Corresponding author: Hendrik Freitag (
Academic editor: L. Penev
The genus
Publications dealing with molecular data of
The first
The larvae examined were partly collected by standardized methods for ecological studies such as colonization and drift sampling during a survey in 2000 / 2001 (
All materials from more recent surveys have been retrieved by means of manual collection from submerged wood debris or through the use of a fine-meshed hand net. This material was preserved in absolute ethyl alcohol and thus, it was suitable for genetic sequencing. The best manual sampling was possible in permanent small to medium sized rivers in forested area (
The label codes for the sampling sites of the first author are arbitrary. They do not follow any temporal or spatial pattern, except for the fact, that eventually varying small letters following a common code number refer to different sections of the same water system.
Small to medium sized, permanent forest streams with heterogeneous morphology and rich in submerged woods and root packs are usually inhabited by the highest numbers of
DNA was extracted from 15 whole specimens (
Phylogram of the consensus tree of the Bayesian analysis with branch lengths measured in expected substitutions per site. Posterior probability values (printed when > 0.5) at respective branches. Sample labels with island of origin and code of the collection site or genbank code.
Additionally, a central part of the cytochrome b apoenzyme (
Scanning electron microscope (SEM) images were obtained using a ZEISS EVO 50 XVP at SMTD. The specimens were coated with gold using two samples each if enough material was available. Those taxa of which only a single or few specimens have been available were only vacuum dried, but not gold-coated prior to scanning. This resulted in lower quality of the micrographs, but has kept the specimen´s surface natural.
Digital habitus photographs were taken with a NIKON SMZ800 stereo microscope with digital photo adapter NIKON DS-Fi1 (unit in DLSU). These photographs were taken at various focus layers and were subsequently combined using COMBINEZM software (
Examination, biometric measuring and imaging of dissected parts were conducted using a NIKON Eclipse 600 microscope with a ZEISS AxioCam MRc5 digitalcamera (unit in SMTD). The morphological details of larvae are described from external view as they are usually visible without dissection, if not stated opposite. The listing of examined material includes the head width (in mm) of all measured larvae.
Morphological terminology used herein mainly follows the
above sea level (altitude)
colonization sample
calculated length (PL + EL)
Coarse Particulate Organic Matter
drift sample
elytral length
elytral width
exemplar / exemplars of adult specimen
Fine Particulate Organic Matter
head width
interocular distance
larva / larvae
manual collection
maximum pronotal width
pronotal length
H. Freitag private collection, Dresden, Germany
Phyllodrom, Institut und Museum für Regenwaldökologie Leipzig, Germany
Natural History Museum Vienna, Austria
Museo Nacional de Ciencias Naturales Madrid, Spain
Palawan Council for Sustainable Development, Philippines
Senckenberg Museum für Tierkunde Dresden, Germany
University of the Philippines Los Baños, Museum of Natural History, Entomological Collection, Philippines
Zoological Museum of the University Copenhagen, Denmark
Zoological State Collections Munich, Germany
Alignment of the
The sequence divergence between
All adults and larvae could be matched unambiguously. Sequences of adult and larva of the same species from the same locality or island showed low difference of positions in most species (2–3 positions different, 0.2–0.4% divergence).
The sequencing of
All sequences were submitted to GENBANK. Accession numbers and curatory information are listed in
Genbank accession numbers of DNA sequences, geographical origins, collectors, collection sites and organismic sample references of specimens used for molecular-genetic analyses.
Species | Stage | Locality | Site | Collector | Voucher |
|
|
adult | Palawan | 16h | Freitag | NMW FR 037 | HE588174 | - | |
larva | Palawan | 16h | Freitag | ZSM FR 038 | HE588175 | - | |
adult | Palawan | 154 | Freitag | ZSM FR 001 | HE588179 | - | |
larva | Palawan | 159 | Freitag | ZSM FR 025 | HE588180 | - | |
adult | Palawan | 154 | Freitag | ZSM FR 008 | HE588169 | - | |
larva | Palawan | 154 | Freitag | ZSM FR 002 | HE588170 | - | |
adult | Palawan | 16f | Freitag | ZSM FR 003 | HE588172 | - | |
larva | Palawan | 16b | Freitag | ZSM FR 040 | HE588173 | - | |
adult | Busuanga | 165 | Freitag | ZSM FR 032 | HE588178 | - | |
adult | Palawan | 20 | Freitag | ZSM FR 005 | HE588176 | - | |
larva | Palawan | 20 | Freitag | ZSM FR 006 | HE588177 | - | |
adult | Busuanga | 169 | Freitag | ZSM FR 013 | HE588168 | - | |
adult | Palawan | CR4 | Freitag | ZSM FR 007 | HE588167 | HE588183 | |
larva | Palawan | CR4 | Freitag | ZSM FR 061 | - | HE588184 | |
adult | Borneo | Čiampor | MNCN FC-B05 | DQ266500 | DQ266511 | ||
larva | Busuanga | 169 | Freitag | ZSM FR 014 | HE588171 | HE588182 | |
adult | Sri Lanka | 1 | Freitag | ZSM FR 035 | HE588181 | - |
2L (0.21, 0.24) (PCSD) “PHIL.: Palawan, P. Princesa Panaguman R., Marufinas
Habitus of
Colour as in
HW c. 0.27 mm; entire larva about 3.0 mm long. Body torpedo-like elongate, subsemicircular in cross section, dorsally vaulted, ventrally almost flat. Posterolateral margins of abdominal segments I–VIII moderately produced (
Head (
Pro-, meso- and metathorax (
Legs (
Abdomen (
Almost the entire material studied belongs to the final instar stage. The two presumably prefinal instar specimens do not vary conspicuously from the description above. The legs appear slightly broader and shorter in relation to the body and their setae patterns are slightly different.
Only known from Palawan and Mindoro (Freitag & Pangantihon, 2010).
1♀ (ZMUC) “PHIL.: Palawan, P.Princesa; Bgy. Binduyan, Olanguan Falls/Brdg., Nat. Highway km 29; 2.5 km upstr.,rocks, gravel boulder, CPOM; degr. prim. veget.,
Colour (
HW 0.32 mm; entire larva about 3.0 mm long. Body elongate, broader than that of
Posterolateral projections (
Head (
Pro-, meso- and metathorax (
Legs (
Abdomen (
Operculum (
The few available prefinal instar specimens vary from the description above by relatively slenderer thoracic and abdominal segments, the relatively longer posterolateral projections, the distinctly shorter and broader legs and fewer setae on tibiae and femora.
The species can most easily be distinguished from
Known from Palawan island.
11L (2 × 0.17, 2 × 0.22, 0.23, 0.24, 3 × 0.27, 0.28, 2 × 0.29) (NMW, ZMUC, CFP) “PHIL.: Palawan, P. Princesa Panaguman R.
This species was originally described based on a single male exemplar. As additional material including females have been collected since then we will provide a short diagnosis of the female characters and sexual dimorphisms. Furthermore, the aedeagus (
Ovipositor as in
Sternite VIII in female (
Colour (
HW c. 0.29 mm; entire larva about 3.0 mm long. Body shape as in
All abdominal posterolateral projections (
Head (
Pro-, meso- and metathorax (
Sublateroposterior portions and anteriomedian sclerites of thoracic venters with setiferous tubercles (
Legs (
Abdomen (
For this species, quite a number of prefinal instar specimens is available. They vary from the final instar description by the overall paler colour, relatively longer posterolateral projections, relatively shorter and broader legs and fewer setae on tibiae and femora.
The species can most easily be distinguished from its congeners
Only known from few rivers in central Palawan.
1♂ (ZSM [FR032]) PHIL.: Busuanga, Coron, San Nicolas/Borac, “7Falls" mount. creek; riffle & pool; gravel, boulders, CPOM, sec. forest/rural; c.50m asl,
Colour as in
HW 0.32 mm; entire larva up to 3.5 mm long. Body shape as in
Head as in
Pro-, meso- and metathorax as in
Abdomen (
Specimens of 4th and 5th instar vary only little from the final larval instar. The species typical dorsosagittal carinae of the posterior abdominal segments are very distinct. However, the emargination of the abdominal segment IX` apex is not conspicuous, the overall colour is paler, the thoracic and abdominal segments are relatively narrower, the legs are slightly shorter and broader and the setae on tibiae and femora are fewer and not well arranged in rims.
The species can clearly be distinguished from other Palawan species by the lack of pale dorsal colour patches and its highly elevated dorsosagittal carinae which is most conspicuous at the posterior portion of abdominal segment VIII where it appears as a drop-shaped protuberance.
Only known from Palawan and Busuanga.
Body 1.82–1.88 mm long (CL), 0.79–0.83 mm broad (EW), 2.2–2.4 times as broad as wide (CL/EW). Body form elongate, moderately convex dorsally.
Colouration (
Head (
Pronotum (
Scutellum subpentagonal, anteriorly slightly impressed, glabrous. Elytra (
Legs slightly longer than body; pro- and mesocoxae large, globular; metacoxae only slightly protruding laterally; femora, tibiae, and tarsi (except distal tarsal segment) covered with elongate setiferous tubercles; tibiae distally with a distinct rim of setae; claws (
Ventrites 1–4 almost glabrous, posteromedially punctate, reticulate anteriorly and laterally; ventrite 5 (
Sternite IX (spiculum gastrale) as in
Aedeagus (
Ovipositor (
Secondary sexual characters. Sternite VIII in female (
Colour as in
HW 0.37–0.41 mm; entire larva up to 3.3 mm long. Body shape as in
Head (
Pro-, meso- and metathorax as in
Abdomen (
The two specimens of prefinal instar stage vary most conspicuously from the above description by the overall paler brown colour, the more conspicuous dorsal setiferous tubercles that let the pronotal signa appear clearly as well as the slightly crested (in cross-section subtriangular) abdominal segment IX.
The species is easily distinguishable from other Palawan species by the lack of any dorsosagittal carinae or elevations, the dark, shiny dorsal colour, its rather large size and the rather shallow dorsal tubercles bearing comparably long, lanceolate to trichoid setae.
Only known from the type locality in central Palawan and one site in northern Palawan (
The species is named in reference to the remote mountainous river habitats where it was exclusively recorded from.
1 L (0.61) (ZSM [FR014]) “PHIL.: Busuanga, Coron; Guadelupe, Balolo R./Brdg. Nat.Rd. km 14; lowld. creek; sec.veget.; run, gravel, CPOM, c.10m asl,
Colour (
HW c. 0.62 mm, entirely c. 3.7 mm long. Body flattened dorsoventrally, moderately vaulted dorsally, almost flat ventrally, with sagittal line (longitudinal groove from prothorax at least up to 5th abdominal segment). Dorsal side moderately densely covered with setiferous tubercles (
Head (
Prothorax subquadrate, almost as long as broad, with round signa (glabrous areas) in posterior half and near depressed sagittal line. Meso- and metathorax subtrapezoidal, distinctly broader than long, distinctly shorter than prothorax (
Legs (
Abdomen (
The two final instar specimens available do not allow to draw conclusions about variations between the instars.
The species resembles the previous ones only in very general characters, such as the presence of posterolateral appendages and the distribution of spiracles. This larva is, however, not torpedo-like elongate and subsemicircular in cross section, but dorsoventrally somewhat depressed, only slightly vaulted dorsally. By this it rather resembles the species
Known from Busuanga, Philippines (recent study;
21 exs., 1L (0.45) (PCSD, SMTD, ZMUC, IMRL, CFP, NMW, ZSM [FR007, FR061]) “PHIL.: Palawan, P. Princesa S Manturon, Cabayugan R.
Colour as in
HW c. 0.50 mm, entire larve up to 4.5 mm long. Body shape somewhat similar to that of
Head (
Prothorax slightly broader than long; tergum with irregularly shaped signa in posterior half. Venter of pro-, meso and metathorax (
Legs as in
Abdomen (
Specimens of the 3rd to the 6th (final) instar stage are available for study. Within this range it is obvious that the younger the specimens the paler they are, the longer are all kinds of setae (in relation to the body) and the fewer setiferous asperities are present. Additionally, the limb setae are varyingly arranged and the terminal abdominal segment is relatively shorter and broader as well as the legs. The latter is most obvious between the final and prefinal instar stages.
The larvae of
Only known from Palawan and Busuanga.
Collecting sites of the
Body form subsemicircular in cross-section or dorsoventrally flattened (depressed), with posterolateral projections or processes on abdominal segments I–VIII and lateral spiracles on mesothorax and abdominal segments I–VIII. Ventral prothorax with five sclerites (including one posteromedial sternellum), meso- and metathorax with six sclerites. Abdomen with pleura on segments I–VIII.
1 | Body torpedo-like elongate, subsemicircular in cross section. Posterolateral abdominal projections small, lobate with tip posteriad directed ( |
2 |
– | Body flattened dorsoventrally (depressed), only slightly vaulted. Posterolateral abdominal projections large, conical with tip posterolaterad directed ( |
6 |
2 | At least terminal segment, but usually several posterior abdominal segments with dorsosagittal carina or elevation ( |
3 |
– | All abdominal segments without dorsosagittal carinae or elevations ( |
|
3 | Some peripheral areas of dorsal pronotum with obvious pale colour patches ( |
4 |
– | Entire dorsal surface colour without obvious pale patches ( |
|
4 | Pronotum anteriorly with small transverse yellow band (less broad than 1/4 of pronotum, |
5 |
– | Pronotum anteriorly with broad transverse, rather pale band (extended over c. 1/3 of pronotum, |
|
5 | Anterior yellowish band of pronotum slightly extended posteriad along the midline ( |
|
– | Anterior yellowish pronotal band not extended posteriad along the midline ( |
|
6 | Frons with one median pointed projection (“horn”) ( |
|
– | Frons without median pointed projection ( |
|
1.
2.
3.
4.
5.
6.
7.
8.
9.
10.
We successfully used mitochondrial DNA sequencing to associate different life stages of beetles with each other, substantiated by morphological description of larvae.Use of DNA sequences has helped to avoid potential pitfalls, as for
The morphological species groups suggested by Freitag & Jäch (2007) were supported here by morphological characters of the larvae. Two main types are recognized. The
The female and the larva of
The first author wishes to express his deep gratitude to the curator of the World Water Beetle Collection & Research Centre at the Natural History Museum Vienna, Austria, Dr. Manfred A. Jäch who provided excellent working conditions and uncomplicated access to the