Sunday, 27 September 2020

Bhavania annandalei: The identity and distribution of a a Hillstream Loach endemic to the Western Ghats of India.

The Hillstream Loach, Bhavania annandalei, was described by Sunda Lal Hora in 1920 from Tenmalai, in what was then Travancore State (current day southern Kerala), and suggested that the species occurs throughout the southern Western Ghats in the Nilgiris, Malabar, and Travancore. Hora diagnosed Bhavania annandalei from its only known congener, Bhavania australis by a combination of characters; the most prominent of which included a broad snout (rather than a pointed one), interrupted lower lip (as opposed to a continuous one), caudal-lobes equal (rather than a longer lower lobe), and presence of a pair of papillae on the lower lip (rather than thier absence). Hora’s description of Bhavania annandalei was however, based on a single adult female specimen collected by Thomas Nelson Annandale from Travancore, Kerala. Though, Hora (1920) seemed to have access to additional juvenile specimens collected by Captain Robert Sewell from the Nilgiris (Cherambadi) and Wayanad (Nellimunda, Mananthavady, and near Vythiri), he did not examine them or provide other details. Subsequently, Hora extended the distribution of the species to Mysore, based on four specimens collected by M.S. Bhimachar from a stream between Kottigehar and Balehonnur (in what was then Mysore State, i.e. the current day Tunga River System in Karnataka). No details of the specimens were provided. 

In his review on ‘Homalopterid fishes from Peninsular India’, Hora synonymized Bhavania annandalei with Bhavania australis, after examining specimens from throughout its distribution range including Kallar/South Travancore (current day Vamanapuram River, Kerala); Pampadumpara/North Travancore (current day Periyar River, Kerala); Sethumadai Hills/Mysore (current day Anamalai hills near Pollachi, Tamil Nadu); and Kottigehar/Mysore (current day Tunga River, Karnataka), and realising that his description of Bhavania annandalei was based mainly on immature specimens. This synonymy was subsequently adopted by Ambet Gopalan Kutty Menon in his review of the Homalopterid Loaches of India, but without examining the type (or fresh topotypes) of Bhavania annandalei, or the topotypes of Bhavania australis. Later workers followed this synonymy and considered Bhavania to be monotypic. ‘Bhavania arunachalensis’, described in 2007 from Naodhing drainage in Arunachal Pradesh, is considered to be a species of doubtful identity and uncertain placement, and is most likely a species of the genus Balitora

Given their hill-stream adaptations (widespread paired fins, flattened ventral surfaces with body suckers and rasping mouths on their ventral surface allowing them to firmly grasp rock or gravel surfaces necessary in the mountain torrents), and the fact that the type locality of Bhavania annandalei (Tenmalai) and Bhavania australis (Walayar) are at least 300 km apart and separated by two significant biogeographic barriers - the Palghat Gap and the Shencottah Gap, it is highly unlikely that the two are conspecific.

In a paper published in the journal Threatened Taxa on 26 July 2020, Remya Sundar of the Center for Aquatic Resource Management and Conservation at the Kerala University of Fisheries and Ocean Studies, VK Anoop and Arya Sidharthan of the School of Ocean Science and Technology, also at the Kerala University of Fisheries and Ocean Studies, Neelesh Dahanukar of the Indian Institute of Science Education and Research and the Zoo Outreach Organization, and Rajeev Raghavan, also of the Center for Aquatic Resource Management and Conservation and School of Ocean Science and Technology, and of the Department of Fisheries Resource Management, at the Kerala University of Fisheries and Ocean Studies, present a re-description of Bhavania annandalei, based upon newly collected specimens.

Six specimens of putative topotypic Bhavania annandalei were collected from Palaruvi falls at Tenmala (Kallada River), Kerala, and six specimens of putative topotypic Bhavania australis were collected from near the Kavarakund falls, upstream of Malampuzha Reservoir, Kerala, India. Samples were collected using a hand net/scoop net during early morning hours, fixed in 10% formalin and transferred to 70% ethanol for permanent voucher storage in the museum collections of the Kerala University of Fisheries and Ocean Studies. Gill tissues were obtained from fresh specimens and preserved in absolute ethanol.

Collection localities of putative topotyes of Bhavania annandalei and Bhavania australis. Sundar et al. (2020).

Bhavania annandalei is distinguished from its only known congener Bhavania australis by a combination of characters: low density and sparsely distributed tubercles on dorsal surface of head, especially on operculum, (whereas Bhavania australis has a high density of tubercles on dorsal surface of head and operculum); gape of mouth comparatively farther from snout tip, as a result the rostral barbels reaching anterior border of upper lip, (in Bhavania australis the gape of mouth is closer to snout tip, and rostral barbels reaching posterior border of upper lip); rostral flaps between the rostral barbels fleshier than in Bhavania australis; fewer post-dorsal scales (34–36 compared to 38–41); fewer scales above the lateral line (11–12 compated to 14–15); and caudal peduncle stout with its depth to width ratio 1.8–2.3 (Bhavania australis has a laterally compressed caudal peduncle with depth to width ratio 2.8–3.6).

Putative topotypes: (a) Bhavania annandalei; (b) Bhavania australis in life (specimens not preserved). Sundar et al. (2020).

The body of Bhavania annandalei is elongate, dorso-ventrally depressed anteriorly, laterally compressed posteriorly; dorsal profile convex, deepest at dorsal-fin origin. Body wider than its depth at dorsal-fin origin, deeper than wide at anus. Head small, rounded, less than one-fourth of standard length; depressed, longer than broad, with minute sparsely distributed indistinct tubercles on dorsal surface of head. Eyes small, dorso-laterally positioned, not visible from underside of head. Snout pointed in lateral view, round in dorsal view. Nostrils positioned dorsally, closer to anterior border of eye than to snout tip, anterior nostril situated inside a skin flap covering the posterior nostrils. Mouth inferior. Lips fleshy. Gape of mouth less than three times maximum head width. Barbels three pairs, two rostral: outer rostral barbels shorter than inner ones; one pair of maxillary barbels, situated slightly anterior to the angle of mouth. Three fleshy rostral flaps interspaced between rostral barbels. Gill opening small, restricted above the base of the pectoral fin.

Dorsal, lateral, and ventral images of putative topotypes: (a) Bhavania annandalei; (b) Bhavania australis. Sundar et al. (2020).

Body with scales except chest and belly. Lateral line complete, with 68–72 small scales. Caudal peduncle slender, its length almost three times its depth. Dorsalfin originating slightly behind the pelvic-fin origin, closer to tip of snout than to caudal-fin base; with two unbranched, followed by seven branched and a simple ray. Pectoral fin elongated, longer than head, with six unbranched, followed by 10 branched and a simple ray. Pelvic-fin length almost equal to head length; fin origin closer to snout tip than to end of caudal peduncle, its posterior end not reaching anus, with two unbranched and eight branched rays. Anal fin with two unbranched and five branched rays. Caudal fin forked, with 19 principal rays. 

Dorsal and ventral view of head: (a), (c) Bhavania annandalei; (b), (d) Bhavania australis. Sundar et al. (2020).

In life Bhavania annandalei is body is chestnut brown on dorsal and lateral sides, creamish-white on chest and belly; 3–4 prominent broad dark brown ventral bands; two broad ventral bands on the dorsalfin base. There are three black-coloured bands on the dorsal fin, 6–7 bands on the pectoral, three bands on the pelvic, 1–2 bands on the anal, and four bands across the caudal fin.

Morphometric analysis is a tool used by palaeontologists, archaeologists, anthropologists and forensic pathologists to analyse and compare specimens. It relies on taking numerous measurements of an object such as a bone or shell, and comparing both these measurements and ratios between measurements to those obtained from other specimens in order to establish relationships between them. 

Morphometric measurements were taken for 37 characters (measured to the nearest 0.1mm using digital calliper) and meristic values were determined for 10 characters using a stereo-zoom microscope. For meristic counts, values in parenthesis after the count respresent its frequency. For statistical analysis of morphometric data, subunits of body were taken as percentage of standard length and subunits of head were taken as percentage of head length. Principal component analysis was performed to check whether the two species formed distinct clusters in multivariate space using correlation matrix. Null hypothesis that the clusters are not significantly different from each other was tested using analysis of similarities employing Euclidian distances and 9999 permutations. Statistical analysis was performed in PAST 4.02.

Using size-adjusted characters, the two species clustered separately on the first two principal component analysis axes. The clusters were significantly different from each other, indicating that the species formed distinct clusters in multivariate space. While lengthlength relationships for most characters showed similar trends for both the species, there were two relationships that showed marked differences. Length-length relationship between caudal peduncle depth and width suggested that width increased rapidly with increasing depth in the case of Bhavania annandalei compared to Bhavania australis. Similarly, length-length relationship between head length and head depth at nape suggested that head depth increased rapidly with increasing head length in the case of Bhavania annandalei compared to Bhavania australis.

Genetic sequences of mitochondrial partial cytochrome oxidase subunit 1 (cox1) of topotypic Bhavania annandalei and Bhavania australis were obtained from the collected specimens. Additional sequences were downloaded from GenBank database. Gene sequences were aligned using MUSCLE and raw genetic distance was estimated using MEGA 7. Data were partitioned into three codon positions of cox1 gene. Partition analysis (a statistical method) and ModelFinder were used to find the right partitioning scheme and nucleotide substitution model for the partition scheme employing minimum Bayesian information criterion. Maximum likelihood analysis was performed in IQ Tree with best partition scheme and ultrafast bootstrap support for 1000 iterations. Phylogenetic tree was edited in FigTree v1.4.2.

Partition analysis and model selection identified separate nucleotide substitution models for all three codon positions, TNe+I for first codon, F81+F for second codon, TN+F+G4 for third codon position of cox1 gene. Maximum likelihood phylogenetic tree based on best partition scheme and model selection recovered Bhavania annandalei and Bhavania australis as a clade sister to Southeast Asian congeners of Balitoridae. Topotypic Bhavania annandalei (MT002520) differed from topotypic Bhavania australis (MT002518) with a raw genetic distance of 6.4% in the cox1 gene.

Maximum likelihood phylogenetic tree based on mitochondrial cytochrome oxidase subunit 1 gene using best partition scheme and model selection (lnL of consensus tree = -2631.97). Indoreonectes keralensis (Nemacheilidae) is used as an outgroup. Values along the nodes are percentage bootstraps based on 1,000 iterations. Sundar et al. (2020).

Bhavania annandalei is known with certainty from the Kallada, Vamanapuram, and Neyyar river systems in southern Kerala, India. These river systems drain the western slopes of the Agasthyamalai Hill ranges, south of the Shencottah Gap. It is highly likely that the species also occurs on the eastern slopes of the Agasthyamalai Hills particularly in the Tambaraparini River system in Tamil Nadu, but detailed surveys and voucher specimens are required to confirm this. In this context, Sundar et al. believe that previous records of Bhavania australis from several tributaries of the Tambaraparini, Manimuthar, and Chittar draining the eastern slopes of the Agasthyamalai, could most likely represent Bhavania annandalei.

The density of chromatophores in Bhavania is likely to be dependent on the micro-habitat as well as the colour and type of substratum it inhabits. Other ecological factors that may influence body colour are forest/canopy cover, intensity of light, turbidity, water flow and water temperature. This is reflected in the different body colours shown by the two species in different habitats and locations, an observation which was also made by Sunda Lal Hora. 

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