Research Article |
Corresponding author: Matthias Glaubrecht ( matthias.glaubrecht@uni-hamburg.de ) Academic editor: Andreas Schmidt-Rhaesa
© 2018 Nuanpan Veeravechsukij, Duangduen Krailas, Suluck Namchote, Benedikt Wiggering, Marco T. Neiber, Matthias Glaubrecht.
This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Citation:
Veeravechsukij N, Krailas D, Namchote S, Wiggering B, Neiber MT, Glaubrecht M (2018) Molecular phylogeography and reproductive biology of the freshwater snail Tarebia granifera in Thailand and Timor (Cerithioidea, Thiaridae): morphological disparity versus genetic diversity. Zoosystematics and Evolution 94(2): 461-493. https://doi.org/10.3897/zse.94.28981
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The freshwater thiarid gastropod Tarebia granifera (Lamarck, 1816), including taxa considered either congeneric or conspecific by earlier authors, is widespread and abundant in various lentic and lotic water bodies in mainland and insular Southeast Asia, with its range extending onto islands in the Indo-West-Pacific. This snail is, as one of the most frequent and major first intermediate host, an important vector for digenic trematodes causing several human diseases. As a typical thiarid T. granifera is viviparous and parthenogenetic, with various embryonic stages up to larger shelled juveniles developing within the female’s subhemocoelic (i.e non-uterine) brood pouch. Despite the known conchological disparity in other thiarids as well as this taxon, in Thailand Tarebia has been reported with the occurrence of one species only. In light of the polytypic variations found in shell morphology of freshwater snails in general and this taxon in particular, the lack of a modern taxonomic-systematic revision, using molecular genetics, has hampered more detailed insights to date, for example, into the locally varying trematode infection rates found in populations of Tarebia from across its range in Thailand as well as neighboring countries and areas. Here, we integrate evidence from phylogeographical analyses based on phenotypic variation (shell morphology, using biometry and geometric morphometrics) with highly informative and heterogeneous mtDNA sequence data (from the gene fragments cytochrome c oxidase subunit 1 and 16 S rRNA). We evaluate both the morphological and molecular genetic variation (using several phylogenetic analyses, including haplotype networks and a dated molecular tree), in correlation with differences in the reproductive biology among populations of Tarebia from various water bodies in the north, northwest, central, and south of Thailand, supplementing our respective analyses of parasite infections of this thiarid by cercaria of 15 trematode species, reported in a parallel study. Based on the comparison of topotypical material from the island of Timor, with specimens from 12 locations as reference, we found significant, albeit not congruent variation of both phenotype and genotype in Tarebia granifera, based on 1,154 specimens from 95 Thai samples, representing a geographically wide-ranging, river-based cross-section of this country. Our analyses indicate the existence of two genetically distinct clades and hint at possible species differentiation within what has been traditionally considered as T. granifera. These two lineages started to split about 5 mya, possibly related to marine transgressions forming what became known as biogeographical barrier north of the Isthmus of Kra. Grounded on the site-by-site analysis of individual Tarebia populations, our country-wide chorological approach focussing on the conchologically distinct and genetically diverse lineages of Tarebia allows to discuss questions of this either reflecting subspecific forms versus being distinct species within a narrowly delimited species complex. Our results, therefore, provide the ground for new perspectives on the phylogeography, evolution and parasitology of Thai freshwater gastropods, exemplified here by these highly important thiarids.
chorology, conchological variation, biometry, geometric morphometrics, molecular genetics, viviparity, parthenogenesis, Isthmus of Kra
Thailand is situated in one of the most biodiverse areas of the world (e.g.
In addition, Thailand can be divided into geographical regions based on distinct drainage basins; with those in the north, for example, forming the Chao Phraya drainage flowing into the Gulf of Thailand, those in the northeast as part of the Mekong river basin which eventually drains into the South China Sea, or the north-western region as part of the Salween river system. In contrast to these and other major river systems, in the south there are shorter rivers that either run east to the Gulf of Thailand or west to the Andaman Sea. These water bodies in Thailand form hotspots of aquatic biodiversity with various local endemism.
Among the aquatic biota, limnic molluscs are diverse, and include about 280 species of fresh and brackish water gastropods (
Accordingly, non-marine molluscs in Thailand should receive more attention and focus on studies looking into species diversity and contributing to solving fundamental questions and the evolution of faunal diversity. However, biological information on gastropods in Thai river systems and lakes is generally scarce and often lacks recently collected material or available former museum collections which hampers more in-depth studies. This is problematic, as several freshwater snails with their main occurrence in Southeast Asia have a considerable importance as first intermediate hosts for infections in humans and animals. Despite their proven medical importance, in particular the faunistic and systematic knowledge on cerithioidean freshwater snails of the various families acting as one of the most important vectors for digenic human pathogens, is precarious. The Cerithioidea is an ecologically and phylogenetically important, albeit essentially marine caenogastropod group, with its freshwater members in Southeast Asia acting as first intermediate hosts of a wide array of diverse trematodes (see details and references e.g. in
Cerithioidean freshwater taxa were long subsumed under the historical concept of “melaniids”, which was later uncritically replaced by the family assignment to the Thiaridae (see e.g.
Thiaridae which are found mostly in tropical to subtropical regions worldwide, inhabit virtually all freshwater and brackish-water bodies, both in lotic (including springs, creeks, rivers and streams) and lentic habitats (lakes and ponds). They are essentially, and presumably originally, widely distributed throughout Southeast Asia and in Australia (see
To complicate matters, Thiaridae are both parthenogenetic, with many populations essentially representing clones of individual females, and viviparous, with various typical embryonic stages developing within the female’s non-uterine, i.e. subhemocoelic brood pouch, and with distinct reproductive strategies to be found, viz. eu-viviparous vs. ovo-viviparous modes that are correlated with the amount of nourishment provided by the female (
In Thailand, the Thiaridae are represented by several described species, mostly being conchologically highly variable, such as e.g. Melanoides tuberculata (O. F. Müller, 1774), Mieniplotia scabra (O. F. Müller, 1774) or Tarebia granifera (Lamarck, 1816), the latter being commonly referred to as the “Quilted Melania” in the aquarium industry. Accordingly, as is typical in thiarids, a plethora of species names has been applied, irrespective of the fact that their known polymorphic phenotype, in combination with their viviparity and mainly parthenogenetic reproduction, renders unequivocal species delimitation quite problematic; see for detailed discussion e.g.
This holds true especially for species assigned to Tarebia H. & A. Adams, 1854, which are found in rivers, streams and lakes as well as canals and ponds throughout its authochthonous distributional range. It extends, according to literature records (e.g.
Distribution of the freshwater thiarid snail Tarebia granifera (Lamarck, 1816) across its range in Southeast Asia, with the focus on occurrences in Thailand, contrasted with type and topotypical material from the island of Timor. Asteriks: type locality of “Melania” granifera Lamarck, 1816, reconstructed to originate from near Kupang in western Timor (see text for more details); black dots: sequenced material used in this study; white dots: shell material from museum collections analysed and literature records; white dots with black dot inside: wet material preserved in ethanol.
In addition, this snail has become widely invasive in the tropics outside its native range, the spreading being attributed to the aquarium trade. As early as the 1950s, though,
That way, this snail exhibits its potency as neozoon, in combination with its role as important vector for several diseases, supporting the life cycles of digenic parasites infecting humans as well as other animals. Throughout Southeast Asia and in particular in Thailand, T. granifera is known as major first intermediate host and thus transmission vector for trematode parasites dangerous to humans, livestock and wild animals; among which are most prominently several species of the Heterophyidae and Opisthorchiidae reported as causing opportunistic infections in people (e.g.
Therefore, being able to ecologically adopt apparently to a broad range of different freshwater habitats, Tarebia is highly diverse, with quite polymorphic shells, which are mostly elongately ovate, turreted and strongly sculptured, with both spiral grooves and ridges formed by nodules or tubercles, resulting in a plethora of named shell phenotypes (see Fig.
Shells of Tarebia granifera (Lamarck, 1816) from Timor and Thailand. a. Syntypes (MHNG 1093/72/1-4) from Timor. b–g. Morph A, i.e. specimens from Thailand corresponding to T. granifera (SUT 0514044, SUT 0516123, SUT 0515088, SUT 0515068, SUT 0515059, SUT 0516144). h–m. Morph B, i.e. specimens from Thailand corresponding to named T. lineata (Gray, 1828) (SUT 0515081, SUT 0514046, SUT 0516129, SUT 0515092, SUT 0515095, SUT 0516143). n–s. Morph C from Thailand (SUT 0515079, SUT 0516126, SUT 0515055, SUT 0515091, SUT 0516147, SUT0516142). t–y. Shells of T. granifera from Timor Leste (ZMH 119364, ZMH 119359, ZMH 119357, ZMH 119353, ZMH 119363, ZMH 119361). For locality data, see the material list in the main part of the text. Scale bar: 10 mm.
In light of these phenotypical variations found in the shell morphology of Tarebia, a modern taxonomic-systematic revision, utilizing evidence from molecular phylogenetics and phylogeographical analyses, becomes desirable. However, as it is the case for most thiarids this taxon also has not found more attention yet as to intra- and interspecific species diversity, neither in Thailand nor elsewhere in adjacent regions. Here, we present results from our study of the morphological and molecular genetic variation in combination with the distributional and phylogenetic relationships as well as differences in the reproductive biology of thiarids, in particular in populations from the North, Central, Northeast and South of Thailand. We focus on the two phylogenetically highly informative and heterogeneous mitochondrial gene fragments cytochrome c oxidase subunit 1 and 16 S rRNA genes. In addition, we have studied the progeny and ontogeny of representatives from populations throughout the geographical distribution in Thailand, i.e. the frequency of various ontogenetic stages of embryos and shelled juveniles in the females’ brood pouch. Combining the study of morphological variation (using biometry and geometric morphometrics) with molecular genetic variation and reproductive biology analyses, we compared the populations of Thailand as our special focus to topotypical samples recently collected from the type locality Timor as reference.
Viewed from the background of a molecular backbone phylogeny we are, finally, able to analyse a suite of questions concerning the nature of cladogenesis, phylogeography and reproductive biology in these snails, in context with the infections by various trematodes, eventually hoping to elucidate the interrelationship and co-existence of human-infectious trematode parasites and their first intermediate snail hosts.
The National Committee on Hydrology separates Thailand into 25 distinct hydrological units or river basins, which are used in this study as an established geographical reference. These units comprise the following rivers and drainage systems: Salween, Mekong, Kok, Shi, Moon, Ping, Wang, Yom, Nan, Chao Phraya, Sakaekrang, Pasak, Tha Chin, Mae Klong, Prachinburi, Bang Pakong, Tonle Sap, Peninsular East Coast, Phetchaburi, Peninsular West Coast, Southeast Coast, Tapi, Songkhla Lake, Pattani and Southwest Coast. These catchment and drainage systems are re-grouped here into seven areas, each with specific characteristics; refer to Figs
1 Central area: This is the most important area for Thailand, as it is an area without large water sources. The region, therefore, depends heavily on water from river basins upstream, such as Chao Phraya River as the main river of Thailand. The Chao Phraya begins at the confluence of the Ping and Nan rivers (Northern area) at Nakhon Sawan province. It flows from north to south from the central plains through Bangkok to the Gulf of Thailand.
2 Northern area: This area is a rich source of water for the central area (see above). For example, water of the Wang River flowing from north to south has its source in the Chiang Rai province. One of the principal cities along the river is Lampang, which is on the north bank of a curve in the river. From Lampang, the river flows southwards passing into Tak province. It joins the Ping River near Mae Salit north of the town of Tak. The Ping River originates in the Chiang Mai province, flowing through the provinces of Lamphun, Tak, and Kamphaeng Phet. The Nan River originates in the Nan province, subsequently draining the provinces Uttaradit, Phisanulok and Phichit. The Yom River joins the Nan River in the Chumsaeng district, Nakhon Sawam province. When the Nan River joins the Ping River it forms the Chao Phraya.
3 North-western area: This is a part of the drainage system of the Salween River, which flows into the neighbouring country of Myanmar.
4 Western area: This is part of the basin formed by the Me Klong River, which runs into the Gulf of Thailand.
5 North-eastern area: This is part of the Mekong river basin’s catchment area, which drains into the South China Sea.
6 Eastern area: An area characterized by many short rivers.
7 Southern area: Many short rivers and high annual rainfall characterize this area. There are a number of large water reservoirs.
Specimens of Tarebia granifera were collected throughout Thailand. For reference, we compare with samples available to MG from Timor Leste through the courtesy of Vince Kessner, who collected there recently. All samples were preserved in 95 % ethanol. Voucher specimens are kept in the collection of the Center of Natural History (CeNak), Zoological Museum, Universität Hamburg, Germany (ZMH) and the collection of the Parasitology and Medical Malacology Research Unit, Department of Biology, Faculty of Science, Silpakorn University, Thailand (SUT).
To reconstruct in detail the distributional range, in addition to own collecting activities in most parts of the region, material was analysed in several major museum collections, as well as literature records which were sufficiently verifiable as to the species identity (in general documented by descriptions and, even better, figures of shells collected).
Geographic coordinates of newly collected material were taken with a GPS device at the sampling site (WGS84 datum). Where GPS data for sampling sites were unavailable, coordinates were determined as accurately as possible from a map. Localities of the samples were mapped on a dot-by-dot basis on a public domain map (NaturalEarth, www.naturalearthdata.com) with ArcMap 10.4.1 (Esri Inc., Redlands, CA, USA). Final maps were compiled using Photoshop CS6 (Adobe Systems Inc., San José, CA, USA). The spelling of localities (whenever possible) follows GeoNames (http://www.geonames.org).
For climatic data, we used information from the climate of the world database (https://www.weatheronline.co.uk/reports/climate/Thailand).
The snails identified as belonging to Tarebia were grouped according to their morphological characteristics and geographic origin in four preliminary classes or morphs (see Fig.
The following biometrical parameters of the adult shells were taken with a digital calliper (accuracy: 0.1 mm): height of shell (h), width of shell (w), length of aperture (la), width of aperture (wa), height of body whorl (hbw), height of the last three whorls (l3w) and number of whorls (nw) (Fig.
For the two different mitochondrial clades, the Shapiro-Wilk-test was performed on all measured variables for each group individually to test for normal distribution. If at least one group was not normally distributed, we conducted a Wilcoxon signed rank test with continuity correction, to test for significant differences between clades. If the data for both groups were normally distributed, a Levene-test based on absolute deviations from the mean was performed to check for homoscedasticity. In case homoscedasticity was detected, we tested different groups using a two-sample t-test. Otherwise, we performed Welch’s heteroscedastic t-test.
All available type specimens and the other examined material was photographed by remote shooting with EOS Utility 2.12.2.1 for Windows (Canon Inc., Tokyo, Japan) and Digital Photo Professional 3.12.51.2 for Windows (Canon Inc.) using a digital camera (EOS 5D MKII with Canon macro photograph lens MP-E 65 mm and compact macro lens EF 50 mm, Canon Inc.). Shell orientation was adjusted so that the apertural plane of the shell was perpendicular in relation to the optical axis of the camera and the shell’s columella parallel to the background. Photo stacks were assembled in Helicon Focus 5.3.14.2 for Windows (Helicon Soft Ltd., Kharkiv, Ukraine). The images were then edited with Photoshop CS6 (Adobe Systems Inc.).
A total of 1,169 standardized images of adult, unbroken shells could be included in our geometric morphometrics data set. Using tpsUtil version 1.74 (
The content of the brood pouch was counted as best proxy for differences in the thiarid reproductive strategy following the method described in
Sequences from 131 specimens of T. granifera from 95 populations in Thailand and 12 specimens from 11 populations in Timor Leste were generated (see Table
Collection voucher numbers, geographic coordinates of sampling sites and GenBank accession numbers for specimens of Tarebia granifera (Lamarck, 1816) used in the molecular analyses.
Voucher Number | Latitude | Longitude | GenBank accession number | |
---|---|---|---|---|
cox1 | 16 S rRNA | |||
SUT 0514050 | 18°17’08.5”N, 098°39’16.9”E | MK000303 | MK025577 | |
SUT 0514051 | 18°17’04.4”N, 098°39’15.0”E | MK000304 | – | |
SUT 0514054 (A) | 18°17’23.0”N, 098°39’03.6”E | MK000307 | MK025580 | |
SUT 0514054 (B) | 18°17’23.0”N, 098°39’03.6”E | – | MK025581 | |
SUT 0514052 (B) | 18°16’26.1”N, 098°38’54.0”E | MK000305 | MK025578 | |
SUT 0514052 (C) | 18°16’26.1”N, 098°38’54.0”E | MK000306 | MK025579 | |
SUT 0515081 (B1) | 19°28’33.6”N, 098°07’02.4”E | MK000331 | – | |
SUT 0515081 (B9) | 19°28’33.6”N, 098°07’02.4”E | – | MK025609 | |
SUT 0515077 | 19°25’31.1”N, 097°59’27.2”E | – | MK025606 | |
SUT 0515083 | 19°22’19.6”N, 098°26’35.9”E | MK000332 | MK025610 | |
SUT 0515078 | 19°21’54.8”N, 097°58’10.7”E | MK000329 | MK025607 | |
SUT 0515079 (C3) | 19°15’31.6”N, 097°54’44.6”E | MK000330 | – | |
SUT 0515079 (C5) | 19°15’31.6”N, 097°54’44.6”E | – | MK025608 | |
SUT 0516119 | 18°51’22.2”N, 100°11’09.1”E | MK000350 | MK025628 | |
SUT 0514045 (B1) | 18°56’00.5”N, 099°38’54.6”E | MK000300 | – | |
SUT 0514045 (B2) | 18°56’00.5”N, 099°38’54.6”E | – | MK025574 | |
SUT 0514044 (A) | 18°52’47.5”N, 099°40’01.0”E | MK000298 | MK025572 | |
SUT 0514044 (B1) | 18°52’47.5”N, 099°40’01.0”E | MK000299 | – | |
SUT 0514044 (B2) | 18°52’47.5”N, 099°40’01.0”E | – | MK025573 | |
SUT 0514046 | 18°46’39.8”N, 099°38’38.7”E | MK000301 | MK025575 | |
SUT 0516124 | 18°42’14.8”N, 099°35’31.7”E | MK000353 | MK025631 | |
SUT 0515090 | 19°11’30.4”N, 101°12’13.2”E | MK000336 | MK025614 | |
SUT 0516114 | 18°51’45.1”N, 100°28’37.1”E | MK000348 | MK025625 | |
SUT 0516108 | 18°05’03.1”N, 100°13’00.1”E | – | MK025621 | |
SUT 0516113 (B) | 18°00’50.6”N, 100°08’22.6”E | MK000346 | MK025623 | |
SUT 0516113 (C1) | 18°00’50.6”N, 100°08’22.6”E | – | MK025624 | |
SUT 0516113 (C2) | 18°00’50.6”N, 100°08’22.6”E | MK000347 | – | |
SUT 0516112 (B2) | 17°52’19.5”N, 100°18’02.1”E | MK000345 | – | |
SUT 0516112 (B3) | 17°52’19.5”N, 100°18’02.1”E | – | MK025622 | |
SUT 0513019 (A) | 17°52’29.5”N, 100°18’25.6”E | MK000292 | – | |
SUT 0513019 (B) | 17°52’29.5”N, 100°18’25.6”E | – | MK025563 | |
SUT 0513023 | 17°52’51.3”N, 100°16’14.9”E | – | MK025564 | |
SUT 0516109 | 17°43’42.3”N, 099°58’49.6”E | MK000344 | – | |
SUT 0515075 (B1) | 17°13’23.4”N, 098°13’34.2”E | MK000327 | – | |
SUT 0515075 (B2) | 17°13’23.4”N, 098°13’34.2”E | – | MK025604 | |
SUT 0515076 (B1) | 17°26’04.8”N, 098°03’33.3”E | MK000328 | – | |
SUT 0515076 (B2) | 17°26’04.8”N, 098°03’33.3”E | – | MK025605 | |
SUT 0516126 (C1) | 16°52’29.3”N, 099°07’13.6”E | MK000355 | – | |
SUT 0516126 (C2) | 16°52’29.3”N, 099°07’13.6”E | – | MK025633 | |
SUT 0515073 | 16°42’38.5”N, 098°30’22.2”E | MK000326 | MK025602 | |
SUT 0515072 | 16°41’39.3”N, 098°31’04.4”E | MK000325 | MK025601 | |
SUT 0515074 | 16°40’58.4”N, 098°31’06.9”E | – | MK025603 | |
SUT 0516103 (B1) | 17°33’16.2”N, 099°29’48.2”E | MK000343 | – | |
SUT 0516103 (B2) | 17°33’16.2”N, 099°29’48.2”E | – | MK025620 | |
SUT 0515086 (A1) | 17°01’07.6”N, 100°55’36.0”E | MK000333 | – | |
SUT 0515086 (A2) | 17°01’07.6”N, 100°55’36.0”E | – | MK025611 | |
SUT 0515087 | 16°57’21.3”N, 100°55’31.0”E | MK000334 | MK025612 | |
SUT 0516118 (A) | 16°52’13.1”N, 100°50’17.4”E | MK000349 | MK025626 | |
SUT 0516118 (B) | 16°52’13.1”N, 100°50’17.4”E | – | MK025627 | |
SUT 0515067 | 16°50’36.3”N, 100°45’16.1”E | MK000319 | MK025595 | |
SUT 0516130 | 16°39’46.3”N, 101°08’09.8”E | – | MK025637 | |
SUT 0516121 | 16°37’23.8”N, 100°54’00.5”E54’00.5”E | – | MK025629 | |
SUT 0516120 | 16°36’01.3”N, 100°54’29.9”E | MK000351 | – | |
SUT 0516123 | 16°34’24.1”N, 100°59’23.6”E | MK000352 | MK025630 | |
SUT 0515088 (A1) | 16°32’51.7”N, 100°54’03.2”E | MK000335 | – | |
SUT 0515088 (A2) | 16°32’51.7”N, 100°54’03.2”E | – | MK025613 | |
SUT 0516129 (B2) | 16°32’25.6”N, 101°04’58.4”E | MK000358 | – | |
SUT 0516129 (B3) | 16°32’25.6”N, 101°04’58.4”E | – | MK025636 | |
SUT 0514041 | 15°47’54.2”N, 101°14’08.1”E | – | MK025570 | |
SUT 0514042 | 15°47’52.2”N, 101°13’54.4”E | MK000296 | – | |
SUT 0514040 (B) | 15°47’29,7”N, 101°13’30,7”E | – | MK025568 | |
SUT 0514040 (C) | 15°47’29,7”N, 101°13’30,7”E | – | MK025569 | |
SUT 0514043 (B1) | 15°47’19.3”N, 101°15’07.4”E | MK000297 | – | |
SUT 0514043 (B2) | 15°47’19.3”N, 101°15’07.4”E | – | MK025571 | |
SUT 0515068 | 17°23’24.7”N, 101°22’27.3”E | MK000320 | MK025596 | |
SUT 0516125 | 17°04’38.0”N, 101°29’20.6”E | MK000354 | MK025632 | |
SUT 0516128 (B4) | 17°03’03.9”N, 101°31’38.7”E | MK000357 | – | |
SUT 0516128 (B5) | 17°03’03.9”N, 101°31’38.7”E | – | MK025635 | |
SUT 0515064 (B4) | 16°34’45.6”N, 102°50’22.5”E | MK000317 | – | |
SUT 0515064 (B5) | 16°34’45.6”N, 102°50’22.5”E | – | MK025593 | |
SUT 0516131 (B) | 14°35’32.3”N, 101°50’30.1”E | MK000359 | MK025638 | |
SUT 0516131 (C) | 14°35’32.3”N, 101°50’30.1”E | MK000360 | MK025639 | |
SUT 0516135 | 12°37’50.0”N, 101°20’35.0”E | MK000362 | MK025642 | |
SUT 0516127 (B1) | 15°40’59.6”N, 100°14’59.3”E | MK000356 | – | |
SUT 0516127 (B2) | 15°40’59.6”N, 100°14’59.3”E | – | MK025634 | |
SUT 0516132 | 14°55’12.3”N, 101°13’10.9”E | MK000361 | MK025640 | |
SUT 0516133 | 14°44’06.4”N, 101°11’31.4”E | – | MK025641 | |
SUT 0515055 (C1) | 13°49’01.2”N, 100°02’27.9”E | MK000308 | – | |
SUT 0515055 (C2) | 13°49’01.2”N, 100°02’27.9”E | – | MK025582 | |
SUT 0515091 (C1) | 14°37’25.9”N, 098°43’40.5”E | MK000337 | – | |
SUT 0515091 (C2) | 14°37’25.9”N, 098°43’40.5”E | – | MK025615 | |
SUT 0515092 (B1) | 14°26’03.0”N, 098°51’14.7”E | MK000338 | – | |
SUT 0515092 (B2) | 14°26’03.0”N, 098°51’14.7”E | – | MK025616 | |
SUT 0515093 | 14°14’27.6”N, 099°03’55.9”E | MK000339 | – | |
SUT 0515061 (B) | 13°54’18.1”N, 099°23’07.8”E | – | MK025591 | |
SUT 0515061 (C) | 13°54’18.1”N, 099°23’07.8”E | MK000316 | MK025592 | |
SUT 0515060 (B1) | 13°51’17.7”N, 099°22’58.9”E | MK000315 | – | |
SUT 0515060 (B2) | 13°51’17.7”N, 099°22’58.9”E | – | MK025590 | |
SUT 0515059 (A1) | 13°46’44.8”N, 099°25’26.7”E | MK000313 | – | |
SUT 0515059 (A2) | 13°46’44.8”N, 099°25’26.7”E | – | MK025588 | |
SUT 0515059 (B) | 13°46’44.8”N, 099°25’26.7”E | MK000314 | MK025589 | |
SUT 0515058 | 13°45’00.5”N, 099°26’27.4”E | MK000312 | MK025587 | |
SUT 0515057 (B1) | 13°41’28.1”N, 099°29’08.1”E | MK000311 | – | |
SUT 0515057 (B2) | 13°41’28.1”N, 099°29’08.1”E | – | MK025586 | |
SUT 0515056 (A) | 13°37’00.15”N, 099°24’36.9”E | – | MK025583 | |
SUT 0515056 (B) | 13°37’00.15”N, 099°24’36.9”E | MK000309 | MK025584 | |
SUT 0515056 (C) | 13°37’00.15”N, 099°24’36.9”E | MK000310 | MK025585 | |
SUT 0515070 (B1) | 13°32’54.2”N, 099°21’42.3”E | MK000322 | – | |
SUT 0515070 (B2) | 13°32’54.2”N, 099°21’42.3”E | – | MK025598 | |
SUT 0515070 (C) | 13°32’54.2”N, 099°21’42.3”E | MK000323 | MK025599 | |
SUT 0515069 | 13°32’52.2”N, 099°17’33.7”E | MK000321 | MK025597 | |
SUT 0515071 | 13°32’07.4”N, 099°20’31.8”E | MK000324 | MK025600 | |
SUT 0515066 | 13°19’29.2”N, 099°14’22.0”E | MK000318 | MK025594 | |
SUT 0513032 | 12°48’02.7”N, 099°58’53,2”E | MK000293 | MK025565 | |
SUT 0516146 (B3) | 11°55’29.1”N, 099°42’40.9”E | MK000372 | – | |
SUT 0516146 (B7) | 11°55’29.1”N, 099°42’40.9”E | – | MK025652 | |
SUT 0516146 (C) | 11°55’29.1”N, 099°42’40.9”E | MK000373 | MK025653 | |
SUT 0514037 (A1) | 11°36’50.0”N, 099°40’07.9”E | – | MK025566 | |
SUT 0514037 (A7) | 11°36’50.0”N, 099°40’07.9”E | MK000294 | – | |
SUT 0514038 | 11°26’14.4”N, 099°26’33.0”E | MK000295 | MK025567 | |
SUT 0511149 | 10°44’28,8”N, 099°12’54.9”E | MK000291 | MK025562 | |
SUT 0516137 (B1) | 08°48’06.9”N, 099°26’45.1”E | MK000363 | – | |
SUT 0516137 (B2) | 08°48’06.9”N, 099°26’45.1”E | – | MK025643 | |
SUT 0514048 | 08°52’18.8”N, 099°25’59.1”E | MK000302 | MK025576 | |
SUT 0516147 | 09°12’39.8”N, 099°11’55.7”E | MK000374 | MK025654 | |
SUT 0516148 | 09°12’25.7”N, 099°12’25.7”E | MK000375 | MK025655 | |
SUT 0516142 (B) | 09°08’07.2”N, 099°40’31.6”E | MK000367 | MK025647 | |
SUT 0516142 (C) | 09°08’07.2”N, 099°40’31.6”E | MK000368 | MK025648 | |
SUT 0516139 | 08°47’23.0”N, 099°38’13.2”E | MK000365 | MK025645 | |
SUT 0516145 (B1) | 08°43’17.3”N, 099°40’14.8”E | MK000371 | – | |
SUT 0516145 (B2) | 08°43’17.3”N, 099°40’14.8”E | – | MK025651 | |
SUT 0515097 (A1) | 08°10’20.8”N, 098°47’37.6”E | MK000341 | – | |
SUT 0515097 (A2) | 08°10’20.8”N, 098°47’37.6”E | – | MK025618 | |
SUT 0515098 | 08°09’49.2”N, 098°47’50.9”E | MK000342 | MK025619 | |
SUT 0515095 | 07°22’11.0”N, 099°40’47.9”E | MK000340 | MK025617 | |
SUT 0516138 | 07°42’48.3”N, 099°51’33.6”E | MK000364 | MK025644 | |
SUT 0516144 (A1) | 07°13’36.6”N, 100°31’41.8”E | – | MK025650 | |
SUT 0516144 (A2) | 07°13’36.6”N, 100°31’41.8”E | MK000370 | – | |
SUT 0516141 (B1) | 06°52’29.3”N, 100°19’48.4”E | MK000366 | – | |
SUT 0516141 (B2) | 06°52’29.3”N, 100°19’48.4”E | – | MK025646 | |
SUT 0516143 | 06°49’29.5”N, 100°19’49.7”E | MK000369 | MK025649 | |
ZMH 119364 | 08°31’32.3”S, 125°58’50.0”E | – | MK025664 | |
ZMH 119359 | 09°00’30.6”S, 126°03’45.0”E | MK000382 | MK025661 | |
ZMH 119358 | 09°00’44.8”S, 126°03’49.2”E | MK000381 | MK025660 | |
ZMH 119354 | 09°01’11.4”S, 126°03’58.3”E | MK000377 | MK025656 | |
ZMH 119357 | 08°26’36.3”S, 126°28’11.4”E | MK000380 | MK025659 | |
ZMH 119356 | 08°20’32.1”S, 127°01’07.9”E | MK000379 | MK025658 | |
ZMH 119353 | 08°25’34.6”S, 126°41’42.5”E | MK000376 | – | |
ZMH 119362 | 08°56’47.1”S, 124°58’28.4”E | MK000385 | – | |
ZMH 119355 | 08°44’36.4”S, 126°22’49.7”E | MK000378 | MK025657 | |
ZMH 119360 | 08°47’05.0”S, 126°22’32.0”E | MK000383 | MK025662 | |
ZMH 119363 | 08°47’05.0”S, 126°22’32.0”E | MK000386 | – | |
ZMH 119361 | 09°01’59.6”S, 125°59’35.9”E | MK000384 | MK025663 |
Forward and reverse strands were assembled using the program Geneious (Biomatters Limited, Auckland, New Zealand) and corrected by eye. The protein coding cox1 sequences were aligned with MUSCLE (
For information on vouchers and GenBank accession numbers, see Table
Bayesian Inference (BI), Maximum likelihood (ML) and maximum parsimony (MP) approaches were used to reconstruct the phylogenetic relationships. The sequence data set was initially divided into four partitions for the nucleotide model-based ML and BI approaches: 1.) 1st codon positions of cox1, 2.) 2nd codon positions of cox1, 3.) 3rd codon positions of cox1, and 4.) 16S. To select an appropriate partitioning scheme and/or evolutionary models for the mitochondrial sequences, the data set was analysed with PartitionFinder 2.1.1 (
The BI analysis was performed using MrBayes 3.2.6. Metropolis-coupled Monte Carlo Markov chain (MC3) searches were run with four chains in two separate runs for 50,000,000 generations with default priors, trees and parameters sampled every 1,000 generations under default heating using the best-fit model as suggested by PartitionFinder. Diagnostic tools in MrBayes, including estimated sample size (ESS) values ≥ 200, were used to ensure that the MC3 searches had reached stationarity and convergence. The first 5,000,000 generations were discarded as burn-in.
Heuristic ML analysis was performed with Garli using the best-fit models as suggested by PartitionFinder. Support values were computed by bootstrapping with 1,000 replications.
Heuristic MP searches were carried out with PAUP v4.0b10 (
Bayesian posterior probabilities (PP) values ≥ 0.95 and bootstrap (BS) values ≥ 70% and were interpreted as significant/meaningful support. BS values from the ML and MP analyses were mapped onto the Bayesian 50% majority-rule consensus tree with SumTrees 3.3.1, which is part of the Dendropy 3.8.0 package (
We used the General Mixed Yule-coalescent (
We dated the divergence times for the main clades of Tarebia included in this study using the Bayesian algorithm implemented in Beast 2 based on the concatenated mitochondrial data assuming a strict molecular clock as the test implemented in MEGA 7 (
4 syntypes (MHNG 1093/72/1-4).
Originally given as “Timor” by
In addition, in the past some authors employed “Melania” lineata for shells found to exhibit spiral ridges and/or dark bands on its body whorls. Accordingly,
The distributional range of Tarebia granifera (Fig.
In Thailand, this species occurs in most lentic and lotic water bodies ranging throughout the various regions, provinces and river systems. There, T. granifera was found in both natural and artificial water bodies on a variety of substrata, such as e.g. sand, mud, rock (and, alternatively, concrete bridge foundations, concrete walls), on bottoms of reservoirs, irrigation canals and ornamental ponds. This species is usually found together with other thiarids, most often with M. tuberculata and Mieniplotia scabra. We were not able to correlate any consistent ecological features that clearly distinguish either at particular locations or specific habitat and/or populations where T. granifera was found to occur. Thus, the ecological requirements of this taxon, in particular contrasting those to that of other thiarids, remain insufficiently known.
In the following we document here in detail the geographical origin of material studied from Thailand, in comparison with the syntypes as well as topotypical material from Timor as reference (see above). Data on other localities indicated in Fig.
Thailand:
Pai drainage (Salween river system): Mae Hong Son province: Pang Mapha district, Huai Pa Hung, 19°22’20”N, 098°26’36”E, 435 m (SUT 0515083, 03. V. 2015); Mueang Mae Hong Son district, Huay Nam Kong, 19°28’34”N, 098°07’02”E, 425 m (SUT 0515081, 03. V. 2015); Tham Pla, 19°25’31”N, 097°59’27”E, 300 m (SUT 0515077, 02. V. 2015); Pai River, 19°21’55”N, 097°58’11”E, 215 m (SUT 0515078, 02. V. 2015); Huay Sua Tao, 19°15’32”N, 097°54’45”E, 235 m (SUT 0515079, 02. V. 2015).
Moei drainage (Salween river system): Tak province: Tha Song Yang district, check point near Moei River, 17°13’23”N, 098°13’34”E, 130 m (SUT 0515075, 02. V. 2015); Mae Salit Luang harbour, 17°26’05”N, 098°03’33”E, 110 m (SUT 0515076, 01. V. 2015); Mae Sot district, Ban Wang Takhian, 16°42’39”N, 098°30’22”E, 195 m (SUT 0515073, 30. IV. 2015); Thong Dee harbour, 16°41’39”N, 098°31’04”E, 205 m (SUT 0515072, 30. IV. 2015); Ban Huay Muang, 16°40’58”N, 098°31’07”E, 200 m (SUT 0515074, 30. IV. 2015).
Ping drainage (Chao Phraya river system): Chiang Mai province: Chom Thong district, Mae Soy bridge, 18°17’23”N, 098°39’04”E, 270 m (SUT 0514054, 24. VI. 2014); Ban Huay Phang, 18°17’09”N, 098°39’17’’ E, 260 m, SUT 0514050, 25. VI. 2014; Ban Mae Suai Luang, 18°17’04”N, 098°39’15”E, 270 m (SUT 0514051, 25. VI. 2014); Ban Mai Saraphi, 18°16’26”N, 098°38’54”E, 275 m (SUT 0514052, 25. VI. 2014); Tak province: Mueang Tak district, Ban Pak Huay Mae Tho, 16°52’29”N, 099°07’14”E, 105 m (SUT 0516126, 10. III. 2016).
Wang drainage (Chao Phraya river system): Lampang province: Chae Hom district, Wang river, 18°56’01”N, 099°38’55”E, 375 m (SUT 0514045, 23. IV. 2014); Ban Thung Hang stream, 18°52’48”N, 099°40’01”E, 375 m (SUT 0514044, 23. IV. 2014); Huay MaeYuak, 18°46’40”N, 099°38’39”E, 350 m (SUT 0514046, 22. IV. 2014); km. 40 + 075 bridge, 18°42’15”N, 99°35’32”E, 330 m (SUT 0516124, 09. III. 2016).
Yom drainage (Chao Phraya river system): Phayao province: Chiang Muan district, Thansawan waterfall, 18°51’22”N, 100°11’09”E, 230 m (SUT 0516119, 08. III. 2016); Phrae province: Mueang Phrae district, Mae Nam Saai km 9/457 bridge, 18°05’03”N, 100°13’00”E, 170 m (SUT 0516108, 07. III 2016); Sung Men district, Mae Marn reservoir, 18°00’51”N, 100°08’23”E, 205 m (SUT 0516113, 07. III 2016); Sukhothai province: Si Satchanalai district, Tat Duen waterfall, 17°33’16”N, 099°29’48”E, 135 m (SUT 0516103, 06. III. 2016).
Nan drainage (Chao Phraya river system): Nan province: Bo Kluea district, Wa river, 19°11’30”N, 101°12’13”E, 715 m (SUT 0515090, 11. VI. 2015); Ban Luang district, Huay Si Pun reservoir, 18°51’45”N, 100°28’37”E, 430 m (SUT 0516114, 08. III. 2016); Uttaradit province: Tha Pla district, Kaeng Sai Ngam, 17°52’20”N, 100°18’02”E, 255 m (SUT 0516112, 07. III 2016); Kaeng Wang Wua, 17°52’30”N, 100°18’26”E, 230 m (SUT 0513019, 28. VI. 2013); Huai Nam Re Noi, 17°52’51”N, 100°16’15”E, 270 m (SUT 0513023, 28. VI. 2013); Laplae district, Mae pool waterfall, 17°43’42”N, 099°58’50”E, 125 m (SUT 0516109, 07. III. 2016).
Khek drainage (Chao Phraya river system): Phitsanulok province: Nakhon Thai district, Huai Nam Sai, 17°01’08”N, 100°55’36”E, 215 m (SUT 0515086, 20. V. 2015); Ban Kaeng Lat, 16°57’21”N, 100°55’31”E, 325 m (SUT 0515087, 20. V. 2015); Wang Thong district, Kaeng Sopha, 16°52’13”N, 100°50’17”E, 415 m (SUT 0516118, 08. III. 2016); Poi waterfall, 16°50’36”N, 100°45’16”E, 200 m (SUT 0515067, 08. II. 2015); Khao Kho district, Kaeng Wang Nam Yen, 16°37’24”N, 100°54’01”E, 710 m (SUT 0516121, 09. III. 2016); Rajapruek resort, 16°36’01”N, 100°54’30”E, 705 m (SUT 0516120, 09. III. 2016); Phetchabun province: Khao Kho district, Huai Sa Dao Pong, 16°34’24”N, 100°59’24”E, 320 m (SUT 0516123, 10. III. 2016); Kaeng Bang Ra Chan, 16°32’52”N, 100°54’03”E, 600 m (SUT 0515088, 21. V. 2015).
Pa Sak drainage (Chao Phraya river system): Phetchabun province: Lom Sak district, Than Thip waterfall, 16°39’46”N, 101°08’10”E, 375 m (SUT 0516130, 11. III. 2016); Khao Kho district, Samsipkhot waterfall, 16°32’26”N, 101°04’58”E, 385 m (SUT 0516129, 11. III. 2016); Wichian Buri district, Ban Wang Ta Pak Moo 13, 15°47’54”N, 101°14’08”E, 120 m (SUT 0514041, 27. VI. 2014); Huai Leng, 15°47’52”N, 101°13’54”E, 115 m (SUT 0514042, 27. VI. 2014); Ban Wang Tian, 15°47’30”N, 101°13’31”E, 120 m (SUT 0514040, 27. VI. 2014); Huay Range reservoir at Ban Wang Ta Pak, 15°47’19”N, 101°15’07”E, 140 m (SUT 0514043, 27. VI. 2014); Lop Buri province: Phatthan Nikhom district, Suanmaduea waterfall, 14°55’12”N, 101°13’11”E, 135 m (SUT 0516132, 26. IV. 2016); Sara Buri province: Muak Lek district, Dong Phaya Yen waterfall, 14°44’06”N, 101°11’31”E, 155 m (SUT 0516133, 26. IV. 2016). Nakhon Sawan province: Mueand Nakhon Sawan district, Bungboraped, 15°41’00”N, 100°14’59”E, 30 m (SUT 0516127, 10. III. 2016).
Loei drainage (Mekong river system): Loei province: Phu Ruea district, Pla Ba waterfall, 17°23’25”N, 101°22’27”E, 665 m (SUT 0515068, 07. II. 2015); Phu Luang district, km. 50/350 at Loei River, 17°04’38”N, 101°29’21”E, 675 m (SUT 0516125, 10. III. 2016); Tatkoktup waterfall, 17°03’04”N, 101°31’39”E, 690 m (SUT 0516128, 10. III. 2016).
Chee drainage (Mekong river system): Khon Kaen province: Mueang Khon Kaen district, Bueng Thung Sang, 16°34’46”N, 102°50’23”E, 170 m (SUT 0515064, 05. II. 2015).
Moon drainage (Mekong river system): Nakhon Ratchasima province: Pak Thong Chai district, Lamphraphloeng reservoir, 14°35’32”N, 101°50’30”E, 260 m (SUT 0516131, 22. III. 2016).
Khwae drainage (Mae Klong river system): Kanchanaburi province: Thong Pha Phum district, Hindad hot spring, 14°37’26”N, 098°43’41”E, 160 m (SUT 0515091, 27. VI. 2015); Sai Yok district, Sai Yok Yai waterfall, 14°26’03”N, 098°51’15”E, 105 m (SUT 0515092, 27. VI. 2015); Sai Yok Noi waterfall, 14°14’28”N, 099°03’56”E, 115 m (SUT 0515093, 27. VI. 2015).
Phachi drainage (Mae Klong river system): Kanchanaburi province: Dan Makham Tia district, Ban Thung Makham Tia, 13°54’18”N, 099°23’08”E, 45 m (SUT 0515061, 17. III. 2015); Ban Ta Pu, 13°51’18”N, 099°22’59”E, 55 m (SUT 0515060, 17. III. 2015); Ban Nong Phai, 13°46’45”N, 099°25’27”E, 70 m (SUT 0515059, 17. III. 2015); Ratchaburi province: Chom Bueng district, Phachi River bridge, 13°45’01”N, 099°26’27”E, 65 m (SUT 0515058, 17. III. 2015); Ban Dan Thap Tako, 13°41’28”N, 099°29’08”E, 80 m (SUT 0515057, 17. III. 2015); Ban Pa Wai, 13°37’00”N, 099°24’37”E, 75 m (SUT 0515056, 17. III. 2015); Suan Phueng district, Lum Nam Phachi, 13°32’54”N, 099°21’42”E, 110 m (SUT 0515070, 23. I. 2015); Huai Ban Bor, 13°32’07”N, 099°20’32”E, 135 m (SUT 0515071, 23. I. 2015); Huay Nueng, 13°32’52”N, 099°17’34”E, 155 m (SUT 0515069, 23. I. 2015); Suan Phueng district, Ban Purakom, 13°19’29”N, 099°14’22”E, 275 m (SUT 0515066, 23. I. 2015).
Mae Klong river system: Nakhon Pathom province: Mueang Nakhon Pathom district, pond on campus of Silpakorn University, 13°49’01”N, 100°02’28”E, 80 m (SUT 0515055, 13. I. 2015).
Gulf of Thailand: Rayong province: Mueang Rayong district, Mae Rumphueng beach (Mae Rumphueng canal), 12°37’50”N, 101°20’35”E, 10 m (SUT 0516135, 28. IV. 2016); Phetchaburi province: Cha-am district, Khlong Cha-am (Cha-am canal), 12°48’03”N, 099°58’53”E, 20 m (SUT 0513032, 16. X. 2013); Prachuap Khiri Khan province: Mueang Prachuap Khiri Khan district, Khlong Bueng reservoir, 11°55’29”N, 099°42’40.9” E, 70 m (SUT 0516146, 11. V. 2016); Huai Yang district, Khlong Huai Yang (Yang canal), 11°36’50”N, 099°40’08”E, 55 m (SUT 0514037, 23. XI. 2014); Bang Saphan district, Kar on waterfall, 11°26’14”N, 099°26’33”E, 55 m (SUT 0514038, 23. XI. 2014); Chumphon province: Tha Sae district, Krapo waterfall, 10°44’29”N, 099°12’55”E, 75 m (SUT 0511149, 2. VII. 2011); Surat Thani province: Tha Chang district, Khlong Tha Sai (Takhoei canal), 09°12’40”N, 099°11’56”E, 10 m (SUT 0516147, 04. VI. 2016); Phunphin district, Ban Tung Ao (Ta Khoei canal), 09°12’26”N, 099°12’26”E, 5 m (SUT 0516148, 04. VI. 2016); Don Sak district, Vibhavadi waterfall (Tha Thong canal), 09°08’07”N, 099°40’32”E, 25 m (SUT 0516142, 09. V. 2016); Ban Na San district, Dat Fa waterfall, 08°52’19”N, 099°25’59”E, 80 m (SUT 0514048, 22. XI. 2014); Khlong Klai (Nong Noi canal), 08°48’07”N, 099°26’45”E, 110 m (SUT 0516137, 9. V. 2016); Nakhon Si Thammarat province: Nopphitam district, Khlong Prong (Klai canal), 08°47’23”N, 099°38’13”E, 100 m (SUT 0516139, 09. V. 2016); Krung Ching waterfall, 08°43’17”N, 099°40’15”E, 195 m (SUT 0516145, 09. V. 2016); Phatthalung province: Si Banphot district, Khlong Tha Leung (Tha Nae canal), 07°42’48”N, 099°51’34”E, 70 m (SUT 0516138, 08. V. 2016); Songkhla province: Singhanakhon district, Khlong Sathing Mo (Songkhla lake), 07°13’37”N, 100°31’42”E, 10 m (SUT 0516144, 08. V. 2016); Khlong Hoi Khong district, Khlong La reservoir, 06°52’29”N, 100°19’48”E, 60 m (SUT 0516141, 07. V. 2016); Khlong Cham Rai reservoir, 06°49’30”N, 100°19’50”E, 55 m (SUT 0516143, 07. V. 2016).
Andaman Sea: Krabi province: Mueang Krabi district, Khlong Sai (Khlong Sai canal), 08°10’20.8’’ N, 098°47’38’’ E, 25 m (SUT 0515097, 30. X. 2015); Wang Than Thip (Wang Than Thip canal), 08°09’49”N, 098°47’51”E, 20 m (SUT 0515098, 30. X. 2015); Trang province: Yan Ta Khao district, Khlong Palian (Palian canal), 07°22’11”N, 099°40’48”E, 20 m (SUT 0515095, 29. X. 2015).
Timor Leste: Manatuto district, W bank of Laclo river near Condae, ca. 4 km WSW of Manatuto, 08°31’32”S, 125°58’50”E, 35 m (ZMH 119364, 21 VI. 2012); south coast, 3.8 km N of Nancuro beach, 4.7 km SE of Natarbora, 09°00’31”S, 126°03’45”E, 20 m (ZMH 119359, 13. XI. 2011); 3.4 km N of Nancuro beach, 5 km SE of Natarbora, 09°00’45”S, 126°03’49”E, 20 m (ZMH 119358, 13. XI. 2011); 2.5 km N of Nancuro beach, 5.7 km SE of Natarbora, 09°01’11”S, 126°03’58”E, 15 m (ZMH 119354, 13. XI. 2011); Baucau district, NE of Baucau, Watabo beach, 08°26’36”S, 126°28’11”E, 20 m (ZMH 119357, 9. XI. 2011); Lautem district, Ira-Ara village, Lutu-Ira, 08°20’32”S, 127°01’08”E, 100 m (ZMH 119356, 23. V. 2011); near the Baucau/Lautem district border marker, 11.8 km NE of Laga, 08°25’35”S, 126°41’43”E, 5 m (ZMH 119353, 10. XI. 2011); Bobonaro district, north coast, 0.5 km from the mouth, Large seasonal stream in Batugade, 08°56’47”S, 124°58’28”E, 10 m (ZMH 119362, 20. V. 2012); Viqueque district, Ossu subdistrict, near village Usu Decima, Wai-eu-Lau, 08°44’36”S, 126°22’50”E, 670 m (ZMH 119355, 13. V. 2011); spring in the village, Loihuno, 08°47’05”S, 126°22’32”E, 255 m (ZMH 119360, 11. XI. 2011); spring in the village, Loihuno, 08°47’05”S, 126°22’32”E, 255 m (ZMH 119363, 17.V. 2012); Manufahi district, south coast, Fatuhcahi village, Wetetefuik creek, 09°02’00”S, 125°59’36”E, 30 m (ZMH 119361, 12 XI. 2011).
The final alignment of the cox1 sequences had a length of 658 base pairs (bp) and that of the 16S sequences 781 bp. Genetic p-distances for cox1 sequences of specimens determined as T. granifera from Thailand ranged from 0% to 14.7%, whereas all cox1 sequences obtained from specimens from Timor Leste were identical.
For 16S sequences, p-distances among specimens from Thailand ranged from 0% to 10.4% and for Timor Leste, pairwise p-distance between specimens were very low, ranging from 0% to 0.1%.
All three phylogenetic analyses recovered two deeply divergent clades of specimens assigned to T. granifera (clades A and B, Fig.
Bayesiam 50% majority-rule consensus tree showing two major mitochondrial clades in Tarebia granifera (Lamarck, 1816). Numbers at the nodes correspond to posterior probabilities (left), maximum likelihood (middle) and maximum parsimony (right) bootstrap values. At the tips of the tree voucher numbers (see material list in the main part of the text), country codes (THA: Thailand; TIM: Timor Leste; IDN: Indonesia) and the river where specimens were collected are indicated. The inset map shows the distribution of mitochondrial clades in Thailand (clade A: blue dots; clade B: magenta dots) and major river systems. The letters a–c in the map refer to localities, for which climatic data were available (see also Fig.
All specimens from Timor Leste were included in clade A together with specimens mostly from the southern to southern-central parts of Thailand (Fig.
In contrast, specimens of T. granifera assigned to clade B were more frequent in the northern part of Thailand, i.e. the majority of specimens in this clade originate from the northern to north east Thai provinces, such as Chang Mai, Mueang Mae Hong Son, Phayao, Lampang, Nan, Uttaradit, Tak, Sukhothai, Phitsanulok, Phetchabun and Loei, while only few specimens in this clade are from the southern-central Thai provinces Phatthalung, Nakhon Si Thammarat, Surat Thani, Ratchaburi, Kanchanaburi and Lop Buri. Almost all specimens assigned to clade B were placed in a polytomy in the tree shown in Fig.
When analysed by drainage systems, we found that all specimens from the north-western part of Thailand, which is drained through the Salween river system into the Andaman Sea, were included in clade B. Likewise, specimens from the headwaters of the Ping, Wang, Yom and Nan rivers belonging to the Chao Phraya system, with few exceptions, were assigned to clade B in the phylogenetic analyses. In the lower courses of northern to northern-central Thai drainages, such as e.g. the Chao Phraya and Mae Klong drainages that run into the Gulf of Thailand, specimens assigned to both clades are present.
Similarly, specimens belonging to both mitochondrial clades are present in the Mekong drainage, whereas specimens assigned to clade A predominate in the smaller rivers in the Thai parts of the Malay Peninsula to the north and south of the Isthmus of Kra that either drain into the Gulf of Thailand or the Andaman Sea (Fig.
In contrast to this geographical pattern in Tarebia granifera, with broadly speaking an essentially southern clade A and an essentially northern clade B, we found no correspondence of specimens from the three morphotypes with the two genetically differentiated clades as outlined above as all morphs were present in both clades (data not shown).
Evolutionary relationships among haplotypes were inferred applying a median-joining network approach that showed the two mitochondrial clades A and B to be separated by > 60 steps (cox1 and 16S; Fig.
Molecular analysis of Tarebia. a–b. Median-joining haplotype networks based on 16S (a) and cox1 (b) sequence data of Tarebia granifera (Lamarck, 1816). The size of each circle represents the frequency of a haplotype and the colour refers to main mitochondrial clades obtained from the phylogenetic analyses (Fig.
The ABGD approach suggested that the T. granifera clades A and B could be classified as two species for prior intraspecific divergences (d) of the combined cox1 and 16S data set of d ≥ 0.0077. The bGMYC analysis (Fig.
The results of the BEAST analysis assuming a strict molecular clock and a divergence rate of 1% per million years (Fig.
The shells of Tarebia granifera (Fig.
As shown in Fig.
Starting off from the type series of T. granifera from Timor (Fig.
We were not able to find any correlation of shell morphology with molecular genetic clusters as described above, or any other geographical or ecological factor matching these distinct phenotypes in Tarebia granifera.
For ranges and mean values of measured shell parameters for the different predefined groups, i.e. shell morphs/geographic groups or genetic clades, see Table
Biometric data for different shell morphs/geographic groups (see also Figs
Min | Max | Mean | Median | Standard deviation | |
---|---|---|---|---|---|
Height | |||||
Morph A | 9.29 | 29.83 | 18.93 | 18.90 | 3.69 |
Morph B | 8.56 | 32.38 | 19.73 | 20.06 | 3.87 |
Morph C | 10.53 | 26.88 | 19.03 | 18.94 | 3.02 |
Morph B+C | 8.56 | 32.38 | 19.62 | 19.81 | 3.76 |
Timor | 11.67 | 28.53 | 19.68 | 19.69 | 3.75 |
Clade A | 8.56 | 32.38 | 19.24 | 19.52 | 3.86 |
Clade B | 9.45 | 30.67 | 19.66 | 19.68 | 3.64 |
Width | |||||
Morph A | 3.73 | 13.28 | 8.26 | 8.44 | 1.71 |
Morph B | 3.49 | 14.46 | 8.35 | 8.55 | 1.61 |
Morph C | 4.39 | 11.58 | 7.94 | 7.98 | 1.32 |
Morph B+C | 3.49 | 14.46 | 8.28 | 8.39 | 1.58 |
Timor | 5.04 | 12.18 | 8.15 | 8.20 | 1.42 |
Clade A | 3.73 | 13.28 | 8.05 | 8.13 | 1.51 |
Clade B | 3.49 | 14.46 | 8.46 | 8.61 | 1.64 |
Aperture height | |||||
Morph A | 4.38 | 14.39 | 9.23 | 9.31 | 1.84 |
Morph B | 4.23 | 15.35 | 9.31 | 9.46 | 1.75 |
Morph C | 4.94 | 18.96 | 9.06 | 8.98 | 1.72 |
Morph B+C | 4.23 | 18.96 | 9.27 | 9.38 | 1.74 |
Timor | 5.16 | 13.6 | 9.13 | 9.12 | 1.62 |
Clade A | 4.38 | 14.39 | 9.06 | 9.10 | 1.72 |
Clade B | 4.23 | 18.96 | 9.41 | 9.50 | 1.77 |
Aperture width | |||||
Morph A | 1.63 | 8.92 | 4.30 | 4.29 | 0.87 |
Morph B | 1.68 | 8.91 | 4.25 | 4.25 | 0.90 |
Morph C | 2.42 | 8.41 | 4.31 | 4.23 | 1.01 |
Morph B+C | 1.68 | 8.91 | 4.26 | 4.25 | 0.92 |
Timor | 2.40 | 6.07 | 4.09 | 4.14 | 0.71 |
Clade A | 1.63 | 8.92 | 4.18 | 4.21 | 0.87 |
Clade B | 1.68 | 8.91 | 4.32 | 4.29 | 0.91 |
Last whorl height | |||||
Morph A | 5.91 | 19.55 | 12.48 | 12.47 | 2.44 |
Morph B | 5.77 | 20.37 | 12.60 | 12.78 | 2.35 |
Morph C | 6.81 | 15.83 | 11.99 | 11.92 | 1.87 |
Morph B+C | 5.77 | 20.37 | 12.50 | 12.65 | 2.29 |
Timor | 6.53 | 17.78 | 12.35 | 12.48 | 2.22 |
Clade A | 5.91 | 17.81 | 12.25 | 12.38 | 2.29 |
Clade B | 5.77 | 20.37 | 12.69 | 12.81 | 2.33 |
Last three whorls height | |||||
Morph A | 7.93 | 26.43 | 16.84 | 17.04 | 3.29 |
Morph B | 7.73 | 28.74 | 16.93 | 17.13 | 3.28 |
Morph C | 9.20 | 21.34 | 15.97 | 15.95 | 2.49 |
Morph B+C | 7.73 | 28.74 | 16.77 | 16.85 | 3.19 |
Timor | 9.46 | 23.89 | 16.56 | 16.40 | 3.12 |
Clade A | 7.93 | 26.22 | 16.49 | 16.54 | 3.22 |
Clade B | 7.73 | 28.74 | 17.01 | 17.13 | 3.17 |
H/W | |||||
Morph A | 1.77 | 2.95 | 2.31 | 2.29 | 0.25 |
Morph B | 1.22 | 3.13 | 2.37 | 2.38 | 0.22 |
Morph C | 1.50 | 3.05 | 2.41 | 2.41 | 0.24 |
Morph B+C | 1.22 | 3.13 | 2.38 | 2.39 | 0.22 |
Timor | 1.92 | 2.87 | 2.41 | 2.41 | 0.18 |
Clade A | 1.50 | 3.13 | 2.39 | 2.41 | 0.22 |
Clade B | 1.22 | 2.93 | 2.34 | 2.35 | 0.23 |
Last three whorls/width | |||||
Morph A | 1.27 | 2.54 | 2.05 | 2.04 | 0.13 |
Morph B | 1.22 | 2.53 | 2.03 | 2.03 | 0.15 |
Morph C | 1.39 | 2.65 | 2.02 | 2.03 | 0.16 |
Morph B+C | 1.22 | 2.65 | 2.03 | 2.03 | 0.15 |
Timor | 1.66 | 2.28 | 2.03 | 2.04 | 0.13 |
Clade A | 1.27 | 2.65 | 2.05 | 2.06 | 0.16 |
Clade B | 1.22 | 2.38 | 2.02 | 2.01 | 0.13 |
Results of biometric (a–d) and geometric morphometrics study (e), for four different morphs (A,B,C,Timor) of Tarebia granifera (Lamarck, 1816). Boxplots of (a) shell height, (b) shell width, (c) height of the last three whorls and (d) index of height of last three whorls agaianst shell width. Significant differences between groups are indicated by bars above the boxplots (e) Relative variance in shell shape along PC1 and PC2. Colour corresponding planes indicate the spread of each morph in the data set.
Between genetic clades at least one of the groups was found to be not normally distributed (Shapiro-Wilk-test, p < 0.05) for shell width and l3w/w. By contrast normal distribution was found for lw3 and shell height. Subsequent Levene-testing identified the height and l3w data sets as homoscedastic (p > 0.05), hence a two-sample t-test was performed, identifying significant differences (p < 0.025) between the means for the two clades for lw3 and no significant differences for shell height. For shell width and l3w/w a Wilcoxon signed rank test was performed, revealing significant differences (p < 0.025) for the mean of both shell parameters. However, similar to the situation when comparing the different shell morphs/geographical groups, it has to be noted that the ranges of all measured shell parameters widely overlap and, therefore, do not allow to derive diagnostic characteristics for the two main clades found in the phylogenetic analyses (see boxplots in Fig.
Results of biometric (a–d) and geometric morphometrics study (e), for the two mitochondiral clades of Tarebia granifera (Lamarck, 1816) found in this study. Boxplots of (a) shell height, (b) shell width, (c) height of the last three whorls and (d) index of height of last three whorls agaianst shell width. Significant differences between groups are indicated by bars above the boxplots (e). Relative variance in shell shape along PC1 and PC2. Colour corresponding planes indicate the spread of each morph in the data set.
A principal component analysis (PCA) identified the first six major axes to account for a relevant proportion of variance (p > 0.05) (PC1: 0.303; PC2: 0.181; PC3: 0.117; PC4: 0.090; PC5: 0.058; PC6: 0.052), explaining a cumulative proportion of 0.801 of variance.
Principal components (PC) 1–6 had all at least one group that proved to be not normally distributed (Shapiro-Wilk-test, p < 0.05). Subsequent Kruskal-Wallis-testing was significant (p < 0.05) in PC1–5 and not significant in PC6. Hence, no further testing was done for PC6. The Bonferroni-corrected Dunn-test identified the mean value for specimens from Timor to be significantly different (p > 0.025) from all other morphs on PC1. By contrast, examining PC2 and PC4 with the same test, proved morph A and B to be the only groups not significantly different (with regard to mean values) from one another. Finally, on PC3 and PC5 the Bonferroni-corrected Dunn-Test revealed the mean value of morph C not to be significantly different from all other groups, but the means of morph A and B to be significantly different to that of the specimens from Timor.
Finally, when morph C was integrated into morph B (since these were only differentiated on the basis of slight differences in banding pattern), PC1–5 supported only the group consisting of specimens from Timor to have significantly different means from all other specimens (data not shown). The scatter plot in Fig.
For PC1 and PC3–6 at least one of the groups (clade A/clade B) was not normally distributed (Shapiro-Wilk-test, p > 0.05). Hence, we conducted Wilcoxon singed rank tests for all these PC, with none showing significant differences between groups (p > 0.05). By contrast, in PC2 both groups showed normally distributed data. Therefore, Levene-testing based on deviations from the mean followed and was found significant (p < 0.05). Accordingly, we conducted Welch’s two sample t-test, revealing significant differences between the means of the two clades on PC2. The scatter plot in Fig.
Females of Tarebia granifera were found to contain embryos and shelled juveniles in their “marsupium”, or subhemocoelic brood pouch, situated in the neck region as in other thiarids studied so far. They usually release crawling juveniles with shells comprising several whorls that are built before hatching from the brood pouch. In this study, we found the snails to possess brood pouches filled with all ontogenetic stages, ranging from early to late embryos and six additional size classes of juveniles, with shells measuring between less than 0.5 to more than 3 mm (see Figs
Frequency of ontogenetic stages in the subhemocoelic brood pouches of female Tarebia granifera (Lamarck, 1816) (morph B) depending on occurrence in Thailand. Blue dots: mitochondrial clade A; pink dots: mitochondrial clade B. Size classes are assigned different colours in the pie charts (see legend) and rivers are coloured according to drainage systems; numbers at the pie charts refer to the total number of dissected specimens and the number of gravid females (in parentheses). The small letters refer to the stations Chiang Mai (a), Ko Samui (b) and Phuket (c) for which meteorological data representing the different climatic regions of Thailand were analysed (see Fig.
The frequency of these different size classes in the subhemocoelic brood pouch of the total of n = 1,007 dissected females of Tarebia granifera from a total of 107 populations from Thailand (n = 95) and Timor Leste (n = 12) is shown as to their geographic occurrence for the two mitochondrial clades A (n = 42) and B (n = 53) as well as the predefined morphs A, B and C in Figs
Frequency of ontogenetic stages in the subhemocoelic brood pouches of female Tarebia granifera (Lamarck, 1816) depending on occurrence in Thailand and Timor Leste. a. Morph A in Thailand; b. Morph C in Thailand; c. Timor Leste. Blue dots: mitochondrial clade A; pink dots: mitochondrial clade B. Size classes are assigned different colours in the pie charts (see legend) and rivers are coloured according to drainage systems; numbers at the pie charts refer to the total number of dissected specimens and the number of gravid females (in parentheses).
In all examined populations, the number of early and late embryonic stages was above 50%, in most cases even above 75%; see Fig.
Composition of contents of the subhemocoelic brood pouches of female Tarebia granifera (Lamarck, 1816) (a, c) and proportions of gravid animals, i.e. those with filled brood pouch, versus non-gravid specimens (b, d) from Thailand and Timor Leste. a. Composition of contents of the brood pouches for morph A, B and C from Thailand (THA) and specimens from Timor Leste (see Figs
When considering the overall distribution of different size classes in the different morphs/geographic clusters or mitochondrial clades, the resulting histograms (Fig.
We also compared the size class composition of offspring in the subhemocoelic brood pouches of Tarebia populations from different drainage systems. Although considerable variation was present among the rivers and streams of the 17 drainage systems in Thailand (Fig.
Composition of contents of the subhemocoelic brood pouches of female Tarebia granifera (Lamarck, 1816) (a) and proportions of gravid animals, i.e. those with brood pouch containing juveniles or other stages, and non-gravid specimens (b) from Thailand grouped according to rivers. For colour coding, see the inset legends.
The distribution of gravid vs. non-gravid specimens according to the 17 rivers systems exhibits some variation (Fig.
Whether reproduction is seasonal, or whether there is any influence of the month of collecting on our data, can currently not be answered with certainty. In an attempt to correlate reproduction (i.e. the frequency of gravid vs. non-gravid females) with climatic effects such as, for example, rainy season resulting in high water levels in rivers and streams, we have used published meteorological data (e.g. minimum/maximum temperature and precipitation) for stations representing the different climatic regions of Thailand, viz. Chiang Mai for northern inland region, Ko Samui for the Gulf of Thailand and Phuket for the Andaman Sea localities (see map in Fig.
Proportions of gravid vs. non-gravid specimens of Tarebia granifera (Lamarck, 1816) collected in different months within a given year, plotted on climate charts for localities that are representative for different climatic regimes in Thailand. (a) Chiang Mai for inland locations; (b) Ko Samui for the Gulf of Thailand; (c) Phuket for the Andaman Sea (see also Fig.
As evolutionary biologists working with molluscs, we should aim at testing the universality of known and disputed speciation mechanisms, and it is with a clear focus on these mechanisms we should choose our molluscan models to increase their frequency as a source of data in order to decipher the underlying mechanisms of biodiversity.
The combination of molecular genetics and phenotypic analyses in concert with information on the geographical occurrence and additional data, e.g. on biological properties such as reproductive strategies, provides a powerful tool for the study of species differentiation, or diversification indicating speciation. It allows truly biological species to be distinguished, not only as perceivable taxonomic or even genetic units, but also as natural entities of evolutionary significance; if we want to make here the careful distinction between a species taxon (with identifying characteristics) and species entity (as a group of coevolving populations); see for the theoretical background of applications of species concepts in freshwater molluscs
In freshwater gastropods high levels of morphological disparity and taxonomic diversity are frequently correlated, but often only because traditionally disparity was equated with diversity, as has been exemplified for limnic Cerithioidea, such as e.g. the Mediterranean melanopsids (
As has been discussed by the latter author with focus on freshwater gastropods, the widely adopted typological practice during the 19th and way into the 20th century of naming allopatric populations, in isolated fashion and often based on single specimens only, as if representing putatively distinct (morpho-) species, has led to a plethora of species and subspecies names. Freshwater gastropods were found to exhibit a pronounced individual conchological variability, which has been attributed to the environmental conditions of their habitats that widely fluctuate on a temporal and spatial scale (e.g.
In the course of the systematic revision of these thiarids, based on an evolutionary systematic approach (see
However, an assessment of the significance of distinct phenotypic traits is in general lacking, as is an understanding of the genetic basis of phenotypical variation in particular for gastropods. For the limnic pomatiopsid Oncomelania hupensis,
Owing to the earlier typological approach that resulted in the traditional overestimation of taxonomical diversity due to conchological disparity, but also in context of the genetically apparently closely related but morphologically highly distinct thiarids found across the distributional ranges throughout Southeast Asia and Australasia, we have to ask whether we are indeed dealing with actually many diverse species as separate evolutionary entities rather than only few, though highly polymorphic species with maybe several sympatric morphs exhibiting different ecophenotypical adaptations in shell response to the many variable environments where thiarids are usually to be found.
In the present study, we examined phenotypically distinguishable shell morphs of yet another thiarid from Thailand, traditionally assigned to Tarebia granifera, in reference to samples from Timor Leste as known type locality of the nominal species, using biometry and geometric morphometrics in combination with phylogeographical analyses of molecular genetics and reproductive strategy.
We found Tarebia to be widespread in almost all freshwater bodies throughout Thailand, with a wide range of conchological variants or morphs, of which some closely resemble the types and topotypical material of granifera collected on Timor. While in Thailand Tarebia has been reported with only one species by
Applying a drainage-based phylogeographical as well as a biometrical approach, we were unable to find for the populations in Thailand any correlation of the morphs distinguished in this study based on discernable shell features as well as overall “Gestaltwahrnehmung” with any criteria deducible from our observations given above, neither with geographical occurrence or preferred habitat and substrate nor with the molecular genetic substructuring detected (see below). So, all available evidence points at the coexistence of different morphologies or disparate phenotypes in Tarebia granifera in this part of mainland Southeast Asia. However, in the absence of any of the discussed parameters or factors to be causally correlated with these morphological differences we are left with the hypothesis that they either qualify for reflecting phenotypical plasticity correlated with ecological variables in the habitat of the individual populations studied, and/or, alternatively, being correlated with the parthenogenetic reproduction discussed further below.
Biometric analyses are found useful tools for the study of characteristics that shape morphologically distinct entities, thus allowing to look into evolutionary pattern (e.g.
Although there are some differences in the biometric parameters and in the geometric morphometrics of Thai Tarebia, it is generally impossible to delimit distinct entities (in the sense of being at least indicative of the existence as biological species) based on these features, as all of them largely overlap (Figs
The measurement of shell height of T. granifera showed that they are within the size range previously reported as to vary between 6 to 44 mm (e.g.
The results of geometric morphometrics revealed the overall shell shape of T. granifera from Thailand to be very similar to, and virtually undistinguishable from, conspecifics from Timor Leste (Fig.
In contrast to shell morphology (morphs A–C, or lineata vs. granifera phenotypes), we found based on molecular genetics strong indication as to the distinction of at least two natural entities within Tarebia in Thailand. As our analyses revealed, there is a most pronounced separation of two distinct mtDNA clades in this taxon, marked on the one hand by long branches in the resulting phylogenetic tree connecting these two clades, and on the other hand by very shorter branches within each of them (Figs
Therefore, our analyses would potentially allow for a more narrow species delimitiation within what has been to date traditionally treated in Thailand as T. granifera only (
However, the p-distance of 13.8 % for cox1 and 10 % for 16S sequences has to be considered relatively high, hinting potentially at the existence of two genetically distinct species. However, a definite decision as to this species question in Tarebia in Thailand should remain open until the geographical distribution of genetically characterized populations of T. granifera and other congeneric forms is completely resolved and better understood within the entire autochthonous range in the Oriental region. Thus, it should be the privilege of a more comprehensive and in-depth analysis of the biogeographical situation based on an ongoing molecular genetic study (Glaubrecht unpubl. data).
While we found representatives of clade A in the northern tributaries of rivers such as the Chao Phraya and Mae Klong that run into the Gulf of Thailand, with only few others occurring at some localities in the south of Thailand (Figs
However, although being more frequent in the northern provinces, some representatives of clade B also occur in more southern locations, such as in the provinces Surat Thani (SUT 0516137), Nakhon Si Thammarat (SUT 0516139) and Phatthalung (SUT 0516138). We anticipate that this might reflect occurrences of passive dispersal, potentially via aquatic plant or other material or even transport by birds, rather than vicariance via the influence of sea level or tidal flows in drainage systems. The results of the median-joining haplotypes network and bGMYC analysis (Fig.
As we found in our molecular analyses this major split of clade A and B in Thai Tarebia to be as old as most likely c. 5.32 million years ago (Fig.
For the distributional pattern found in Tarebia in Thailand, a vicariant hypothesis can be formulated using a major biogeographic transition zone between the Sundaic and Indochinese biota, located just north of the Isthmus of Kra. It is interpreted as the result of Neogene marine transgressions that breached this isthmus in two locations for prolonged periods of time, i.e. more than 1 million year duration, as was shown e.g. by phylogeographic analyses of a freshwater decapod crustacean, the giant freshwater prawn Macrobrachium rosenbergii (cf.
While the separation of Tarebia and Thiara hint at a Late Miocene splitting event (anticipated to have occurred somewhere in the Indo-Malayan insular region of the Sunda and Sahul shelfs), our molecular and distributional data on T. granifera (Figs
The fact that today the distributional boundaries of the two Tarebia populations in clade A and B do not coincide exactly with the position of the Isthmus of Kra, but are instead placed further to the north, could in this case be attributed to later palaeo-drainage differentiation in connection with orogenesis or other tectonic events in the mountainous central and northern regions of Thailand, as it was discussed using relevant geological and available biogeographical data, for example, from fishes and gastropods in
Tarebia snails are all viviparous, i.e. they incubate embryos and later ontogenetic stages in an extra-uterine structure, called the subhemocoelic brood pouch, located at the back of the head in the female’s body running alongside and below, but being independent of the pallial organs (e.g. the gonoduct), and formed apparently by an invagination of the genital grove found in other oviparous cerithioidean gastropods (
In the Thai populations of Tarebia, as well as those from Timor, we found most if not all ontogenetic stages contained at the same time in the female’s marsupium, from early embryos to late embryos and shelled juveniles, in all morphs (A–C), both molecular genetic clades (A and B) and specimens from all drainage systems, without a clear-cut differentiation of this reproductive strategy. In particular, the ontogeny of T. granifera in Thailand is not obviously correlated to specific drainage systems, no matter where these water bodies eventually drain. Therefore, we conclude that Tarebia throughout its distributional range covered here is eu-viviparous, with only very few representatives in some populations (see Figs
As in this later case, it could be hypothesized that any environmental factor might affect the reproductive strategy also in Tarebia. However, our analysis of representative climatic charts for the two parameters temperature and precipitation revealed no clear regional pattern of brood pouch content, as no correlation with the various ontogenetic stages were found across all locations in Thailand where T. granifera was sampled (see Figs
As in most (if not all) thiarids, Tarebia apparently lacks males in most populations, as we failed to find positive evidence for their existence. Parthenogenetic reproduction has gained much interest in the past in evolutionary biology, not only with respect to the origin of sex. Clonal reproduction in natural populations has obviously many advantages over sexual modes, with growth rates in the former often being much accelerated over the latter, as all individuals within the population are able to contribute (Maynard Smith 1978). In addition, these clones are considered instrumental in fast colonization of new habitats and areas, as even a single female can give rise to a new population (
Also in malacology there are some classical case studies, such as the New Zealand freshwater hydrobiid Potamopyrgus antipodarum (
It would be tempting to anticipate a similar phenomenon of T. granifera in Thailand and Timor Leste here from the varying frequencies (with up to 17.40 %) of non-gravid specimens. However, none ad hoc feature such as e.g. shell morphology between male and female could be differentiated in these aphallic cerithiodeans. So, in the present study we assumed not only any brood pouch-bearing snail to be female but also those without brood pouch as being non-gravid females rather than being rare males, for the reasons discussed above in connection with regional and/or climatic differences.
Given the prediction supported here that thiarid gastropods reproduce largely (if not completely) via parthenogenesis, the application in particular of the biological species concept is not made easy in case of thiarids.
In case of the thiarids it remains to be seen in how far they are actually prone exclusively to parthenogenesis. For example, for populations of Melanoides tuberculata in Israel
In view of the pronounced phenotypic plasticity reported herein for the Thai Tarebia granifera, it should be asked, in addition or alternatively to environmental factors, in how far this conchologically expressed variation is correlated to or even caused by these, at least frequently, parthenogenetically reproducing thiarids. Resulting in monoclonal lineages, populations of morphologically varying freshwater snails with partly or potentially completely parthenogenetic females hitherto have erroneously been treated as species under the traditional typological approach (not only in malacology). However, this simplistic and often non-comprehensive approach has most likely underestimated natural variation and intraspecific disparity by, at the same time, overestimating taxonomic diversity, resulting in taxonomic redundancy as an underrated phenomenon in evolutionary biology.
The development of an accurate and rapid method for the detection of males in aphallic thiarids, in order to evaluate the frequency of parthenogenesis in individual populations and species or higher-level taxa, respectively, remain an essential desideratum in biosystematics research on these snails. In addition, it remains to be analysed thoroughly whether and in how far there is a correlation of partially or completely parthenogenetic populations with parasite infections by digenic trematodes, for example, in the thiarids Melanoides tuberculata (see
Our preliminary analyses of the brood pouch content in the latter species under study here revealed that infected females tend to have fewer embryos than non-infected specimens, which might be a hint to the influence of parasite load on the reproductive mode of this major intermediate host. Therefore, given the human infection aspects of these trematode-carrying gastropods, our study not only has implication for human health in Thailand. We also hope that with studying trematode infections in the various conchologically disparate and molecular genetically distinct lineages of Tarebia we will eventually gain deeper insights into the complex evolutionary interplay of various trematode parasites and their snail hosts mediating infections in the human population.
This research was supported by the Thailand Research Fund through the Royal Golden Jubilee Ph. D. Program (Grant No. PHD/0093/2556) to Nuanpan Veeravechsukij and Duangduen Krailas. Both and Matthias Glaubrecht also thank the Deutsche Akademische Austauschdienst (DAAD) and the Deutsche Forschungsgemeinschaft (DFG; grant GL 297/29-1) for financial support of this study. We are grateful to the Department of Biology, Faculty of Science, Silpakorn University for support. We also thank Vince Kessner (Adelaide River, Australia) very much for collecting thiarids in Timor Leste and for providing material of Tarebia to one of us (MG) for study. Cennet Gerstage (CeNak, Hamburg) helped with statistics. We are indebted to two anonymous reviewers for their instructive comments and suggestions to the manuscript version.