Asymmetric Reproductive Isolation between Two Sympatric Annual Killifish with Extremely Short Lifespans

BACKGROUND: Interspecific reproductive isolation is typically achieved by a combination of intrinsic and extrinsic barriers. Behavioural isolating barriers between sympatric, closely related species are often of primary importance and frequently aided by extrinsic factors causing spatial and temporal interspecific separation. Study systems with a severely limited role of extrinsic factors on reproductive isolation may provide valuable insights into how reproductive isolation between sympatric species is maintained. We used no-choice experimental set-up to study reproductive barriers between two closely related sympatric African killifish species, Nothobranchius furzeri and Nothobranchius orthonotus. These fish live in small temporary savannah pools and have complete spatial and temporal overlap in reproductive activities and share a similar ecology. PRINCIPAL FINDINGS: We found that the two species display largely incomplete and asymmetric reproductive isolation. Mating between N. furzeri males and N. orthonotus females was absent under standard experimental conditions and eggs were not viable when fish were forced to mate in a modified experimental setup. In contrast, male N. orthonotus indiscriminately mated with N. furzeri females, the eggs were viable, and offspring successfully hatched. Most spawnings, however, were achieved by male coercion and egg production and embryo survival were low. Behavioural asymmetry was likely facilitated by mating coercion from larger males of N. orthonotus and at relatively low cost to females. Interestingly, the direction of asymmetry was positively associated with asymmetry in post-mating reproductive barriers. SIGNIFICANCE: We showed that, in fish species with a promiscuous mating system and multiple matings each day, selection for strong mate preferences was relaxed. This effect was likely due to the small proportion of resources allocated to each single mating and the high potential cost to females from mating refusal. We highlight and discuss the fact that males of rarer species may often coercively mate with females of a related, more abundant species

Nothobranchius pienaari Shidlovskiy, Watters & Wildekamp, 2010, sp. nov.

<i>Nothobranchius pienaari</i> sp. nov. <p>(Figs. 6–8)</p> <p> <i>Nothobranchius rachovii,</i> non Ahl, 1926: Pienaar, 1968: 64</p> <p> <b>Holotype.</b> MRAC A4-039-P-0081, adult male, 36.4 mm SL, Mozambique, Chokwe to Massingir road, 101 km SE of Massingir, Mazimechopes River system, 24° 24.7' S, 32° 46.6' E, B. Watters, R. Wildekamp, B. Cooper, J. Jordaan, J. Bornman and J. Ippel, 10 April 2004.</p> <p> <b>Paratypes.</b> MRAC A4-039-P-0082-0083, two females, 35.8–38.5 mm SL, collected with holotype; MRAC A4-039-P-0022, one male, 30.8 mm SL; Mozambique, EN1 road from Vilankulo to Beira, SE of Mexungue, Gorongose River system, 20° 42.97' S, 34° 07.86' E; B. Cooper and R. Wildekamp, 31 March 2004; MRAC A4-039-P-0066-0072, 2 males, 5.3–38.0 mm SL, 5 females, 21.8–30.3 mm SL, Mozambique, road Nhamatanda (Vila Machado) to Inchope, 8 km SE of Dundo, Pungwe River system, 19° 40.7' S, 34° 46.7' E, B. Watters, R. Wildekamp, B. Cooper, J. Jordaan, J. Bornman and J. Ippel, 4 April 2004.</p> <p> <b>Diagnosis.</b> Morphology similar to <i>N. rachovii</i> and <i>N. krysanovi</i> but distinguished from those by a slightly deeper body in males (see Table 3). <i>Nothobranchius pienaari</i> is distinguished from other species of the genus by the following combination of characters: male with colouration consisting of alternating blue-green to grey-green and dark red-brown to almost black scale margins and oblique bars on body and dorsal and anal fins, dark brown spotted basal half of caudal fin, and orange subterminal and black terminal band. It can be distinguished from the other species of the subgenus <i>Nothobranchius</i> as follows: from <i>N. furzeri</i> by male colouration and a slightly higher number of dorsal fin rays (14–16 vs. 14 or 15); from <i>N. orthonotus</i> by male colouration, lower number of scales on the mid-longitudinal series (26–31 vs. 28–33) and from <i>N. kadleci</i> by male colouration and shape of the frontal region, higher number of dorsal and anal fin rays (14–16 vs. 13 or 14 and 15–17 vs. 13 or14). Female distinguished by faint brown oblique bars on posterior part of flanks. <i>Nothobranchius pienaari</i> can also be distinguished from <i>N. rachovii</i> and <i>N. krysanovi</i> by a much higher number of diploid chromosomes (2n = 34 vs. 2n = 16 for <i>N. rachovii</i> and 2n = 18 for <i>N. krysanovi</i>). It can also be distinguished from the karyotypes of <i>N. furzeri</i> and <i>N. orthonotus</i> that both have with 2n = 38 a higher number of diploid chromosomes (Scheel, 1981).</p> <p> <b>Description.</b> Morphometric data for <i>N. pienaari</i> are presented in Table 3. Robust <i>Nothobranchius</i> species of medium size. Snout slightly pointed, mouth terminal, slightly upward directed. Body laterally compressed and deep. Dorsal profile concave on head and convex from nape to end of dorsal fin, especially in older males. Upper and lower profile of caudal peduncle straight.</p> <p>Dorsal fin rays 14–16; anal fin rays 15–17. Pelvic fins short, not reaching first anal fin ray. Pectoral fin reaching past first pelvic fin ray.</p> <p>Scales on median longitudinal line 26–31+2 or 3 on caudal fin base, most with shallow pit in centre with one neuromast. Transverse row of scales above pelvic fin 11 or 12. circumpeduncular scales 12 or 13. Supraorbital squamation G-type. Anterior scales partly covered with epidermal tissue. Frontal and central supraorbital neuromast systems fused, forming two distinct, shallow grooves, both lined with three shallow lobes on both sides. Posterior cephalic neuromast system in two curved grooves.</p> <p>Male: maximum observed size 38.5 mm SL. Body laterally compressed and deep. All unpaired fins rounded, dorsal and anal fin covered with thin layer of epidermal tissue. Papillae on all rays of dorsal and anal fins, projecting slightly from epidermis. Opercular membrane projecting from opercle, distal end wrinkled.</p> <p>Female: smaller than male, maximum observed size 35.8 mm SL. Body less deep than male. Dorsal and caudal fin rounded, anal fin more posteriorly situated than in male, triangular, tip rounded, rays 3–8 longer, more rigid and branched only near tip. No epidermal tissue on dorsal and anal fins, and no papillae on rays.</p> <p> <b>Colouration.</b> Live male (see Figs. 6 & 7): body colour light to dark blue-green to green scale centres. Scales with cherry-red to dark red-brown, sometimes almost black margins. These margins form narrow oblique bars, lower end forward. Bars on every second row of scales. Males of some populations appear very dark to almost black. Head grey-brown. Abdomen lighter grey-brown to pale pink, back olive-brown. Operculum with 3-4 dark red-brown oblique stripes. Throat pink to silver-grey depending on population. Projecting part of branchiostegal membrane brick-red or red-brown; may have narrow light blue edge. Iris bronze or golden with incomplete vertical dark bar. Proximal part of caudal fin silver-grey, with bronze sheen in some cases, and small dark brown markings between rays. In some populations these markings can merge and form irregular cross bars. Caudal fin margin black with, in most populations, a red to orange submarginal band. Width of this band varies between populations from broad to narrow or barely discernable. Dorsal fin light blue-grey with bronze hue and numerous dark red-brown or brown spots and stripes which may merge to form irregular bands across the fin rays. Submarginal part of dorsal fin green-grey or blue-grey with small dark spots. Dorsal fin with narrow white-blue margin. Anal fin blue-grey with irregular red-brown spots that may merge to bands. Markings are larger in proximal part of fins. Submarginal parts darker; may have narrow blue-white margin. Ventral fins blue-grey with red-brown markings in basal third. Pectoral fins colourless with blue-white margin. Iris bronze to golden with dark cross-bar. <i>Nothobranchius pienaari</i> is polymorphic in the colouration of its caudal fin. The orange submarginal band ranges from wide and very distinct to almost absent, depending on the population.</p> <p>Live female (see Fig. 8): body colour mainly grey-brown, darker on the back and lighter on abdomen. Posterior part of flanks with several darker but faintly developed bars, inclined to the rear. Fins colourless. Iris bronze or golden with incomplete vertical dark bar.</p> <p> <b>Distribution and habitat.</b> <i>Nothobranchius pienaari</i> occupies a vast area in the coastal lowlands of central and southern Mozambique and the adjacent part of northeastern RSA (see Fig. 3). It is known from temporary pools and swamps in the floodplains of several rivers flowing into the Indian Ocean south of the lower Zambezi River to the basins of the Chefu, Limpopo, and Mazimechopes Rivers in southwestern Mozambique and northeastern RSA. The Kruger National Park population in the RSA has a unique elevated position but is also within the drainage system of the Limpopo River (Watters, 2006, Watters <i>et al.</i>, 2009). Usually found in water-filled depressions in the floodplains of rivers and bordering swamps. Water depth is variable with the progress of the season. Aquatic vegetation may consist of <i>Nymphea</i>, <i>Ottelia</i>, <i>Lagarosiphon</i> and <i>Utricularia</i> species. Other fish species that may occur with <i>N. pienaari</i> are <i>N. furzeri, N. kadleci, N. orthonotus</i>, as well as non-annual fishes like <i>Clarias gariepinus</i>, <i>Protopterus annectens</i> and unidentified <i>Barbus</i> species (Watters <i>et al.</i>, 2009, Reichard <i>et al.</i>, 2009, Reichard, 2010).</p> <p> <b>Etymology.</b> Named after Dr. U. de V. Pienaar, retired biologist of the Kruger National Park, Republic of South Africa, and discoverer of <i>Nothobranchius</i> species, in honour of his efforts to protect and conserve these populations. The species name is pronounced “pea-nahri”.</p>Published as part of <i>Shidlovskiy, Konstantin M., Watters, Brian R. & Wildekamp, Rudolf H., 2010, Notes on the annual killifish species Nothobranchius rachovii (Cyprinodontiformes; Nothobranchiidae) with the description of two new species, pp. 37-57 in Zootaxa 2724</i> on pages 49-51, DOI: <a href="http://zenodo.org/record/199938">10.5281/zenodo.199938</a&gt