Research Article |
Corresponding author: Teddy Angarita-Sierra ( tgangaritas@unal.edu.co ) Academic editor: Uwe Fritz
© 2022 Teddy Angarita-Sierra, Sergio Daniel Cubides-Cubillos, Juan Pablo Hurtado-Gómez.
This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Citation:
Angarita-Sierra T, Cubides-Cubillos SD, Hurtado-Gómez JP (2022) Hidden in the highs: Two new species of the enigmatic toadheaded pitvipers of the genus Bothrocophias. Vertebrate Zoology 72: 971-996. https://doi.org/10.3897/vz.72.e87313
|
Bothrocophias microphthalmus (Cope, 1875) currently comprises most mid- to highland populations of the genus Bothrocophias in the eastern versant of the Andes. We describe two new species of Bothrocophias from the highlands of the Colombian Andes previously referred to as B. microphthalmus based on morphological and genetic evidence. Our phylogenetic analysis revealed that B. micropthalmus as currently recognized is paraphyletic with respect to B. hyoprora, and the two new taxa are sister lineages. These new toadheaded pitvipers can be morphologically distinguished from their congeners based on the presence of contact between the internasal scales, the number of prefoveal scales, the presence of a lacunolabial scale, the arrangement of supralabial scales, ventral scale counts, the color pattern of the dorsal and ventral surfaces of the body and tail, and hemipenial morphology. We discuss some possible taxonomic scenarios for the lineages found within the B. microphthalmus species complex but refrain from making additional taxonomic changes given our reduced sampling of the southern lineages.
Cryptic species, high Andean snakes, integrative taxonomy, medically important snakes, mtDNA, South America, Viperidae
Some of the most enigmatic and poorly known South American viperids are the toadheaded pitvipers of the genus Bothrocophias (
A few taxonomic studies of Bothrocophias species have been conducted since the generic recognition of Bothrocophias and the description of B. myersi by
Bothrocophias microphthalmus has the broadest distribution in the Andes among toadheaded pitviper species; it ranges from the eastern versant of the Cordillera Oriental of Colombia in the departments of Boyacá, Casanare, and Cundinamarca (1,700–2,400 m asl); the Amazonian slopes of the Ecuadorian Andes in the provinces Morona-Santiago, Pastaza, Tungurahua, and Zamora-Chinchipe (850–1,650 m asl); the Amazonian slopes of Brazil in the state of Rondônia (ca. 150 m asl); the eastern versant of the Peruvian Andes in the departments of Cuzco, San Martin, Huanaco, and Sira (800–2,100 m asl); and across the Bolivian Andean highlands in the departments of Beni, Cochabamba, La Paz, Pando, and Santa Cruz (600–1,600 m asl) (Nicéforo-María 1975;
Given the high diversity of medically important snakes in Colombia, the National Health Institute (INS, Spanish acronym) of Colombia has put much effort over the last decade into making collections of venomous snakes to enhance the production and neutralization capacity of their polyvalent viperid snake antivenom for the treatment of serious snakebite accidents caused by viperid snake species such as Bothrocophias microphthalmus (
Live Bothrocophias specimens were obtained as donations from Colombia’s Environmental Regional Autonomous Corporations (CARs, Spanish acronym). When snake specimens died in captivity, they were fixed and preserved in 10% formalin and 70% ethanol, respectively, and deposited in the INS zoological collection (INSZ). The procedures used and environmental conditions during the captivity period and fixing process were approved by the Animal Ethical Use Committees of the Instituto Nacional de Salud de Colombia (protocol INT-R04.0000–001), abided by the Colombian animal welfare law (
There remains much debate over the utility of different species concepts in light of the recognition that no single concept can simultaneously account for the diverse ways in which life evolves (
We believe the various populations of B. microphthalmus to be a species complex based on previously discussed taxonomic challenges (
We followed the species delimitation approaches proposed by
To evaluate the genetic distinctiveness and phylogenetic position of Colombian populations historically assigned to B. micropthalmus, we obtained genetic sequences from five individuals deposited in the Venomous Animal Tissue Bank of the National Institute of Health (INSBT), Bogotá, Colombia (Table
Genomic DNA was extracted using the phenol-chloroform method (
Cytochrome b (MT-CYB) and NADH subunit 4 (MT-ND4) sequences of Bothrocophias species used in the present study.
Bothrocophias species | Locality | Genbank accession numbers (CYTB-ND4) | Voucher | Source |
B. myrringae sp. nov. | La Calera, Cundinamarca. Colombia | OP082447–OP082452 | INS268 | This study |
B. myrringae sp. nov. | El Calvario, Meta. Colombia | OP082446–OP082451 | INS099 | This study |
B. tulitoi sp. nov. | Garagoa, Boyacá. Colombia | OP082448–OP082453 | INS100 | This study |
B. tulitoi sp. nov. | Garagoa, Boyacá. Colombia | OP082450–OP082455 | INS148 | This study |
B. tulitoi sp. nov. | Garagoa, Boyacá. Colombia | OP082449–OP082454 | INS169 | This study |
B. cf. microphthalmus | Zamora, Ecuador | AF292577.1–AF292615.1 | FHGO2566 | Wüster et al. (2002) |
B. cf. microphthalmus | Zamora, Ecuador | FR691570.1–FR691538.1 | QCAZ6016 | Melaun et al. (2010) |
B. microphthalmus | Pasco, Peru | AY223594–AY223638.1 | LS9372 | Parkinson et al. (2003) |
B. cf. microphthalmus | Bolivia | FR691567.1–FR691540.1 | MNKR4983 | Melaun et al. (2010) |
B. cf. microphthalmus | Bolivia | FR691568.1–FR691541.1 | MNKR4988 | Melaun et al. (2010) |
B. cf. microphthalmus | Bolivia | FR691569.1–FR691542.1 | MNKR4989 | Melaun et al. (2010) |
B. cf. microphthalmus | Bolivia | FR691565.1–FR691539.1 | SMF86334 | Melaun et al. (2010) |
B. hyoprora | Leticia, Amazonas. Colombia | AY223593.1–FN431781.1 | CLP | Parkinson et al. (2003) |
B. hyoprora | Morona, Ecuador | AF292576.1–AF292614.1 | FHGO4005 | Wüster et al. (2002) |
B. hyoprora | Orellana, Ecuador | FR691571.1–FR691537.1 | QCAZ5577 | Melaun et al. (2010) |
B. lojanus | Loja, Ecuador | FR691566.1–FR691536.1 | QCAZ6018 | Melaun et al. (2010) |
B. campbelli | Chimborazo, Ecuador | AF292584.1–AF292622.1 | INHMT | Wüster et al. (2002) |
Azemiops feae | China | KJ872487 |
|
|
Agkistrodon piscivorus | EEUU | EF669477 | Zhi et al. (2007) | |
Bothrops pubescens | Brazil | MN37992 |
|
The obtained sequences were aligned with sequences of the genus Bothrocophias deposited in Genbank. We used Azemiops feae, Agkistrodon piscivorus, and Bothrops pubescens as outgroups (Table
The best partitioning scheme and evolutionary model were obtained using ModelFinder (
We examined specimens of Bothrocophias microphthalmus housed in the following biological collections in Colombia: Museo de la Universidad La Salle (
Terminology used in the diagnosis, comparisons, and description sections followed
We carried out a multidimensional scaling (MDS) test to assess whether the meristic characters and color pattern allow two candidate species to be distinguished among Colombian populations currently assigned to B. microphthalmus. This statistical technique does not require a priori grouping of specimens or linearity assumptions of the dataset (
Over the last decade (2010–2020), the Colombian surveillance health system (SIVIGILA Spanish acronym) has reported a total of 336 bothropic-like envenomations across known and new localities within Colombia, and 240 of these snakebites have been clearly caused by Bothrops species. Determining the incidence of snakebites of the two new species is a major challenge because Bothrocophias might be misidentified as Bothrops asper and B. atrox. In addition, Bothrocophias is not listed in the list of possible genera that cause snakebites in Colombia on the SIVIGILA notification form for reporting snakebite accidents. Thus, there are either no official reports of Bothrocophias microphthalmus envenomation, or snakebites attributed to Bothrops species might have been caused by Bothrocophias species. We reviewed all snakebite records (Instituto Nacional de Salud 2020) attributed to Botrhocophias and retained those most likely caused by the two new species described herein using elevation and the presence of B. atrox records at the site of reported snakebites as exclusion criteria and the match of reported symptoms of snakebite victims with those described in the literature as inclusion criteria (Campbell and Lamar, 2004).
Our final concatenated alignment of the two mitochondrial fragments comprised 1616 bp. The topology of the BI and ML trees was concordant, and clade support was similar among the two trees (Fig.
Kimura 2-p genetic distances between the Bothrocophias clades ranged from 14.4% to 1.76% (Table
Our MDS results showed conspicuous distance or dissimilarity between the two Colombian candidate species of the B. microphthalmus complex. Both candidate species can be distinguished by quantitative differences in traits, such as the presence/absence of lacunolabial scale, postfoveal scale, presence/absence of internasal scales separated by one small scale, prelacunal scales, subnasal scales and preventral scale counts, and number of dorsal bands. The MDS goodness-of-fit between the fitted and observed distances shows an excellent stress value (S) = 0.019, and the candidate species showed low overlap in the morphospace, indicating that there is an underlying structure based on the set of the meristic and color pattern characters used that allow the two groups to be clearly differentiated (Fig.
Maximum likelihood tree obtained in IQ-tree based on the concatenated alignment of the two mitochondrial fragments (1616 bp), depicting the phylogenetic relationships of the genus Bothrocophias. Outgroups were removed for clarity. Values above branches indicate Ultrafast Bootstrap support of the ML tree; values below branches indicate posterior probabilities of the Bayesian tree.
Multidimensional scaling test of the merisitic characters between Bothrocophias tulitoi sp. nov. (cyan squares) and Bothrocophias myrringae sp. nov. (magenta dots). Vectors (black arrows) denote the discrimination capacity of each merisitic variable. A Dorsal scale count after head. B Dorsal scale count midbody. C Dorsal scale count before cloaca. D Dorsal bands. E Ventral scale counts. F Preventral scale counts. G Subcaudal scale counts. H Subnasal scales. I Presence/absence of internasal scales separated by one small scale. J Prefoveal scales. K Subfoveal scale. L Posfoveal scale. M Prelacunal scales. N Presence or absence of lacunolabial scale. O Sublacunal scales. P Supralacunal scales. Q Interoculabial scales. R Intersupraocular scales. S Cantal scales. T Intercantal scales. U Interrictial scales. V Sinsfisial scales. W Supralabial scales. X Infralabial scales. Y Preocular scale.
In general, the independent lines of evidence assessed (Table
Hence, the combination of independent lines of evidence showed a congruent pattern of divergence among the several taxonomic characters assessed, indicating lineage separation between candidate species from Colombia, Ecuador, and Bolivia. Although only four of the five lines of evidence assessed were congruent within Colombian populations, there was clear evidence of lineage separation, including in hemipenial morphology, which is a proxy of reproductive isolation. Therefore, the available data of the present study, including the observed phylogenetic positions and morphological distinctiveness, support the recognition of the two Colombian clades as new taxa, which are described as follows.
Estimates of the evolutionary divergence between sequences of Bothrocophias microphthalmus populations expressed as percentages (averages). Values below the diagonal represent between-lineage divergences using 1270 bp of the Cytochrome b (MT-CYB) and NADH subunit 4 (MT-ND4) genes.
Bothrocophias species/clade | 1 | 2 | 3 | 4 | 5 | 6 | |
1 | B. lojanus | ||||||
2 | B. microphthalmus-Bolivia | 13.06% | |||||
3 | B. microphthalmus-Ecuador | 13.51% | 9.52% | ||||
4 | B. microphthalmus-Peru | 13.26% | 8.64% | 7.02% | |||
5 | B. hyoprora | 14.44% | 9.72% | 8.21% | 9.31% | ||
6 | B. myrringae sp. nov. | 13.88% | 9.96% | 7.94% | 8.04% | 6.49% | |
7 | B. tulitoi sp. nov. | 12.78% | 8.91% | 7.30% | 7.88% | 6.00% | 1.76% |
Species delimitation for Bothrocophias microphthalmus populations hypothesized as candidate species using the integrative taxonomic approach of
Species hypotheses for Bothrocophias microphthalmus complex | mtDNA tree (UFB/Bp) | mtDNA genetic distinctiveness | Color pattern | Meristic characters | Hemipenial morphology |
B. microphthalmus – Bolivia | Distinguishable lineage highly supported (100/1) | Highly distinct (8.64%) | Clearly diagnosable | Clearly diagnosable | Unknown |
B. microphthalmus – Ecuador | Distinguishable lineage highly supported (97/1) | Highly distinct (7.02%) | Clearly diagnosable | Clearly diagnosable | Clearly diagnosable |
B. microphthalmus – Peru | Distinguishable lineage poorly supported (>95/>0.95) | Highly distinct (7.02%) | Clearly diagnosable | Clearly diagnosable | Unknown |
B. tulitoi sp. nov. | Distinguishable lineage highly supported (95/1) | Highly distinct (6–12.78%) from non-Colombian populations, but short distance from B. microphthalmus Colombia 1 (<2%) | Clearly diagnosable | Clearly diagnosable between non-Colombian populations, and moderate within Colombian populations* | Clearly diagnosable |
B. myrringae sp. nov. | Distinguishable lineage highly supported (99/1) | Highly distinct (6.49–13.88%) from non-Colombian populations, but short distance from B. microphthalmus Colombia 2 (<2%) | Clearly diagnosable | Clearly diagnosable between non-Colombian populations, and moderate within Colombian populations* | Clearly diagnosable |
Generic placement. The new species are recognized as members of the genus Bothrocophias based on their placement in phylogenetic trees and according to the following combination of morphological characters defined by
Bothrocophias microphthalmus. (
(Fig.
COLOMBIA (n=20; Fig.
Bothrocophias tulitoi sp. nov. can be distinguished from all its congeners by the following combination of characters: (1) 150–172 ventral scales in females, 153–162 ventral scales in males; (2) internasal scales in contact or separated by a single small scale; (3) absence of canthorostral scales; (4) absence of lacunolabial scale; (5) one prelacunal scale; (6) hemipenial lobes subconical and ornamented toward the apex by large and dense calyces with spinulate edges; (7) bifurcation point of hemipenial lobes about 3–6 sudcaudal scales; (8) hemipenial body ornamented by numerous dense, large, and strongly calcified mesial spines arranged in oblique lines; (9) in sulcate view, lateral and mesial spines of the hemipenial body homogeneous in size; (10) body surface with less than 28 dark-brown bands dorsally and/or juxtaposed trapezoid-shaped blotches with paler centers; and (11) ventral surface of the tail uniformly bright reddish or orange-reddish speckles with black spots without a regular pattern (Figs
Bothrocophias tulitoi sp. nov. can be distinguished from all Bothrocophias species by having a creamy white or pale yellow ventral ground color with ventral scales heavily marked with black pigment towards the edges contacting the paraventral scales, and the presence of spots without a regular pattern on the mesial surface turning heavily mottled with dark brown pigment toward the tail (versus homogeneously dark brown to black in B. campbelli; mottled heavily with dark brown pigment, with the pale interspaces between the ventrolateral blotches encroaching on the lateral edges of ventral scales in B. colombianus; greyish brown medially, becoming paler laterally, with or without alternating dark brown spots in B. hyoprora; yellow mottled with pale to dark brown, darker posteriorly in B. lojanus; and pale pink to almost white in B. myersi); ventral surface of the tail with bright reddish or orange-reddish speckles with black spots without a regular pattern, and heavily marked with dark pigment towards the base (versus proximally dark brown and distally yellow or yellow-green in B. andianus; bright yellow to tan with diffuse grayish or brown pigment in B. campbelli; cream or pale yellow with a sparse peppering of brown in B. colombianus; and whitish with a moderate suffusion of grey in B. myersi). Comparisons of meristic and hemipenial characters with all its congeners of toadheaded pitvipers are summarized in Table
Additionally, Bothrocophias tulitoi sp. nov. can be distinguished from Ecuadorian, Peruvian, and Bolivian populations of toadheaded pitvipers currently classified as B. microphthalmus by having ventral surface of tail with bright reddish or orange-reddish speckles with black spots without a regular pattern and heavily marked towards the base (versus heavily marked with black or dark brown pigment proximally, mottled medially with pale to dark brown, and pale diffuse mottling or pale with interspaces mottled distally in Bolivian and Peruvian populations). Comparisons of meristic and hemipenial characters with the Ecuadorian, Peruvian, and Bolivian populations of toadheaded pitvipers currently classified as B. microphthalmus are summarized in Table
Comparisons of meristic and hemipenial characters between B. myrringae sp. nov., B. tulitoi sp. nov. and all its congeners of toadheaded pitvipers.
Trait | B. andianus | B. campbelli | B. colombianus | B. hyoprora | B. lojanus | B. myersi | B. myrringae sp. nov. | B. tulitoi sp. nov. |
Snout | No upturned | No upturned | No upturned | Upturned | No upturned | No upturned | No upturned | No upturned |
Intersupraoculars | 3–10 | 3–8 keeled | 6–10 Tuberculate | 2–9 smooth | 3–5 slightly keeled | 3–6 smooth | 7–9 slightly keeled | 5–9 slightly keeled |
Lacunolabial | Present | Absent | Absent | Often present | Absent | Present | Present | Absent |
Dorsal scale surface | Keeled | Keeled | Tuberculate | Keeled | Keeled | Keeled | Keeled | Keeled |
Ventral scale counts | 154–179 | 152–177 | 162–173 | 118–142 | 144–145 | 139–151 | 152–161 | 150–168 |
Subcaudal scale counts | 49–63 | 48–64 | 51–54 | 44–52 | 37–46 | 44–52 | 41–54 | 46–58 |
Shape of the hemipenial lobes | Robust and cylindrical | Robust and cylindrical | Conical | Slim and cylindrical | Conical | |||
Walls of the Sulcus spermaticus | Robust and well defined | Robust and well defined | Robust and well defined | Weakly defined | Robust and well defined | |||
Size of the lateral and mesial spines of the hemipenis | Variable | Variable | Variable | Variable | Invariable | |||
Ornamentation of the hemipenial lobes apex | Large and dense calyces | Large and dense calyces | Large and dense calyces | Large and scarce calyces | Large and dense calyces |
Meristic variation in Bothrocophias tulitoi sp. nov. and B. myrringae sp. nov. Bold numbers represent mode or mean. Numbers within parenthesis represent minimum and maximum values observed. * = meristic characters undifferenced by sex.
Trait | B. tulitoi sp. nov. | B. myrringa sp. nov. | B. cf. microphthalmus – Bolivia. | B. cf. microphthalmus – Ecuador. | B. microphthalmus – Peru |
---|---|---|---|---|---|
Sample size | Male (n=10), Female (n=17) | Male (n=3), Female (n=3) |
|
Male (n=10), Female (n=12) | Male (n=2), Female (n=1) |
Upturned snout | |||||
Male | Absent | Absent | Moderately | Moderately | Moderately |
Female | Absent | Absent | Moderately | Moderately | Moderately |
Snout | |||||
Male | Prognathous | Prognathous | Prognathous | Prognathous | Truncate |
Female | Prognathous | Prognathous | Prognathous | Prognathous | Truncate |
Dorsal scales | 21(21–23) /23 (21–23) /19 (17–19) | * | |||
Male | 23 (22–24) / 23 (21–23) / 19 (17–19) | 23 (21–23) /23 /19 | 23 (21–25) / 23 (21–25) / 18 (15–20) | ||
Female | 23 (21–24) / 23 (22–25) /19 (17–19) | 23 (21–23) / 23 (21–23) / 19 (17–19) | 23 (22–24) / 23 (21–23) / 19 (17–19) | ||
Dorsal bands | 16–18 * | ||||
Male | 24 (20–29) | 30 (28–34) | 21 (19–23) | 18 | |
Female | 26 (19–29) | 30 (25–34) | 16 (15–17) | 20 | |
Preventral scales | 2 (1–3) * | ||||
Male | 4 (3–6) | 4 (4–5) | 2 (1–2) | 4 (3–6) | |
Female | 5 (3–5) | 5 (3–5) | 2 (1–2) | 5 (3–5) | |
Ventral scales | |||||
Male | 157.2 (153–162) | 153.7 (152–155) | 142–152 | 148 (139–155) | 144–146 |
Female | 162.2 (150–172 | 159.3 (157–161) | 143–147 | 147.3 (138–151) | 155–159 |
Subcaudal scale | |||||
Male | 51.5 (46–58) | 52.7 (52–54) | 50 | 49.7 (46–53) | 52–55 |
Female | 50.2 (45–55) | 48 (41–52) | 42–45 | 55.3 (50–60) | 51–53 |
Anal scale | |||||
Male | Single | Single | Single | Single | Single |
Female | Single | Single | Single | Single | Single |
Rostral | |||||
Male | 1 | 1 | 1 | 1 | 1 |
Female | 1 | 1 | 1 | 1 | 1 |
Cantorostral | |||||
Male | Absent | Absent | Present | Present | Present |
Female | Absent | Absent | Present | Present | Present |
Subnasal | 1 | ||||
Male | 0 (0–1) | 0 (0–1) | 1 | ||
Female | 0 (0–1) | 0 (0–1) | 1 | ||
Prefoveal | 4–8 * | 2 * | |||
Male | 2 (2–4) | 2 | 2 (2–7) | ||
Female | 3 (2–4) | 2 (3–4) | 4 (5–4) | ||
Subfoveal | 1–2 * | 2 * | |||
Male | 0 | 0 | 1 (0–2) | ||
Female | 0 (0–1) | 0 | 1 (0–2) | ||
Postfoveal | 2 | ||||
Male | 1 (0–1) | 0 (0–1) | 1 (0–2) | ||
Female | 1 (0–2) | 1 | 1 (0–2) | ||
Prelacunal | 1 * | 4 * | |||
Male | 1 | Absent | 1 | ||
Female | 1 | Absent | 1 (1–2) | ||
Lacunolabial | |||||
Male | Absent | Present | Absent | Absent | Absent |
Female | Usually absent | Present | Absent | Absent | Absent |
Supralacunal | |||||
Male | 1 | 1 | 1 | 1 | 1 |
Female | 1 (1–2) | 1 (1–2) | 1 | 1 | 1 |
Sublacunal | 2 | ||||
Male | 1 | 1 | 1 | 1 | |
Female | 1 (1–2) | 1 | 1 | 1 | |
Internasals | Usually in contact, rarely separated by one small scale | Usually in contact, rarely separated by one small scale | Separated by 2–3 scales | Usually separated by 2–3 scales | Usually separated by one large scale |
Interoculolabial | 3–4 * | 6–9 * | |||
Male | 5 (5–9) | 7 (6–7) | 8 (8–10) | ||
Female | 8 (6–8) | 7 (6–8) | 8 (8–9) | ||
Intrasupraoculars | 5–9 * | ||||
Male | 7 (6–10) | 8 (7–9) | 8 (8–9) | ||
Female | 8 (5–12) | 7 (7–8) | 8 (8–9) | ||
Intercantals | 5–4 * | 3 * | |||
Male | 4 (2–4) | 3 | 2 | ||
Female | 4 (3–5) | 4 (3–4) | 2 | ||
Interrrictals | 28 (26–31) * | 26 (23–31) * | 26 * | ||
Male | 25 (23–28) | 27 (27–28) | |||
Female | 25 (23–29 | 25 (25–26) | |||
Supralabials | 7–8 * | 7 * | |||
Male | 7 (7–8) | 6 (6–7) | 7 (7–8) | ||
Female | 7 (6–8) | 6 | 7 (7–8) | ||
Infralabials | 8–10 * | 10 * | |||
Male | 9 (9–10) | 8 (9–10) | 8 (8–10) | ||
Female | 9 (8–11) | 9 (8–10) | 9 (8–10) | ||
Preoculars | 2 | 2 * | |||
Male | 3 (2–3) | 2 (2–3) | 2 (2–3) | ||
Female | 3 (2–3) | 2 (2–3) | |||
Postoculars | |||||
Male | 2 | 2 | 2 | 2 | 2 |
Female | 2 | 2 | 2 | 2 (1–2) | 2 |
Hemipenial lobes shape | Conical | Cylindrical | Cylindrical | ||
Hemipenial ornamentation | Numerous dense, large, and strongly calcified mesial spines; hemipenial lobes distally ornamented by dense and small calyces | Few large and strongly calcified mesial spines arranged in oblique rows, with lateral and mesial spines of the hemipenial body variable in size; hemipenial lobes distally ornamented by large and weakly developed calyces | Few large and strongly calcified mesial spines arranged in oblique rows, with lateral and mesial spines of the hemipenial body variable in size; hemipenial lobes distally ornamented by dense and small calyces | ||
Walls of the Sulcus spermaticus | Well defined | Weakly defined | Well defined | ||
SVL (mm) | 327–724 * | ||||
Male | 377.4 (157–554) | 491.7 (388–570) | |||
Female | 459.9 (177–820) | 595 (436–754) | |||
TL (mm) | 49–108 * | ||||
Male | 70.7 (32–105) | 89.0 (68–103) | 780 | ||
Female | 71.4 (27–119) | 83.7 (67–102) | 908 |
(Figs
Color in life of Bothrocophias tulitoi sp. nov. A, C Lateral and dorsal view of a male neonate (paratype INSZ 0128). B, D Lateral and dorsal view of an adult female (paratype INSZ 0144). All specimens from vereda Cienaga La Valvanera, municipality of Garagoa, department of Boyacá, Colombia. Coordinates: N 5.106535941, W –73.25888414.
Hemipenial architecture. A, B, C Sulcate, lateral, and asulcate views of the hemipenis of the holotype of Bothrocophias tulitoi sp. nov. (INSZ 073) from vereda Ciénaga La Valvanera, municipality of Garagoa, department of Boyacá, Colombia. Coordinates: N 5.106535941, W –73.25888414; elevation 1894 m. asl. D, E, F Sulcate, lateral, and asulcate views of the hemipenis of the holotype of Bothrocophias myrringae sp. nov. (INSZ 0268) from vereda Mundo Nuevo, municipality of La Calera, department of Cundinamarca, Colombia. Coordinates: N 4.660602778, W –73.88491667; elevation 2,761 m asl.
Ground color of the dorsal surface of the head is pale brown to brown with diffuse marks dark-brown or grey without a distinctive pattern. Ground color of the lateral surfaces of the head is scattered grey-brown from the snout to anterior edge of the eye; a conspicuous dark-brown postocular stripe running obliquely from the posterior edge of the eye to the angle formed by the quadrate and jaw bone joint encompasses the temporal scales, the last two supralabial scales, the last infralabial scale, and the mesial scales located between the preventral and infralabial scales; conspicuous tricolored ocelli in the third to fifth supralabial scales and third to seventh infralabial scales with center white or white-cream, followed by an internal edge dark-brown or black, and external broad circle or edge yellow or yellow-reddish. Ventral surfaces of the head are dark orange-gold and peppered with brown with conspicuous tricolored ocelli as described above in the first, third to seventh infralabial scales and first pair of gular scales. Ground color of the dorsal body surfaces is yellow-tan to brown mottling with dark brown-reddish pigment and weak orange speckles; 27 dark-brown bands and/or opposite or juxtaposed trapezoid-shaped blotches with pale center ornamented with or without brown spots. Ground color of body ventral surfaces is creamy white or yellow with ventral scales heavily marked with black pigment towards the edges contacting paraventral scales and spotted without a regular pattern on the mesial surface turning heavily mottled with dark brown pigment toward the tail; edges of the spots of the mesial surfaces yellow reddish turning dark brown toward the tail. Dorsal surfaces of the tail covered by nine broad dark brown bands separated by four narrow pale bands that fuse toward the distal end of the tail; tail ventral surface with bright reddish or orange-reddish speckles with black spots without a regular pattern, and heavily marked with dark pigment towards the base.
(Fig.
(Fig.
(Table
(n=7, Fig.
We dedicate this species to the late Colombian educator Tulio Manuel Angarita Serrano (1941–2021, father of the first author), known as Tulito (employing the diminutive Spanish suffix “ito”) by his colleagues, friends, and relatives. The specific epithet tulitoi represents the Latin translation of the nickname from the Spanish name Tulito. Professor Angarita-Serrano was a pioneer of the modern Colombian education model that helped catalyze the development of the theoretical and practical tools needed to implement institutional educational projects in Colombian public and private schools (see Angarita-Serrano 1990; Angarita-Serrano 1994; Angarita-Serrano and Chaves 1995; Angarita-Serrano 1996; Angarita-Serrano, 2000). He was also known for being a big thinker, a passionate advocate for the rights to education and free thought, and the development of educational paradigms that have helped Colombians overcome the new social, socioeconomic, and environmental challenges of the third millennium.
(Fig.
A total of 40 snakebite events over the last decade might have been caused by B. tulitoi sp. nov. Both mild and moderate envenomation have been noted in 50% of patients, and no severe cases nor fatalities were reported. Local symptoms reported included oedema (92.5%), pain (87.5%), erythema (47.5%), ecchymosis (20%), paresthesia (17.5%), phlyctens (15%), paresthesia (17.5%), and bruises (7.5%); systemic symptoms included sickness (45%), vomiting (15%), vertigo (12.5%), bradycardia (7.5%), gingivorrhea (7.5%), muscular weakness (5%), hematuria (5%), hypotension (5%), abdominal pain (5%), and altered vision (5%).
Bothrocophias microphthalmus.
[Figs
COLOMBIA [n=5; Fig.
Bothrocophias myrringae sp. nov. can be distinguished from all its congeners by the following combination of characters: (1) 157–161 ventral scales in females, 152–155 ventral scales in males; (2) internasal scales in contact or separated by a single small scale (3) absence of canthorostral scales; (4) lacunolabial scale usually present; (5) hemipenial lobes slim and cylindrical, moderately capitate distally, weakly ornamented toward the apex with large and scarce calyces with weakly spinulate edges; (8) bifurcation point of the hemipenial lobes about 2–4 sudcaudal scales; (9) hemipenial body ornamented by numerous mesial spines that increase in size from the center to periphery of the hemipenial body and arranged in oblique lines; (10) in sulcate view, lateral and mesial spines of the hemipenial body variable in size; (11) sulcus spermaticus walls weakly defined; (12) usually more than 28 dark-brown bands and/or opposite or juxtaposed trapezoid-shaped blotches with paler centers dorsally; and (13) ventral surfaces of the tail with bright reddish or orange-reddish speckles with black spots without a regular pattern and heavily marked with dark pigment towards the base (Fig.
Bothrocophias myrringae sp. nov. can be distinguished from all its congeners by its creamy yellow ventral surfaces and ventral scales mottled with dark brown pigment, becoming creamy white toward the edges in contact with the paraventral scales, forming a white-cream paraventral stripe which proximally fuses with the final edges of the postocular stripe, and distally is interrupted by dark spots without a regular pattern; mesial ventral surfaces become heavily mottled with dark brown pigment toward the tail (versus homogeneously dark brown to black in B. campbelli; heavily mottled with dark brown pigment, with the pale interspaces between the ventrolateral blotches encroaching on the lateral edges of the ventral scales in B. colombianus; greyish brown medially, becoming paler laterally, with or without alternating dark brown spots in B. hyoprora; yellow mottled with pale to dark brown, darker posteriorly in B. lojanus; pale pink to almost white in B. myersi); ventral tail surface bright reddish or orange-reddish speckles with black spots in an irregular pattern, and the base of tail heavily marked with dark pigment (versus base of tail dark brown and distally yellow or yellow-green in B. andianus; bright yellow to tan with diffuse grayish or brown pigment in B. campbelli; cream or pale yellow with a sparse peppering of brown in B. colombianus; whitish with a moderate suffusion of grey in B. myersi. Comparisons of meristic and hemipenial characters with all its congeners of toadheaded pitvipers are summarized in Table
Bothrocophias myrringae sp. nov. can be distinguished from Ecuadorian, Peruvian, and Bolivian populations of toadheaded pitvipers currently classified as B. microphthalmus by ventral surface of tail with uniformly bright reddish or orange-reddish speckles with black spots without a regular pattern (versus heavily marked proximally with black or dark brown pigment, medially mottled with pale to dark brown, and distally with pale diffuse or pale mottling with interspaces in Bolivian and Peruvian populations). Comparisons of meristic and hemipenial characters with the Ecuadorian, Peruvian, and Bolivian populations of toadheaded pitvipers the complex are summarized in Table
(Figs
(Fig.
(Fig.
Comparison of the head pholidosis among the holotypes of Bothrocophias microphthalmus, Bothrocophias tulitoi sp. nov. and Bothrocophias myrringae sp. nov. A, B, C Dorsal, lateral, and ventral view of the head of B. microphthalmus ANSP 11515. D, E, F Dorsal, lateral, and ventral view of the head of B. tulitoi sp. nov. INSZ 073. G, H, I Dorsal, lateral, and ventral view of the head of B. myrringae sp. nov. Scales: dark green = second/third supralabial scale. yellow = lacunolabial/prelacunal. Blue = internasal scales. Fuchsia = scale separating internasal scales.
(Fig.
Color in life of Bothrocophias myrringae sp. nov., and sexual dimorphism. A, B Lateral and dorsal view of the holotype INSZ 0268. C Male exhibiting melanic coloration on dorsal body and head surfaces, as well as conspicuous tricolored ocelli on the infralabial scales (paratype INSV-SR009). D Female exhibiting creamish-yellow to tan coloration on the dorsal body and head surfaces, without tricolored ocelli on the infralabial scales. Both C and D specimens are from vereda de Coasavistá, municipality of Fómeque, Cundinamarca, coordinates N 4.495001, W –73.852056. Pictures by Ronald A. Díaz-Flores.
(Table
(n=2, Fig.
The specific epithet myrringae is the Latin translation of the Spanish nickname “Mirringa,” which means “pinch” or something very small. The word “Mirringa” was popularized by Rafael Pombo (1833–1912), a Colombian poet and writer who wrote a popular fable titled “Mirringa Mirronga.” Given the popularity of the fable, as well as the homophonic similarity of “Mirringa” and the name “Myriam,” the nickname “Myrringa” began to be used as a term of endearment. The name of the new species is in honor of the educator Myriam Sierra Guerrero (mother of the first author). She was the philosophical and conceptual advisor of professor Tulio Manuel Angarita Serrano and contributed to the development of the modern Colombian education model that all schools within Colombia currently employ. Professor Sierra Guerrero also helped develop the theoretical framework for the implementation of institutional educational projects in Colombian public and private schools (see Angarita-Serrano 1990; Angarita-Serrano 1994; Angarita-Serrano and Chaves 1995; Angarita-Serran 1996; Angarita-Serrano 2000).
The known localities of Bothrocophias myrringae sp. nov. are from 1754 to 2761 m a.s.l. in both the central mountains and eastern slopes of the Cordillera Oriental of Colombia in the municipalities of La Calera, Choachí, Fómeque, and Guayabetal (Cundinamarca), and El Calvario and San Juanito (Meta, Fig.
A total of three snakebite events in the last decade might have potentially been caused by B. myrringae sp. nov., all of which were from the municipality of El Calvario (Meta). Each case was categorized as mild, moderate, and severe, respectively, and one fatality was reported. Local symptoms reported included oedema (100%), pain (100%), erythema (66%), and phlyctens (33%); systemic symptoms included respiratory failure (33%) and muscular weakness (33%). Symptoms such as paresthesia ecchymosis, bruising, sickness, vomit, vertigo, gingivorrhagia, hematuria, and altered vision were not reported.
Our phylogenetic trees returned Bothrocophias microphthalmus as non-monophyletic and support the hypothesis that it is a species complex as suggested by
Bothrocophias hyoprora and B. microphthalmus (sensu lato) are consistently recovered as monophyletic, and the genus Bothrocophias is consistently inferred to be monophyletic according to several molecular and morphological phylogenetic analyses (
The low genetic distances found between B. tulitoi sp. nov. and B. myrringae sp. nov., could be construed as evidence against the recognition of these lineages as distinctive taxa. However, low genetic divergence is to be expected among recently diverged lineages, which is common among Andean lineages (
Nevertheless, according to
Some meristic abnormalities were observed in captive-born neonate specimens of B. tulitoi sp. nov. (INSZ 130), such as canthorostral scales that were present on one side and absent on the other. Three neonate specimens (INSZ 128, 130, 136) possessed high interoculobial counts. The fact that such abnormalities were only observed on captive-born specimens, might suggest an effect of environmental changes during development due to the captive conditions (
Our data support the speculation of
The Cordillera Oriental is a younger bivergent contractional belt that reactivated a Mesozoic rift system in a combination of thin-skinned ramp-flat thrust systems and thick-skinned basement-involved structures (
The estimated diversification time of the genus Bothrocophias (11 Mya) coincides with the most intense peaks of Andean Mountain building following the late middle Miocene ~12 Ma (
An integrative taxonomic approach is still needed to delimit the several lineages within Bothrocophias microphthalmus (sensu lato). The distribution of the Ecuadorian populations is broad (from Pastaza to Zamora Chinchipe provinces), spanning various types of habitats and different Andean regions, and they exhibit pronounced morphological variation, as well as unusual arboreal behaviors not reported in any other population of B. cf. microphthalmus (
To determine the actual distribution of B. microphthalmus (sensu lato), we compared the type specimen (ANSP 11515, Fig.
Envenomation by B. tulitoi sp. nov., and B. myrringae sp. nov. appear to have effects on humans like other species in the genus and other bothropoid taxa (
Resolution of the taxonomy of the Bothrocophias microphthalmus species complex has significant implications for the public health of Andean countries that continually face the challenges of snakebite accidents. The first step in treating snakebites is the accurate identification of snakes causing envenomation. Following an accurate identification, medical practitioners can administer an appropriate treatment, including antivenom therapy regimen, as well as anticipate possible clinical complications. Accurate identification of the venomous snakes causing snakebite accidents also facilitates the detection, quantification, and characterization of envenoming events by epidemiological surveillance systems, which aids the development of preventive health strategies and epidemiological efforts to reduce the incidence of snakebite accidents. The new taxonomic insights provided by our study thus significantly contribute to achieving the World Health Organization’s goal of reducing the neglected disease of snakebite envenoming (
We thank Francisco Javier Ruiz, Carlos Antonio Castro, Juan José Torres-Ramirez, and Monica Sarmiento (INS staff) for assistance, help during lab procedures, and allowing us to examine specimens of Bothrocophias under their care. We are grateful to Prof. Mario Vargas Ramirez, for allowing the use of the molecular laboratory at the Genetics Institute, National University, Bogota. We thank Oscar Ramirez-Ruiz for making the line drawings in Fig.
Specimens examined or compared with literature descriptions:
Bothrocophias microphthalmus (sensu lato)
BOLIVIA (n=5,
ECUADOR (n=22): Morona Santiago: Cantón Morona: Parroquia Sinai. Locality: Banks of the Jurumbuno River, QCAZR13300, coordinates N 2.08617, W –78.1501. Cantón: Santiago. Parroquia: Patuca. Locality: Puchimi. In a creek near the camp, QCAZR15974, coordinates N 2.78075, W –78.15412. Cantón San Juan Bosco: Parroquia: San Carlos de Limón. Locality: Comunidad Shuar Kunkuk, foothills of the Cordillera del Cóndor, QCAZR16144-5, coordinates N 3.321027, W –78.2121; Parroquia: Pan de Azucar. Referencia: Sector Siete Iglesias-San Juan Bosco. Locality: upper part of the transverse ravine to the Pan de Azúcar River, QCAZR17021 coordinates N 3.13196, W –78.55739. — Pastaza: Cantón: Mera; Parroquia: Mera. Locality: Llanganates National Park, Comunidad Zarentza. Line October 9 of Llanganates National Park, 400m from the Zarentza community school, QCAZR13730–36, coordinates N 1.35935, W –78.05774. — Zamora Chichipe: Catón Nangaritza: Reserva Maycú, QCAZR12637, coordinates N 4.233086, W –78.61119. Cantón Zamora: Parroquia Zamora. Locality: Main trail, gate to Podocarpus National Park, Bombuscaro, QCAZR13858 coordinates N 4.114639, W –78.96702; Locality: Higuerones trail, sector of the Avecillas farm QCAZR13858 coordinates N 4.133359, W –78.98547. Cantón: Nangaritza. Parroquia: Zurmi. Locality: Maycu Nature Reserve, trail in terra-firme forest, surroundings of creek QCAZR15498 coordinates N 4.24706, W –78.65253; Locality: Trail parallel to the Nangaritza River and tributary streams; Locality: via Las Orquídeas-Nuevo Paraiso. Tepuy Lookout Trail QCAZR15500, coordinates N 4.25707, W –78.68095; Locality: trail ladera baja QCAZR15501 coordinates N 4.21895, W –78.62946. Parroquia: Nuevo Paraiso. Locality: Camp next to the Nangaritza River, near the tarabita. Trail from camp upstream QCAZR15770-72 coordinates N 4.45291, W –78.81508.
PERU (n=3): San Martin: Municipalities of Balsa Puerto and Moyabamba. Locality: unknown, pictures of the holotype provided by Dr. Ned Gilmore ANSP 11515 coordinates N 5.952166667, W –76.76213889 (approximate coordinates taken from midpoint between Puerto and Moyabamba municipalities). — Pasco: Oxapampa. Locality: unknown, pictures shared by Juan Timms from a single specimen housed in Oxapampa hospital, coordinates N 10.5775, W –75.40167 (see Supplement material). Pozuzo. Locality: unknown, picture shared by Juan Timms from a single specimen housed in Pozuzo hospital, coordinates N –10.23767, W –75.69016.
Bothrocophias tulitoi sp. nov.
COLOMBIA (n=2): Cundinamarca: Municipality of Gutiérrez. Locality: unknown, IAvH-R6879–80, coordinates N 4.25472, W –74.0025 (approximate to the town).
Table
Data type: .docx
Explanation note: Partitions and evolutionary models for the concatenated alignment obtained using Model Finder (MF) and bModelTest (bMT).
Table
Data type: .docx
Explanation note: Meristic and mensural (in mm) characters of holotype specimens of Bothrocophias tulitoi sp. nov., B. myrringae sp. nov., and B. microphthalmus.