Karenia papilionacea, A. I. Haywood et K. A. Steidinger, 2004

Al-Kandari, Research Article Manal, Saburova, Maria, Polikarpov, Igor, Larsen, Jacob, Lundholm, Nina & Hussain, Sumaiah, 2025, Morphological and molecular characterization of Kareniaceae (Dinophyceae, Gymnodiniales) in Kuwait’s waters, Botanica Marina (Warsaw, Poland) 68 (2), pp. 155-173 : 166-167

4.2 Karenia papilionacea

The taxonomic survey by Al-Kandari et al. (2009) revealed the second morphotype of Karenia in Kuwait ’ s waters. It was distinguished from K. selliformis by its cell shape, being wider rather than oblong. The small globular nucleus of this morphotype was located in the left lobe of the hypocone versus the larger, transversely elongated, and posteriorly located nucleus in K. selliformis (Table 3). While the morphology of this taxon resembled K. brevis , its unambiguous identification remained uncertain for a long time. In taxonomic and ecological phytoplankton studies in Kuwait ’ s waters, this taxon has been variously referred to as either Gymnodinium breve (Al-Yamani et al. 2004) , Karenia cf. brevis ( Al-Kandari et al. 2009, plate 8A, LM), K. brevis ( Devlin et al. 2019) , or K. papilionacea ( Al-Yamani and Saburova 2019; Al-Yamani et al. 2012, plate 60, LM; Polikarpov et al. 2020) ( Table 4).

Morphologically, K. papilionacea is similar to K. brevis , and these two species can be distinguished only by minor details. The cells of K. papilionacea are wider (24 – 36 µm vs 18 – 48 µm), possess pointed rather than bulbous apical protrusions (carina), and have slightly deeper antapical excavations ( Haywood et al. 2004). Given the high morphological variability observed in both species, even within the same strain ( Persson et al. 2013; Stuart 2011), it is doubtful whether species discrimination based on morphological criteria alone can be considered reliable. Nevertheless, these two species are genetically distinct ( Haywood et al. 2004).

K. papilionacea may have previously been misidentified as K. brevis worldwide. The geographic range of K. brevis appears to be limited to the Gulf of Mexico ( Brand et al. 2012; Magaña et al. 2003; Steidinger 2009). Previous reports of fish mortality caused by K. brevis in Asia and the Mediterranean Sea require molecular analysis to verify if they are actually misidentifications of K. papilionacea or other Karenia species (e.g., Tsikoti and Genitsaris 2021; Yamaguchi et al. 2016; Yeung et al. 2005).

The seven strains of K. papilionacea examined in this study displayed similarity to each other, and the morphological characters observed under LM were consistent with previous reports of this species (e.g., Al-Yamani and Saburova 2019; Kim et al. 2023; Yamaguchi et al. 2016), including its original description ( Haywood et al. 2004). Despite some morphological variability within and among strains in cell size and shape, the typical cells were wide, dorsoventrally flattened, with short apical grooves, pointed apices, and round nuclei in the left lobe of the hypocone ( Figure 2 ; Table 3).

In the phylogenetic tree based on partial LSU rDNA sequences, the strains from Kuwait were grouped together within the monophyletic clade of K. papilionacae among strains isolated from New Zealand (the type locality) and Japan with strong statistical support (Figure 5). The sequences obtained from Kuwait ’ s strains were identical, sharing 99.7 % similarity (2 bp differences) to that of the type strain of K. papilionacea ( U92252 ), and were clearly genetically distinct from the K. brevis clade. Therefore, our results provide the first unambiguous molecular identification of K. papilionacea from Kuwait ’ s coastal waters and resolve the previous regional taxonomic uncertainty for this species. This implies that earlier records of K. brevis -like species in Kuwait ’ s waters need to be reconsidered and attributed to K. papilionacea . Furthermore, the genetic similarity among the strains isolated seven years apart (first in 2014 and then in 2021) and from different sampling sites ( Kuwait Bay and semi-enclosed marina, Figure 1C ; Table 1) suggests the persistent presence of the same population of this species throughout the study area.

Similar to K. selliforms , recent phylogenetic analyses based on both LSU and ITS have revealed genetic divergence among the K. papilionacea strains, resulting in the distinction of at least two phylotypes within this species ( Kim et al. 2023; Yamaguchi et al. 2016). No morphological differences were observed between phylotypes, while different physiological growth traits were assumed to distinguish them ( Yamaguchi et al. 2016). In our LSU-based phylogeny, the topology of the K. papilionacea clade was almost the same as previously reported by Yamaguchi et al. (2016) and Kim et al. (2023) and consisted of three distinct, well-supported subclades (Figure 5). All of Kuwait ’ s strains were placed in a large, diverse, and strongly supported clade (89/0.99) among strains from Australia, New Zealand, Japan, China, and Korea, corresponding to the original phylotype (as per Yamaguchi et al. 2016). The second distinct subclade, with high nodal support (85/0.99), solely comprised K. papilionacea strains restricted to the western Japanese coast ( Yamaguchi et al. 2016), representing phylotype I. Two strains originating from the French Atlantic and north-western Mediterranean Sea, Spain, were genetically divergent from those of the original phylotype and phylotype I, clustering separately into a well-supported subclade (86/0.99).

Figures

Figure 1: Area of investigation: (A) general location of the sampling area; (B) map of Kuwait’s marine area; (C) satellite image of Kuwait Bay and adjacent waters showing location of sampling sites.

Figure 2: Light micrographs of Karenia papilionacea cells (Figure 2A–C: strain KW-F3- 015;Figure 2D–G: strain KW-E1-014; Figure 2N: strain KW-G4-062; Figure 2H–M, O-R: isolates from field samples; arrows point to the apical groove; arrowheads point to the open sulcal extension onto the epicone; n marks the nucleus): (A, B) ventral view showing apical groove, open-ended sulcus onto the epicone, and nucleus in the left lobe of hypocone; (C) epifluorescence microscopy under bluelight illumination showing numerous kidneyshaped chloroplasts; (D, E) ventral and dorsal views of the same cell showing apical groove, sulcus, and nucleus; (F) apical view in deep focus showing dorso-ventral compression; (G) epifluorescence microscopy showing several elongated chloroplasts; (H) two cells in ventral and dorsal view; (I–K) cells in left lateral, apical, and antapical views showing dorso-ventral compression; (L) ventral view showing apical groove and open-ended sulcus onto the epicone; (M) dorsal view showing apical groove over epicone; (N) Lugol’s-fixed cell in ventral view; (O–R) contracted cells during butterfly-like movement. Scale bars: 10 µm.