Trichomycterus illuvies, Reis & Pinna, 2023

Reis, Vinícius & Pinna, Mário De, 2023, Diversity and systematics of Trichomycterus Valenciennes 1832 (Siluriformes: Trichomycteridae) in the Rio Doce Basin: iterating DNA, phylogeny and classical taxonomy, Zoological Journal of the Linnean Society 197, pp. 344-441 : 394-405

TRICHOMYCTERUS ILLUVIES SP. NOV.

( FIG. 24)

Z o o b a n k r e g i s t r a t i o n: u r n: l s i d: z o o b a n k. org:act: 734919BD-6C8A-4AA5-B2A4-B1EFE8F83059

Holotype: MZUSP 112750, 45 mm SL; Brazil, state of Minas Gerais, Ferros, Rio Santo Antônio (tributary of Rio Doce ) at bridge in town of Ferros (19°13’34.50”S

43°1’9.50”W); col. O. T. Oyakawa & T. F. Teixeira, 16 August 2012 .

Paratypes: All from Brazil, state of Minas Gerais. MZUSP 126758 , 10 , 32.2–43.7 mm SL, 2c&s, 31.9, 40.8 mm SL; collected with holotype; MZUSP 110720 , 1 , 37.6 mm SL; Bom Jesus do Galho; Ribeirão Sacramento Creek , tributary of Rio Doce Basin (19°47’50.00”S 42°18’9.00”W); col. A. Netto-Ferreira & R. Pádua, 3 August 2010 .

Diagnosis: The combination of the following traits distinguishes T. illuvies from congeners: (1) colour pattern consisting of a broad dark stripe running along mid-lateral line of body from immediately posterior to opercle to base of caudal fin; (2) pectoral-fin rays I + 7 (vs. I + 5, I + 6 or I + 8); (3) two lateral-line pores (vs. three); (4) first dorsal- and anal-fin pterygiophores positioned anteriorly to 18 th and 23 rd vertebrae, respectively; (5) six branchiostegal rays (vs. seven or more). Among congeners in south-eastern South America, character 1 distinguishes T. illuvies from all congeners except for T. giganteus , T. itatiayae , T. nigroauratus and the T. reinhardti species complex ( Costa & Katz, 2021); character 2 from the T. brasiliensis and T. reinhardti species complexes ( Barbosa & Costa, 2010; Costa, 2021; Costa & Katz, 2021), plus T. trefauti (all preceding with I + 6 or fewer pectoral-fin rays) and from T. astromycterus , T. caipora , T. giganteus , T. immaculatus , T. lauryi , T. nigricans , T. pradensis and T. tantalus (with I + 8 or more); character 3 from T. alternatus , T. astromycterus , T. aff. caipora , T. ipatinga , T. tantalus and T. vinnulus ; character 4 from all congeners except T. maculosus Barbosa & Costa, 2010 , T. melanopygius , T. nigricans , T. quintus Costa, 2020 ; character 5 from all congeners except for T. alternatus , T. astromycterus , T. aff. caipora and T. immaculatus . Among congeners in the Rio Doce Basin , T. illuvies is most similar to T. alternatus . In addition to the characters above, T. illuvies can be further distinguished from T. alternatus by the absence of a large fenestra between the orbitosphenoid and the frontal (vs. presence), by the caudal-fin procurrent rays beginning anteriorly at 34 th vertebrae (vs. 28 th to 32 nd vertebrae) and by the more numerous opercular odontodes (18–24 vs. 12–16).

Description: Morphometric data for specimens examined is presented in Table 10. Body long and almost straight, trunk roughly round in cross-section near head, then slightly deeper than wide and softly compressed to caudal peduncle, tapering to caudal fin. Dorsal profile of body gently convex to dorsal-fin origin, then straight or slightly concave along caudal peduncle to caudal-fin origin. Ventral profile convex from gular region to vent, due partly to abdominal distension, then straight or slightly concave along anal-fin origin to caudal-fin base. Caudal peduncle as deep as body at end of anal-fin base.

Head approximately one-fifth of SL, pentagonal, longer than wide and depressed. Mouth sub-terminal. Upper jaw slightly longer than lower. Upper lip wider than lower lip, and laterally continuous with base of maxillary barbel. Lower lip small, approximately 2/3 width of upper one, partly divided into right and left portions by median concavity. Lower lip with uniform covering of tiny villi, resulting in velvet-like surface and not clustered into large papillae. Region between upper and lower lips with slender fleshy lobe.

Dentary and premaxillary teeth similar to each other in shape. Dentary teeth conical, arranged in four irregular rows extending from symphysis to slightly up into coronoid process, with size of individual teeth increasing markedly towards symphysis and from posterior to anterior rows. Total area of premaxillary teeth slightly smaller than that of dentary teeth, with conical teeth arranged irregularly in four rows over entire ventral surface of premaxilla.

Eye medium sized, protruding, positioned dorsally on head, without free orbital rim and covered with transparent skin. Eye located on anterior half of HL, closer to lateral border of head than to the midline in dorsal view. Anterior naris surrounded by tube of integument directed anterolaterally, continuous posterolaterally with nasal barbel. Posterior naris closer to anterior naris than to eyes, surrounded by tube of integument incomplete posteriorly. Maxillary barbel narrowing markedly towards fine tip, reaching lateroposterior border of interopercle. Rictal barbel inserted immediately ventral to maxillary barbel, its tip reaching anterolateral margin of interopercle. Nasal barbel originating on posterolateral region of anterior naris, its tip surpassing eye, but not reaching anterior margin of opercle. Interopercular patch of odontodes small compared to head length, oval in shape and with well-developed odontodes, prominent in ventral aspect of head. Interopercular patch of odontodes extending from vertical through ventroposterior border of eye to ventroanterior to opercle patch of odontodes. Odontodes arranged in two or three irregular series, with those on mesial series much longer than those on lateral one; odontodes gradually larger posteriorly in both series, with those posteriorly on mesial row largest. Interopercular odontodes 30–44. Opercular patch of odontodes on dorsolateral surface of posterior part of head, positioned anterodorsally to pectoral-fin base, roundish to oval in shape in dorsal aspect of head and with same size as eye diameter. Opercular odontodes 18–24, sunk in individual slits of integument, progressively larger posteriorly, all with fine tips, with largest ones curved distally and claw-like. Entire patch surrounded by fleshy fold rim of integument.

Pectoral fin with its base immediately posterior and ventral to opercular patch of odontodes. Pectoral-fin rays I + 7 or rarely I + 8. First pectoral-fin ray (unbranched) longest, prolonged as short filament beyond fin margin. Other rays progressively less long, their tips following continuous line along fin margin. Pelvic fin with convex distal margin, its origin slightly posterior to middle of SL and anterior to vertical through dorsal-fin origin, its tip variably reaching anywhere between anterior and posterior margins of urogenital opening. Bases of pelvic fins close to each other. Pelvic-fin rays I + 4. Dorsal fin long, its distal margin sinusoidal. Dorsal-fin origin closer to base of caudal fin than to tip of snout. Dorsal-fin rays iii + II + 7 (3). Anal fin slightly smaller than dorsal fin, its distal margin gently convex. Anal-fin origin posterior to vertical through end of dorsal-fin base. Anal-fin rays iii + II + 5 (3). Caudal fin round, with 6 + 7 principal rays. Adipose fin absent or modified into low integumentary fold extending between end of dorsal fin and caudal-fin origin. Post-Weberian vertebrae, 37 (2) or 38 (1). First dorsal-fin pterygiophore immediately anterior to neural spine of 18 th (3) vertebra, first anal-fin pterygiophore immediately anterior to neural spine of 23 rd (3) vertebra. Caudal-fin procurrent rays plus one segmented non-principal ray dorsally and ventrally. Procurrent caudal-fin rays, 12–14 (3) dorsally and 11–13 (3) ventrally, beginning anteriorly at 34 th (2) vertebra. Ribs 13 (3). Branchiostegal rays 6 (3). Dorsal-fin pterygiophores 8. Anal-fin pterygiophores 6.

Cephalic lateral line canals with simple, non-dendritic tubes ending in single pores. Supraorbital canal mostly in frontal bone. Supraorbital pores invariably present: s1 mesial to nasal-barbel base and autopalatine, s3 mesial to posterior nostril and anterior to frontal, single or paired s6, closer to mesial line than to eyes, posteromedial to eye and at midlength of frontal. Infraorbital latero-sensory canal incomplete with four pores, i1 and i3 anteriorly and i10 and i11 posteriorly. This canal extending from sphenotic posteriorly to terminal pore located ventroposteriorly to eye. Infraorbital pore i1 located ventrolateral to nasal-barbel base and autopalatine, i3 ventrolateral to posterior nostril and anterior to frontal, i10 and i11 posterior to eye. Otic canal without pores. Postotic pores po1, anteromedial to opercular patch of odontodes, and po2, mesial to opercular patch of odontodes. Lateral line of trunk anteriorly continuous with postotic canal and reduced to short tube. Lateral line pores ll1 and ll2 dorsomedial to pectoral-fin base.

Coloration in ethanol: Dark chromatophores distributed into inner and outer skin layers. Those on inner skin layer forming broad dark stripes or faded maculae responsible for main colour features of body. Basic arrangement composed of two stripes and one row of maculae. First stripe along mid-dorsal line from occiput to base of caudal fin. Row of maculae ventrolateral to that first stripe, extending from base of head through upper part of flanks, dorsal portion of caudal peduncle and upper part of caudal-fin base. In some specimens, row fusing with the first stripe. Second stripe running along mid-lateral line, from immediately posterior to opercle to middle of caudal-fin base. In some specimens, second short row ventral to main one, extending from mid-length of abdomen through ventral margin of caudal peduncle to base of caudal-fin. Dorsal stripes and rows disrupted by fusions (mostly along anterior part of body). Second stripe never fusing with first one. Head darkest on region corresponding to neurocranium, outlined by brain pigment seen by transparency. Unpigmented circular area extending from posterior margin of eye to base of opercular patch of odontodes. Base of nasal barbels surrounded with concentration of dark pigment, extending posteriorly as elongate dark field to anterior margin of eyes. Margin of integumentary fold of opercular patch of odontodes darkly pigmented. Interopercular patch of odontodes white. Ventral side of the body lacking dark pigment. Fins with small brownish spots randomly distributed on fin rays. Caudal fin with vertical dark stripe crossing bases of principal rays.

Remarks: Although T. illuvies seems to be part of the T. alternatus metaspecies due to its general morphology and colour pattern of some specimens, it differs from the latter in many traits as listed in the diagnosis. One of those is the the absence of a large fenestra between the orbitosphenoid and frontal bones, a conspicuous feature of the skull in T. alternatus (cf. Reis et al., 2019: fig. 5C) and its close relatives such as T. immaculatus and T. melanopygius and its exospecies (see Remarks on T. alternatus ), such as T. astromycterus and T. vinnulus relative to T. alternatus . Currently there is no DNA sample of T. illuvies .

Etymology: The name is derived from Latin illuvies , meaning filth, dirt or flood, alluding to the environmental mayhem caused by the mining company Samarco SA on the Rio Doce. This is a long-term reminder of the catastrophe suffered by that hydrographic basin.

Geographical distribution: Trichomycterus illuvies is found in two drainages belonging to Rio Doce Basin , the Santo Antônio River (holotype locality) and Ribeirão Sacramento (paratype locality) ( Fig. 25). The two localities are separated by 97.9 km in straight line and the species is expected to occur in some of the intervening areas.

TRICHOMYCTERUS IMMACULATUS (EIGENMANN & EIGENMANN, 1889)

( FIGS 8–9, 26)

Pygidium immaculatum Eigenmann & Eigenmann, 1889: 52 (syntypes from Juiz de Fora , Parahybuna River ; São Matheos River ; Juiz de Fora, Paraíba do Sul River; Goiás, Brazil; syntypes: MCZ 8266 , 8300 , 8302 , 8305 , 8307 ; lectotype herein designated as MCZ 8302 , from Rio São Matheus); Gosline, 1945: 60 (date of authorship mistakenly cited as Eigenmann, 1918).

Trichomycterus immaculatum ; Burgess, 1989: 322 (list).

Trichomycterus immaculatus ; Caramaschi & Caramaschi, 1991: 1 (diagnosis); Bizerril, 1994: 623 (list); Costa & Bockmann, 1994: 717 (comparative material); Braga, 2004: 35 (anatomy); Triques & Vono, 2004: 82 (list, relationships); Bockmann & Sazima, 2004: 71 (comparisons); Bockmann et al., 2004: 227 (comparisons); Lima & Costa, 2004: 3 (comparisons); Fernandez & Vari, 2004: 881 (comparative material); Diogo et al., 2004: 264 (comparative material); Sarmento-Soares et al., 2005: 209 (comparisons); Wosiacki & Oyakawa, 2005: 471 (comparisons); Wosiacki, 2005: 52 (comparisons); Diogo, 2006: 17 (comparative material); Ingenito & Buckup, 2007: 1177 (biogeography); Ferraris 2007: 419 (check list); Lima et al., 2008: 316 (comparisons); Barbosa & Costa, 2010: 121 (comparisons); Pereira et al., 2010: 5 (barcoding); Roldi et al., 2011: 2 (comparisons); Sarmento-Soares et al., 2011: 262 (comparisons); Ferrer & Malabarba, 2011: 66 (comparative material); DoNascimiento et al., 2014a: 709 (comparisons); García-Melo et al., 2016: 237 (comparisons); Ochoa et al., 2017: 75 (relationships); Sales et al., 2018: 3 (citation); Katz et al., 2018: 559 (comparative material); Reis et al., 2019: 12 (citation); Reis & de Pinna, 2019: 118 (comparisons); Donin et al., 2020: 1 (comparisons); Costa et al., 2020a: 1 (relationships); Costa et al., 2020b: 1 (relationships); Reis et al., 2020: 1 (relationships); DoNascimiento & Prada-Pedreros, 2020: 981 (comparative material); Ochoa et al., 2020: 3 (relationships); Fernandez et al., 2021 (relationships).

Trichomycterus pradensis Sarmento-Soares et al., 2005 (Jucuruçu, Rio Jucuruçu , 2 km before the city of Jucuruçu on road Itamaraju-Jucuruçu, middle of Rio Jucuruçu Basin , 16°50’10”S, 40°08’40”W, Bahia, Brazil); Ferraris, 2007: 422 (comparisons); Barbosa & Costa, 2011: 308 (comparisons); Schaefer & Férnandez, 2009: 518 (comparisons); DoNascimiento et al., 2014a: 709 (comparisons); García-Melo et al., 2016: 237 (comparisons); Ochoa et al., 2017: 75 (c o m p a r i s o n s); Vo l p i, 2 0 1 7 (m o l e c u l a r d a t a; comparisons); Sales et al., 2018: 6 (citation); Katz et al., 2018: 559 (comparative material); Donin et al., 2020: 9 (comparisons); Ochoa et al., 2020: 3 (relationships); Costa et al., 2020b: 2912 (relationships); Costa et al., 2020a: 6 (comparisons); DoNascimiento & Prada-Pedreros, 2020: 978 (comparisons); Reis et al., 2020: 1 (relationships).

Trichomycterus inmaculatus ; Fernández et al., 2015:

11 (misspelling).

Diagnosis: The combination of the following traits distinguishes T. immaculatus from congeners: (1) most common colour pattern uniform greyish to nearly black over entire body, with less frequent colour variants with round dark maculae randomly distributed on dorsum; caudal fin dusky in adults or with slender dark horizontal stripe from base to tip of fin in some juveniles; (2) pectoral-fin rays I + 8 (vs. I + 5, I + 6 or I + 7); and (3) two lateral-line pores (vs. three or more). Among congeners in south-eastern South America, character 1 distinguishes T. immaculatus from all congeners (although the plain colour pattern is present also in T. jacupiranga , T. melanopygius , T. nigricans , T. tantalus and T. trefauti , the variable colour pattern including round dark maculae randomly distributed on dorsum distinguishes T. immaculatus from all latter); character 2 distinguishes T. immaculatus from all congeners except for T. astromycterus , T. caipora , T. giganteus , T. immaculatus , T. lauryi , T. nigricans and T. tantalus ; character 3 distinguishes T. immaculatus from T. astromycterus , T. aff. caipora , T. nigricans , T. ipatinga , T. tantalus and T. vinnulus . Among congeners in the Rio Doce Basin , T. immaculatus is most similar to T. melanopygius and T. tantalus . It can be distinguished from those two species, in addition to characters aforementioned, from T. melanopygius by the fewer post-Weberian vertebrae (36–37 vs. 38–40) and by a dusky caudal fin in adults (vs. a well delimited horizontal dark stripe in the middle of caudal fin); and from T. tantalus by fewer opercular odontodes (15–25 vs. 25 to 33) and a truncated caudal fin (vs. forked).

Description: Morphometric data for specimens examined is presented in Table 11. Body long and nearly straight, trunk roughly round in cross-section near head, then slightly deeper than wide and gently compressed to caudal peduncle, tapering to caudal fin. Dorsal profile of body gently convex to dorsal-fin origin, then straight or slightly concave along caudal peduncle to caudal-fin origin. Ventral profile convex from gular region to vent, due partly to abdominal distension, then straight or slightly concave along anal-fin origin to caudal-fin base. Caudal peduncle varying from nearly as deep as body at the beginning of dorsal-fin base to as deep as body at the end of anal-fin base.

Head length approximately one-fifth of SL, roughly pentagonal, longer than wide and depressed. Mouth subterminal. Upper jaw slightly longer than lower. Upper lip wider than lower one and laterally continuous with base of maxillary barbel. Lower lip small, approximately 2/3 width of upper one, partly divided into right and left portions by median concavity. Lower lip with uniform covering of tiny villi, not clustered into large papillae, resulting in velvet-like surface. Region between upper and lower lips with slender fleshy lobe.

Dentary and premaxillary teeth similar to each other in shape and size. Dentary teeth conical, arranged in four irregular rows, first row with 9–12 teeth extending from symphysis to slightly up of coronoid process, with size of individual teeth increasing markedly towards symphysis and from posterior to anterior rows. Total area of premaxillary teeth smaller than that of the dentary, with conical teeth arranged irregularly in four rows, first row with 10–16 teeth over entire ventral surface of premaxilla.

Eye medium sized, slightly protruding, positioned dorsally on head, without free orbital rim and covered with transparent skin. Eye located on anterior half of HL, closer to lateral border of head than to the midline in dorsal view. Anterior naris surrounded by tube of integument directed anterolaterally, continuous posterolaterally with nasal barbel. Posterior naris closer to anterior naris than to eye, surrounded by tube of integument incomplete posteriorly. Length of all barbels variable. Maxillary barbel narrowing markedly towards fine tip, variably reaching from middle of interopercle to the base of pectoral fin. Rictal barbel inserted immediately ventral to maxillary barbel, its tip reaching from anterolateral to posterolateral border of interopercle. Nasal barbel originating on posterolateral region of anterior naris, reaching anywhere from middle of eye to anterior border of opercle. Interopercular patch of odontodes large compared to head length, oval in shape and with well-developed odontodes, prominent in ventral aspect of head. Interopercular patch of odontodes extending from vertical through posterior border of eye to anterior margin of opercle patch of odontodes. Interopercular odontodes arranged in two or three irregular series, with those on mesial series much longer than those on lateral one; odontodes gradually larger posteriorly in both series, with those posteriorly on mesial row largest. Interopercular odontodes 37–59. Opercular patch of odontodes on dorsolateral surface of posterior part of head, positioned anterodorsally to pectoral-fin base, roundish in shape and larger than eye in dorsal aspect of head. Opercular odontodes 15–25, sunk in individual slits of integument, progressively larger posteriorly, all with fine tips, with largest ones curved distally and claw-like. Entire patch surrounded by well-differentiated fleshy fold rim of integument.

Pectoral fin with its base immediately posterior and ventral to opercular patch of odontodes. Pectoral-fin rays invariably I + 8. First pectoral-fin ray, unbranched, longer and prolonged as filament beyond fin margin. Other rays progressively less longer, their tips following continuous line along fin margin. Pelvic fin with convex distal profile, its origin slightly posterior to middle of SL and anterior to vertical through dorsal-fin origin, its length varying from not reaching to reaching (but never entirely covering) anal and urogenital openings in adults. Base of pelvic fins positioned close to each other. Pelvic-fin rays I + 4, first ray unbranched. Dorsal fin long, its distal margin sinusoidal. Dorsal-fin origin closer to base of caudal fin than to tip of snout. Dorsal-fin rays iii + II + 7 (12), iii + III + 6 (1) or iv + II + 7 (2). Anal fin slightly smaller than dorsal fin, its distal margin gently convex. Anal-fin origin posterior to vertical through end of dorsal-fin base.Anal-fin rays ii + II + 5 (2) or iii + II + 5 (13). Caudal fin from subtruncate to gently concave or emarginate, with 6 + 7 (14) or 5 + 7 (1) principal rays. Adipose fin absent or modified into low integumentary fold extending between end of dorsal fin and caudal-fin origin. Post-Weberian vertebrae 34 (2), 35 (5) or 36 (8). First dorsal-fin pterygiophore immediately anterior to neural spine of 16 th (9) or 17 th (6) vertebra, first anal-fin pterygiophore immediately anterior to neural spine of 20 th (1), 21 st (9) or 22 nd (4) vertebra. Procurrent caudal-fin rays, 13–17 dorsally and 12–14 ventrally, beginning anteriorly at 31 st or 32 nd vertebrae plus one posterior segmented non-principal ray in each lobe. Ribs 10 (5), 11 (9) or 12 (1). Branchiostegal rays 6 (1) or 7 (14). Dorsal-fin pterygiophores 8. Anal-fin pterygiophores 6.

Cephalic lateral line canals with simple, non-dendritic tubes ending in single pores. Supraorbital canal mostly in frontal bone. Supraorbital pores invariably present: s1 mesial to nasal-barbel base and autopalatine, s3 mesial to posterior nostril and anterior to frontal, s6 single or, more commonly, paired, posteromedial to eye and at midlength of frontal. Infraorbital latero-sensory canal incomplete with four pores, i1 and i3 anteriorly and i10 and i11 posteriorly. Canal extending from sphenotic posteriorly to terminal pore located ventroposteriorly to eye. Infraorbital pore i1 located ventrolateral to nasal-barbel base and autopalatine, i3 ventrolateral to posterior nostril and anterior to frontal, i10 and i11 posterior to eye. Otic canal without pores. Postotic pores po1, anteromedial to opercular patch of odontodes, and po2, mesial to opercular patch of odontodes. Lateral line of trunk anteriorly continuous with postotic canal and reduced to short tube, with one lateral line bone partially ossified in some specimens. Lateral line pores ll1 and ll2 dorsomedial to pectoral-fin base.

Coloration in ethanol: Trichomycterus immaculatus is variable in pigmentation pattern, with two extreme colour morphs bridged by intermediate conditions ( Figs 8–9). At one extreme, entire body covered with uniform scattering of dark chromatophores in inner and outer integument layers, not forming maculae or spots. Specimens with this colour pattern usually with dusky caudal fin with transparent margin. At other extreme, chromatophores usually uniformly distributed on inner skin layer. On outer layer, chromatophores darker and coalescing to form round maculae, as large as or larger than, eye diameter. Maculae covering dorsum to the mid-lateral line, completely absent ventral to that limit. Maculae randomly distributed in adults, but in young specimens, maculae more evident and arranged in rows along midline of body, from dorsoposteriorly to opercle to base of caudal-fin resulting in pattern similar to that of T. alternatus . Fins mostly transparent, except for caudal dusky in adults and, in some juveniles, dark stripe from base of caudal-fin to its tip. Specimens intermediate between two extreme colour morphs with faint maculae, only slightly darker than background colour and restricted to dorsum, never extending ventrally beyond lateral midline. Colour pattern of head, caudal fin and ventral part of body not correlated with spotted or plain morphs. Head lacking maculae or blotches, with area corresponding to neurocranium darker than rest of head. Integument surrounding opercular odontodes darkly pigmented. Interopercular patch of odontodes white. Cheeks less darkly pigmented than rest of head. Caudal fin ranging from uniformly dusky with hyaline margins to darkly pigmented along middle caudal-fin rays, forming dark longitudinal stripe in young specimens. Stripe gradually less evident in larger specimens, due to fading or masking by additional caudal-fin pigmentation. Ventral part of body without dark pigmentation.

Remarks: Trichomycterus immaculatus stands out, along with T. alternatus , as one of the most complex taxonomic cases in this paper and requires a comparatively detailed discussion. A number of factors contribute to such situation, such as a nonmonospecific series of syntypes, a substantial degree of intraspecific variation, the existence of synonyms, and pseudocryptic species (in sympatry). Below we address each of those points in sequence.

Trichomycterus immaculatus was described on the basis of 14 syntypes from three different basins, mostly collected in association with the Thayer Expedition to Brazil (1865–1866): Rio Paraíba do Sul (MCZ 8300, 10 ex.; MCZ 8305, 1 ex.; MCZ 8307, 1 ex.), Rio São Mateus (an isolated coastal basin north of the Rio Doce ; MCZ 8302, 1 ex.) and an unspecified locality in the state of Goiás, probably in the Rio Tocantins-Araguaia system (MCZ 8298, 1 ex.). Localities and dates of Thayer Expedition collections follow Higuchi (1996). The latter lot contains a single small specimen which differs in key characteristics from all remaining syntypes. It has I + 7 pectoral-fin rays (vs. I + 8 in remaining syntypes), body depth 12% SL (vs. 14–17.9%); caudal peduncle depth 9.2% SL (vs. 11.3–14.9%) and paired S6 cephalic laterosensory pore (vs. single). Its specific identity cannot be determined at this point because of broader uncertainties in Trichomycterus taxonomy, the resolution of which extrapolate the scope of the present study. It is certain however that this specimen from Goiás is not conspecific with remaining syntypes and, given morphological differences, is not closely related to them. Syntypes from Rio Paraíba do Sul differ from the one from Rio São Mateus in number of interopercular odontodes (59 vs. 66–75) and in some body proportions such as the head length (15.7% SL vs. 17.5–19.3%), body depth (14.1% SL vs. 15.8–17.9%) and position of pelvic fin (55.9% SL vs. 60.6–63.3%). Recent molecular analyses have repeatedly indicated that forms referrable to T. immaculatus from the Rio Doce and Rio Paraíba do Sul are not conspecific and not closely related ( Ochoa et al., 2020; Costa et al., 2020b). The problem has been impervious to proper resolution so far because of incomplete representation from the Rio Doce and Rio São Mateus. Herein we have included samples of T. immaculatus and possibly related forms from all relevant sites, including Eigenmann’s locality in the Rio São Mateus (S127 and S146 on Fig. 1), and Rio Doce (S142 on Fig. 1) and from localities in the Paraíba do Sul (S194 to S196 on Fig. 1). The resulting picture makes it clear that the form in the Rio Doce is a single species throughout the basin, also occurring, with little differentiation, in adjacent basins to the north ( Rio Jucuruçu , Rio Peruípe , Rio Itanhaém , Rio Itaúnas and Rio São Mateus). On the other hand, large genetic and phylogenetic distances between T. immaculatus -like taxa from the Doce and the one from the Paraíba do Sul, as found in previous studies ( Katz et al., 2018; Costa et al., 2020b), are unabiguously corroborated, strongly supporting their separate specific status.

The biological situation of the entities involved is therefore clear in terms of general patterns in major basins. However, the nomenclatural problem is aggravated because the type series includes syntypes from both the São Mateus (a costal basin near the lower Rio Doce Basin ) and Rio Paraíba do Sul, in addition to the distantly-related one from Goiás. This renders the applicability of available names unstable, even more so because there are potential synonyms in both basins. In order to avoid taxonomic uncertainties and to stabilize the associated nomenclature, it is necessary to designate a lectotype. We discard the Goiás specimen from consideration because it belongs to a taxon markedly distinct from all other syntypes, not matching the concept of T. immaculatus in virtually the entire literature (including the original description of the species). Its small size and outlying provenance relative to other syntypes also make it inadequate as a name-bearer. Among remaining syntypes, the specimen from the Rio São Mateus is well preserved and allows observation of all characters mentioned in the original description. Additionally, a non-type specimen from the Rio Doce was also subsequently chosen to illustrate the species in Eigenmann (1918: pl. 52), further demonstrating that the authors considered the taxon in the Rio Doce as conforming to their conception of T. immaculatus . Therefore, we herein designate specimen MCZ 8302 ( Fig. 26) as the lectotype of T. immaculatus . The specimen was collected in the Rio São Mateus, an isolated coastal basin near the Rio Doce in the state of Espírito Santo. Our decision has nomenclatural ramifications because it implies that T. pradensis is a synonym of T. immaculatus and that the taxon from Rio Paraíba do Sul must bear the next available name [in this case Trichomycterus paquequerensis (Miranda Ribeiro, 1943) ; see below]. Our decision is based on the state of knowledge of the taxon in the Rio Doce and adjacent basins, represented in this study by hundreds of specimens and 33 molecular samples from throughout its geographic distribution. This database constitutes a solid basis on which to associate the name T. immaculatus with a well-delimited entity, both taxonomically and geographically. The taxon in the Rio Paraíba do Sul is not as well known and is represented by comparatively limited samples from few localities in this and other studies. An alternative decision to designate the name T. immaculatus to the Paraíba do Sul taxon runs the risk of once again perpetuating nomenclatural instability. As further corollary of lectotype designation, the type locality of T. immaculatus is now restricted to the Rio São Mateus, state of Espírito Santo, Brazil. With that act, the name T. immaculatus now applies to the taxon occurring in the Rio Doce and adjacent basins, but not to the form in the Rio Paraíba do Sul. Also, the remaining 13 syntypes, MCZ 8298, MCZ 8300, MCZ 8305 and MCZ 8307 are now paralectotypes.

With the issue of type determination resolved, it is possible to proceed to other problems about the taxonomic situation of T. immaculatus . The first of which is the status of potentially closely related nominal forms. Trichomycterus pradensis was described from three adjacent but independent basins in southern Bahia and northern Minas Gerais, Brazil: Jucuruçu (locality of the holotype), Peruípe and Itanhém. The three basins are sequentially north of the Rio Doce. The original description ( Sarmento-Soares et al., 2005) states that T. pradensis is distinguished from all congeners by a combination of three characters: closely-set S6 pores, a small patch of robust opercular odontodes (with 8 to 10 odontodes) and eight branched pectoral-fin rays. Examination of the holotype and 52 paratypes of T. pradensis , along with other data obtained for the present paper, shows that those traits need reevaluation. First, the number and spacing between S6 pores is variable in the type series, a situation similar to that in several other species of Trichomycterus from the Rio Doce. Second , the number of opercular odontodes actually varies between 13 and 25 in the type series of T. pradensis , with no specimens in the 8–10 range. Finally, eight branched pectoral-fin rays can be found in other Trichomycterus species (e.g. T. immaculatus , T. tantalus and in some rare cases in T. astromycterus also in the Rio Doce Basin ). The combination of all three characteristics (considering the corrected values for opercular odontode number) is matched in T. immaculatus , and we could not find additional distinguishing traits once all evidence is taken into consideration. The spotted colour pattern of the holotype of T. pradensis (cf. Sarmento et al., 2005: fig. 1) might superficially seem to distinguish it from the uniform-coloured T. immaculatus . However, the colour pattern varies in the entire range between those two extremes in the type series of T. pradensis (as mentioned in the original description; Sarmento-Soares et al., 2005: 296) ( Fig. 8), with such variation seen also in T. immaculatus ( Fig. 9). This variation may even be seasonal or environmentally induced, considering that a uniformly-coloured specimen from the Rio Doce reportedly changed to a spotted pattern in the course of 2 months in aquarium conditions ( Costa et al., 2020a). Careful examination of specimens referable to both T. pradensis and T. immaculatus , including type material of the two species and data from both internal and external anatomy, fails to reveal any other possibly diagnostic phenotypic characteristics. The same conclusion is reflected in DNA sequences, with little genetic barcoding divergence (1.2%) between specimens referable to T. pradensis (including topotypic material) and forms referable to T. immaculatus from throughout the Rio Doce and the Rio São Mateus. Finally, results of our phylogenetic analysis place samples of T. pradensis from the type locality in a clade including remaining specimens from the Rio Doce and adjacent basins, with divergence values insufficient to demontrate species differentiation (an observation previously made by Volpi, 2017). With both phenotypic and molecular evidence failing to demonstrate species-level divergence between forms referable to T. pradensis and T. immaculatus , plus the determination of the lectotype of the latter, the former species is proposed as a junior synonym of the latter.

Our analyses have shown that sequences of T. immaculatus from the Rio Doce are highly divergent from those of the Paraíba do Sul, fitting in widely disjunct branches of the phylogenetic tree obtained ( Fig. 1). This indicates that the populations identified as T. immaculatus from those two basins are actually two different species, cryptic or semicryptic. Because the lectotype is from the Rio São Mateus and demonstrably conspecific with the form in the Rio Doce , then the species in the Rio Paraíba do Sul must bear another name. Costa et al. (2020a) has recently demonstrated that T. paquequerensis (Miranda Ribeiro, 1943) is conspecific with the form then known as T. immaculatus in the Rio Paraíba do Sul. Their conclusion is based on both molecular and morphological data, and on recently collected topotypes of T. paquequerensis . Such conclusions led Costa et al. (2020a) to propose the latter species as a junior synonym of T. immaculatus . The biological situation is correctly inferred and we concur with their conclusion. However, with the determination of the lectotype situation implemented here, the name T. immaculatus no longer applies to the Rio Paraíba do Sul form. Therefore, it follows that T. paquequerensis is the oldest available name and the valid name for the species in the Rio Paraíba do Sul previously identified as T. immaculatus . This scenario solves the apparent conundrum of T. immaculatus coming out as two separate (and phylogenetically disjunct) terminals in the trees of Katz et al. (2018) and Costa et al. (2020b) (identified as T. immaculatus and T. cf. immaculatus in the former and as T. immaculatus and T. ‘immaculatus’ in the latter), a result corroborated here. The pairs of homonimous species each refer to the Rio Doce on the one hand and the Rio Paraíba do Sul on the other. In both papers, the sequences of T. immaculatus from the Rio Doce were originally from Ochoa et al. (2017), collected in the middle course of the basin and well within the sampling coverage of the present analysis. Although the situation seems clear nomenclaturally and biologically on the basis of sequence data, the morphological differentiation between the two species is subtle. We have found that among specimens examined for this study, T. immaculatus differs from the Paraíba do Sul taxon ( T. paquequerensis ) only by the fewer interopercular odontodes (37–59 vs. 66–75 in adult specimens) and by a shorter prepelvic length (50.9–58.0% SL vs. 60.6–63.3%). The holotype of T. paquequerensis is small (34 mm SL) and in poor preservation condition ( Fig. 27), but a count of 40 and 43 interopercular odontodes (on each side) can still be determined. The number of odontodes in species of Trichomycterus increases with growth, so that similar-sized specimens should be used for meaningful comparisons. Specimens 35–50 mm SL of T. immaculatus (sensu stricto) have 30–36 interopercular odontodes, therefore confirming that the species still has fewer odontodes than T. paquequerensis at those sizes, although the difference is expectedly less pronounced than in large specimens. Such differences still await confirmation on additional specimens of T. paquequerensis .

Two species are particularly prone to confusion with T.immaculatus in the Rio Doce , namely T. melanopygius and T. tantalus . Trichomycterus melanopygius is the species most similar to T. immaculatus , especially due to its homogeneous greyish colour pattern and to the presence of a dark horizontal stripe along the middle caudal-fin rays, visible in all specimens of the former and in small specimens (rarely adults) of the latter (in large specimens the trait is usually concealed by additional pigmentation). The two species look so similar in external aspect that they may easily be mistaken in superficial examination and have in fact been mixed together in museum collections. They have been proposed as pseudocryptic species ( Reis et al., 2020), which means that despite such similarities they can be readily distinguished by examination of details (given here in their respective diagnoses). This conclusion is corroborated by their highly significant COI genetic distance of 3.9% (cf. molecular results, Table 2).

Trichomycterus tantalus (described below) is another species that is similar to T. immaculatus , although not to the same degree as T. melanopygius . Most specimens of T. tantalus are different from T. immaculatus in colour pattern, because of an abrupt fading of dark coloration ventral to the lateral midline. However, such difference is not absolute, and some specimens are within the range of colour patterns seen in T. immaculatus ( Fig. 6A), but the two species are still easily distinguishable on the basis of several other traits, including an evident concave or forked caudal fin and a hypertrophied opercular patch of odontodes in T. tantalus (vs. a truncate caudal fin and regular-sized opercle in T. immaculatus ), among other characters described in more detail in their diagnoses. Their species-level differentiation is also corroborated by a barcoding divergence of 3.8% ( Table 2).

Geographical distribution: Trichomycterus immaculatus is widely distributed in the entire Rio Doce and adjacent basins (Rios Jucuruçu, Peruípe and Itanhaém, Itaúnas and São Mateus) ( Fig. 28). As defined here, T. immaculatus does not occur in the Rio Paraíba do Sul Drainage (see Remarks above). The species has a broad range of habitats, including both the main channel of large rivers (such as that of the Rio Doce ) and smaller tributaries and headwaters (also observed in the Rio Doce Basin ). In this regard, it differs from similar-looking relatives, T. melanopygius and T. tantalus , the former being restricted to small tributaries and headwaters and the latter occurring exclusively in main channels.

Type material examined (localities and respective dates of Thayer Expedition material follow Higuchi, 1996): type material of Pygidium immaculatum ; lectotype (designated herein): MCZ 8302, 1, 121.4 mm SL; Sao Matheos (state of Espírito Santo, Rio São Mateus at São Mateus); col. C.F. Hartt & E. Copeland in Thayer Expedition to Brazil, November-December 1865. Paralectotypes: MCZ 8300, 5, paralectotypes, 123.2– 165.0 mm SL; Juiz de Fora, Parahybuna (state of Minas Gerais, Rio Paraibuna at Juiz de Fora); col. H.W. Halfeld, 1854. MCZ 8305, 1, paralectotype, 117.6 mm SL; Juiz de Fora and Parahyba (state of Minas Gerais, Juiz de Fora and environs in the Paraiba Valley); col. L. Agassiz & J. Whitaker, in Thayer Expedition to Brazil, 21–27 June 1865. MCZ 8307, 1, paralectotype, 116.2 mm SL; same data as MCZ 8305. MCZ 8296, 1, paralectotype, 51.6 mm SL, Goyaz (state of Goiás or Tocantins, probably somewhere in the Tocantins-Araguaia Basin); col. Senhor Honorio (A.H. Ferreira), 1867. Type material of Trichomycterus pradensis ; MNRJ 28484, 09, paratypes, 39.4–109.0 mm SL; Jucuruçu, Rio Jucuruçu. MNRJ 28485, 5, paratypes, 40.0– 66.5 mm SL; Bahia, riacho a 500 m do ponto anterior, bacia do Rio Peruípe ; MNRJ 28488, 7, paratypes, 49.1–99.0 mm SL; Palmópolis, Rio Dois de Abril , bacia do Rio Jucuruçu. MNRJ 28490, 9, paratypes, 46.7–74.1 mm SL; Itanhém, Córrego Água Fria, Rio Itanhém.

Additional material studied: All following from Brazil, state of Minas Gerais; MZUSP 58479, 29, 47.5– 115.2 mm SL; Joanésia; Guacho Creek, tributary of Santo Antônio River Basin (19 ° 7 ’ 37.86 ” S 42°39’48.93”W); col. F.A. Bockmann & P.M.C. Araujo, 5 October 1997. MZUSP 69333, 1, 63 mm SL; Coroaci, Suaçuí Pequeno River (18°36’45.93”S 42°16’52.91”W); col. A.M. Zanata, 28 April 2001. MZUSP 69359, 1, 77.8 mm SL; Coroaci; Suaçuí Pequeno River (18°37’20.29”S 42°16’16.34”W); col. A.M. Zanata, 29 April 2001. MZUSP 69367, 1, 83.5 mm SL; Coroaci, Suaçuí Pequeno River (18°41’38.00”S 42°12’50.00”W); col. A.M. Zanata, 29 April 2001. MZUSP 75034, 2, 67 mm SL; Sarduá Tronqueiras River, tributary of Suaçuí River Basin (18°46’44.29”S 42°23’41.00”W); col. F. Di Dario & B. Di Dario, 8 December 2001. MZUSP 75059, 8, 78.6– 48.8 mm SL; Coroaci, Suaçuí River Basin (18°36’45.57”S 42°16’18.73”W); col. F. Di Dario & B. Di Dario, 6 December 2001. MZUSP 81028, 9, 133.8– 55.8 mm SL; Manhuaçu, Manhuaçu River (20°15’24.00”S 42°7’2.00”W); col. Carlos B.M. Alves, 23 April 2002. MZUSP 81029, 6, 59–114 mm SL; Manhuaçu, Manhuaçu River (20 °15’ 24.00 ”S 42°7’2.00”W); col. Carlos B.M. Alves, 22 April 2002. MZUSP 81032, 8, 37.1–79.7 mm SL; São Luiz, Manhuaçu River (20°20’12.00”S 42°4’48.00”W); col. Carlos B.M. Alves, 21 April 2002. MZUSP 94488, 2, 65.9–82.3 mm SL; Alto Rio Doce, Xopotó River, Piranga River Basin (21°4’4.00”S 43°27’50.00”W); col. Oyakawa, Baena e Loeb, 11 July 2007. MZUSP 123334, 45, 38.5– 85.3 mm SL; Bom Jesus do Manhuaçu, Manhuaçu River (20°17’34.24”S 42°8’50.88”W); col. T. Pessali, 7 September 2015. MZUSP 123335, 10, 45.3–55.9 mm SL; Bom Jesus do Manhuaçu, Manhuaçu River, Rio Doce Basin (20°17’34.24”S 42°8’50.88”W); col. T. Pessali, 22 April 2015. MZUSP 123336, 22, 53.1–105.3 mm SL; Realeza, Manhuaçu River, Rio Doce Basin (20°14’49.13”S 42°5’2.18”W); col. T. Pessali, 21 April 2015. MZUSP 123337, 92, 52.2–98.3 mm SL; Realeza, Manhuaçu River, tributary of Rio Doce Basin (20°14’49.13”S 42°8’26.17”W); col. T. Pessali, 8 September 2015. MZUSP 123342, 1, 67.5 mm SL; Naque, main channel of Rio Doce River (19°14’19.42”S 42°18’22.65”W); col. T. Pessali, 19 July 2017. MZUSP 123344, 7, 45.6–84.6 mm SL; Açucena, Corrente Grande River, Rio Doce Basin (18°57’9.86”S 42°21’38.20”W); col. T. Pessali, 31 July 2017. MZUSP 123345, 123, 48.4– 112.9 mm SL; Baguari, main channel of Rio Doce River (19°1’3.06”S 42°7’16.31”W); col. T. Pessali, 12 December 2016. MZUSP 123347, 12, 39.2–77.7 mm SL; Açucena, Corrente Grande River, Rio Doce Basin (18°57’9.86”S 42°21’38.20”W); col. T. Pessali, 31 May 2017. MZUSP 123348, 83, 50.2–111.4 mm SL; Baguari, main channel of Rio Doce River (19°1’27.83”S 42°7’34.40”W); col. T. Pessali, 20 December 2016. MZUSP 123349, 74, 43.7–76.7 mm SL, Itambacuri, Suaçui Grande River, tributary of Rio Doce Basin (18 ° 5’ 42.17” S 41°41’22.16”W); col. T. Pessali, 9 September 2015. MZSUP 123352, 4, 40–67.3 mm SL; Açucena, Corrente Grande River, Rio Doce Basin (18° 57’ 9.86”S 42°21’38.20”W); col. T. Pessali, 31 May 2017. MZUSP 123353, 23, 40.3–70.3 mm SL; Baguari, São Mateus Creek, tributary of Corrente Grande River (18°59’24.54”S 42°18’57.98”W); col. T. Pessali & V. Reis, 17 November 2017. MZUSP 123354, 1, 43.7 mm SL; Açucena, São Mateus Creek, tributary of Corrente Grande River (18°59’24.54”S 42°18’57.98”W); col. T. Pessali, August 2013. MZUSP 123356, 15, 46.9– 79.4 mm SL; Açucena, São Mateus Creek, tributary of Corrente Grande River (18°57’10.39”S 42°21’39.81”W); col. T. Pessali, 2 September 2017. MZUSP 123357, 3, 69.3–80.2 mm SL; Baguari, Suaçui Pequeno, tributary of Rio Doce Basin (18°56’2.50”S 42°5’2.18”W); col. T. Pessali & V. Reis, 16 November 2017. MZUSP 123367, 1, 51.8 mm SL; Bom Jesus do Manhuaçu, Manhuaçu River, Rio Doce Basin (20°17’34.24”S 42°8’50.88”W); col. T. Pessali, 27 April 2015. MZUSP 123368, 18, 45.6– 85.7 mm SL; Baguari, main channel of Rio Doce River (19°1’14.68”S 42°7’6.81”W); col. T. Pessali, 13 December 2016. MZUSP 123370, 8, 44.5–56.0 mm SL; Itambacuri, Suaçui Grande River, Rio Doce Basin (18°5’42.17”S 41°41’22.16”W); col. T. Pessali, 9 September 2015; Minas Gerais MZUSP 126408, 1, 91 mm SL, Rio Doce Municipality , Rio Doce Basin, Córrego dos Borges , stream flowing into left bank of Risoleta Neves Reservoir, (20°12’21.63”S 42°52’56.24”W); col. V.J.C. Reis, M.C.C. de Pinna, G.F. de Pinna & G. Ballen, 24 June 2018. LBP 1019, 2, 68.8–114.9 mm SL; between Capela Nova and Caranaíba, Piranga River, tributary of Rio Doce Basin (20°58’10.26”S 43°42’19.86”W); col. J.C. Oliveira, A.L. Alves & L.R. Sato, 13 October 2001. MZUFV 3335, 3, 109.4– 118.3 mm SL; Guaraciaba, UHE Brecha, Piranga River, Rio Doce Basin (20°29’42.01”S 43°0’15.59”W); col. J.A. Dergam & Tottola, 23 March 2003. MZUFV 3348, 2, 100.7– 105.5 mm SL; Ponte Nova, UHE Brito, Rio Piranga River (20°23’22.43”S 42°54’9.23”W); col. J.A. Dergam & M.R. Tottola, 24 March 2003. MZUFV 3378, 3, 82.5– 99.7 mm SL; Guaraciaba, UHE Brecha, Piranga River, Rio Doce Basin (20°29’34.47”S 43°0’24.18”W); col. J.A. Dergam & M.R. Tottola, 10 April 2003. MZUFV 3393, 10, 9.0– 110.5 mm SL; Ponte Nova, UHE Brito, Rio Piranga River (20°23’42.38”S 42°54’9.36”W); col. J.A. Dergam & M.R. Tottola, 12 April 2003. MNRJ 22453, 6, 41.8–81.3 mm SL; Manhuaçu, Rio Doce , tributary of Rio Manhuaçu , São Luiz Stream; col. P.A. Buckup, A.T. Aranda, F.A.G. Melo, 8 August 2001. MNRJ 37629, 3, 67.4–129.7 mm SL; Baixo Guandu, Mutum Preto River, locality between a village located at the KM14 and Alto Mutum; col. M. Britto, F. Pupo, L. Sarmento-Soares, 1 August 2010. Brazil, state of Espírito Santo: MZUSP 41738, 2, 48.4–71.6 mm SL; Ibatiba; São João River Basin (20°23’60.00”S 41°25’0.00”W); col. C.A.S. Lucena & P.V. Azevedo, 6 September 1989. MBML 55, 1, 49.7 mm SL; Colatina, Santa Maria do Rio Doce Basin (19°36’14.00”S 40°37’13.00”W); col. R.L. Teixeira & J.A.P. Scheneider, 29 August 1996. MBML 438, 12, 34.2–53.2 mm SL; Santa Teresa, Rúdio Waterfall, Santa Maria do Rio Doce Basin (19°50’11.00”S 40°41’27.00”W); col. R.L. Teixeira, 30 June 2000. MBML 636, 3, 70.0– 100.5 mm SL; Itaguaçu, Fazenda Coser, Santa Joana Basin (19°48’6.00”S 40°51’20.00”W); col. R.L.Teixeira & P.S.Miller, 8 September 2000. MBML 648, 5, 66.2– 85.3 mm SL; Itarana, Jatibocas Stream, Santa Joana Basin (19°52’26.00”S 40°52’31.00”W); col. R.L.Teixeira & P.S.Miller, 19 April 2001. MBML 672, 2, 66.6–74.8 mm SL; Itaguaçu, Fazenda Coser, Santa Joana Basin (19°48’6.00”S 40°51’20.00”W); col. R.L.Teixeira & P.S. Miller, 21 May 2001. MBML 695, 24, 44.4–104.6 mm SL; Itarana, Jatibocas Stream, Santa Joana Basin (19°52’26.00”S 40°52’31.00”W); col. R.L. Teixeira & P.S. Miller, 10 August 2000. MBML 701, 13, 36.5–71.9 mm SL; Itarana, Santa Joana Basin (19°52’26.00”S 40°52’31.00”W); col. R.L. Teixeira & P.S. Miller, 19 August 2000. MBML 753, 10, 62.1–117.5 mm SL; Itarama, Jatiboca Stream, Santa Joana Basin (19°52’26.00”S 40°52’31.00”W); col. R.L. Teixeira & P.S. Miller, 18 October 2000. MBML 765, 1, 94.7 mm SL; Itarana, Santa Joana River (19° 52 ’ 26.00 ”S 40°52’31.00”W); col. R.L. Teixeira & P.S. Miller, 8 February 2002. MBML 788, 1, 79.6 mm SL; Itarana, Jatibocas Stream, Santa Joana Basin (19°52’26.00”S 40°52’31.00”W); col. R.L. Teixeira, 12 October 2000. MBML 805, 4, 71.6–93.1 mm SL; Itarana, Jatibocas Stream, Santa Joana Basin (19 ° 52 ’ 26.00 ”S 40°52’31.00”W); col. R.L. Teixeira & P.S. Miller, 21 June 2001. MBML 807, 7, 65.5–80.5 mm SL; Itarana, Jatibocas Stream, Santa Joana Basin (19°52’26.00”S 40°52’31.00”W); col. R.L. Teixeira & P.S. Miller, 8 February 2001. MBML 999, 4, 40.4–51.1 mm SL; Itarana, Santa Joana River, tributary of Rio Doce Basin (19°52’26.00”S 40°52’31.00”W); col. R.L. Teixeira & P.C.M. Mili, 21 June 2001. MBML 1014, 1, 43.8 mm SL; Itarana, Limoeiro Creek, Santa Joana River (19°52’26.00”S 40°52’31.00”W); col. R.L. Teixeira, 19 April 2001. MBML 1357, 3, 43.4–66.4 mm SL; Colatina, Cachoeira do Oito Waterfall, Pancas River Basin (19°27’21.00”S 40°37’29.00”W); col. R.L. Teixeira & G.I. Almeida, 8 December 2005. MBML 1395, 1, 57.7 mm SL; Colatina, Santa Maria do Rio Doce Basin (19°32’55.00”S 40°38’7.00”W); col. R.L. Teixeira & Borlute, 11 October 2005. MBML 2243, 1, 71.7 mm SL; Laranja da Terra e Serra Pelada, Timbuva Stream, Gandu River Basin (19°54’55.00”S 41°5’18.00”W); col. L.M. Sarmento-Soares, R.F. Martins-Pinheiro, A.T. Aranda, R.L. Teixeira, M.M.C. Roldi & M.M. Lopes, 12 June 2009. MBML 2274, 3, 50.2–118.9 mm SL; Laranja da Terra, Lagoa Stream, Gandu River Basin (19°59’26.00”S 41°3’55.00”W); col. L.M. Sarmento-Soares, R.F. Martins-Pinheiro, A.T. Aranda, R.L. Teixeira, M.M.C. Roldi & M.M. Lopes, 12 June 2009. MBML 2290, 11, 27.6–85.6 mm SL; Afonso Cláudio, Rio do Cobre River, tributary of Guandu Basin (20° 9’19.00”S 41°8’31.00”W); L.M. Sarmento-Soares, R.F. Martins-Pinheiro, A.T. Aranda, R.L. Teixeira, M.M.C. Roldi & M.M. Lopes, 14 June 2009. MBML 2295, 2, 34.7–38.2 mm SL; Afonso Claudio Rio do Peixe River, Gandu River Basin (20°7’39.00”S 41°8’9.00”W); col. A.T. Aranda, R.F. Martins-Pinheiro, R.L. Teixeira & M.M.C. Roldi, 14 June 2009. MBML 2304, 18, 34.2–58.1 mm SL; Afonso Cláudio, Rio da Cobra River , tributary of Guandu River (20°12’4.00”S 41°8’7.00”W); col. A.T. Aranda, R.F. Martins-Pinheiro, R.L. Teixeira & M.M.C. Roldi, 14 June 2009. MBML 2312, 13, 33.1–76.5 mm SL; Afonso Cláudio, Rio da Cobra River, Guandu Basin (20°10’57.00”S 41°4’50.00”W); col. A.T. Aranda, R.F. Martins-Pinheiro, R.L. Teixeira & M.M.C. Roldi, 14 June 2009. MBML 2974, 10, 50.9–99.2 mm SL; Colatina, Santa Maria do Rio Doce Basin (19°32’21.00”S 40°37’50.00”W); col. R.L. Teixeira & E.C. Perrone, 6 November 1988. MBML 3465, 4, 53.1–103.5 mm SL; Baixo Gandu, Mutum Preto River, Mutum Preto River (19°23’7.00”S 40°53’52.00”W); col. L.M. Sarmento-Soares, R.F. Martins-Pinheiro, M.R. Britto & F.M.R.S. Pupo, 1 October 2010. MBML 3571, 9, 31.2–61.5 mm SL; Águia Branca, Águas Claras Stream, São José River (18°57’17.00”S 40°45’19.00”W); col. L.M. Sarmento-Soares, R.F. Martins-Pinheiro, M.R. Britto & F.M.R.S. Pupo, 5 October 2010. MBML 3878, 1, 47 mm SL; Colatina, São João Pequeno River (19°28’45.00”S 40°44’15.00”W); col. M.M. Martinelli, 27 September 2010. MBML 4274, 3, 32.3–42.1 mm SL; Águia Branca, Braço Azul River, São José Basin (19°3’44.00”S 40°34’21.00”W); col. J.L. Helmer, 12 August 2011. MBML 4385, 1, 49.35 mm SL; Santa Teresa, Santa Maria do Rio Doce River (19°46’21.10”S 40°38’2.70”W); col. R.B. Soares, J. Gurtler & V.R. Bada, 24 September 2011. MBML 4431, 1, 39.91 mm SL; Santa Teresa, Santa Maria do Rio Doce River (19°46’21.10”S 40°38’2.70”W); col. R.B. Soares, J. Gurtler & V.R. Bada, 24 September 2011. MBML 6139, 1, 66.2 mm SL; Santa Teresa, Cinco de Novembro River, Santa Maria do Rio Doce Basin (19°49’42.90”S 40°38’18.40”W); col. C.J. Cunha, J.P. Silva & R.B. Soares, 20 August 2012. MBML 6153, 8, 23.7–84.4 mm SL; Santa Teresa, Cinco de Novembro River, Santa Maria do Rio Doce River (19°50’26.00”S 40°37’47.30”W); col. C.J. Cunha, J.P. Silva & R.B. Soares, 20 August 2012.