ArticlePDF Available

First record of heterospecific amplexus behaviour between Pelophylax plancyi (Lataste, 1880) and P. nigromaculatus (Hallowell, 1861) from Nanjing, China


Abstract and Figures

This publication reports the first documented record of heterospecific amplexus between Pelophylax plancyi and P. nigromaculatus from Nanjing, China. Herpetology Notes, volume 14: 773-774 (2021) (published online on 16 May 2021)
Content may be subject to copyright.
During mating, adult frogs generally recognise
conspecifics through visual displays, audible calls
or chemical signals (Belanger and Corkum, 2009).
However, communication cues are not always efficient
and mixed-species amplexus are not uncommon
(Haddad et al., 1994). Non-conspecific male and female
amplexus include not only heterospecific amplexus
but also multiple individual amplexus (Oliveira et al.,
2014), amplexus between two males (Theis and Caldart,
2015), amplexus by anurans with caudates (Simović, et
al., 2014), and even amplexus with reptiles (Shin et al.,
In natural ecosystems, cases of heterospecific
amplexus have been documented in various amphibian
species (Sodré et al., 2014, Vásquez Cruz et al., 2019).
Within the Pelophylax genus, Marsh frogs P. ridibundus
(Pallas, 1771), Pool frogs P. lessonae (Camerano,
1882) and their hybrids the Edible frogs P. esculentus
(Linnaeus, 1758), represents a suitable model to study
hybridisation (Spolsky and Uzzell, 1984). However,
relatively small populations are at risk of losing genetic
information due to introgressive hybridization (Zhang
et al. 2018).
Chinese species of the genus Pelophylax include the
Black-spotted Pond frog, P. nigromaculatus (Hallowell,
1861) and the Gold-striped Pond frog, P. plancyi (Lataste,
1880), two common species that generally share their
breeding habitats in spring (Liu et al., 2010). While
heterospecific amplexus have not been reported yet,
genetic analyses suggest their occurrence as hybrids are
present in large numbers in some populations (Komaki
et al., 2015). Here we present a case of heterospecific
amplexus between P. plancyi and P. nigromaculatus.
On 1 May 2019, around 19:15 h, we observed a
male P. plancyi in axillary amplexus with a female P.
nigromaculatus (Fig. 1) at Xuanwu Lake in Nanjing,
People’s Republic of China (32.0583°N, 118.8194°E).
At this site, air temperature and relative humidity were
30 °C and 76% respectively. The pair was observed
in amplexus for about 15 minutes before they left
towards a nearby rocky pool. We captured both frogs
before they could lay eggs, and we measured their
snout-vent length (SVL) using a digital calliper to the
nearest 0.01mm. The SVL of P. plancyi was 36.50
mm and for P. nigromaculatus it was 62.21 mm. At
the same site, we also observed several P. plancyi in
amplexus. However, we did not observe any amplexus
of P. nigromaculatus despite being within the breeding
season. This heterospecific amplexus has the potential
to result in the hybridization.
Herpetology Notes, volume 14: 773-774 (2021) (published online on 16 May 2021)
First record of heterospecific amplexus behaviour between
Pelophylax plancyi (Lataste, 1880) and P. nigromaculatus
(Hallowell, 1861) from Nanjing, China
Hina Amin1, Kevin R. Messenger1,*, and Amaël Borzée1,*
1 College of Biology and the Environment, Nanjing Forestry
University, Jiangsu, Nanjing 210037, People’s Republic of
* Corresponding authors. E-mail:;
© 2021 by Herpetology Notes. Open Access by CC BY-NC-ND 4.0.
Figure 1. Heterospecific amplexus between a male Gold-
striped pond frog (Pelophylax plancyi) and a female Black-
spotted pond frog (Pelophylax nigromaculatus) observed at
Xuanwu Lake, Nanjing, China in May 2019 (Photo by Hina
Hina Amin et al.
Acknowledgments. We are grateful to Dr. Ming-Feng Chuang
for comments on an earlier version of the manuscript
Belanger, R.M., Corkum, L.D. (2009): Review of aquatic sex
pheromones and chemical communication in anurans. Journal
of Herpetology 43: 184–191.
Haddad, C.F.B., Pombal-Júnior, J.P., Batistic, R.F. (1994): Natural
hybridization between diploid and tetraploid species of leaf-
frogs, genus Phyllomedusa (Amphibia). Journal of Herpetology
28: 425–430.
Komaki, S., Igawa, T., Lin, S.-M., Tojo, K., Sumida, M. (2015):
Robust molecular phylogeny and palaeodistribution modelling
resolve a complex evolutionary history: Glacial cycling drove
recurrent mtDNA introgression among Pelophylax frogs in East
Asia. Journal of Biogeography 42: 2159–2171.
Liu, K., Wang, F., Chen, W., Tu, L., Min, M.-S., Bi, K. , Fu, J. (2010):
Rampant historical mitochondrial genome introgression between
two species of green pond frogs, Pelophylax nigromaculatus
and P. Plancyi. BMC Evolutionary Biology 10: 201.
Oliveira, V.F., Dias, T.M., Santos, T.G. (2014): Multiple amplexus
and spawning in the leaf frog Phyllomedusa iheringii (Hylidae,
Phyllomedusinae). Herpetology Notes 7: 119–120.
Spolsky, C., Uzzell, T. (1984): Natural interspecies transfer of
mitochondrial DNA in amphibians. Proceedings of the National
Academy of Sciences of the United States of America 81: 5802–
Simović, A., Anderson, N., Marko, A., Sladjana, G., Nikolić, S.
(2014): Unusual amplexuses between anurans and caudates.
Herpetology Notes 7: 25–29.
Sodré, D., Martins, A.A.V., Vallinoto, M. (2014): Heterospecific
amplexus between the frog Leptodactylus macrosternum
(Anura: Leptodactylidae) and the toad Rhinella cf. granulosa
(Anura: Bufonidae). Herpetology Notes 7: 287–288
Shin, Y., Ambu, J., Borzée, A. (2020): Observations on heterospecific
amplexus in asiatic toads (Anura: Bufonidae: Bufo gargarizans)
in the republic of korea. Herpetology Notes 13: 411–413.
Theis, T.F., Caldart, V.M. (2015): Multiple interspecific amplexus
between a male of the invasive Bullfrog Lithobates catesbeianus
(Ranidae) and two males of the Cururu toad Rhinella icterica
(Bufonidae). Herpetology Notes 8: 449–451.
Vásquez Cruz, V., Canseco-Márquez, L., Pérez-Gámez, E. (2019):
Two incidents of heterospecific amplexus involving mexican
treefrogs (Smilisca baudini), a rio grande leopard frog (Lithobates
berlandieri), and a morelet’s leaf frog (Agalychnis moreletii)
(Amphibia: Anura: Hylidae, Ranidae, and Phyllomedusidae).
IRCF Reptiles & Amphibians 26: 140-–141.
Zhang, Q.-P., Hu, W.-F., Zhou, T.-T., Kong, S.-S., Liu, Z.-F.,
Zheng, R.-Q. (2018): Interspecies introgressive hybridization
in spiny frogs Quasipaa (family Dicroglossidae) revealed by
analyses on multiple mitochondrial and nuclear genes. Ecology
and Evolution 8: 1260–1270.
Accepted by Kanto Nishikawa
... This can lead to unusual amplexus, such as with dead individuals (Bettaso et al., 2008;Müller, 2016), with inanimate objects (Díaz-Ricaurte & Serrano, 2022;Mollov et al., 2010) or more often with individuals of other species-'interspecific amplexus' (e.g. Amin et al., 2021;Luría-Manzano & Vázquez-Córzas, 2011). This behaviour is still poorly understood, but it is likely based on hasty responses to chemical and visual cues and promoted by low densities of female conspecifics and strong intraspecific male competition (Hettyey & Pearman, 2003;Marco & Lizana, 2002). ...
Full-text available
Even though amphibians are diverse and abundant in most habitats, data on the life history of many species are still scarce. This is especially evident for African amphibians. Amplexus, the mating position in which a male grasps a female, is an important aspect of anuran reproduction. Herein, we report five new cases of interspecific amplexus among frogs in South Africa and compile previously reported events in sub-Saharan Africa.
Full-text available
Amphibians are one of the most species‐rich vertebrate taxa, with diverse and complex reproductive behaviors. Even though mate recognition plays a vital role in reproductive success, unusual amplexus with non‐suitable mates (misdirected amplexus) have been reported to occur in the wild. Misdirected amplexus may decrease fitness thus likely having ecological and evolutionary consequences and their frequency might increase with human‐induced changes in habitat. However, it is still unclear what promotes this unusual behavior and how widespread it is at spatial and phylogenetic scales. To fill this gap, we compiled a global database of misdirected amplexus events from literature and personal observations, with detailed information on environmental, geographical and behavioral aspects of reported events. Furthermore, we discuss its spatial, temporal and phylogenetic patterns. We provide a data set of 378 misdirected amplexus events for 156 amplectant species distributed across 69 genera and 18 families distributed in 52 countries in all continents except Antarctica. We collected data published or collected during the last 100 years, from 1920 to 2020, with information on the hour, month and year of the misdirected amplexus events. We recorded a total of 282 interspecific amplexus, 46 necrophiliac amplexus and 50 amplexus with objects or non‐amphibian species, with USA and Brazil being the countries with the highest number of records. Misdirected amplexus did not occur equally in all zoogeographical realms, with most events occurring in the Neotropical and Nearctic realms. Interspecific amplexus was especially represented in the Neotropical and Mediterranean regions. Most events involved three species‐rich globally‐distributed families (Bufonidae, Ranidae and Hylidae) but misdirected amplexus was widespread in the phylogeny. We provide a comprehensive data set of misdirected amplexus for anurans, with our results showing that it occurs extensively in the wild and across the globe. This data set provides a baseline for understanding misdirected amplexus and their spatial, temporal and phylogenetic patterns. Likewise, this data set offers a baseline to test the environmental, ecological and ethological drivers behind this reproductive behavior and we encourage researchers to report detailed observations of misdirected amplexus to better understand this process and its potential costs at the individual and population levels. The data are not copyright restricted; this data paper should be cited when data are used for publication, and the authors would appreciate being notified of research projects or teaching purposes when these data are used.
Full-text available
Introgression may lead to discordant patterns of variation among loci and traits. For example, previous phylogeographic studies on the genus Quasipaa detected signs of genetic introgression from genetically and morphologically divergent Quasipaa shini or Quasipaa spinosa. In this study, we used mitochondrial and nuclear DNA sequence data to verify the widespread introgressive hybridization in the closely related species of the genus Quasipaa, evaluate the level of genetic diversity, and reveal the formation mechanism of introgressive hybridization. In Longsheng, Guangxi Province, signs of asymmetrical nuclear introgression were detected between Quasipaa boulengeri and Q. shini. Unidirectional mitochondrial introgression was revealed from Q. spinosa to Q. shini. By contrast, bidirectional mitochondrial gene introgression was detected between Q. spinosa and Q. shini in Lushan, Jiangxi Province. Our study also detected ancient hybridizations between a female Q. spinosa and a male Q. jiulongensis in Zhejiang Province. Analyses on mitochondrial and nuclear genes verified three candidate cryptic species in Q. spinosa, and a cryptic species may also exist in Q. boulengeri. However, no evidence of introgressive hybridization was found between Q. spinosa and Q. boulengeri. Quasipaa exilispinosa from all the sampling localities appeared to be deeply divergent from other communities. Our results suggest widespread introgressive hybridization in closely related species of Quasipaa and provide a fundamental basis for illumination of the forming mechanism of introgressive hybridization, classification of species, and biodiversity assessment in Quasipaa.
Full-text available
Sympatric populations of the leaf-frogs Phyllomedusa distincta (2n = 26) and P. tetraploidea (4n = 52) were studied in southeastern Brazil. In a region of sympatry, seven of 15 leaf-frogs were triploid hybrids (3n = 39). The advertisement calls of both species are similar, and may not function adequately as a premating isolation mechanism. Triploids apparently exhibit low fertility or sterility, supporting the assertion that diploid and tetraploid populations are valid species. We suggest that P. tetraploidea originated by autopolyploidy of P. distincta, based on the following evidence: (1) the indistinguishable vocalizations of diploid and tetraploid species, and (2) the geographic distribution of species in the Phyllomedusa burmeisteri group.
Full-text available
Chemical communication is important in aquatic environments, particularly where visual and acoustical signals are limited. Both larval (tadpole) and adult anurans (frogs and toads) use waterborne chemical signals for many activities. Adult anurans commonly rely on acoustical communication for sex recognition and mating; however, a growing body of evidence suggests that anurans also may use aquatic sex pheromones for localization of potential mates. We provide an overview of how the anuran nasal cavity reorganizes during metamorphosis from the larval to the adult stage. Also, we focus on the behavior of reproductive anurans in response to chemical information detected by olfaction of waterborne chemical cues. Overall, we synthesize the current literature on anuran sex pheromones and chemical communication in the aquatic environment.
Full-text available
Mitochondrial introgression may result in the mitochondrial genome of one species being replaced by that of another species without leaving any trace of past hybridization in its nuclear genome. Such introgression can confuse the species genealogy estimates and lead to absurd inferences of species history. We used a phylogenetic approach to explore the potential mitochondrial genome introgression event(s) between two closely related green pond frog species, Pelophylax nigromaculatus and P. plancyi. DNA sequence data of one mitochondrial and two nuclear genes from an extensive sampling of the two species were collected, and the genealogies of the three genes were constructed and compared. While the two nuclear genes congruently showed mutual reciprocal monophyly of both species, the mitochondrial phylogeny separated a Korean P. nigromaculatus clade, a paraphyletic central China P. plancyi assemblage, and a large well-supported introgression clade. Within the introgression clade, the mitochondrial haplotypes of the two species were mixed together. This reticulated pattern can be most parsimoniously explained by an ancient mitochondrial introgression event from P. plancyi to P. nigromaculatus that occurred at least 1.36 MYA, followed by multiple recent introgression events from P. nigromaculatus back to P. plancyi within the last 0.63 MY. The re-constitution of previously co-adapted genomes in P. plancyi may be responsible for the recent rampant introgression events. The Korean P. nigromaculatus clade likely represents the only surviving "true" mitochondrial lineage of P. nigromaculatus, and the central China P. plancyi assemblage likely represents the "original" P. plancyi mitochondrial lineage. Refugia in the Korean Peninsula and central China may have played a significant role in preserving these ancient lineages. The majority of individuals in the two species have either introgressed (P. nigromaculatus) or reclaimed (P. plancyi) mitochondrial genomes while no trace of past hybridization in their nuclear genomes was detected. Asymmetrical reproductive ability of hybrids and continuous backcrossing are likely responsible for the observed mitochondrial introgression. This case is unique in that it includes an ancient "forward" introgression and many recent "backward" introgressions, which re-constitutes the original nuclear and mitochondrial genomes of P. plancyi. This hybrid system provides an excellent opportunity to study cyto-nuclear interaction and co-adaptation.
Full-text available
mtDNAs of two Central European water frog species, Rana ridibunda and Rana lessonae, were examined by electrophoresis of restriction enzyme fragments. Two types of mtDNA occur in R. ridibunda. One shares with mtDNA of R. lessonae 25.8% of 132 fragments generated by 19 enzymes, corresponding to a nucleotide sequence divergence of 8.1%; the other has diverged from R. lessonae mtDNA by only 0.3%. This latter type is a variant R. lessonae mtDNA that has been transferred into R. ridibunda; the introgression may have occurred via the hybridogenetic hybrid lineages collectively known as Rana esculenta. Of 37 R. ridibunda from Poland, 59% had the typical R. ridibunda mtDNA; 41% had the modified R. lessonae mtDNA as did a single individual from Switzerland (introduced). A single R. ridibunda from Turkey, outside the present range of R. lessonae, had the typical R. ridibunda mtDNA phenotype. Discordancies between inheritance of mitochondrial and nuclear genomes point up the danger of relying on a single molecular feature in reconstructing phylogeny. In addition, studies of mtDNA provide otherwise inaccessible information on complex evolutionary histories of closely related species. A knowledge of these complexities is important to an understanding of phylogenetic relationships and of the genetic processes that underlie the evolution of clonal taxa.
Pelophylax frogs in East Asia provide an opportunity to explore the impact of glacial cycling on demographic and genetic dynamics, because it has been suggested that they experienced distribution shifts and subsequent mtDNA introgression from Pelophylax plancyi to Pelophylax nigromaculatus in association with climatic oscillations. However, their evolutionary history, including the pattern of introgression, is incompletely understood. We used phylogenetic analyses based on multiple markers to address their evolutionary history, and palaeodistribution modelling to test whether the predicted distribution can explain the pattern of introgression suggested by molecular phylogenetics.