Introduction
The suborder Tintinnina Kofoid & Campbell, Reference Kofoid and Campbell1929 (Ciliophora, Spirotrichea, Choreotrichida) serves as an effective model for studying planktonic species diversity and biogeography, emphasising their roles in marine trophodynamics and biogeochemical cycles (Dolan and Pierce, Reference Dolan, Pierce, Dolan, Montagnes, Agatha, Coats and Stoecker2013; Pierce and Turner, Reference Pierce and Turner1993; Santoferrara and McManus, Reference Santoferrara, McManus, M.A and A.B2020). Tintinnid ciliates are unique in possessing a proteinaceous lorica that currently serves as the essential foundation for species determination in biodiversity assessments, despite their frequent interspecific similarities and polymorphism (Agatha and Simon, Reference Agatha and Simon2012; Dolan et al., Reference Dolan, Yang, Lee and Kim2013; Santoferrara et al., Reference Santoferrara, Bachy, Alder, Gong, Kim, Saccà, Da Silva Neto and Strüder-Kypke2016). However, recent cytological and molecular studies suggest that lorica-based species identifications are partly incorrect and might be misleading for defining species boundaries (Santoferrara et al., Reference Santoferrara, Tian, Alder and McManus2015, Reference Santoferrara, Bachy, Alder, Gong, Kim, Saccà, Da Silva Neto and Strüder-Kypke2016; Xu et al., Reference Xu, Sun, Shin and Kim2012). However, the majority of the known tintinnids are still based on loricae traits, underscoring the relevance of lorica-based identification given the current state of knowledge (Agatha and Tsai, Reference Agatha and Tsai2008; Santoferrara and McManus, Reference Santoferrara, McManus, M.A and A.B2020).
Tintinnid ciliates exhibit high diversity, classified into 14 families and 77 genera with ∼1000 species (Santoferrara and McManus, Reference Santoferrara, McManus, M.A and A.B2020). A checklist of tintinnids reported from the Indian coasts includes a total of 151 species belonging to 33 genera (Elangovan and Gauns, Reference Elangovan and Gauns2018). In contrast, the biodiversity in the oceanic realm, mainly the deeper euphotic regions, is inadequately explored. Additionally, studies corroborated with descriptions and morphometrics of tintinnid species from Indian waters are rare, except for some recent studies focusing on the morphological features of lorica and distributional records (Anandavelu et al., Reference Anandavelu, Asha Devi, Singh and Chowdhury2024; Sarun and Patil, Reference Sarun and Patil2024). The tintinnid genera have been classified as cosmopolitan, neritic, warm-temperate, boreal, or austral taxa (Dolan et al., Reference Dolan, Pierce, Yang and Kim2012; Pierce and Turner, Reference Pierce and Turner1993). Among them, the warm–temperate genus Codonaria Kofoid & Campbell, Reference Kofoid and Campbell1939, which includes one of the oldest species, Codonaria cistellula (Fol, Reference Fol1883) Kofoid & Campbell, Reference Kofoid and Campbell1939, has been reported from the Mediterranean Sea, the Atlantic Ocean, and the western tropical Pacific Ocean (Balech, Reference Balech1959; Brandt, Reference Brandt1907; Fernandes, Reference Fernandes2004; Fol, Reference Fol1883; Gold and Morales, Reference Gold and Morales1977; Jörgensen, Reference Jörgensen1924; Kofoid and Campbell, Reference Kofoid and Campbell1929, Reference Kofoid and Campbell1939; Paulmier, Reference Paulmier1997), but not from the Indian Ocean. The occurrence of C. cistellula is reported for the first time from the Indian Ocean, along with details of its lorica structure.
Materials and methods
Regular monitoring studies were conducted between 2018 and 2019 onboard FORV Sagar Sampada and ORV Sagar Kanya to understand the marine ecosystem process of the Eastern Arabian Sea (∼6–22°N and 77–67°E), with further details available in Sherin et al. (Reference Sherin, Gupta, Sudheesh, Ramu, Reddy, Harikrishnachari and Vijayan2023). As part of the study, 5–8 l of seawater collected during March and May 2018 was concentrated by gravity settling, and the microzooplankton were obtained by siphoning through a 20 µm bolting silk (Hydro-Bios). The samples were then fixed with 3–8% acid Lugol’s iodine and stored for approximately 6 years until analysis in 2024. The samples were analysed and measured at 400× magnification using an inverted LEICA DMi8 microscope equipped with a LEICA DMC4500 camera. A total of nine specimens of Codonaria cistellula were recorded from the Mumbai (69.65° E, 19.06° N and 69.46° E, 19.07° N), Veraval (69.54° E, 20.02° N), and Okha (67.90° E, 21.40° N) transects, and restricted between 75 and 100 m depth along offshore contours (100–2000 m). Since most specimens (five individuals) were empty loricae or contained resting cysts, and loricae containing ciliate cells were scarce, all are thus considered in this study. Consequently, the species identification was lorica-based with reference to the original description of Fol (Reference Fol1883) and compared with subsequent reports (Balech, Reference Balech1959; Brandt, Reference Brandt1907; Fernandes, Reference Fernandes2004; Gold and Morales, Reference Gold and Morales1977; Jörgensen, Reference Jörgensen1924; Kofoid and Campbell, Reference Kofoid and Campbell1929, Reference Kofoid and Campbell1939; Paulmier, Reference Paulmier1997). The morphological terminology followed by Kofoid and Campbell (Reference Kofoid and Campbell1939) and the classification followed by Santoferrara and McManus (Reference Santoferrara, McManus, M.A and A.B2020). Among the individuals, two were mounted on separate glass slides in a solution of Lugol’s iodine and glycerol (1:10 ratio), subsequently sealed with coverslips, using DPX mounting media (Voucher ID: IO/SS/TIN/00003A & B), and four individuals were stored in 0.2 ml PCR tubes (Axygen™). The specimens were deposited at the Centre for Marine Living Resources and Ecology (CMLRE), Kerala, India (Voucher ID: IO/SS/TIN/00004-00006).
Results
Suborder TINTINNINA Kofoid & Campbell, Reference Kofoid and Campbell1929
Family Dictyocystidae Haeckel, 1873
Genus Codonaria Kofoid & Campbell, Reference Kofoid and Campbell1939
Codonaria cistellula (Fol, Reference Fol1883) Kofoid & Campbell, Reference Kofoid and Campbell1939 (Figure 1, Table 1)
1883 Cyttarocylis cistellula (n. sp.) – Fol, Arch. Sci. Phys. Nat. 9: 570: Pl. XVIII, figure 8.
Description of lorica from the Indian Ocean
Morphometrics: Lorica length 75–88 µm (84 ± 4 µm, n = 9) and bowl width 59–69 µm. Collar 26–29 µm high and 46–50 µm wide (opening diameter), 30–34% of the lorica length. Ratio of lorica length to opening diameter 1.66–1.77 (Figure 1; Table 1).
Codonaria cistellula (Fol, Reference Fol1883) Kofoid & Campbell, Reference Kofoid and Campbell1939: (A) C. cistellula from original description by Fol (Reference Fol1883); (B) schematic representation of morphometric measurements by following Kofoid and Campbell (Reference Kofoid and Campbell1939) (SC: subapical cone; SL: subapertural ledge; TL: total length; BL: bowl length; CL: collar length; BW: bowl width; NW: neck width; OD: opening diameter; LD: ledge diameter; a: constriction angle; b: collar apex angle); (B–J) C. cistellula in the Indian Ocean from the present study.
Morphometric data on the lorica of Codonaria cistellula (Fol, Reference Fol1883) Kofoid & Campbell, Reference Kofoid and Campbell1939 from the Indian Ocean
Note: Length measurements in µm and angles in degrees (°). X̅, arithmetic mean; M, median; n, number of individuals investigated; SD, standard deviation; SE, standard error; CV, coefficient of variation; Min, minimum; Max, maximum.
Lorica is composed of a funnel-shaped collar separated by a constriction from a globular, ellipsoidal, or slightly elongated globular bowl, merging at an angle of 119–129° into a posterior process. The constriction between the collar and bowl is distinct. Collar subdivided into the obconical main part with a wide subapertural ledge and the subapically inserting conical inner portion. The obconical collar proper with apex angle 41–49°, 0.30–0.34 total length, or 0.54–0.58 opening diameter. Subapical cone short 5.6–6.4 µm high, 0.11–0.13 opening diameter, often edged by a minutely, irregularly notched hyaline border. Subapertural ledge 62–71 µm wide, 1.31–1.43 opening diameter, or 0.95–1.14 bowl width, margin flaring and irregular. Neck 41–56 µm wide, distinctly constricted to 0.84–0.96 opening diameter or 0.62–0.75 bowl width. Bowl slightly elongated to subspheroidal, bowl length 50–60 µm high, 0.85–0.96 bowl width or 1.1–1.24 opening diameter, bowl width slightly lower or almost equal to the ledge diameter. Posterior process bluntly pointed or rounded with a closed end.
Distribution
Out of nine specimens recovered, four specimens of lorica containing ciliate cells were recorded at a water temperature of 25–26 °C and a salinity of 36‰, and were found at depths of 75–100 m along the offshore contours (100–2000 m) of the North-Eastern Arabian Sea (NEAS).
Discussion
Morphological variability and remarks from the Indian Ocean
Cyttarocylis cistellula Fol, Reference Fol1883 is the type species of Codonaria Kofoid & Campbell, Reference Kofoid and Campbell1939, delineated from Codonella Haeckel, 1873 based on its cylindrical lorica cone projecting beyond the obconical collar (Agatha, Reference Agatha2010; Bachy et al., Reference Bachy, Gómez, López-García, Dolan and Moreira2012; Kofoid and Campbell, Reference Kofoid and Campbell1939). Although some morphological forms were initially recorded by Brandt (Reference Brandt1907) as varieties of C. cistellula, Kofoid and Campbell (Reference Kofoid and Campbell1929, Reference Kofoid and Campbell1939) later elevated these varieties to distinct species (Codonaria angusta, C. australis, C. benguelensis, C. mucronata, and C. oceanica) without a clear distinction from the parent species (Balech, Reference Balech1959, Reference Balech1962; Xu et al., Reference Xu, Agatha and Dolan2024). The variabilities assumed in the lorica morphology include the shape of the bowl, the ratio of subapertural ledge to bowl width, the degree of depth of the constriction, and the posterior process (Kofoid and Campbell, Reference Kofoid and Campbell1929, Reference Kofoid and Campbell1939). The specimens from the Indian Ocean belong to Codonaria cistellula (Fol, Reference Fol1883) Kofoid & Campbell, Reference Kofoid and Campbell1939 (lorica length: 75–88 µm; bowl width: 59–69 µm; lorica opening diameter [LOD]: 46–50 µm; n = 9; Figure 1B–J), although the population exhibits considerable variability in lorica morphology. In some specimens, the posterior lorica portion differs in shape, the length is about 1/5 shorter, and the bowl width is somewhat smaller as compared to the original description (lorica length: 100 µm; bowl width: 70 µm) (Fol, Reference Fol1883). Furthermore, the lorica size of Indian Ocean specimens falls well within the recorded range reported from other regions (lorica length 70–120 μm, bowl width 63–83 μm, and LOD 49–59 μm), including the Mediterranean Sea (Balech, Reference Balech1959; Jörgensen, Reference Jörgensen1924; Paulmier, Reference Paulmier1997), the north Atlantic Ocean (Brandt, Reference Brandt1907; Paulmier, Reference Paulmier1997), subtropical waters of Southern Brazil (Fernandes, Reference Fernandes2004), and the western tropical Pacific Ocean (Gold and Morales, Reference Gold and Morales1977).
Only five loricae conform to Fol’s illustration of lorica with a broadly rounded posterior portion (Figure 1C–F, I), while the others closely resemble other Codonaria species, for which Balech (Reference Balech1959, Reference Balech1962) demonstrated the pronounced polymorphism (Fernandes, Reference Fernandes2004). However, it is worth noting that Fol’s observations were likely based on live specimens, whereas most subsequent records were derived from fixed or empty loricae, which may have undergone distortion or alteration over time. In the present study, three loricae have an ellipsoidal bowl with a distinct, posterior blunt protuberance (Figure 1B, G, H), and the subapertural ledge diameter is slightly lower than or equal to bowl width, which is greater in all other individuals. Nevertheless, they match those of Fernandes (Reference Fernandes2004) (see figure 59: lorica length: 90 µm; LOD: 55 µm; ledge to bowl width, ratio: 0.96, inference from micrograph) and Gold and Morales (Reference Gold and Morales1977) (see figure 3: lorica length: 89 µm; LOD: 48 µm; ledge to bowl width, ratio: 1.1, inference from micrograph), in terms of lorica shape other than the ratio. This ratio apparently varies within the morphotype and thus is rather an insignificant feature for species circumscription, as indicated for the posterior protuberance (Balech, Reference Balech1959). In addition, a single empty lorica of the smallest dimensions (lorica length: 75 µm; bowl width: 59 µm; LOD: 46 µm), composed of a globose bowl with a bluntly pointed posterior process (Figure 1J), closely resembles the micrograph by Paulmier (Reference Paulmier1997), except for the presence of the posterior process. However, the latter exhibits comparatively smaller dimensions (see Plate 4, figure 7: lorica length: 63 µm; bowl width: 45 µm; LOD: 38 µm), as inference from the micrograph (Paulmier, Reference Paulmier1997). Given the morphological variability observed and as the cytological and molecular characteristics of Codonaria forms are unknown, it remains uncertain whether the observed morphological variations reflect intraspecific variability or represent distinct, closely related species.
The specimens of C. cistellula were widespread in the warmer parts of the Atlantic Ocean (Brandt, Reference Brandt1907; Fernandes, Reference Fernandes2004; Paulmier, Reference Paulmier1997), with the largest being found in the Mediterranean (Balech, Reference Balech1959; Fol, Reference Fol1883; Jörgensen, Reference Jörgensen1924). However, lorica length is known to exhibit plasticity due to varying environmental conditions or life stages within populations (Agatha and Tsai, Reference Agatha and Tsai2008; Dolan et al., Reference Dolan, Yang, Lee and Kim2013; Laval-Peuto and Brownlee, Reference Laval-Peuto and Brownlee1986; Santoferrara et al., Reference Santoferrara, Tian, Alder and McManus2015). Among the morphotypes, a gradual increase in lorica length is observed, which falls within the size range of C. cistellula; furthermore, the most conservative diagnostic feature, the LOD (Balech, Reference Balech1959; Laval-Peuto and Brownlee, Reference Laval-Peuto and Brownlee1986), remains confined to a narrow range (46–50 µm). Although the LOD was not reported by Fol (Reference Fol1883), an approximate value of 50 µm can be inferred from his illustration using proportional relationships to the lorica length (100 µm), which is comparable to the Indian Ocean specimens. However, this estimation must be treated with caution, as the lorica opening rim is indistinctive in the illustration, and precise measurements from historical figures are inherently difficult. Moreover, the circumscription of the species is complicated by lorica polymorphism, warranting further genetic analyses. The identical features across all forms include the lorica with the subapical cone, edged with minutely irregularly notched hyaline border (Balech, Reference Balech1959; Brandt, Reference Brandt1907; Fernandes, Reference Fernandes2004; Paulmier, Reference Paulmier1997). Moreover, Codonaria cistellula has been considered a polymorphic species with several regional forms or potential subspecies (Balech, Reference Balech1959; Fernandes, Reference Fernandes2004), highlighting C. oceanica and C. mucronata (Kofoid and Campbell, Reference Kofoid and Campbell1929, Reference Kofoid and Campbell1939). For instance, C. cistellula from the type locality (Bay of Villefranche-sur-Mer) exhibited different degrees of collar agglutination, yet their molecular sequences (based on SSU-rDNA) were identical (Bachy et al., Reference Bachy, Gómez, López-García, Dolan and Moreira2012). Admittedly, in the present material, loricae containing ciliate cells were limited to a single morphological form. Consequently, a comparative investigation of morphotypes was not feasible, further constrained by methodological limitations (preservation and prolonged storage), highlighting the need for future studies with abundant material from the region. For the present, the occurrence of Codonaria cistellula in the Indian Ocean is thus confirmed from NEAS.
Data availability
The specimens were deposited in the Centre for Marine Living Resources and Ecology (CMLRE), Kochi (Voucher ID: IO/SS/TIN/00003-00006).
Acknowledgements
The authors are grateful to the Secretary, Ministry of Earth Sciences, New Delhi, and Head, CMLRE, Kochi, for supporting the work and for providing the facilities onboard FORV Sagar Sampada for in situ measurements. The authors express their gratitude to Dr. G.V.M. Gupta, Project Investigator of the project “Marine Ecosystem Dynamics of Eastern Arabian Sea (MEDAS)” under the Marine Living Resources Programme. The authors sincerely acknowledge the reviewers’ insightful and constructive comments, especially the valuable suggestions of Dr. Sabine Agatha, Department of Environment and Biodiversity, Paris Lodron University of Salzburg, Austria, which improved the manuscript. All the participants of MEDAS cruises who rendered help in sampling are acknowledged. This is the CMLRE contribution number 214.
Author contributions
I.A.: Species identification, visualisation, writing – original draft, and review; C.R.A.D.: Conceptualisation, validation, reviewing, and editing; P.M.H.: Sample analysis and review.
Funding
Ministry of Earth Sciences (MoES), India.
Competing interests
The authors declare no conflict of interests.
