Geoplana baptistae, Amaral, Silvana Vargas Do, Oliveira, Simone Machado De & Leal-Zanchet, Ana Maria, 2012

Amaral, Silvana Vargas Do, Oliveira, Simone Machado De & Leal-Zanchet, Ana Maria, 2012, Three new species of land flatworms and comments on a complex of species in the genus Geoplana Stimpson (Platyhelminthes: Continenticola), Zootaxa 3338, pp. 1-32 : 19-28

Geoplana baptistae sp. nov. Leal-Zanchet & Oliveira

Geoplana sp. 3: Castro & Leal-Zanchet, 2005

Etymology. The specific name honours MSc. Vanessa dos Anjos Baptista and her collaboration in collecting several specimens of land planarians which were deposited in the scientific collection of the Instituto de Pesquisas de Planárias (UNISINOS).

Type material. Holotype: MZUSP PL.01148: K. A. Kopp, leg. 04.VII.2001, Santa Maria, RS, Brazil—ante-rior region in three parts: transverse sections on 140 slides; pre-pharyngeal region: transverse sections on 13 slides; pharynx: sagittal sections on 46 slides; copulatory apparatus: sagittal sections on 30 slides.

Paratypes: MZU PL.00116: D. Cechin, leg. 19.XI.2000, Santa Maria, RS, Brazil—preserved in ethanol 70o; MZU PL.00117: V. S. Lemos, leg. 13.I.2001, Santa Maria, RS, Brazil—pre-pharyngeal region: transverse sections on 18 slides; pharynx: sagittal sections on 17 slide; copulatory apparatus: sagittal sections on 52 slides; MZU PL.00118: R. A. Castro leg. 13.I.2001, Santa Maria, RS, Brazil—preserved in ethanol 70o; MZU PL.00119: A. L. R. Seitenfus leg. 13.I.2001, Santa Maria, RS, Brazil—preserved in ethanol 70o; MZU PL.00120: R. A. Castro, leg. 31.III.2001, Santa Maria, RS, Brazil—anterior tip: transverse sections on 41 slides; anterior region at the level of the ovaries: transverse sections on 23 slides; pre-pharyngeal region: transverse section on 13 slides; pharynx: transverse sections on 40 slides; coppulatory apparatus: sagittal sections on 22 slides; MZU PL.00121: R. A. Castro leg. 31.III.2001, Santa Maria, RS, Brazil—preserved in ethanol 70o; MZU PL.00122: I. Fick, leg. 21.VII.1999, Santa Maria, RS, Brazil—preserved in ethanol 70o. MZU PL.00123 R. A. Castro, leg. 26.VII.2001, Santa Maria, RS, Bra-zil—preserved in ethanol 70o; MZU PL.00124: V. A. Baptista, leg. 29.IX.2001, Santa Maria, RS, Brazil—pre-pha-ryngeal region: transverse sections on 51 slides; pharynx: sagittal sections on 80 slides; copulatory apparatus: sagittal sections on 142 slides; MZU PL.00125: V. M. Dias leg. 06.IV.2002, Santa Maria, RS, Brazil—anterior tip: transverse sections on 71 slides; anterior region at the level of the ovaries: transverse sections on 26 slides; pre-pharyngeal region: transverse section on 16 slides; pharynx: sagittal section on 31 slides; copulatory apparatus: transverse sections on 15 slides; MZU PL.00126: L. B. Matos leg. 15.XII.2002, Santa Maria, RS, Brazil—preserved in ethanol 70o.

Type locality. Santa Maria, state of Rio Grande do Sul (RS), Brazil.

Distribution. Rio Grande do Sul (Santa Maria), Brazil.

Diagnosis. Body rather elongate, with almost parallel margins, broad and flat, anterior end pointed, posterior end gradually narrowed. Live specimens with dorsum dark-brown to the naked eye and venter orange to light brown. Eyes dorsal, with inconspicuous clear halos; sensory pits restricted to the proximity of anterior tip; poorly developed glandular margin with three types of secretory cells; mc:h, 6%–11%; pharynx bell-form; very short esophagus; esophagus: pharynx length, 7% to 9%; anteriormost testes anterior to ovaries, most posterior ones near to root of pharynx; sperm ducts open laterally into proximal part of diverticula of prostatic vesicle; prostatic vesicle, mainly extrabulbar, tubular, consisting of two portions, a laterally expanded T-shaped ental portion, ventrally located, and a sinuous, mainly horizontally located distal portion which enters the bulbar muscular coat; male atrium relatively short, with the entire cavity occupied by the conical penis papilla; straight ejaculatory duct opens through the tip of the symmetrical papilla; ovovitelline ducts emerge dorsally from end of anterior third or median third of ovaries, and ascending behind gonopore; common glandular ovovitelline duct short; vagina as a dorsally curved ental portion of female atrium; female atrium oval-elongate, with an ample lumen, a little narrower in the ental third, lined by an high columnar to pseudostratified epithelium with lacunae; male atrium shorter than female atrium; length of female atrium, 164% that of male one; straight gonopore canal; no folds separating male and female atria.

Description. External morphology. Body rather elongate, with almost parallel margins, broad and flat, anterior end pointed, posterior end gradually narrowed. When crawling, maximum length reaches 120 mm (Table 5). Mouth distance from anterior tip varies from 58% to 72%; gonopore between 71% and 83%, relative to body length (Table 5). Live specimens with dark brown dorsum to the naked eye ( Fig. 26 ). In preserved specimens dorsal colour fades, becoming light brown with darker margins; anterior extremity lighter than the rest of the dorsum. Under the stereomicroscope, dorsal ground-colour brown or gray covered by dense dark-brown pigmentation. Venter orange to light brown, becoming grayish in preserved specimens.

Eyes uniserially surround anterior tip, becoming pluriserial after 1 mm from the tip in paratype MZU PL.00124 (about 1% of body length). Initially, they form two series, the more dorsal comprising the largest eyes. Between 4 mm and 12 mm (approximately 5% and 15% of body length) behind anterior end, these form three to four series near to body margins. After that, they become more dorsal and surrounded by inconspicuous clear halos ( Figs. 27, 28 ). Approximately after 28 mm from anterior tip (about 35% of body length), they form a lateral band on each side of the dorsum, occupying the maximum width of 2 mm on each side of the body (18% of body width on each side of the body). Towards posterior end, they become less numerous ( Fig. 27 ).

Internal morphology. Anterior region: Sensory pits, as simple invaginations, about 30 µm to 47 µm deep, contouring anterior tip, occurring initially at intervals of about 25 µm, posteriorly becoming gradually sparser until they disappear approximately at 10 mm from anterior tip (approximately 14% of body length in the holotype). Eyes (20 µm to 38 µm) contour the anterior tip in a single row; after that run along both sides of the body (two or three on each side). Cutaneous musculature tripartite, similar to that of pre-pharyngeal region (see next section), poorly developed close to the tip. Ventral cutaneous longitudinal muscles gradually become more developed laterally, reaching the same thickness than medially, about 0.4 mm after the tip. Mesenchymal musculature poorly developed in anterior tip with fibers in various directions. Transversal sub-intestinal mesenchymal layer comprising oblique fibres, together with some traverse fibers aggregate ventrally, about 1.4 mm after the anterior tip. Openings of rhabditogen cells and erythophil cells are absent on first 100 µm of body length. Weakly stained cyanophil cells with amorphous secretion open throughout the anterior tip. After that, three types of secretory cells open through dorsal and ventral epidermis: weakly stained cyanophil cells with amorphous secretion; rhabditogen cells, and erythrophil cells with granular secretion. There are only openings of cyanophil cells through body margins (sensorial border). Small concentrations of erythrophil cells, representing the beginning of the glandular margin, appear approximately 10 mm from the anterior tip (about 14% of body length in the holotype).

Epidermis and musculature at pre-pharyngeal region ( Fig. 29 ): Creeping sole broad, occupying 94% to 99% of body width (Table 5). Three types of secretory cells open through dorsal epidermis and body margins: (1) abundant rhabditogen cells with xanthophil secretion and less frequent (2) cyanophil cells with fine granular secretion, (3) erythrophil cells with fine granular secretion, and (4) xanthophilous cells with coarse granular secretion. Creeping sole receives abundant cells with cyanophil coarse granular secretion, as well as less numerous rhabditogen cells and erythrophil cells with fine granular secretion. Poorly developed glandular margin ( Fig. 29 ) comprising cells with coarse granular, densely packed xanthophil secretion and cells with coarse granular, loosely disposed erythrophil secretion, plus a few cells with coarse granular cyanophil secretion.

Cutaneous musculature with the usual three layers, longitudinal layer with thick bundles, being approximately at least double the height of the other two. Laterally to the sagittal plane, the cutaneous musculature remains thick, and close to the body margins, it becomes progressively lower. Mc:h 6% to 11% ( Table 6 ).

Mesenchymal musculature mainly composed of transversal, oblique, and dorsoventral fibres, the former ones constitute a supra-intestinal transversal (3–4 fibers thick) and a sub-intestinal transversal layer (2–3 fibers thick). Oblique fibers constitute a dorsal, subcutaneous layer (4–6 fibers thick). Longitudinal fibers were indiscernible.

Pharynx ( Fig. 30 ): Pharynx bell-form with folded walls; dorsal insertion approximately at the same transverse level as the mouth, both located in the middle third of pharyngeal pouch. Esophagus short, partially projected ventrally to intestine, lined with ciliated cuboidal epithelium mainly presenting insunk nuclei and with some erythrophil secretory cells with granular secretion and cell bodies in the mesenchyme; esophageal muscularis comprises a circular muscle layer with longitudinal interposed fibers. Esophagus: pharynx length, 7% to 9%. Pharynx and pharyngeal lumen lined with ciliated columnar epithelium exhibiting insunk nuclei. Pharyngeal glands with cell bodies located in mesenchyme, mainly anteriorly to pharyngeal pouch. Three secretory cell types: (1) cells with fine granular erythrophil secretion; 2) cells with mixed cyanophil and erythrophil fine granular secretion and (3) less numerous cells with cyanophil amorphous secretion. Outer musculature of pharynx (ca. 23 µm thick) with a thin longitudinal subepithelial layer, followed by a thicker circular one, mixed internally with few longitudinal fibers. Towards pharyngeal tip, circular layer becomes as thin as longitudinal one. Inner pharyngeal musculature (ca. 50 µm thick) of thick circular subepithelial layer, mixed with some longitudinal fibers. Inner musculature gradually diminishes towards pharyngeal tip.

Reproductive apparatus: Testes beginning anteriorly to ovaries and extending up to the same transverse level as the ventral insertion of the pharynx (Table 5), comprising one or two irregular rows, dorsal to the intestinal branches on each side of the body ( Fig. 29 ). Pre-pharyngeally, sperm ducts, subdivided in two or three ductules, dorsal to ovovitelline ducts, at least one ductule laterally displaced. Laterally to the posterior third of pharynx, these form spermiducal vesicles, opening laterally into proximal part of diverticula of prostatic vesicle ( Figs 31, 32 ). Prostatic vesicle, mainly extrabulbar, tubular, of two portions, a laterally expanded ental portion, ventrally located, and a mainly horizontally located distal portion which enters the bulbar muscular coat, continuing into the ejaculatory duct. The latter traverses the conical and symmetrical penis papilla to open through its tip. Male atrium oval-elongate, relatively short, with few, low folds and with the entire cavity occupied by the conical penis papilla which may extend into the female atrium (Table 5; Figs. 31–36, 38 ).

Sperm ducts lined with ciliated cuboidal epithelium; thin muscularis (ca. 3 µm thick) mainly comprised of circular fibers. Prostatic vesicle lined with irregular ciliated columnar to ciliated pseudostratified epithelium, gradually diminishing its height towards ejaculatory duct. Erythrophil cells with granular secretion and bodies lying in mesenchyme, mainly around and anteriorly to the vesicle, show numerous openings into vesicle. Muscularis of proximal and distal portions of vesicle 20–25 µm thick and 15 µm thick in diverticula comprising interwoven circular, oblique, and longitudinal fibers.

Ejaculatory duct lined with ciliated columnar epithelium, receiving numerous openings from secretory cells with amorphous, cyanophil secretion and subepithelial bodies. It is coated with a muscularis (ca. 5 µm thick) of circular fibers interposed with some longitudinal ones. Penis papilla lined with non-ciliated columnar epithelium. Three types of secretory cells run longitudinally in the papilla, with numerous openings through its lining epithelium: (1) cells with fine densely arranged granular heavily stained erythrophil secretion; (2) cells with granular xanthophil secretion; and (3) cells with cyanophil amorphous secretion. Erythrophil and cyanophil cells present cell bodies external to common muscle coat; xanthophil cells, subepithelial cells bodies. Muscularis (8–25 µm) mainly of circular layer with some mixed longitudinal fibers; thinner towards the tip of the papilla. Longitudinal, radial and oblique muscle fibers cross the papilla.

Epithelial lining of male atrium, columnar (about 40–50 µm), non-ciliated, showing xanthophil apical secretion. Two types of secretory cells, empty through the epithelium: abundant cells with cyanophil amorphous secretion and cells with fine granular xanthophil secretion, both with cell bodies internal to common muscle coat. In addition, there are some cells with fine granular erythrophil secretion and cell bodies external to common muscle coat. Muscularis well developed (40–80 µm) throughout male atrium, comprising circular subepithelial fibers and subjacent longitudinal ones.

Ovaries oval-elongate in shape, 0.5 mm long anterior-posteriorly and approximately 0.2 mm dorso-ventrally in the holotype. Ovovitelline ducts emerging dorsally from end of anterior third or median third of ovaries, then leading backwards immediately dorsal to nerve plate. Behind gonopore, ovovitelline ducts ascend posteriorly and medially inclined, to unite dorsally to the female atrium (proflex condition with dorsal approach), thus forming the common glandular ovovitelline duct. A conspicuous common glandular ovovitelline duct opens into ental portion of female atrium. Female atrium oval-elongate in shape, with a spacious lumen, a little narrower in the ental third, with elongate lateral folds arising from its floor. Length of female atrium, 164% of male atrium length (Table 5). Ental portion presents a dorsally directed diverticulum (vagina) (Table 5, Figs. 31, 32 , 34–36 ).

Paired ovovitelline ducts lined by ciliated cuboidal to columnar epithelium; muscle coat of paired and common ovovitelline ducts mainly comprising circular fibers and some interposed longitudinal fibers. Shell glands with xanthophil secretion opening into distal ascending portion of paired ovovitelline ducts, and also into common glandular ovovitelline duct ( Figs. 31 – 33, 35 ).

Lining epithelium of vagina and female atrium columnar (approximately 50 µm) with lacunae, irregular in height, becoming pseudodostratified in various parts of female atrium (100–160 µm) ( Figs. 33, 35 , 37 ); non-ciliated in female atrium and ciliated in vagina. Three types of secretory cells empty through the epithelium: abundant cells with cyanophil amorphous secretion and cell bodies external to common muscle coat; cells with fine granular xanthophil secretion and cell bodies internal to common muscle coat; and cells with fine granular erythrophil secretion and cell bodies external to common muscle coat. Female atrium with muscularis of interwoven circular and longitudinal fibers, thinner (15–25 µm) than in male atrium. Muscularis of vagina mainly of circular fibers with some interposed longitudinal fibers (5–8 µm).

Gonopore canal approximately vertical in sagittal plane. Male and female atria with ample communication, without folds separating them ( Figs. 31 – 36, 38 ). Gonopore canal lined with ciliated columnar epithelium with numerous openings of cells with xanthophil amorphous secretion, rhabditogen cells, and cells with granular erythrophil secretion. Muscularis of circular fibers with some interposed as well as subjacent longitudinal fibers.

Common muscle coat thin with circular, longitudinal and oblique fibers. Between atrial muscularis and common muscle coat, a poorly developed stroma with muscle fibers variously oriented.

Vitellaria, situated between intestinal branches, open into the ovovitelline ducts.

Remarks. Vitellaria were well developed in paratypes MZU PL.00124, MZU PL.00125, and MZU PL.00117, and in maturation in the holotype. Paratype MZU PL.00120 was incompletely mature, thus presenting small testes, and inconspicuous vitellaria and shell glands. Apart from the abundant apocrine secretion produced by cells of the epithelial lining of the female atrium in paratype MZU PL.00117, there were also xanthophil granules and cyanophil amorphous secretion in both lacunae of this epithelium and the atrial lumen ( Fig. 37 ). A small amount of secretion was also present in the male atrium. The holotype, from an open area in a native field, was darker than specimens from forested areas, the dark-brown colour due to dense, dark pigmentation covering the dorsal ground colour. It was directly fixed after falling into a pitfall, and showed an everted pharynx, and a very distended copulatory apparatus. In the paratype MZU PL.00125, the female atrium is relatively shorter (53%, see table 5) than is the male, when compared to other mature specimens.

Figures

FIGURE 26. Geoplana baptistae sp. nov.: photograph of a live specimen (paratype MZU PL. 00124) in dorsal view. Scale bar: 30 mm.

FIGURES 27 – 28. Geoplana baptistae sp. nov.: (27) colour pattern of a preserved specimen (paratype MZU PL. 00122) in dorsal view; (28) detail of eyes pattern of a preserved specimen (paratype MZU PL. 00124) in dorsal view. (e) eyes. Scale bar: 5 mm.

FIGURES 29 – 30. Geoplana baptistae sp. nov.: (29) pre-pharyngeal region (paratype MZU PL. 00124) in transverse section; (30) pharynx of paratype MZU PL. 00120 in sagittal section. (di) dorsal insertion, (es) esophagus, (i) intestine, (im) internal musculature, (lu) pharyngeal lumen, (m) mouth, (mm) mesenchymatic muscles, (n) nerve plate, (om) outer musculature, (sc) secretory cells, (t) testes, (v) vitellaria, (vi) ventral insertion. Scale bar: 1 mm.

FIGURES 31 – 32. Geoplana baptistae sp. nov.: (31) diagrammatic sagittal composite reconstruction of the copulatory apparatus of the holotype; (32) diagrammatic horizontal composite reconstruction of the copulatory apparatus of paratype MZU PL. 00125. (cmc) common muscle coat, (cov) common glandular ovovitelline duct, (ej) ejaculatory duct, (fa) female atrium, (go) gonopore, (ma) male atrium, (ov) ovovitelline ducts, (pp) penis papilla, (pv) prostatic vesicle, (sc) secretory cells, (sv) spermiducal vesicle, (va) vagina. Scale bar: 1 mm.

FIGURES 33 – 35. Geoplana baptistae sp. nov.: (33) copulatory apparatus of paratype MZU PL. 00124 in sagittal section; (34) copulatory apparatus of paratype MZU PL. 00125 in horizontal section; (35) copulatory apparatus of the holotype in sagittal section. (cmc) common muscle coat, (cov) common glandular ovovitelline duct, (ej) ejaculatory duct, (fa) female atrium, (go) gonopore, (ma) male atrium, (pp) penis papilla, (pv) prostatic vesicle, (sc) secretory cells, (sg) shell glands, (va) vagina. Scale bar: 1 mm.

FIGURES 36 – 38. Geoplana baptistae sp. nov.: (36) copulatory apparatus of paratype MZU PL. 00120 in sagittal section; (37) detail of epithelial lining of the female atrium of paratype MZU PL. 00120 in sagittal section; (38) copulatory apparatus of paratype MZU PL. 00117. (cmc) common muscle coat, (cov) common glandular ovovitelline duct, (ej) ejaculatory duct, (fa) female atrium, (go) gonopore, (ma) male atrium, (pp) penis papilla, (pv) prostatic vesicle, (sc) secretory cells, (sv) spermiducal vesicle, (va) vagina. The arrow indicates apocrine secretion in the female atrium. Scale bars: 1 mm (Figs. 36; 38); 0.1 mm (Fig. 37).

Tables

TABLE 6. Cutaneous musculature and body height, in micrometer, in the median region of a transverse section of the pre-pharyngeal region, and ratio of the height of cutaneous musculature to the height of the body (mc: h index) of specimens of Geoplana baptistae sp. nov. * Specimen collected in a pitfall trap and directly fixed in formaldehyde.

	Holotype* Paratype MZU PL.00117	Paratype MZU PL.00124	Paratype MZU PL.00125
Dorsal musculature	49 35	49	31
Ventral musculature	80 46	58	53
Body height	1140 1340	1660	1250
Mc:h (%)	11 6	6	7