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The phylogenetic position of the unusual diplostomoidean genus Schwartzitrema Vigueras, 1941 (Digenea, Diplostomoidea) and description of 2 new species from Neotropics

Published online by Cambridge University Press:  14 November 2025

T.J. Achatz Open the ORCID record for T.J. Achatz [Opens in a new window] ,
L.L. Ho ,
M. Montes Open the ORCID record for M. Montes [Opens in a new window] ,
J. Barneche ,
A. Fecchio Open the ORCID record for A. Fecchio [Opens in a new window]  and
V.V. Tkach Open the ORCID record for V.V. Tkach [Opens in a new window]
T.J. Achatz
Affiliation:
Department of Natural Sciences, Middle Georgia State University , Macon, GA, USA
L.L. Ho
Affiliation:
Department of Natural Sciences, Middle Georgia State University , Macon, GA, USA
M. Montes
Affiliation:
Centro de Estudios Parasitológicos y Vectores (CEPAVE), Consejo Nacional de Investigaciones Científicas y Técnicas, Universidad Nacional de La Plata (CCT, CONICET-UNLP), La Plata, Buenos Aires, Argentina
J. Barneche
Affiliation:
Centro de Estudios Parasitológicos y Vectores (CEPAVE), Consejo Nacional de Investigaciones Científicas y Técnicas, Universidad Nacional de La Plata (CCT, CONICET-UNLP), La Plata, Buenos Aires, Argentina
A. Fecchio
Affiliation:
Department of Ornithology, The Academy of Natural Sciences of Drexel University, Philadelphia, PA, USA
V.V. Tkach*
Affiliation:
Department of Biology, University of North Dakota , Grand Forks, ND, USA
*
Corresponding author: V.V. Tkach; Email: vasyl.tkach@und.edu
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Abstract

Schwartzitrema Vigueras, 1941, is a small, rarely reported genus of diplostomoideans that parasitize avian definitive hosts, predominantly anhingas, on several continents. No DNA sequences were previously available for any Schwartzitrema spp. In the present study, we describe adults of 2 new Schwartzitrema spp. from anhingas in Brazil. We also provide description of metacercaria of one of the new species from Argentina. Schwartzitrema macrotesticulata n. sp. lacks a clear separation between its testes, appearing almost as a contiguous, irregularly shaped, lobular mass. A highly characteristic feature of Schwartzitrema macroholdfastus n. sp. is a strongly enlarged lobe of the holdfast organ. These features, along with distribution of vitellarium and nature of the preputial folds surrounding the genital cones, separate the new species from their congeners. We generated partial sequences of the large ribosomal subunit (28S) rDNA and cytochrome c oxidase subunit I (cox1) mtDNA genes from adult forms of both species and cox1 of metacercarial stages. The 28S sequences were used for phylogenetic inference. The pairwise sequence divergence between the 2 species is 4.0–4.6% in cox1 gene. Intraspecific variability in the same cox1 region did not exceed 0.8% despite the long geographic distance between collecting sites in Brazil and Argentina.

Keywords

DiplostomidaeSchwartzitremanew speciesAnhingaCrenicichla

Information

Type
Research Paper
Information
Journal of Helminthology , Volume 99 , 2025 , e120
Creative Commons
Creative Common License - CCCreative Common License - BY
This is an Open Access article, distributed under the terms of the Creative Commons Attribution licence (http://creativecommons.org/licenses/by/4.0), which permits unrestricted re-use, distribution and reproduction, provided the original article is properly cited.
Copyright
© The Author(s), 2025. Published by Cambridge University Press

Introduction

Schwartzitrema Vigueras, 1941 is a small genus of diplostomoidean digeneans parasitic as adults in various piscivorous birds, primarily anhingas, in the Nearctic, Neotropics, Australasia, Indomalaya, and Afrotropics (Vigueras, Reference Vigueras1940; Chandler, Reference Chandler1951; Dubois and Fain, Reference Dubois and Fain1956; Gupta, Reference Gupta1964; Dubois, Reference Dubois1968). Members of this genus share a bipartite body with cup-shaped prosoma as well as strongly developed pseudosuckers with auricular expansions in the prosomal cup (Dubois, Reference Dubois1968; Niewiadomska, Reference Niewiadomska, Gibson, Jones and Bray2002). Vigueras (Reference Vigueras1940) erected the genus for Schwartziella schwartzi Vigueras, 1940, collected from anhinga (Anhinga anhinga (L.)) in Cuba. Later, the genus name was changed to Schwartzitrema as the original genus name was preoccupied (Chandler, Reference Chandler1951). Prior to this study, the genus included 7 species: Schwartzitrema schwartzi (Vigueras, 1940) from A. anhinga in Cuba; Schwartzitrema seamsteri Chandler, 1951 from frigate bird (Fregata magnificens Mathews) in Texas, USA; Schwartzitrema anhingae Gupta, 1964 and Schwartzitrema pandubi (Pande, 1939) from oriental darter (Anhinga melanogaster Pennant) in India; Schwartzitrema novaehollandiae Dubois et Pearson, 1967 from Australian darter (Anhinga novaehollandiae Gould) in Australia; Schwartzitrema perezi Gupta, 1962 from Asian openbill (Anastomus oscitans Boddaert) in India; Schwartzitrema truonis (Dubois et Fain, 1956) from reed cormorant (Microcarbo africanus Gmelin) in Rwanda (see Dubois, Reference Dubois1982). No DNA sequences are available from any member of this genus.

In the present study, we collected 2 new Schwartzitrema spp. from anhingas in Brazil as well as metacercariae from fishes in Argentina. We provide morphological descriptions of adult stages of both species as well as the metacercaria of one species. Partial sequences of the large ribosomal subunit (28S) rDNA were used to explore the phylogenetic affinities of the genus among other diplostomoideans. Fragments of the cytochrome c oxidase subunit I (cox1) mtDNA were used for differentiation among species.

Materials and methods

Adult Schwartzitrema spp. specimens were collected from intestines of A. anhinga in the Municipality of Poconé, Mato Grosso State, Brazil. Metacercariae were collected from the meninges of the brains in pike cichlids, Crenicichla iguassuensis Haseman and Crenicichla vittata (Heckel) in Argentina (Table 1). Live digeneans were heat-killed and preserved in 80% ethanol. The adult specimens used for morphological study were stained with an aqueous alum carmine and permanently mounted following Lutz et al. (Reference Lutz, Tkach, Weckstein and Webster2017); metacercariae were relaxed in hot water, fixed in 10% formalin, stained with hydrochloric carmine, dehydrated through a graded series of ethanol solutions, cleared in eugenol, and mounted in Canada balsam for observation under light microscopy. Measurements and drawings of adults were done using an Olympus© BX53 microscope (Olympus America, Center Valley, Pennsylvania, USA) equipped with DIC optics and a drawing tube; drawings of the metacercariae were made based on a series of photographs taken using an Olympus© BX51 microscope equipped with digital camera. All measurements are in micrometers. Upon receiving positive reviews of the manuscript, type specimens of the new species described herein (holotypes and paratypes) were mailed to be deposited in the Museu Paraense Emílio Goeldi (Belém, Pará State, Brazil). Unfortunately, the shipment was lost. Despite every effort to locate the shipment for over 6 months, it was never recovered. We did not have additional remaining specimens of either species to designate new types. At the same time, we have detailed illustrations of both species and photographs of partial paratypes depicting characteristic differentiating features. Besides, we have DNA sequence data of a conserved gene (28S) and a variable gene (cox1). Therefore, we opted to preserve the descriptions of the 2 species of Schwartzitrema and deposited photo vouchers in the collection of the H. W. Manter Laboratory (HWML), University of Nebraska, Lincoln, Nebraska, USA. Metacercariae are deposited in the Helminthological Collection of the Museo de La Plata, Argentina (MLP-He).

Table 1.

Hosts, geographical origin, GenBank accession numbers, and museum accession numbers of Schwartzitrema spp. collected or described in the present study

Abbreviations: MPEG, Collection of the Emílio Goeldi Museum, Belém, Pará, Brazil; MLP-HE, Helminthological Collection of the Museo de La Plata, Argentina.

As discussed by Gudla et al. (Reference Gudla, Orlofske, Brant, Tkach, Dubay, Holtz and Achatz2023), previous descriptions of former strigeids have been primarily based on laterally positioned specimens. While we relaxed and mounted some specimens dorso-ventrally, many of our specimens were mounted laterally to allow comparison with prior descriptions. We use the terminology of Gudla et al. (Reference Gudla, Orlofske, Brant, Tkach, Dubay, Holtz and Achatz2023) to refer to lateral specimen measurements: thickness for the distance between ventral and dorsal margins of the prosoma and opisthosoma; width (dorso-ventral) and width (lateral) for organs that are not round in cross-section; and diameter for organs that are round in cross-section.

DNA extraction, amplification, and sequencing were carried out as described by Achatz et al. (Reference Achatz, Burkman, Fecchio, Pulis and Tkach2023). Only the partial 28S sequences were used for phylogenetic analysis. The newly obtained sequences were aligned with previously published sequences of diplostomoideans using ClustalW as implemented in MEGA7 (Kumar et al., Reference Kumar, Stecher and Tamura2016). For our analysis we chose sequences of matching length that represented each genus of the Diplostomidae Poirier, 1886 (per Achatz and Tkach, Reference Achatz and Tkach2025); only 2 members of the former Proterodiplostomidae Dubois, 1936 were included as this diplostomid lineage was proven to be monophyletic. We chose Suchocyathocotyle crocodili (Yamaguti, 1954) as the outgroup based on the topology presented by Achatz et al. (Reference Achatz, Pulis, Junker, Binh, Snyder and Tkach2019). MEGA7 identified the general time-reversible model with estimates of invariant sites and gamma distributed among-site variation (GTR + I + G) as the best-fitting nucleotide substitution model for the 28S alignment. The phylogenetic analysis was conducted using Bayesian Inference (BI) as implemented in MrBayes Ver. 3.2.6 software (Ronquist and Huelsenbeck, Reference Ronquist and Huelsenbeck2003). The BI analysis was performed using MrBayes software as follows: Markov chain Monte Carlo (MCMC) chains were run for 3,000,000 generations, log-likelihood scores were plotted, and only the final 75% of trees were used to produce the consensus tree. This number of generations was considered sufficient because the SD stabilized below 0.01 by the end of the run. MEGA7 was used to calculate pairwise distances.

Results

Molecular analysis

The 28S alignment was 1,088 bp long upon trimming to the length of the shortest sequence; 36 sites were excluded due to ambiguous homology. The phylogenetic analysis (Fig. 1) resulted in a tree with topology similar to that demonstrated in several prior studies (Blasco-Costa and Locke, Reference Blasco-Costa and Locke2017; Hernández-Mena et al., Reference Hernández-Mena, García-Varela and Pérez-Ponce de León2017; Locke et al., Reference Locke, Van Dam, Caffara, Pinto, López-Hernández and Blanar2018; Achatz et al., Reference Achatz, Pulis, Junker, Binh, Snyder and Tkach2019, Reference Achatz, Chermak, Martens, Pulis, Fecchio, Bell, Greiman, Cromwell, Brant, Kent and Tkach2021, Reference Achatz, Martens, Kostadinova, Pulis, Orlofske, Bell, Fecchio, Oyarzún-Ruiz, Syrota and Tkach2022a, Reference Achatz, Martens, Kudlai, Junker, Boe and Tkachb, Reference Achatz, Burkman, Fecchio, Pulis and Tkach2023, Reference Achatz, Chacko, Prasadan and Tkach2024, 2025; Queiroz et al., Reference Queiroz, López-Hernández, Locke, Pinto and Anjos2020). Schwartzitrema spp. were placed within a weakly supported clade with several other taxa (Fig. 1). Within this weakly supported clade, Icthyocotylurus erraticus Rudolphi, 1809 + Cotylurus spp. formed a separate subclade (100% supported) from a subclade of Cardiocephaloides spp. + Schwartzitrema spp. (99% supported). Within the latter, Cardiocephaloides spp. formed a 98% supported clade separate from both Schwartzitrema spp. (100% supported clade; Fig. 1).

Figure 1.

Phylogenetic interrelationships among 55 diplostomoids based on Bayesian Inference (BI) analysis of partial 28S rDNA gene sequences. The BI posterior probability values lower than 80% are not shown. Schwartzitrema spp. are within the shaded boxes. The new sequences generated in this study are in bold. The scale bar indicates the number of substitutions per site. GenBank accession numbers are provided after names of taxa.

Pairwise sequence comparisons have demonstrated 0.1% divergence between the 2 species from Brazil in the sequenced region of the 28S gene. The interspecific divergence between these species in cox1 was 4.0–4.6% while intraspecific variability in cox1 was 0.0–0.7% in Schwartzitrema macroholdfastus n. sp. and 0.8% in Schwartzitrema macrotesticulata n. sp. Interestingly, some specimens of S. macroholdfastus n. sp. did not show any variability between Brazil and Argentina.

Descriptions of new taxa

Schwartzitrema macrotesticulata n. sp. (Figs 2–4)

Figure 2.

Schwartzitrema macrotesticulata n. sp. holotype, ventral view (a) vitellarium shown; (b) vitellarium omitted. Abbreviations: F, prepucial fold; P, pseudosuckers.

Figure 3.

Schwartzitrema macrotesticulata n. sp. (a) Paratype, lateral view, vitellarium shown; (b) Paratype, lateral view, vitellarium omitted; (c) posterior end of paratype, lateral view; (d) posterior end of holotype, ventral view. Abbreviations: F, prepucial fold; P, pseudosuckers.

Figure 4.

Schwartzitrema macrotesticulata n. sp. metacercaria (a) entire specimen, ventral view with excretory granules omitted; (b) holdfast organ and ventral sucker, ventral view with excretory granules omitted; (c) anterior part of prosoma with excretory granules shown. Abbreviations: P, pseudosuckers; G, gonad primordium.

Taxonomic summary

Type host: Anhinga anhinga L. (Suliformes: Anhingidae).

Second intermediate host: Crenicichla iguassuensis Haseman (Cichliformes: Cichlidae).

Site of infection in definitive host: Small intestine.

Site of infection in second intermediate host: Meninges of brain.

Type locality: Pantanal, Fazenda Retiro Novo, Municipality of Poconé, Mato Grosso State, Brazil (16°21′53″S 56°17′31″W).

Other locality: Parque Nacional Iguazu, Misiones State, Argentina, upstream from the waterfalls. (25°40′34″S 54°27′08″W).

Photo paratype deposited: one photograph of paratype HWML 218112; labeled ex. A. anhinga, small intestine, Fazenda Retiro Novo, Municipality of Poconé, Mato Grosso State, Brazil, 8 June 2017, coll. A. Fecchio. One damaged voucher specimen deposited: HWML 218114, labeled identically to the photo paratype.

Metacerariae deposited: Voucher series consists of 8 specimens deposited in the Helminthological Collection of the Museo de La Plata. Vouchers: MLP-He 8202.

Representative DNA sequences of adult digeneans: GenBank PV454642–PV454644 (28S), PV453983–PV453984 (cox1).

Representative DNA sequence of metacercariae: GenBank PV453985 (cox1).

Etymology: The species name refers to its large testes that appear to be a single structure.

ZooBank registration: urn:lsid:zoobank.org:act:789A8965-EA9D-4DFF-AA82-4A000EAE94A5.

Description of adult. Based on 8 specimens; measurements of dorso-ventral holotype given in text; measurements of entire series given in Table 2. Body 1,340 long, with distinct separation of prosoma and opisthosoma. Prosoma cup-shaped, 608 × 514. Opisthosoma elongated, 732 × 442. Prosoma:opisthosoma length ratio 0.8. Forebody 26% of body length. Tegument unarmed. Oral sucker nearly terminal, rounded, 107 × 112. Ventral sucker in middle of prosoma, well-developed, rounded, 129 × 150. Oral sucker:ventral sucker width ratio 0.7. Pseudosuckers well-developed, elongated, positioned in prosoma cup posterior to pharynx. Holdfast organ consisting of proximal glandular part (likely incorporating proteolytic gland) near prosoma-opisthosoma junction and distally extending bilobed part, 367 long. Prepharynx short. Pharynx oval, 51 × 54. Esophagus short; cecal bifurcation in anterior half of prosoma. Ceca slender, posterior ends not observed.

Table 2.

Measurements of adult Schwartzitrema spp. described. Range followed by mean in parentheses

a Measurements for the combined testes length and width due to the lack of distinct separation between testes.

Testes not distinctly separated from each other, irregularly shaped, with 2–3 large lobes. Seminal vesicle weakly developed, ventral, near posterior margin of testicular zone; joins metraterm dorsally to form hermaphroditic duct at base of genital cone. Genital cone protrusible, positioned in genital atrium, with prepucial fold surrounding entire circumference, dorsal part more pronounced. Genital pore terminal, strongly muscular.

Ovary 103 × 129, immediately pretesticular, anterior margin of ovary positioned at 47% of opisthosoma length posterior to prosoma-opisthosoma junction, transversely oval. Ootype, Mehlis’ gland and vitelline reservoir near mid-length of testes, difficult to observe. Uterus extends anteriorly beyond level of ovary and posteriorly to genital cone, ventral to gonads and seminal vesicle. Zero to 9 eggs (3 in holotype), 106–131 × 48–81. Vitellarium located in prosoma and opisthosoma, distributed between level of ventral sucker (or slightly anterior) to near the level of genital cone, ventral to gonads.

Excretory vesicle not well observed. Excretory pore terminal.

Description of metacercariae. Based on 9 dorso-ventrally mounted metacercariae; range is followed by mean in parentheses. Body 1,300–1,846 (1,599) long, with distinct separation of prosoma and opisthosoma. Prosoma dorso-ventrally flattened, 1,079–1,484 (1,274) × 372–876 (601). Opisthosoma cylindrical, much shorter than prosoma, 221–362 (323) × 212–287 (245). Prosoma:opisthosoma length ratio 3.3–4.9 (4.0). Forebody 53–58% (55%) of body length. Tegument unarmed. Oral sucker nearly terminal 64–102 (84) × 74–114 (89). Ventral sucker 95–120 (105) × 121–146 (131). Oral sucker:ventral sucker width ratio 0.6–0.9 (0.7). Pseudosuckers well-developed, elongated, positioned in anterior quarter of prosoma. Holdfast organ near prosoma-opisthosoma junction, sucker-like with distinct bilobed part, 279–373 (327) long. Prepharynx not observed. Pharynx oval, 45–57 (52) × 27–45 (36). Esophagus long; cecal bifurcation posterior to level of pseudosuckers; posterior ends of ceca not observed.

Only one early testicular primordium observed. Ovary primordium near midlength of opisthosoma, weakly developed, subspherical, entire, 48–71 (61) × 35–68 (54). Excretory vesicle not well-observed. Excretory granules medium sized, numerous, occupy most of prosoma in some specimens. Excretory pore terminal.

Remarks

Schwartzitrema macrotesticulata n. sp. belongs to the genus based on the bipartite body with cup-shaped prosoma, elongated, well-developed, pseudosuckers, and short hermaphroditic duct that opens at the tip of a genital cone. The new species can be differentiated from S. anhingae, S. pandubi, S. schwartzi, S. seamsteri, and S. truonis based on the anterior extent of the vitellarium. The vitellarium of the new species reaches to the level of, or slightly anterior to, the ventral sucker, while the vitellarium of other species does not extend anteriorly to this level, often not even extending into the prosoma.

The new species can be readily distinguished from S. perezi based on the presence of a preputial or prepuce-like fold that surrounds the genital cone; this structure is absent in S. perezi. Schwartzitrema novaehollandiae was described with a pseudoprepucial fold that may be absent in some specimens (Dubois, Reference Dubois1968). The hermaphroditic duct of S. macrotesticulata n. sp. begins at the base of the genital cone, while the male and female efferent ducts do not join until near the tip of the genital cone of S. novaehollandiae. The new species is from South America, while S. perezi and S. novaehollandiae were described from India and Australia, respectively.

The separation between testes in the new species could not be detected even using high-quality optics. Although we cannot confidently state at this point that this species has only one testis, the contiguous nature of testes represents yet another feature distinguishing S. macrotesticulata n. sp. from other congeners, all of which have clearly separated testes.

Schwartzitrema macrotesticulata n. sp. showed significant level of pairwise sequence divergence (4.0–4.6%) from S. macroholdfastus n. sp. in cox1 gene.

Schwartzitrema macroholdfastus n. sp. Achatz, Ho, Fecchio et Tkach (Fig. 5)

Figure 5.

Schwartzitrema macroholdfastus n. sp. (a) holotype, ventral view, vitellarium shown; (b) holotype, ventral view, vitellarium omitted; (c) paratype, lateral view, vitellarium shown; (d) posterior end of paratype, dorsal view, vitellarium omitted.

Taxonomic summary

Type host: Anhinga anhinga L. (Suliformes: Anhingidae).

Second intermediate host: Crenicichla vittata (Heckel) (Cichliformes: Cichlidae).

Site of infection in definitive host: Small intestine.

Site of infection in second intermediate host: Meninges of brain.

Type locality: Pantanal, Fazenda Retiro Novo, Municipality of Poconé, Mato Grosso State, Brazil (16°21′53″S 56°17′31″W).

Other locality: Laguna Ibera, Colonia Pellegrini, Corrientes, Argentina (28°32′12″S 57°10′17″W).

Photo paratype deposited: one photograph of paratype HWML 218113; labeled ex. A. anhinga, small intestine, Fazenda Retiro Novo, Municipality of Poconé, Mato Grosso State, Brazil, 8 June 2017, coll. A. Fecchio.

Representative DNA sequences of adult digeneans: GenBank PV454645 (28S), PV453986–PV453987 (cox1).

Representative DNA sequence of metacercariae: GenBank PV453988 (cox1).

Etymology: The species name refers to its exceptionally large holdfast organ.

ZooBank registration: urn:lsid:zoobank.org:act:38BBC648-F040-4CF1-92D6-85FD313EDDD0.

Description of adult. Based on 11 specimens; measurements of dorso-ventrally mounted holotype given in text; measurements of entire series given in Table 2. Body 2,196 long, with distinct separation of prosoma and opisthosoma. Prosoma cup-shaped, 813 × 447. Opisthosoma elongated, 1,383 × 503. Prosoma:opisthosoma length ratio 0.6. Forebody 25% of body length. Tegument unarmed. Oral sucker nearly terminal, rounded, 83 × 93. Ventral sucker well-developed, rounded, 173 × 142, just post-equatorial in prosoma. Oral sucker:ventral sucker width ratio 0.7. Pseudosuckers well-developed, elongated, positioned inside cup-shaped prosoma, posterior to pharynx. Holdfast organ consisting of proximal glandular part (likely incorporating proteolytic gland) near prosoma-opisthosoma junction and distally extending bilobed part, one lobe substantially longer than other, 632 long. Prepharynx short. Pharynx oval, 43 × 50. Esophagus short; cecal bifurcation in anterior half of prosoma. Ceca slender, extend posteriorly to level of seminal vesicle.

Testes tandem, anterior testis asymmetrical; posterior testis bilobed. Seminal vesicle ventral to posterior testes; joins metraterm dorsally to form hermaphroditic duct at base of genital cone. Genital cone protrusible, positioned in genital atrium, with dorsal prepucial fold that surrounds about 50–75% of genital cone circumference. Genital pore terminal, strongly muscular.

Ovary 160 × 225, immediately pretesticular, transversely oval, anterior margin of ovary positioned at 38% of opisthosoma length posterior to prosoma-opisthosoma junction. Ootype, Mehlis’ gland, and vitelline reservoir intertesticular. Uterus extends anteriorly beyond level of ovary and posteriorly to genital cone, ventral to gonads and seminal vesicle. Eggs 91–135 × 48–80; egg number varies from 1 to 19 (8 in holotype). Vitellarium located in prosoma and opisthosoma, distributed between level of ventral sucker (or slightly anterior) to near the level of genital cone, ventral to gonads.

Excretory vesicle not well observed. Excretory pore terminal.

Remarks

Schwartzitrema macroholdfastus n. sp. belongs to the genus based on the bipartite body with cup-shaped prosoma, elongated, well-developed, pseudosuckers, and short hermaphroditic duct that opens at the tip of a genital cone. This new species can be differentiated from all congeners based on its massive holdfast organ with one of the lobes extending anteriorly beyond oral sucker. The holdfast organ of all other congeners, except for S. anhingae and S. pandubi, is much smaller.

The vitellarium of S. macroholdfastus n. sp. reaches the level of, or slightly anterior to, the ventral sucker, while the vitellarium of S. anhingae, S. pandubi, S. schwartzi, S. seamsteri, and S. truonis does not reach anteriorly the level of ventral sucker and often does not extend into the prosoma.

The prepucial fold of S. macroholdfastus n. sp. surrounds about 50–75% of genital cone circumference while the prepucial fold of S. macrotesticulata n. sp. surrounds the entire genital cone; Schwartzitrema perezi lacks a prepucial fold. The hermaphroditic duct of S. macroholdfastus n. sp. begins at the base of the genital cone, while in S. novaehollandiae the male and female efferent ducts do not join until near the tip of the genital cone. The new species is from South America, while S. perezi and S. novaehollandiae were described from India and Australia, respectively.

Schwartzitrema macroholdfastus n. sp. showed significant level of pairwise sequence divergence (4.0-4.6%) from S. macrotesticulata n. sp. in cox1 gene.

Discussion

After the present study, Schwartzitrema includes 9 species (Vigueras, Reference Vigueras1940; Dubois and Fain, Reference Dubois and Fain1956; Gupta, Reference Gupta1964; Dubois and Pearson, Reference Dubois and Pearson1967; Dubois, Reference Dubois1982). This is the first study to sequence any member of this genus, and first to report Schwartzitrema spp. in continental South America. Schwartzitrema schwartzi is the congener that also parasitizes A. anhinga, but has only been found in Cuba (Vigueras, Reference Vigueras1940; Dubois, Reference Dubois1968). Schwartzitrema novaehollandiae, S. anhingae, and S. pandubi parasitize other anhinga species in Australia and India (Dubois, Reference Dubois1968).

An additional member of the genus was described by Khan et al. (Reference Khan, Hazing RR and Nisa2009) based on specimens from cattle egret Ardea coromanda (Boddaert) in Pakistan. However, the description was very superficial, the illustrations were poor, and the characteristic morphological features of Schwartzitrema spp. (i.e., the strongly developed pseudosuckers) were not included in the description or illustration. There are several other similar-looking diplostomids, such as Cotylurus Szidat, 1928, and Apatemon Szidat, 1928, which more closely morphologically conform with the described specimens (e.g., see illustrations in Dubois, Reference Dubois1968, and Niewiadomska, Reference Niewiadomska, Gibson, Jones and Bray2002). Therefore, we consider Schwartzitrema bilqeesae Khan, Das, Ghazi et Nisa, 2009, a species inquirenda.

Prior to this study, the intermediate hosts of Schwartzitrema spp. were unknown. We collected metacercariae of both new species from the meninges of the brains of 2 Crenicichla spp. in Argentina. It is probable that other congeners also parasitize fish as second intermediate hosts, and likely infect the meninges of brains. While we did not study pathogenicity of these digeneans, it is extremely likely that Schwartzitrema spp. are fish pathogens similar to other diplostomoideans that utilize fish 2nd intermediate host, such as Bolbophorus Dubois, 1935, Diplostomum von Nordmann, 1832, Posthodiplostomum Dubois, 1936, Crassiphiala Van Haitsma, 1925, Uvulifer Yamaguti, 1934 (Terhune et al., Reference Terhune, Wise, Avery, Khoo and Goodwin2003; Ondračkova et al., Reference Ondračkova, Bartošová, Valová, Jurajda and Gelnar2004; Blasco-Costa & Locke, Reference Blasco-Costa and Locke2017; Charo-Karisa et al., Reference Charo-Karisa, Ali, Marijani, Ibrahim, Trinh, Chadag and Benzie2021; Capasso & Gutiérrez, Reference Capasso and Gutiérrez2023; Tkach and Achatz, Reference Tkach and Achatz2024). Unfortunately, the first intermediate host of Schwartzitrema spp. remains unknown. Future studies should seek to identify the first intermediate host for this genus as well as sequence Schwartzitrema spp. from other biogeographic realms, such as the Afrotropics and Australasia.

Acknowledgments

We are grateful to Dr. João B. Pinho (Universidade Federal de Mato Grosso, Cuiabá, Mato Grosso, Brazil) for his invaluable help in organizing and collecting the specimens used in this work and obtaining collecting permits, and to Jeffrey A. Bell (University of North Dakota) for his assistance in collecting specimens. We extend our gratitude to the Administración de Parques Nacionales for granting permission to work within Iguazú National Park. We also thank the Centro de Estudios Parasitológicos y Vectores (CEPAVE) for providing the infrastructure where part of this work was conducted.

Financial support

This work was supported by the National Science Foundation (grant DEB-1120734) as well as the University System of Georgia Stem Initiative IV (Middle Georgia State University) and the Center for Middle Georgia Studies for TJA.

Competing interests

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

References

Achatz, TJ, Burkman, CA, Fecchio, A, Pulis, EE and Tkach, VV (2023) Description of Anhingatrema n. gen. (Digenea: Diplostomidae) with two new species from New World anhingas (Aves: Anhingidae). Acta Parasitologica 68, 159–171.10.1007/s11686-022-00643-0CrossRefGoogle ScholarPubMed
Achatz, TJ, Chacko, S, Prasadan, PK and Tkach, VV (2024) Proterodiplostomid no longer: Molecular phylogeny reveals the true position of Proalarioides (Digenea: Diplostomoidea). Parasitology International 102, 102917. https://doi.org/10.1016/j.parint.2024.102917CrossRefGoogle ScholarPubMed
Achatz, TJ, Pulis, EE, Junker, K, Binh, TT, Snyder, SD and Tkach, VV (2019) Molecular phylogeny of the Cyathocotylidae (Digenea, Diplostomoidea) necessitates systematic changes and reveals a history of host and environment switches. Zoologica Scripta 48, 545–556.10.1111/zsc.12360CrossRefGoogle ScholarPubMed
Achatz, TJ, Chermak, TP, Martens, JR, Pulis, EE, Fecchio, A, Bell, JA, Greiman, SE, Cromwell, KA, Brant, SV, Kent, ML and Tkach, VV (2021) Unravelling the diversity of the Crassiphialine (Digenea: Diplostomidae) with molecular phylogeny and descriptions of five new species. Current Research in Parasitology and Vector-Borne Diseases 1, 100051. https://doi.org/10.1016/j.crpvbd.2021.100051CrossRefGoogle ScholarPubMed
Achatz, TJ, Martens, JR, Kostadinova, A, Pulis, EE, Orlofske, SA, Bell, JA, Fecchio, A, Oyarzún-Ruiz, P, Syrota, YY and Tkach, VV (2022a) Molecular phylogeny of Diplostomum, Tylodelphys, Austrodiplostomum and Paralaria (Digenea: Diplostomidae) necessitates systematic changes and reveals history of evolutionary host switching events. International Journal for Parasitology 52, 47–63. https://doi.org/10.1016/j.ijpara.2021.06.002CrossRefGoogle Scholar
Achatz, TJ, Martens, JR, Kudlai, O, Junker, K, Boe, NW and Tkach, VV (2022b) A new genus of diplostomids (Digenea: Diplostomoidea) from Nile crocodiles (Crocodylus niloticus) in South Africa with an updated key to diplostomid genera. Journal of Parasitology 108, 453–466.10.1645/22-23CrossRefGoogle Scholar
Achatz, TJ and Tkach, VV (2025) Recent advances and current state of knowledge of phylogenetics and systematics of the Diplostomoidea with a proposal of a new classification system and a key to genera. Journal of Helminthology 99, e87.10.1017/S0022149X25100473CrossRefGoogle Scholar
Blasco-Costa, I and Locke, SA (2017) Life history, systematics and evolution of the Diplostomoidea Poirier, 1886: progress, promises and challenges emerging from molecular studies. Advances in Parasitology 98, 167–225.Google ScholarPubMed
Capasso, S and Gutiérrez, JS (2023) Diplostomum spathaceum. Trends in Parastiology 39, 969–970.10.1016/j.pt.2023.06.002CrossRefGoogle ScholarPubMed
Chandler, AC (1951) Trematodes from the Man-O-War Bird, Fregata magnificens rothschildi, on the Texas coast, with the description of a new species, Schwartzitrema seamsteri. Texas Journal of Science 3, 186–189Google Scholar
Charo-Karisa, H., Ali, SE, Marijani, E, Ibrahim, NA, Trinh, TQ, Chadag, MV and Benzie, JAH (2021) Genetic parameters for black spot disease (diplopstomiasis) caused by Uvulifer sp. infection in Nile tilapia (Oreochromis niloticus L.). Aquaculture 532, 736039.10.1016/j.aquaculture.2020.736039CrossRefGoogle Scholar
Dubois, G (1968) Synopsis des Strigeidae et des Diplostomatidae (Trematoda). Mémoires de la Société Neuchâteloise des Sciences Naturelles 10, 1–258.Google Scholar
Dubois, G (1982) Répertoire des synonymes récents de genres et d’especes de la superfamily des Strigeoidea Railliet, 1919 (Trematoda). Bulletin de la Société Neuchâteloise des Sciences Naturelles 105, 163–183.Google Scholar
Dubois, G and Fain, A (1956) Contribution à l’étude des Strigeida du Congo belge. I. Bulletin de la Société de la Neuchâteloise des Sciences Naturelles 79, 17–38.Google Scholar
Dubois, G and Pearson, JC (1967) Quelques Strigeida (Trematoda) d’Australie. II. Bulletin de la Société des Sciences Naturelles de Neuchâtel 88, 77–99.Google Scholar
Gudla, B, Orlofske, SA, Brant, SV, Tkach, VV, Dubay, S, Holtz, L and Achatz, TJ (2023) Taxonomic re-assessment and morphological redescription of Nematostrigea serpens annulata (Digenea: Strigeidae) from osprey in North America. Journal of Parasitology 109, 550–558.10.1645/23-41CrossRefGoogle Scholar
Gupta, R (1964) Schwartzitrema anhingi sp. nov. from the Indian Darter, Anhinga melanogaster Pennant, with a remark on the taxonomic position of the genus Schwartzitrema (Vigueras, 1940) Vigueras, 1941 (Trematoda: Strigeidae). Revista De Biología Tropical 12, 75–79.Google Scholar
Hernández-Mena, DI, García-Varela, M and Pérez-Ponce de León, G (2017) Filling the gaps in the classification of the Digenea Carus, 1863: systematic position of the Proterodiplostomidae Dubois, 1936 within the superfamily Diplostomoidea Poirier, 1886, inferred from nuclear and mitochondrial DNA sequences. Systematic parasitology 94, 833–848.10.1007/s11230-017-9745-1CrossRefGoogle ScholarPubMed
Khan, A, Das Hazing RR, SN and Nisa, N (2009) A new species of trematode genus Schwartzitrema Perez Vigueras, 1941 (Strigeidae) from a bird (cattle egret, Bubulcus ibis L.) in Sindh, Pakistan. International Journal of Biology and Biotechnology 6, 1–3.Google Scholar
Kumar, S, Stecher, G and Tamura, K (2016) MEGA7: Molecular Evolutionary Genetics Analysis Version 7.0 for Bigger Datasets. Molecular Biology and Evolution 33, 1870–1874.10.1093/molbev/msw054CrossRefGoogle ScholarPubMed
Locke, SA, Van Dam, AR, Caffara, M, Pinto, HA, López-Hernández, D and Blanar, CA (2018) Validity of the Diplostomoidea and Diplostomida (Digenea, Platyhelminthes) upheld in phylogenomic analysis. International Journal for Parasitology 48, 1043–1059.10.1016/j.ijpara.2018.07.001CrossRefGoogle ScholarPubMed
Lutz, HL, Tkach, VV and Weckstein, JD (2017) Methods for specimen-based studies of avian symbionts. In Webster, M (ed), The Role of Collections in Ornithology: The Extended Specimen. Studies in Avian Biology, pp. 127–183. Boca Raton, FL: CRC Press, Boca Raton.Google Scholar
Niewiadomska, K (2002) Family Strigeidae Railliet, 1919. In Gibson, DI, Jones, A, and Bray, RA (eds), Keys to the Trematoda , Volume 1, pp. 231–241. Wallingford, CAB International and The Natural History Museum.10.1079/9780851995472.0231CrossRefGoogle Scholar
Ondračkova, M., Bartošová, Š., Valová, Z, Jurajda, P. and Gelnar, M. (2004) Occurrence of black-spot disease caused by metacercariae of Posthodiplostomum cuticola among juvenile fishes in water bodies in the Morava River basin. Acta Parasitologica 49, 222–227.Google Scholar
Vigueras, IP (1940) Notas sobre algunas especies nuevas de trematodes y sobre otras poco conocidas. Revista de la Universidad de Habana 5, 217–242.Google Scholar
Queiroz, M, López-Hernández, D, Locke, S, Pinto, H and Anjos, L (2020) Metacercariae of Heterodiplostomum lanceolatum (Trematoda: Proterodiplostomidae) found in Leptodactylus podicipinus (Anura: Leptodactylidae) from Brazil: A morphological, molecular and ecological study. Journal of Helminthology 94, e66. https://doi.org/10.1017/S0022149X19000646CrossRefGoogle Scholar
Ronquist, F and Huelsenbeck, JP (2003) MRBAYES 3: Bayesian phylogenetic inference under mixed models. Bioinformatics 19, 1572–1574.10.1093/bioinformatics/btg180CrossRefGoogle ScholarPubMed
Terhune, JS, Wise, DJ, Avery, JL, Khoo, LH and Goodwin, AE (2003) Infestations of the trematode Bolbophorus sp. in channel catfish. Southern Regional Aquaculture Center 1801, 1–4.Google Scholar
Tkach, VV and Achatz, TJ (2024) Crassiphiala bulboglossa. Trends in Parasitology 41, 162–163.10.1016/j.pt.2024.11.013CrossRefGoogle Scholar
Figure 0

Table 1. Hosts, geographical origin, GenBank accession numbers, and museum accession numbers of Schwartzitrema spp. collected or described in the present study

Figure 1

Figure 1. Phylogenetic interrelationships among 55 diplostomoids based on Bayesian Inference (BI) analysis of partial 28S rDNA gene sequences. The BI posterior probability values lower than 80% are not shown. Schwartzitrema spp. are within the shaded boxes. The new sequences generated in this study are in bold. The scale bar indicates the number of substitutions per site. GenBank accession numbers are provided after names of taxa.

Figure 2

Figure 2. Schwartzitrema macrotesticulata n. sp. holotype, ventral view (a) vitellarium shown; (b) vitellarium omitted. Abbreviations: F, prepucial fold; P, pseudosuckers.

Figure 3

Figure 3. Schwartzitrema macrotesticulata n. sp. (a) Paratype, lateral view, vitellarium shown; (b) Paratype, lateral view, vitellarium omitted; (c) posterior end of paratype, lateral view; (d) posterior end of holotype, ventral view. Abbreviations: F, prepucial fold; P, pseudosuckers.

Figure 4

Figure 4. Schwartzitrema macrotesticulata n. sp. metacercaria (a) entire specimen, ventral view with excretory granules omitted; (b) holdfast organ and ventral sucker, ventral view with excretory granules omitted; (c) anterior part of prosoma with excretory granules shown. Abbreviations: P, pseudosuckers; G, gonad primordium.

Figure 5

Table 2. Measurements of adult Schwartzitrema spp. described. Range followed by mean in parentheses

Figure 6

Figure 5. Schwartzitrema macroholdfastus n. sp. (a) holotype, ventral view, vitellarium shown; (b) holotype, ventral view, vitellarium omitted; (c) paratype, lateral view, vitellarium shown; (d) posterior end of paratype, dorsal view, vitellarium omitted.