cyriac, V. P. & Kodandaramaiah, U. Digging their own macroevolutionary grave: fossoriality as an evolutionary dead end in snakes. J. Evolution. Biol. 31, 587–598 (2018).CAS 

Google Scholar 
Zamudio, K. R., Bell, R. C., Nali, R. C., Haddad, C. F. B. & Prado, C. P. A. Polyandry, predation, and the evolution of frog reproductive modes. Am. Nat. 188, S41–S61 (2016).PubMed 

Google Scholar 
Lion, M. B. et al. Global patterns of terrestriality in amphibian reproduction. Glob. Ecol. Biogeogr. 4, 679–13 (2019).
Google Scholar 
Müller, H. et al. Forests as promoters of terrestrial life-history strategies in East African amphibians. Biol. Lett. 9, 20121146–20121146 (2013).PubMed 
PubMed Central 

Google Scholar 
Velo-Antón, G., García-París, M., Galán, P. & Cordero Rivera, A. The evolution of viviparity in Holocene islands: ecological adaptation versus phylogenetic descent along the transition from aquatic to terrestrial environments. J. Zool. Syst. Evol. Res. 45, 345–352 (2007).
Google Scholar 
Liedtke, H. C. AmphiNom: an amphibian systematics tool. Syst. Biodivers. 17, 1–6 (2019).
Google Scholar 
R Core Team. R: a language and environment for statistical computing (R Foundation for Statistical Computing, Vienna, Austria, 2020). http://www.R-project.org/.IUCN. IUCN 2020. The IUCN Red List of Threatened Species. Version 2020-2. https://www.iucnredlist.org (2019).AmphibiaChina. The database of Chinese amphibians. Electronic Database (Kunming Institute of Zoology (CAS), Kunming, Yunnan, China, 2019) http://www.amphibiachina.org/.Ron, S. R., Yanez-Muñoz, M. H., Merino-Viteri, A. & Ortiz, D. A. Anfibios del Ecuador. Version 2019.0 (Museo de Zoología, Pontificia Universidad Católica del Ecuador, 2019). https://bioweb.bio/faunaweb/amphibiaweb.Greven, H. In Reproductive Biology and Phylogeny of Urodela 447–475 (Taylor & Francis, 2003).Marks, S. B. & Collazo, A. Direct development in Desmognathus aeneus (Caudata: Plethodontidae): a staging table. Copeia 1998, 637–648 (1998).
Google Scholar 
Müller, H., Loader, S. P., Ngalason, W., Howell, K. M. & Gower, D. J. Reproduction in brevicipitid frogs (Amphibia: Anura: Brevicipitidae)—evidence from Probreviceps m. macrodactylus. Copeia 2007, 726–733 (2007).
Google Scholar 
Velo-Antón, G., Santos, X., Sanmartín-Villar, I., Cordero-Rivera, A. & Buckley, D. Intraspecific variation in clutch size and maternal investment in pueriparous and larviparous Salamandra salamandra females. Evol. Ecol. 29, 185–204 (2014).
Google Scholar 
Beaulieu, J. M., O’Meara, B. C. & Donoghue, M. J. Identifying hidden rate changes in the evolution of a binary morphological character: the evolution of plant habit in campanulid angiosperms. Systematic biology 62, 725–737 (2013).Pupko, T., Pe’er, I., Shamir, R. & Graur, D. A fast algorithm for joint reconstruction of ancestral amino acid sequences. Mol. Biol. Evol. 17, 890–896 (2000).CAS 
PubMed 

Google Scholar 
Bollback, J. P. SIMMAP: stochastic character mapping of discrete traits on phylogenies. BMC Bioinforma. 7, 88 (2006).
Google Scholar 
Plummer, M., Best, N., Cowles, K. & Vines, K. CODA: convergence diagnosis and output analysis for MCMC. R. News 6, 7–11 (2006).
Google Scholar 
Tuffley, C. & Steel, M. Modeling the covarion hypothesis of nucleotide substitution. Math. Biosci. 147, 63–91 (1998).MathSciNet 
CAS 
PubMed 
MATH 

Google Scholar 
Xie, W., Lewis, P. O., Fan, Y., Kuo, L. & Chen, M. H. Improving marginal likelihood estimation for Bayesian phylogenetic model selection. Syst. Biol. 60, 150–160 (2011).PubMed 

Google Scholar 
Nakov, T., Beaulieu, J. M. & Alverson, A. J. Diatoms diversify and turn over faster in freshwater than marine environments. Evolution 73, 2497–2511 (2019).PubMed 

Google Scholar 
Etienne, R. S. et al. Diversity-dependence brings molecular phylogenies closer to agreement with the fossil record. Proc. R. Soc. Lond. B 279, 1300–1309 (2011).
Google Scholar  More

Williams, G. C. Natural selection, the costs of reproduction, and a refinement of lack’s principle. Am. Nat. 100, 687–690 (1966).
Google Scholar 
Stearns, S. C. The evolution of life histories. (Oxford University Press, 1992).Kirkwood, T. B. L. Evolution of ageing. Nature 270, 301–304 (1977).ADS 
CAS 
PubMed 

Google Scholar 
Partridge, L., Prowse, N. & Pignatelli, P. Another set of responses and correlated responses to selection on age at reproduction in Drosophila melanogaster. Proc. R. Soc. B. 266, 255–261 (1999).CAS 
PubMed 
PubMed Central 

Google Scholar 
Metcalfe, N. Growth versus lifespan: Perspectives from evolutionary ecology. Exp. Gerontol. 38, 935–940 (2003).PubMed 

Google Scholar 
Lee, W.-S., Monaghan, P. & Metcalfe, N. B. Experimental demonstration of the growth rate–lifespan trade-off. Proc. R. Soc. B. 280, 20122370 (2013).PubMed 
PubMed Central 

Google Scholar 
Lemaître, J.-F. et al. Early-late life trade-offs and the evolution of ageing in the wild. Proc. R. Soc. B. 282, 20150209 (2015).PubMed 
PubMed Central 

Google Scholar 
Jehan, C., Sabarly, C., Rigaud, T. & Moret, Y. Late-life reproduction in an insect: Terminal investment, reproductive restraint or senescence. J. Anim. Ecol. 90, 282–297 (2021).PubMed 

Google Scholar 
Pawelec, G. Age and immunity: What is “immunosenescence”?. Exp. Gerontol. 105, 4–9 (2018).CAS 
PubMed 

Google Scholar 
Schwenke, R. A., Lazzaro, B. P. & Wolfner, M. F. Reproduction–immunity trade-offs in insects. Annu. Rev. Entomol. 61, 239–256 (2016).CAS 
PubMed 

Google Scholar 
Maklakov, A. A. & Chapman, T. Evolution of ageing as a tangle of trade-offs: Energy versus function. Proc. R. Soc. B. 286, 20191604 (2019).CAS 
PubMed 
PubMed Central 

Google Scholar 
Hamel, S. et al. Fitness costs of reproduction depend on life speed: empirical evidence from mammalian populations: Fitness costs of reproduction in mammals. Ecol. Lett. 13, 915–935 (2010).PubMed 

Google Scholar 
Graham, A. L., Allen, J. E. & Read, A. F. Evolutionary causes and consequences of immunopathology. Annu. Rev. Ecol. Evol. Syst. 36, 373–397 (2005).
Google Scholar 
Sorci, G. & Faivre, B. Inflammation and oxidative stress in vertebrate host–parasite systems. Phil. Trans. R. Soc. B. 364, 71–83 (2009).PubMed 

Google Scholar 
Ashley, N. T., Weil, Z. M. & Nelson, R. J. Inflammation: Mechanisms, costs, and natural variation. Annu. Rev. Ecol. Evol. Syst. 43, 385–406 (2012).
Google Scholar 
Babin, A., Moreau, J. & Moret, Y. Storage of carotenoids in crustaceans as an adaptation to modulate immunopathology and optimize immunological and life history strategies. BioEssays 41, 1800254 (2019).
Google Scholar 
Vasto, S. et al. Inflammatory networks in ageing, age-related diseases and longevity. Mech. Ageing Dev. 128, 83–91 (2007).CAS 
PubMed 

Google Scholar 
Finch, C. E. & Crimmins, E. M. Inflammatory exposure and historical changes in human life-spans. Science 305, 1736–1739 (2004).ADS 
CAS 
PubMed 

Google Scholar 
Licastro, F. et al. Innate immunity and inflammation in ageing: A key for understanding age-related diseases. Immun. Ageing 2, 8 (2005).PubMed 
PubMed Central 

Google Scholar 
Pawelec, G., Goldeck, D. & Derhovanessian, E. Inflammation, ageing and chronic disease. Curr. Opin. Immunol. 29, 23–28 (2014).CAS 
PubMed 

Google Scholar 
Pursall, E. R. & Rolff, J. Immune responses accelerate ageing: Proof-of-principle in an insect model. PLoS ONE 6, e19972 (2011).ADS 
CAS 
PubMed 
PubMed Central 

Google Scholar 
Khan, I., Agashe, D. & Rolff, J. Early-life inflammation, immune response and ageing. Proc. R. Soc. B. 284, 20170125 (2017).PubMed 
PubMed Central 

Google Scholar 
Vigneron, A., Jehan, C., Rigaud, T. & Moret, Y. Immune defenses of a beneficial pest: The mealworm beetle, Tenebrio molitor. Front. Physiol. 10, 138 (2019).PubMed 
PubMed Central 

Google Scholar 
Jehan, C., Chogne, M., Rigaud, T. & Moret, Y. Sex-specific patterns of senescence in artificial insect populations varying in sex-ratio to manipulate reproductive effort. BMC Evol. Biol. 20, 18 (2020).PubMed 
PubMed Central 

Google Scholar 
Jehan, C., Sabarly, C., Rigaud, T. & Moret, Y. Age-specific fecundity under pathogenic threat in an insect: Terminal investment versus reproductive restraint. J. Anim. Ecol. 91, 101–111 (2022).PubMed 

Google Scholar 
Chung, K.-H. & Moon, M.-J. Fine structure of the hemopoietic tissues in the mealworm beetle, Tenebrio molitor. Entomol. Res. 34, 131–138 (2004).
Google Scholar 
Urbański, A., Adamski, Z. & Rosiński, G. Developmental changes in haemocyte morphology in response to Staphylococcus aureus and latex beads in the beetle Tenebrio molitor L.. Micron 104, 8–20 (2018).PubMed 

Google Scholar 
Vommaro, M. L., Kurtz, J. & Giglio, A. Morphological characterisation of haemocytes in the mealworm beetle Tenebrio molitor (Coleoptera, Tenebrionidae). Insects 12, 423 (2021).PubMed 
PubMed Central 

Google Scholar 
Söderhäll, K. & Cerenius, L. Role of the prophenoloxidase-activating system in invertebrate immunity. Curr. Opin. Immunol. 10, 23–28 (1998).PubMed 

Google Scholar 
Siva-Jothy, M. T., Moret, Y. & Rolff, J. Insect immunity: an evolutionary ecology perspective. in Advances in Insect Physiology vol. 32 1–48 (Elsevier, 2005).Nappi, A. J. & Ottaviani, E. Cytotoxicity and cytotoxic molecules in invertebrates. BioEssays 22, 469–480 (2000).CAS 
PubMed 

Google Scholar 
Sadd, B. M. & Siva-Jothy, M. T. Self-harm caused by an insect’s innate immunity. Proc. R. Soc. B. 273, 2571–2574 (2006).PubMed 
PubMed Central 

Google Scholar 
Daukšte, J., Kivleniece, I., Krama, T., Rantala, M. J. & Krams, I. Senescence in immune priming and attractiveness in a beetle: Immunosenescence in a beetle. J. Evol. Biol. 25, 1298–1304 (2012).PubMed 

Google Scholar 
Krams, I. et al. Trade-off between cellular immunity and life span in mealworm beetles Tenebrio molitor. Curr. Zool. 59, 340–346 (2013).
Google Scholar 
Moon, H. J., Lee, S. Y., Kurata, S., Natori, S. & Lee, B. L. Purification and molecular cloning of cDNA for an inducible antibacterial protein from larvae of the coleopteran, Tenebrio molitor. J. Biochem. 116, 53–58 (1994).CAS 
PubMed 

Google Scholar 
Lee, Y. J. et al. Structure and expression of the tenecin 3 gene in Tenebrio molitor. Biochem. Biophys. Res. Comm. 218, 6–11 (1996).CAS 
PubMed 

Google Scholar 
Kim, D. H. et al. Bacterial expression of tenecin 3, an insect antifungal protein isolated from Tenebrio molitor, and its efficient purification. Mol. Cells 8, 786–789 (1998).CAS 
PubMed 

Google Scholar 
Roh, K.-B. et al. Proteolytic cascade for the activation of the insect toll pathway induced by the fungal cell wall component. J. Biol. Chem. 284, 19474–19481 (2009).CAS 
PubMed 
PubMed Central 

Google Scholar 
Park, J.-W. et al. Beetle Immunity. in Invertebrate Immunity (ed. Söderhäll, K.) vol. 708 163–180 (Springer US, 2010).Chae, J.-H. et al. Purification and characterization of tenecin 4, a new anti-Gram-negative bacterial peptide, from the beetle Tenebrio molitor. Dev. Comp. Immunol. 36, 540–546 (2012).CAS 
PubMed 

Google Scholar 
Haine, E. R., Pollitt, L. C., Moret, Y., Siva-Jothy, M. T. & Rolff, J. Temporal patterns in immune responses to a range of microbial insults (Tenebrio molitor). J. Insect Physiol. 54, 1090–1097 (2008).CAS 
PubMed 

Google Scholar 
Dhinaut, J., Chogne, M. & Moret, Y. Immune priming specificity within and across generations reveals the range of pathogens affecting evolution of immunity in an insect. J. Anim. Ecol. 87, 448–463 (2018).PubMed 

Google Scholar 
Hoffmann, J. A., Reichhart, J.-M. & Hetru, C. Innate immunity in higher insects. Curr. Opin. Immunol. 8, 8–13 (1996).CAS 
PubMed 

Google Scholar 
Moret, Y. Explaining variable costs of the immune response: selection for specific versus non-specific immunity and facultative life history change. Oikos 102, 213–216 (2003).
Google Scholar 
Khan, I., Prakash, A. & Agashe, D. Immunosenescence and the ability to survive bacterial infection in the red flour beetle Tribolium castaneum. J. Anim. Ecol. 85, 291–301 (2016).PubMed 

Google Scholar 
Rolff, J. Effects of age and gender on immune function of dragonflies (Odonata, Lestidae) from a wild population. Can. J. Zool. 79, 2176–2180 (2001).
Google Scholar 
Doums, C., Moret, Y., Benelli, E. & Schmid-Hempel, P. Senescence of immune defence in Bombus workers. Ecol. Entomol. 27, 138–144 (2002).
Google Scholar 
Schmid, M. R., Brockmann, A., Pirk, C. W. W., Stanley, D. W. & Tautz, J. Adult honeybees (Apis mellifera L.) abandon hemocytic, but not phenoloxidase-based immunity. J. Insect Physiol. 54, 439–444 (2008).CAS 
PubMed 

Google Scholar 
Moret, Y. & Schmid-Hempel, P. Immune responses of bumblebee workers as a function of individual and colony age: senescence versus plastic adjustment of the immune function. Oikos 118, 371–378 (2009).
Google Scholar 
Armitage, S. A. O. & Boomsma, J. J. The effects of age and social interactions on innate immunity in a leaf-cutting ant. J. Insect Physiol. 56, 780–787 (2010).CAS 
PubMed 

Google Scholar 
Korner, P. & Schmid-Hempel, P. In vivo dynamics of an immune response in the bumble bee Bombus terrestris. J. Invert. Pathol. 87, 59–66 (2004).CAS 

Google Scholar 
Li, T., Yan, D., Wang, X., Zhang, L. & Chen, P. Hemocyte changes during immune melanization in Bombyx Mori infected with Escherichia coli. Insects 10, 301 (2019).PubMed Central 

Google Scholar 
Chase, M. R., Raina, K., Bruno, J. & Sugumaran, M. Purification, characterization and molecular cloning of prophenoloxidases from Sarcophaga bullata. Insect Biochem. Mol. Biol. 30, 953–967 (2000).CAS 
PubMed 

Google Scholar 
Kanost, M. R. & Gorman, M. J. Phenoloxidases in insect immunity. in Insect Immunology 69–96 (Elsevier, 2008).Sadd, B. M. et al. Modulation of sexual signalling by immune challenged male mealworm beetles (Tenebrio molitor L.): Evidence for terminal investment and dishonesty. J. Evol. Biol. 19, 321–325 (2006).CAS 
PubMed 

Google Scholar 
Gálvez, D. & Chapuisat, M. Immune priming and pathogen resistance in ant queens. Ecol. Evol. 4, 1761–1767 (2014).PubMed 
PubMed Central 

Google Scholar 
Armitage, S. A. O. & Siva-Jothy, M. T. Immune function responds to selection for cuticular colour in Tenebrio molitor. Heredity 94, 650–656 (2005).CAS 
PubMed 

Google Scholar 
Armitage, S. A. O., Thompson, J. J. W., Rolff, J. & Siva-Jothy, M. T. Examining costs of induced and constitutive immune investment in Tenebrio molitor. J. Evol. Biol. 16, 1038–1044 (2003).CAS 
PubMed 

Google Scholar 
Kokoza, V. A. et al. Transcriptional regulation of the mosquito vitellogenin gene via a blood meal-triggered cascade. Gene 274, 47–65 (2001).CAS 
PubMed 

Google Scholar 
Isaac, P. G. & Bownes, M. Ovarian and fat-body vitellogenin synthesis in Drosophila melanogaster. Europ. J. Biochem. 123, 527–534 (2005).
Google Scholar 
Hoffmann, J. A. The immune response of Drosophila. Nature 426, 33–38 (2003).ADS 
CAS 
PubMed 

Google Scholar 
Tzou, P. et al. Tissue-specific inducible expression of antimicrobial peptide genes in Drosophila surface epithelia. Immunity 13, 737–748 (2000).CAS 
PubMed 

Google Scholar 
Haine, E. R., Moret, Y., Siva-Jothy, M. T. & Rolff, J. Antimicrobial defense and persistent infection in insects. Science 322, 1257–1259 (2008).ADS 
CAS 
PubMed 

Google Scholar 
Moret, Y. & Siva-Jothy, M. T. Adaptive innate immunity? Responsive-mode prophylaxis in the mealworm beetle, Tenebrio molitor. Proc. R. Soc. B. 270, 2475–2480 (2003).CAS 
PubMed 
PubMed Central 

Google Scholar 
Du Rand, N. & Laing, M. D. Determination of insecticidal toxicity of three species of entomopathogenic spore-forming bacterial isolates against Tenebrio molitor L. (Coleoptera: Tenebrionidae). Afr. J. Microbiol. Res. 5, 2222–2228 (2011).
Google Scholar 
Jurat-Fuentes, J. L. & Jackson, T. Bacterial entomopathogens. In Insect Pathology 2nd edn (eds Kaya, H. & Vera, F.) 265–349 (Elsevier Academic Press, Cambridge, Mass, 2012).
Google Scholar 
Dhinaut, J., Balourdet, A., Teixeira, M., Chogne, M. & Moret, Y. A dietary carotenoid reduces immunopathology and enhances longevity through an immune depressive effect in an insect model. Sci. Rep. 7, 12429 (2017).ADS 
PubMed 
PubMed Central 

Google Scholar 
Moreau, J., Martinaud, G., Troussard, J.-P., Zanchi, C. & Moret, Y. Trans-generational immune priming is constrained by the maternal immune response in an insect. Oikos 121, 1828–1832 (2012).
Google Scholar 
Lee, H. S. et al. The pro-phenoloxidase of coleopteran insect, Tenebrio molitor, larvae was activated during cell clump/cell adhesion of insect cellular defense reactions. FEBS Lett. 444, 255–259 (1999).CAS 
PubMed 

Google Scholar 
Zanchi, C., Troussard, J.-P., Martinaud, G., Moreau, J. & Moret, Y. Differential expression and costs between maternally and paternally derived immune priming for offspring in an insect. J. Anim. Ecol. 80, 1174–1183 (2011).PubMed 

Google Scholar 
Moret, Y. ‘Trans-generational immune priming’: Specific enhancement of the antimicrobial immune response in the mealworm beetle, Tenebrio molitor. Proc. R. Soc. B. 273, 1399–1405 (2006).ADS 
CAS 
PubMed 
PubMed Central 

Google Scholar 
Dubuffet, A. et al. Trans-generational immune priming protects the eggs only against gram-positive bacteria in the mealworm beetle. PLoS Pathog. 11, e1005178 (2015).PubMed 
PubMed Central 

Google Scholar  More

ResultsSoil physiochemical propertiesThe preliminary status of the soil analyzed before the commencement of the field preparatory activities revealed that the soil was subtlety fertile with regard to the physical and chemical properties (Table 1).Table 1 Physicochemical properties of the soil.Full size tableAssessment of disease incidence at sowing dates during each year in the trial studyIn the trial study, very low and statistically significant (p  More

Bacterial density and growth potential in the rearing water were related to the microbial carrying capacityQuantifying the bacterial density in each tank verified that we obtained a higher bacterial load in the systems with added organic material. The bacterial density was, on average, 7.8× higher in the systems with high compared to low bacterial carrying capacity. This difference was particularly evident at 2 (34.8×, Kruskal–Wallis p = 0.0008) and 9 DPH (9.1×, Kruskal–Wallis p = 0.0007) (Fig. 1). The bacterial density increased throughout the experiment for the tanks with low microbial carrying capacity (treatment group MMS−, FTS−), reflecting increased larval feeding and defecation. Contrastingly, the bacterial density was relatively stable over time in the MMS+ treatment and even decreased over time in the FTS+ treatment. When averaging the densities at 11 and 15 DPH within each rearing treatment, we observed that the ‘MMS+ to FTS+’ had a considerable difference in the bacterial density between incoming and rearing water (24.2×). In contrast, this difference was below 8.2× in all other treatment tanks. Such differences in density indicated that some communities were below the microbial carrying capacity of the systems. We thus investigated the growth potential to determine if carrying capacity was reached in the rearing water.Figure 1Bacterial density (million bacterial cells mL−1) at various days post-hatching (DPH) in incoming and rearing tank water. Note that the y-axis is log scaled. Colours indicate the rearing treatment, and shape signifies rearing (filled circle) and incoming water (filled triangle).Full size imageThe bacterial net growth potential in the intake and rearing water was quantified as the number of cell doublings after incubation for 3 days11. Generally, the FTS− and MMS− rearing water had net growth potential with an average of 0.2 and 0.1, respectively (Supplementary Fig. 2). In contrast, the rearing water of the FTS+ and MMS+ had a negative net growth potential with averages of −0.2 and −0.06, respectively. In the case of negative net growth potential, the bacterial density decreased during the incubation. A negative net growth potential suggested that the rearing water bacterial communities were at the tank’s microbial carrying capacity at the time of sampling. Thus, the bacterial communities were at the carrying capacity of the high (+) carrying capacity systems and below in the low (−) systems. To gain a deeper understanding of the bacterial community characteristics the 16S rRNA gene of the bacterial community was sequenced at 1 and 9 DPH.Initial rearing condition did not leave a legacy effect on bacterial α-diversityThe bacterial α-diversity of the rearing water was investigated at 1 and 12 DPH (Fig. 2). At 1 DPH, the richness was comparable between the FTS−, FTS+ and MMS+ treatments, but on average, 1.5× higher for the MMS− treatment (307 vs 205 ASVs, Tukey’s test p  More

Hobbs, R. J. et al. Novel ecosystems: Theoretical and management aspects of the new ecological world order. Glob. Ecol. Biogeogr. 15, 1–7 (2006).
Google Scholar 
Kraft, N. J. B. et al. Community assembly, coexistence and the environmental filtering metaphor. Funct. Ecol. 29, 592–599 (2015).
Google Scholar 
Mayfield, M. M. et al. What does species richness tell us about functional trait diversity? Predictions and evidence for responses of species and functional trait diversity to land-use change. Glob. Ecol. Biogeogr. 19, 423–431 (2010).
Google Scholar 
Zobel, M. The species pool concept as a framework for studying patterns of plant diversity. J. Veg. Sci. 27, 8–18 (2016).
Google Scholar 
Birkhofer, K. et al. Land-use type and intensity differentially filter traits in above- and below-ground arthropod communities. J. Anim. Ecol. 86, 511–520 (2017).PubMed 

Google Scholar 
Temperton, V. M. Assembly Rules and Restoration Ecology: Bridging the Gap Between Theory and Practice (Island Press, 2004).
Google Scholar 
Flynn, D. F. B. et al. Loss of functional diversity under land use intensification across multiple taxa. Ecol. Lett. 12, 22–33 (2009).PubMed 

Google Scholar 
Gascon, C. et al. Matrix habitat and species richness in tropical forest remnants. Biol. Conserv. 91, 223–229 (1999).
Google Scholar 
Filloy, J., Zurita, G. A., Corbelli, J. M. & Bellocq, M. I. On the similarity among bird communities: Testing the influence of distance and land use. Acta Oecol. 36, 333–338 (2010).ADS 

Google Scholar 
Vaccaro, A., Filloy, J. & Bellocq, M. What land use better preserves the functional and taxonomic diversity of birds in a grassland biome?. Avian Conserv. Ecol. 14, 1 (2019).
Google Scholar 
Vaccaro, A. S. & Bellocq, M. I. Diversidad taxonómica y funcional de aves: Diferencias entre hábitats antrópicos en un bosque subtropical. Ecol. Austral 29, 391–404 (2019).
Google Scholar 
Sekercioglu, C. H. Bird functional diversity and ecosystem services in tropical forests, agroforests and agricultural areas. J. Ornithol. 153, 153–161 (2012).
Google Scholar 
Zurita, G. A. & Bellocq, M. I. Bird assemblages in anthropogenic habitats: Identifying a suitability gradient for native species in the Atlantic Forest. Biotropica 44, 412–419 (2012).
Google Scholar 
Azpiroz, A. B. et al. Ecology and conservation of grassland birds in southeastern South America: A review. J. Field Ornithol. 83, 217–246 (2012).
Google Scholar 
Devictor, V. et al. Spatial mismatch and congruence between taxonomic, phylogenetic and functional diversity: The need for integrative conservation strategies in a changing world. Ecol. Lett. 13, 1030–1040 (2010).PubMed 

Google Scholar 
Faith, D. P. Conservation evaluation and phylogenetic diversity. Biol. Conserv. 61, 1–10 (1992).
Google Scholar 
Corbelli, J. M. et al. Integrating taxonomic, functional and phylogenetic beta diversities: Interactive effects with the biome and land use across taxa. PLoS ONE 10, 1–17 (2015).
Google Scholar 
Purschke, O. et al. Contrasting changes in taxonomic, phylogenetic and functional diversity during a long-term succession: Insights into assembly processes. J. Ecol. 101, 857–866 (2013).
Google Scholar 
Srivastava, D. S., Cadotte, M. W., Macdonald, A. A. M., Marushia, R. G. & Mirotchnick, N. Phylogenetic diversity and the functioning of ecosystems. Ecol. Lett. 15, 637–648 (2012).PubMed 

Google Scholar 
Cavender-Bares, J., Kozak, K. H., Fine, P. V. A. & Kembel, S. W. The merging of community ecology and phylogenetic biology. Ecol. Lett. 12, 693–715 (2009).PubMed 

Google Scholar 
Mouquet, N. et al. Ecophylogenetics: Advances and perspectives. Biol. Rev. 87, 769–785 (2012).PubMed 

Google Scholar 
Ackerly, D. D., Schwilk, D. W. & Webb, C. O. Niche evolution and adaptive radiation: Testing the order of trait divergence. Ecology 87, S50–S61 (2006).CAS 
PubMed 

Google Scholar 
Cavender-Bares, J., Ackerly, D. D., Baum, D. A. & Bazzaz, F. A. Phylogenetic overdispersion in Floridian oak communities. Am. Nat. 163, 823–843 (2004).CAS 
PubMed 

Google Scholar 
Losos, J. B. et al. Niche lability in the evolution of a Caribbean lizard community. Nature 424, 542–545 (2003).ADS 
CAS 
PubMed 

Google Scholar 
Stevens, R. D., Gavilanez, M. M., Tello, J. S. & Ray, D. A. Phylogenetic structure illuminates the mechanistic role of environmental heterogeneity in community organization. J. Anim. Ecol. 81, 455–462 (2012).PubMed 

Google Scholar 
García-Navas, V. & Thuiller, W. Farmland bird assemblages exhibit higher functional and phylogenetic diversity than forest assemblages in France. J. Biogeogr. 47, 2392–2404 (2020).
Google Scholar 
Henwood, W. D. Toward a strategy for the conservation and protection of the world’s temperate grasslands. Univ. Neb. Press 20, 121–134 (2010).ADS 

Google Scholar 
Myers, N., Mittermeier, R. A., Mittermeier, C. G., da Fonseca, G. A. B. & Kent, J. Biodiversity hotspots for conservation priorities. Nature 403, 853–858 (2000).ADS 
CAS 
PubMed 

Google Scholar 
Landi, M., Oesterheld, M. & Deregibus, V. A. Manual de especies forrajeras de los pastizales naturales de Entre Ríos (1987).Viglizzo, E. F. et al. Ecological lessons and applications from one century of low external-input farming in the pampas of Argentina. Agric. Ecosyst. Environ. 83, 65–81 (2001).
Google Scholar 
Galindo Leal, C. & de Gusmão Câmara, I. The Atlantic Forest of South America: Biodiversity Status, Threats and Outlook (Island Press, 2003).
Google Scholar 
Oliveira-Filho, A. T. & Fontes, M. A. L. Patterns of floristic differentiation among Atlantic Forests in Southeastern Brazil and the influence of climate. Biotropica 32, 793–810 (2000).
Google Scholar 
DeGraaf, R. M., Geis, A. D. & Healy, P. A. Bird population and habitat surveys in urban areas. Landsc. Urban Plan. 21, 181–188 (1991).
Google Scholar 
Ralph, C. J. et al. Manual de métodos de campo para el monitoreo de aves terrestres. Pacific Southwest Research Station, Forest Service, U.S. Department of Agriculture, Albany, CA 46 http://www.srs.fs.usda.gov/pubs/31462. https://doi.org/10.3145/epi.2006.jan.15 (1996).Bibby, C., Jones, M. & Marsden, S. Expedition field techniques: Bird surveys. in (ed. Society, R. G.) (1998).Zurita, G. A. & Bellocq, M. I. Spatial patterns of bird community similarity: Bird responses to landscape composition and configuration in the Atlantic forest. Landsc. Ecol. 25, 147–158 (2010).
Google Scholar 
Koper, N. & Schmiegelow, F. K. K. A multi-scaled analysis of avian response to habitat amount and fragmentation in the Canadian dry mixed-grass prairie. Landsc. Ecol. 21, 1045 (2006).
Google Scholar 
Xeno-canto-Foundation. Xeno-canto website. https://www.xeno-canto.org (2018).Petchey, O. L. & Gaston, K. J. Functional diversity (FD), species richness and community composition. Ecol. Lett. 5, 402–411 (2002).
Google Scholar 
Jetz, W., Thomas, G. H., Joy, J. B., Hartmann, K. & Mooers, A. O. The global diversity of birds in space and time. Nature 491, 444–448 (2012).ADS 
CAS 
PubMed 

Google Scholar 
Revell, L. J. phytools: An R package for phylogenetic comparative biology (and other things). Methods Ecol. Evol. 3, 217–223 (2012).
Google Scholar 
R Core Team. R: A Language and Environment for Statistical Computing. R Foundation for Statistical Computing. https://www.r-project.org (2018).Kembel, S. W. et al. Picante: R tools for integrating phylogenies and ecology. Bioinformatics 26, 1463–1464 (2010).CAS 
PubMed 

Google Scholar 
Webb, C. O., Ackerly, D. D. & Kembel, S. W. Phylocom: Software for the analysis of phylogenetic community structure and trait evolution. Bioinformatics 24, 2098–2100 (2008).CAS 
PubMed 

Google Scholar 
Pinheiro, J., Bates, D., DebRoy, S. & Sarkar, D. R Core Team. nlme: Linear and Nonlinear mixed effects models. R package version 3.1–117. (2014).Lenth, R. V. Least-Squares Means: The R Package lsmeans. J. Stat. Softw. https://doi.org/10.18637/jss.v069.i01 (2016).Article 

Google Scholar 
Cadotte, M. W. & Tucker, C. M. Should environmental filtering be abandoned?. Trends Ecol. Evol. 32, 429–437 (2017).PubMed 

Google Scholar 
Concepción, E. D. et al. Contrasting trait assembly patterns in plant and bird communities along environmental and human-induced land-use gradients. Ecography 40, 753–763 (2016).
Google Scholar 
Cerezo, A., Conde, M. C. & Poggio, S. L. Pasture area and landscape heterogeneity are key determinants of bird diversity in intensively managed farmland. Biodivers. Conserv. 20, 2649–2667 (2011).
Google Scholar 
Pretelli, M. G., Isacch, J. P. & Cardoni, D. A. Year-round abundance, richness and nesting of the bird assemblage of tall grasslands in the south-east Pampas region, Argentina. Ardeola 60, 327–343 (2013).
Google Scholar 
Molinari, R. L. Biografía de la Pampa: 4 siglos de historia del campo argentino (Fundación Colombina “V Centenario,” 1987).
Google Scholar 
Filloy, J. & Bellocq, M. I. Patterns of bird abundance along the agricultural gradient of the Pampean region. Agric. Ecosyst. Environ. 120, 291–298 (2007).
Google Scholar 
Le Viol, I. et al. More and more generalists: Two decades of changes in the European avifauna. Biol. Lett. 8, 780–782 (2012).PubMed 
PubMed Central 

Google Scholar 
Concepción, E. D., Moretti, M., Altermatt, F., Nobis, M. P. & Obrist, M. K. Impacts of urbanisation on biodiversity: The role of species mobility, degree of specialisation and spatial scale. Oikos 124, 1571–1582 (2015).
Google Scholar 
Emerson, B. C. & Gillespie, R. G. Phylogenetic analysis of community assembly and structure over space and time. Trends Ecol. Evol. 23, 619–630 (2008).PubMed 

Google Scholar 
Morse, N. B. et al. Novel ecosystems in the Anthropocene: A revision of the novel ecosystem concept for pragmatic applications. Ecol. Soc. 19, 12 (2014).
Google Scholar 
Loyn, R. H., McNabb, E. G., Macak, P. & Noble, P. Eucalypt plantations as habitat for birds on previously cleared farmland in south-eastern Australia. Biol. Conserv. 137, 533–548 (2007).
Google Scholar 
Marsden, S., Whiffin, M. & Galetti, M. Bird diversity and abundance in forest fragments and Eucalyptus plantations around an Atlantic forest reserve, Brazil. Biodivers. Conserv. 10, 737–751 (2001).
Google Scholar 
Zurita, G. A., Rey, N., Varela, D. M., Villagra, M. & Bellocq, M. I. Conversion of the Atlantic Forest into native and exotic tree plantations: Effects on bird communities from the local and regional perspectives. For. Ecol. Manag. 235, 164–173 (2006).
Google Scholar 
Flynn, D. F. B., Mirotchnick, N., Jain, M., Palmer, M. I. & Naeem, S. Functional and phylogenetic diversity as predictors of biodiversity ecosystem-function. Ecology 92, 1573–1581 (2011).PubMed 

Google Scholar 
Sol, D. et al. The worldwide impact of urbanisation on avian functional diversity. Ecol. Lett. 23, 962–972 (2020).PubMed 

Google Scholar 
Webb, C. O., Ackerly, D. D., McPeek, M. A. & Donoghue, M. J. Phylogenies and community ecology. Annu. Rev. Ecol. Syst. 33, 475–505 (2002).
Google Scholar 
Palacio, F. X., Ibañez, L. M., Maragliano, R. E. & Montalti, D. Urbanization as a driver of taxonomic, functional, and phylogenetic diversity losses in bird communities. Can. J. Zool. 96, 1114–1121 (2018).
Google Scholar 
Sol, D., Bartomeus, I., González-Lagos, C. & Pavoine, S. Urbanisation and the loss of phylogenetic diversity in birds. Ecol. Lett. 20, 721–729 (2017).PubMed 

Google Scholar 
Luck, G. W., Carter, A. & Smallbone, L. Changes in bird functional diversity across multiple land uses: Interpretations of functional redundancy depend on functional group identity. PLoS ONE 8, e63671 (2013).ADS 
CAS 
PubMed 
PubMed Central 

Google Scholar 
Coetzee, B. W. T. & Chown, S. L. Land-use change promotes avian diversity at the expense of species with unique traits. Ecol. Evol. 6, 7610–7622 (2016).PubMed 
PubMed Central 

Google Scholar  More