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Fig. 5 in Morphological and molecular phylogenetic characterization of Sarcocystis kani sp. nov. and other novel, closely related Sarcocystis spp. infecting small mammals and colubrid snakes in Asia

Fig. 5. Maximum Likelihood (ML) phylogenetic reconstruction of the mitochondrial Cytochrome C oxidase Subunit I (cox1) gene tree of the Sarcocystis spp. under investigation. Where possible, sequences of the same species were used as shown in the 18S rRNA gene tree, including the outgroup. The newly sequenced Sarcocystis spp. are marked with black symbols. A total of 25 nucleotide sequences and 603 sites of the barcode area (all codon positions included) was analyzed with 1000 bootstrap replicates. Branch support from three replicate analyses is shown. The evolutionary history was inferred by using ML based on the HKY model. The tree with the highest log likelihood (– 6438.6551) is shown. A discrete Gamma distribution was used to model evolutionary rate differences among sites (four categories [+G, parameter = 0.9037]). The rate variation model allowed for some sites to be evolutionarily invariable ([+I], 17.4959% sites). All positions with less than 98% site coverage were eliminated. Branch lengths are measured in the number of substitutions per site. Note the long branch lengths of ruminant Sarcocytis spp. in comparison to other members of the Sarcocystidae as well as eimeriid coccidia from phylogenetically diverse hosts.

Fig. 2 in Morphological and molecular phylogenetic characterization of Sarcocystis kani sp. nov. and other novel, closely related Sarcocystis spp. infecting small mammals and colubrid snakes in Asia

Fig. 2. Ultrastructure of Sarcocystis sp.1 from the mangrove snake in abdominal musculature of a Sprague-Dawley rat; a) One μm-thin section through a resin-embedded, toluidine-stained sarcocyst showing densely-packed cystozoites (CZ) that were contained in septate compartments; the arrow indicates the cyst wall with small protrusions, which are clearly visible because host cell (HC) tissue is removed at this position; b) gross view of the cyst wall and cystozoites in a longitudinal section; the black arrow indicates a thin septum that separated the compartments filled with cystozoites; protrusions (PT) were broad, short, and irregularshaped, often with a reticulate base that rested on a thin layer of ground substance (GS); c) cross-section of sarcocyst, showing the primary cyst wall at higher magnification to consist of electron-dense, knob-like structures with intermittent invaginations; it appeared as if the primary wall was fenestrated (asterisk) allowing exchange of fine granular material (arrowheads) between the interior and exterior of the cyst; the exterior space between the protrusions was entirely filled with granular substance. d) metrocytes exclusively divided by endodyogeny as only cells with two developing zoites (asterisks) were observed; the white arrow points at deposits of highly electron-dense matter that could form larger clusters in the GS of the septae.

Fig. 1 in Morphological and molecular phylogenetic characterization of Sarcocystis kani sp. nov. and other novel, closely related Sarcocystis spp. infecting small mammals and colubrid snakes in Asia

Fig. 1. Light microscopic observations on the development of Sarcocystis sp.1 in striated musculature of Sprague-Dawley rats; a) typical sporocysts, here in fecal smear from Boiga dendrophila, that were used for infection of rats; sporocysts contained a granular residual body (asterisk) and four sporozoites (SP), which are all visible in the upper sporocyst; b) full-length micrograph of a typical (native) sarcocyst of Sarcocystis sp.1 in striated belly musculature, the arrows indicating folds of the cysts' body; c) high magnification of the cyst wall of a native sarcocyst, the arrows pointing at the apparently smooth wall; this is the same cyst as depicted in Fig. 2a; the inset shows freshly released, live cystozoites under phase-contrast light microscopy.

Fig. 4 in Morphological and molecular phylogenetic characterization of Sarcocystis kani sp. nov. and other novel, closely related Sarcocystis spp. infecting small mammals and colubrid snakes in Asia

Fig. 4. Predicted secondary structure of helix 38 in domain V7 of the 18S rRNA of Sarcocystis sp.1, Sarcocystis sp.2, S. attenuati, S. scandentiborneensis, and S. zuoi in comparison to S. clethrionomyelaphis (shaded inset). The 7-nt long motif 5′-AAUUCGU-3' (relative to all Apicomplexan taxa examined; nt in italic letters in shaded oval) mapped to a hairpin loop position of helix 38 and was characteristic for all species of the S. zuoi – complex; the motif was one nt (Cytosine) shorter in helix 38 of S. clethrionomyelaphis. Sequences were aligned to a secondary structure model of the Eukarya using SSU-ALIGN. The numbering of nucleotides (bars) and helices is based on the 1881 nt-long structural template. Position 1430 corresponds to position 1357 of the predicted secondary structure of Toxoplasma gondii (RH strain) as published by Gagnon et al. (1996).

Fig. 3 in Morphological and molecular phylogenetic characterization of Sarcocystis kani sp. nov. and other novel, closely related Sarcocystis spp. infecting small mammals and colubrid snakes in Asia

Fig. 3. Bayesian Inference (BI) of the 18S rRNA phylogeny of Sarcocystis spp. infecting colubrid snakes and small mammals in Asia; the S. zuoi – complex of species is highlighted by the shaded box. This complex excludes S. clethrionomyelaphis, which branches off basally. All new sequences, including a new isolate of S. zamani from Sumatra, are highlighted by black symbols. The known definitive and/or intermediate hosts associated with the selected sequences of the S. zuoi-group are indicated. Eimeriid species from phylogenetically diverse hosts served as outgroups. Bayesian posterior probabilities of three independent analyses (and alignments) are indicated behind each node, showing only one value if results of replicates were identical.

Fig. 6 in Morphological and molecular phylogenetic characterization of Sarcocystis kani sp. nov. and other novel, closely related Sarcocystis spp. infecting small mammals and colubrid snakes in Asia

Fig. 6. Mapping of (A) potential heme ligand binding sites and (B) amino acid variability among different lineages of tissue cyst-forming coccidia in the barcode area of the mitochondrial COX1 protein. A) Map of putative heme ligand binding sites (arrowheads) in a protein sequence alignment of selected taxa used in the phylogenetic tree of cox1. Identical/conservative aa positions are highlighted by light background, variable positions and gaps are shown against black background. Helix 1 (H1) is shown partial, starting at position 14 of the global barcode alignment (Pentinsaari et al., 2016); aa sequences of helices 2 (H2) and 6 (H6) are shown in full length, while putative heme binding sites of loop 3–4 (L3-4) were in its anterior part only. Domain boundaries and putative ligand binding sites were derived from COX1 of template organisms Saccharomyces cerevisiae and Bos taurus by sequence alignment against Toxoplasma gondii applying three-dimensional homology modelling of protein structure. The complete alignment of the six helices of the barcode area is shown in Supplementary Fig. S2. Note that aa numbering of the barcode area of the Apicomplexan sequence is different to the global alignment, because the former showed one additional aa and domain boundaries were slightly altered. The nucleotide sequence KC209732 of S. tenella (GenBank) is also registered in the barcode reference database BOLD (accession number JRPAA5858-15; http://boldsystems.org); the aa barcode position 14 (Glycine) shown here corresponds to position 16 of the aa translation of GenBank record KC209732. B) 'Heat map' of aa changes (darker shades of green = more changes, number of changes indicated) in the barcode area among chemically/structurally different aa groups in different lineages of tissue cyst-forming coccidia relative to the COX1 protein sequence of T. gondii. Because helix 1 was truncated, records of aa changes in this area are incomplete.

Fig 10 in A Species-Level Phylogeny of Extant Snakes with Description of a New Colubrid Subfamily and Genus

Fig 10. Phylogenetic tree of Serpentes continued. A) Colubrinae continued. B) Colubrinae continued. doi:10.1371/journal.pone.0161070.g010

Fig 5 in A Species-Level Phylogeny of Extant Snakes with Description of a New Colubrid Subfamily and Genus

Fig 5. Phylogenetic tree of Serpentes continued. A) Homalopsidae, Psammophiinae, Buhoma procterae, Prosymninae, Pseudaspidinae, Atractaspidinae, and Aparallactinae. Bi) Oxyrhabdium leporinum and Lamprophiinae. Bii) Ditypophis sp. + Micrelaps bicoloratus and Pseudoxyrhophiinae. doi:10.1371/journal.pone.0161070.g005

Fig 8 in A Species-Level Phylogeny of Extant Snakes with Description of a New Colubrid Subfamily and Genus

Fig 8. Phylogenetic tree of Serpentes continued. A) Dipsadinae continued. B) Dipsadinae continued. doi:10.1371/journal.pone.0161070.g008

Fig 2 in A Species-Level Phylogeny of Extant Snakes with Description of a New Colubrid Subfamily and Genus

Fig 2. Species-level phylogeny on final dataset of 1652 snake species. Maximum-likelihood phylogenetic estimate based on 10 concatenated genes. Node values represent SHL support values. Seven outgroup taxa are not shown. Colors of clades indicate their position in the overall tree, shown at left. Newly sequenced taxa are highlighted in bold. Skeleton of the species tree is displayed on the left with displayed subfamilies/families highlighted. Letters denoted by i and ii represent parts of the tree where external branches do not connect to the part of the tree immediately preceding it. A) Anomalepididae, Epictinae, Leptotyphlopinae, Gerrhopilidae, Xenotyphlopidae, and Typhlopinae. B) Asiatyphlopinae I, Afrotyphlopinae; Madatyphlopinae, and Asiatyphlopinae II. doi:10.1371/journal.pone.0161070.g002

opennotspecifiedApr 2020View details →

FIGURE 2 in A multilocus molecular perspective on the systematics of the poorly known Northeast Indian colubrid snakes Blythia reticulata (Blyth, 1854), B. hmuifang Vogel, Lalremsanga & Vanlalhrima, 2017, and Hebius xenura (Wall, 1907)

FIGURE 2. Phylogenetic relationships of Blythia reticulata, B. hmuifang, Trachischium spp. and Herpetoreas (Hebius) xenura with other natricine snakes inferred from our 139-leaf dataset. A. ML phylogeny. Bootstrap support is shown at each internal branch. B. BI phylogeny. Posterior probability support values are shown at each internal branch. Samples in orange are those for which data were newly generated in this study. Scale bar represents mean nucleotide substitutions per site.

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FIGURE 1 in A multilocus molecular perspective on the systematics of the poorly known Northeast Indian colubrid snakes Blythia reticulata (Blyth, 1854), B. hmuifang Vogel, Lalremsanga & Vanlalhrima, 2017, and Hebius xenura (Wall, 1907)

FIGURE 1. ML phylogeny showing relationships of newly sampled Blythia, Trachischium, and Herpetoreas (Hebius) xenura with other major lineages of snakes inferred from our 91-leaf dataset. Samples in orange are those for which data were newly generated in this study. Bootstrap support is shown at each internal branch. Scale bar represents the number of nucleotide substitutions per site.

opennotspecifiedAug 2008View details →

FIGURE 5–6 in A new species of the colubrid snake genus Atractus (Reptilia: Serpentes) from the central Amazon of Brazil

FIGURE 5–6. Sulcate (5) and asulcate (6) sides of the left hemipenis of Atractus alatagratiae sp. nov. (MNRJ 7888).

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FIGURE 4 in A new species of the colubrid snake genus Atractus (Reptilia: Serpentes) from the central Amazon of Brazil

FIGURE 4. Dorsal view of the holotype of Atractus altagratiae sp. nov. (MNRJ 7888). SVL 240 mm, CL 35 mm.

opennotspecifiedAug 2008View details →

FIGURES 1–3 in A new species of the colubrid snake genus Atractus (Reptilia: Serpentes) from the central Amazon of Brazil

FIGURES 1–3. Dorsal (1) and lateral (2) views of head, and lateral view of midbody (3) of the holotype of Atractus altagratiae sp. nov. (MNRJ 7888). Scale = 5 mm.

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FIGURE 3 in Three's Company: discovery of a third syntype of Stegonotus lividus, a species of colubrid snake from Pulau Semau, Lesser Sunda Islands, Indonesia, with comments on an unpublished 19 Century manuscript by the naturalist Salomon Müller

FIGURE 3. Unpublished colored drawing by Pieter van Oort (1804–1834) showing the specimen of "Coluber lividus" used by Salomon Müller for producing his description (Fig. 2). The small inset shows a drawing of the head scales.

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FIGURE 1 in Three's Company: discovery of a third syntype of Stegonotus lividus, a species of colubrid snake from Pulau Semau, Lesser Sunda Islands, Indonesia, with comments on an unpublished 19 Century manuscript by the naturalist Salomon Müller

FIGURE 1. An 1841 map created by Salomon Müller (1804–1864), showing approximately the western half of Timor Island in the Lesser Sunda Archipelago. The larger island in the lower left corner of the map is Rote. Semau, the collecting locality of the type series of Stegonotus lividus, is the small offshore island shown just to the left of the southwestern end of the larger island, indicated by the red circle. The chart's title reads: "Kaart van het westelijke gedeelte van Timor bevattende de bezittingen der Nederlanders en van hunne bondgenooten aldaar" [Map of the western part of Timor containing the belongings of the Dutch and their allies there]. The inset at right shows the geological composition of the region (a "geognostic sketch"), which reveals how diverse the geology on tiny Semau ("Samauw") is.

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Fig 6 in A Species-Level Phylogeny of Extant Snakes with Description of a New Colubrid Subfamily and Genus

Fig 6. Phylogenetic tree of Serpentes continued. A) Buhoma depressiceps and Elapidae. B) Elapidae continued. doi:10.1371/journal. pone.0161070. g006

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Fig 4 in A Species-Level Phylogeny of Extant Snakes with Description of a New Colubrid Subfamily and Genus

Fig 4. Phylogenetic tree of Serpentes continued. Ai) Viperinae. Aii) Azemiopinae and Crotalinae. B) Crotalinae continued. doi: 10.1371/journal.pone.0161070.g004

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Fig 3 in A Species-Level Phylogeny of Extant Snakes with Description of a New Colubrid Subfamily and Genus

Fig 3. Phylogenetic tree of Serpentes continued. A) Aniliidae, Tropidophiidae, Calabariidae, Candoiidae, Sanziniidae, Charininae, Ungaliophiinae, Erycidae, and Boidae. Bi) Cylindrophiidae + Anomochilidae, Uropeltidae, Xenopeltidae, Loxocemidae, and Pythonidae. Bii) Bolyeridae, Xenophidiidae, Acrochordidae, Xenodermatidae, and Pareatidae.