Unravelling the anthelmintic bioactives from Jasminum grandiflorum L. subsp. Floribundum adopting in vitro biological assessment.

ETHNOPHARMACOLOGICAL RELEVANCE: Jasminum grandiflorum L. is a medicinal plant widely used in the traditional system of Medicine as an anthelmintic in ringworm infections, for treating ulcers, stomatitis, skin diseases, and wounds. AIM OF THE STUDY: The emergence of resistance by different parasites to currently used chemicals has been reported. There are increasing needs for more effective and safer parasiticides. Therefore, the current study was designed to investigate the methanolic extract of the aerial parts of J. grandiflorum subsp. Floribundum (JGTE) to confirm its traditional uses as anthelmintic through a bioassay-guided fractionation and isolation of the active components with anthelmintic activity. MATERIALS AND METHODS: The JGTE was partitioned into dichloromethane (DCM-F) and n-butanol (BuOH-F) fractions. The JGTE, fractions, and the isolated compounds were tested in vitro for their anthelmintic activity using two nematodes; one larval stage of cestode and one arthropod. Four major compounds were isolated from the most active fraction (BuOH-F) including two flavonoids and two secoirridoid glycosides, identified as kaempferol-3-O-neohesperoside (1), rutin (2), oleuropein (3), and ligstroside (4). RESULTS: Among the isolated compounds from most active fraction (BuOH-F), rutin (2) displayed the highest anthelmintic activity in a dose-dependent activity with IC50 of 41.04 µg/mL against H. muscae adult worm, followed by ligstroside (4) with IC50 of 50.56 µg/mL. CONCLUSIONS: These findings could advocate the traditional use of J. grandiflorum L. and provide further insight into the anthelmintic activity of flavonoids.

Morphological and Molecular Evidence for a New Rhabdochonid Species (Nematoda: Rhabdochonidae) Parasitizing Profundulus oaxacae (Profundulidae) in the State of Oaxaca, Mexico.

The genus Rhabdochona Railliet, 1916 is one of the most speciose group of nematodes, parasites of freshwater fishes, with approximately 103 species described worldwide. Twenty-two species have been recorded in the Americas, 14 of them in Mexico. In this paper we describe a new species of Rhabdochona on the basis of light and scanning electron microscopy and molecular tools. Rhabdochona adentata n. sp. was recovered from the gallbladder of the freshwater Oaxaca killifish, Profundulus oaxacae (Meek, 1902) (Profundulidae) captured in the Río Grande, State of Oaxaca, Mexico. Rhabdochona adentata n. sp. differs markedly from its congeners by possessing an unusual prostom lacking anterior teeth, small simple deirids, and the location of the excretory pore at the level of the union of the muscular and glandular esophagus. Sequences of cytochrome c oxidase subunit I (COI) from mitochondrial deoxyribonucleic acid and the D2 + D3 domains of the large ribosomal subunit (28S) were obtained from 3 specimens and were analyzed using maximum likelihood and Bayesian inference analyses. Phylogenetic analyses using COI and 28S genes recovered 1 new lineage of Rhabdochona. The new species is described on the basis of a detailed morphological study. This parasite represents the first species of Rhabdochona without prostomal teeth and with a different site of infection, the gallbladder.

Atrophic Proventriculitis Associated with Gravid Females of Tetrameres sp. (Nematoda: Tetrameridae) in the Magellanic Penguin (Spheniscus magellanicus) on the Coast of Brazil.

The present study offers the first description of proventriculitis associated with the presence of gravid female nematodes of the genus Tetrameres (Nematoda: Tetrameridae) in 3 juvenile Magellanic penguins (Spheniscus magellanicus) found dead on the coast of Brazil. This study broadens knowledge on parasites associated with these hosts and the real impact of this association.

A New Species of Ascarophis (Nematoda: Cystidicolidae) Parasitizing Clinocottus analis (Pisces: Cottidae) from Baja California, Mexico.

A new species of nematode, Ascarophis morronei n. sp. (Cystidicolidae), is described from the stomach wall of the woolly sculpin Clinocottus analis (Cottidae) collected in the rocky intertidal from northwestern Baja California, Mexico. Collected nematodes were studied using both light and scanning electron microscopy. Sequence fragments for 18S rDNA molecular markers were obtained from the new nematode species, in order to test its position within the family Cystidicolidae under a phylogenetic context. Main characters distinguishing this new species include the reduced labia and the morphology of the eggs, distances of nerve ring and excretory pore from the anterior end, and left spicule of males. The new species described here is the second for the genus Ascarophis reported as adult in the Southern California Bight, and the first one recorded for the fish genus Clinocottus.

Redescription and genetic characterization of Cystidicoloides vaucheri, including first description of male and current status on the phylogeny of Cystidicolidae (Nematoda: Habronematoidea).

Cystidicoloides vaucheri collected in the stomach of the redtail catfish Phractocephalus hemioliopterus from River Acre, State of Acre, Brazil is redescribed, including the first description of males and the first genetic characterization based upon 18S and 28S genes of the rRNA. Newly collected females were biometrically smaller than those reported in the original description, but similar morphology shared by the two samples revealed that they belong to the same species. Scanning electron micrographs showed the accurate structure of the cephalic region, described here in detail. Furthermore, the morphology of males completed the specific diagnosis, strengthening the validity of the species. The three other congeners differ from C. vaucheri mainly as follows: in C. dlouhyi the area rugosa is absent, the cephalic structures in C. fischeri are completely distinct, and in both species the spicules have membranous outgrowths, absent in C. vaucheri. Despite the dubious generic assignment of C. izecksohni, it differs from C. vaucheri in several biometrical and morphological features. Because of data availability, only sequences of the 18S were used for phylogenetic reconstructions. Results showed that the genus Ascarophis and the families Cystidicolidae and Physalopteridae are not monophyletic. Cystidicoloides vaucheri formed an independent branch clustering with representatives of Cystidicolidae, confirming its validity. The inclusion of Salmonema and Spinitectus within Cystidicolidae should be reviewed, since they formed an assemblage with species from Rhabdochonidae. In fact, current classification of some taxa belonging to Habronematoidea, Physalopteroidea and Thelazioidea need to be re-evaluated, mainly based on molecular data from different genes.

Gongylonema Parasites of Rodents: A Key to Species and New Data on Gongylonema neoplasticum.

Specimens of the genus Gongylonema were collected from the gastric mucosa of rodents of Rattus rattus Linnaeus, 1758, and Rattus norvegicus Berkenhout, 1769, collected in urban areas in Belém, Pará, in the eastern Brazilian Amazon. The helminths were processed for analysis using light microscopy and scanning electron microscopy (SEM) techniques and presented taxonomic characteristics of the species Gongylonema neoplasticum. The SEM analyses revealed the presence of 2 developed buccal plates (1 dorsal, 1 ventral), also called interlabia, with a prominent and bifurcated ventral plaque. The occurrence of the bifurcated ventral interlabium had not yet been identified by any other author from G. neoplasticum. As a result of our extensive research on published data on Gongylonema spp., we propose a taxonomic key for species of this genus that parasitize rodents. This is the first record of G. neoplasticum in urban areas of the Brazilian Amazon.

First record of Gongylonema nepalensis in domestic and wild ruminants in Europe.

The gullet worm, Gongylonema pulchrum, is cosmopolitan in distribution, infecting a variety of mammals including domestic and wild ruminants. Gongylonema nepalensis recently collected from the esophageal epithelium of water buffaloes in Nepal was separated from G. pulchrum based on its distinctly shorter left spicule relative to body length and unique nucleotide sequences of the ribosomal RNA gene (rDNA) and the mitochondrial cytochrome c oxidase subunit 1 gene (cox-1). During meat inspections at four abattoirs on Sardinia Island, Italy, 25 Gongylonema worms were collected from one each individual of cattle (n=8), sheep (n=7), goats (n=4), and mouflon (Ovis aries musimon; n=6), and characterized morphologically and genetically. Intriguingly, all of the collected worms from these ruminants were G. nepalensis, exhibiting comparable body lengths to G. pulchrum in cattle from other regions but with significantly shorter left spicules like G. nepalensis (less than 20.9% of the entire body length in contrast to 21.8-65.6%, the reported proportion of G. pulchrum). Furthermore, the rDNA nucleotide sequences of these worms from different ruminant species on Sardinia Island were almost identical to each other and to Nepalese G. nepalensis isolates. With the exception of one worm from a sheep (displaying a single nucleotide substitution), the 369-bp cox-1 nucleotide sequences of all the Sardinian G. nepalensis isolates from the different host sources were absolutely identical, forming a clade with Nepalese G. nepalensis isolates and not G. pulchrum isolates. The present study reveals that G. nepalensis is not a local parasite in the Indian subcontinent (Nepal), but instead has a certain geographical distribution in Europe and takes several ruminant species as a definitive host.

Further study on Physaloptera clausa Rudolphi, 1819 (Spirurida: Physalopteridae) from the Amur hedgehog Erinaceus amurensis Schrenk (Eulipotyphla: Erinaceidae).

In the present study, light and scanning electron microscopy (SEM) were used to further study the detailed morphology of Physaloptera clausa Rudolphi, 1819, based on the material collected from the Amur hedgehog E. amurensis Schrenk in China. The results revealed a few previously unreported morphological features and some morphological and morphometric variability between our specimens and the previous studies. The present supplementary morphological characters and morphometric data could help us to recognize this species more accurately.

SCANNING ELECTRON MICROSCOPIC CHANGES IN HABRONEMA MUSCAE AFTER IN VITRO EXPOSURE TO PLANT EXTRACT (VERBESINA ALTERNIFOLIA).

Scanning electron microscopy (SEM) was used to study the morphological alteration occurring in Habronema muscae adult female worms after in vitro exposure to different doses of Verbesina alternifolia oil extract. The half maximal lethal concentration (LC50) was reached 400 ppm after 24hrs, while LC100 was reached 600 ppm after 48hrs. Irreversible degenerative changes were recorded such as shrinking, detachment and distortion of the cuticle, cephalic and distal region. The cuticular surface had a wrinkled, corrugated appearance with longitudinal ridges and transverse thick folds. The lips and papillae were deformed and aggregated over each other. There is a direct relation between the level of the recorded degenerative changes and the increase in the dose and exposure time. In the same time no degen'erative changes were recorded in the control worm exposed to PBS till the end of the exposure period.

Nematode parasites of four species of Carangoides (Osteichthyes: Carangidae) in New Caledonian waters, with a description of Philometra dispar n. sp. (Philometridae).

Parasitological examination of marine perciform fishes belonging to four species of Carangoides, i.e. C. chrysophrys, C. dinema, C. fulvoguttatus and C. hedlandensis (Carangidae), from off New Caledonia revealed the presence of nematodes. The identification of carangids was confirmed by barcoding of the COI gene. The eight nematode species found were: Capillariidae gen. sp. (females), Cucullanus bulbosus (Lane, 1916) (male and females), Hysterothylacium sp. third-stage larvae, Raphidascaris (Ichthyascaris) sp. (female and larvae), Terranova sp. third-stage larvae, Philometra dispar n. sp. (male), Camallanus carangis Olsen, 1954 (females) and Johnstonmawsonia sp. (female). The new species P. dispar from the abdominal cavity of C. dinema is mainly characterised by the body length (5.14 mm), the lengths of markedly unequal spicules (163 and 96 µm) and gubernaculum (102 µm long) provided with a dorsal protuberance and a small, reflexed dorsal barb on its posterior portion. The finding of C. bulbosus represents the first record of this parasite a century after its discovery; the first study of this species by scanning electron microscopy (SEM) enabled detailed redescription. The finding of Johnstonmawsonia sp. in C. fulvoguttatus is the first record of a rhabdochonid nematode from a host belonging to the Carangidae family. Johnstonmawsonia africana Moravec & Puylaert, 1970 and J. campanae Puylaert, 1973 are transferred to Prosungulonema Roytman, 1963 as P. africanum (Moravec & Puylaert, 1970) comb. n. and P. campanae (Puylaert, 1973) n. comb.

Light and scanning electron microscopical examination of the third stage larva of Physocephalus dromedarii (Nematoda: Spirocercidae)--an abomasal nematode of the one humped camel (Camelus dromedarius).

The life cycle of Physocephalus dromedarii was studied under experimental conditions. Larvae obtained from naturally infected Scarabaeus cristatus and Aphodius sp. (Coleoptera: Scarabaeidae) were measured and examined in light and scanning electron microscopy and used to infect a dromedary as final host as well as chicken, mice and a toad as possible paratenic hosts. Larvae with the same morphology and similar measurements were found in naturally infected reptiles (Trapelus flavimaculatus, Eryx jayakari, Cerastes gasperettii). Body length of examined larvae varied between 1450 and 1700 µm. Dorsal, ventral and lateral lips, peg-like papillae and amphideal pits are located on the cephalic cone. There are two asymmetrical cervical deirids, long simple lateral wings and a knob-like posterior end covered with minute spines. In the camel, patency is reached within 12 weeks after infection while larvae in paratenic hosts migrate into the wall of the alimentary tract and become dormant.

A new species of Rhabdochona Railliet, 1916 (Nematoda: Rhabdochonidae) from cyprinid fishes in the Western Ghats Region, India.

A new nematode species, Rhabdochona (Globochona) puntii n. sp. (Rhabdochonidae), is described based on specimens collected from the intestine of the pool barb Puntius sophore (Hamilton) and Neolissochilus hexastichus (McClelland) (both Cyprinidae) from the Gadhena River, the Western Ghats, Maharashtra State, India. The nematode was also found in Wallago attu (Bloch & Schneider) which probably acts as postcyclic host. Rhabdochona (Globochona) puntii n. sp. differs markedly from its congeners in the body size, the number and distribution of caudal papillae, in the presence of an unpaired papilla-like structure on the anterior cloacal lip, and in having unusual shape and structure of the terminal crown of mucrons. This is the seventh species of the subgenus Globochona Moravec, 1972 reported from freshwater Indian fishes.

Physaloptera bainae n. sp. (Nematoda: Physalopteridae) parasitic in Salvator merianae (Squamata: Teiidae), with a key to Physaloptera species parasitizing reptiles from Brazil.

Physaloptera bainae n. sp. (Nematoda: Physalopteridae) from stomach of the large "tegú" lizard Salvator merianae (Squamata: Teiidae), collected in an ecological park that is part of the Atlantic Rainforest biome in Juiz de Fora, Minas Gerais, Brazil, is described. The new species which has females with didelphic uterus, is the only species of the genus exhibiting external teeth in the form of 4 spines that are organized in a cross-shaped pattern at the anterior apical end, with 2 minute adcloacal papillae on the anterior part of cloacal aperture in males and a large cuticular expansion at the anterior end of females. Moreover, P. bainae n. sp. can be differentiated from the other congeners by the combination of other features, e.g., number (23) and pattern of caudal papillae, spicule size (left 554-600; right 589-617) and vulval position (on the first third of body), and because the specimens are particularly large and robust. Physaloptera monodens is considered a junior synonym of P. obtusissima. In addition, a key to Physaloptera species parasitizing reptiles from Brazil is provided.

Habronema muscae (Nematoda: Habronematidae) larvae: developmental stages, migration route and morphological changes in Musca domestica (Diptera: Muscidae).

The present paper describes the morphological modifications occurring during the larval development of Habronema muscae (Nematoda: Habronematidae) in Musca domestica (Diptera: Muscidae), along with the reactions caused by parasitism and the migration route of the nematodes inside the flies. Houseflies were reared on faeces of a H. muscae-infected horse, then dissected and processed by histology. The experimental part of the study was performed in 1996 in the Parasitological Experimental Station W.O. Neitz, Federal Rural University of Rio de Janeiro, Seropédica, Rio de Janeiro State, Brazil. Three different larval stages of H. muscae were recovered, measured and described. The encapsulation of larval nematodes was found in the third larval instar (L3) of M. domestica and cryptocephalic pupa. The mature capsules were observed in dipteran L3, pupae and mainly adults. In 1day-old or more M. domestica adults an active rupturing of capsules by H. muscae L3 and the migration to the head through the circulatory system and insect hemocoel were observed. Infective H. muscae L3s remained exclusively in the head of adult 5days-old or more M. domestica.

New biogeographical and morphological information on Physaloptera ngoci Le-Van-Hoa, 1961 (Nematoda: Physalopteridae) in South-east Asian rodents.

During a study of the helminth fauna of 1,643 rodents trapped along the Mekong River (Thailand, Lao People's Democratic Republic and Cambodia) in 2008-2011, the spirurid nematode Physaloptera ngoci Le-Van-Hoa, 1961 was recovered with an overall prevalence of 2.8%. Based on the original description, it was identified in nine of 23 different Murinae host species and is here reported for the first time from these three countries. A scanning electron microscopy study provides additional morphological data.

A new species of Spirura Blanchard, 1849 (Nematoda: Spiruridae) parasite of Heliosciurus gambianus and Xerus erythropus (Rodentia: Sciuridae) in Senegal.

A new species of Spirura is described from the stomach of Heliosciurus gambianus and Xerus erythropus (Sciuridae). Considering the number of preanal papillae of males, Babero (1973 ) and Giannetto and Canestri Trotti (1995) proposed the subdivision of the genus into 2 groups; those with 4 pairs of preanal papillae (25 species) and with more than 4 pairs of preanal papillae (4 species). Spirura mounporti n. sp. belongs to the second, with 5 pairs of preanal papillae, and differs from Spirura infundibuliformis (McLeod, 1933) Anderson et al., 1993 , Spirura zapi ( Erickson, 1938 ) Chabaud et al., 1965 , Spirura leiperi Gupta and Trivedi, 1985, and Spirura michiganensis Sandground, 1935 in the number of pairs of pre-cloacal papillae. The new species further differs from other species of the genus in having 21 caudal papillae, in the ratio of spicules:body length, and in its morpho-anatomical characters.

Ultrastructural characterization of male and female Physaloptera rara (Spirurida: Physalopteridae): feline stomach worms.

Physaloptera rara (Spirurida: Physalopteridae) has been found in dogs, coyotes, raccoons, wolves, foxes, cats, and bobcats in North America. The parasites' developmental cycles involve insects, including beetles, cockroaches, and crickets, as intermediate hosts. The nematodes firmly attach to the wall of the stomach and duodenum, where they feed on the mucosa and suck blood. Frequent movement of these nematodes results in erosions and ulcers in the gastrointestinal tract. The present study reports the morphological features of adult P. rara using scanning electron microscopy. Adult worms were removed from the stomach of an infected domestic cat. Male and female worms measured 25-29 and 27-41 mm, respectively. The worms were stout and the cuticle was reflected over the lips to form a large cephalic collarette with fine transverse striations. The worms possessed two large, simple triangular lateral pseudolabia, each armed with one external tooth, three internal teeth, two submedian cephalic papillae, an amphid, and three porous-like circumscribed regions. The internal margins of the lips had a pair of cuticular folds. At the anterior end of both male and female worms, an excretory pore was located on the ventral side and a pair of lateral ciliated cervical papillae was seen. The vulva was anterior to the middle of the body of female worms. The tail ends of the female worms were stumpy, with two large phasmids near their extremities. The males' tails bore large lateral alae. Ventral ornamentation, in male worms, was composed of three different cuticular patterns; coblestone-like formations, longitudinal cuticular ridges, and rows of bead-like structures. The spicules were unequal and dissimilar; the right spicule had a thick end and the left spicule had a sharp tip. At the posterior end of the males, four pairs of stalked precloacal papillae, three pairs of postcloacal papillae, and two phasmids were present. Three and four sessile papillae were seen directly anterior and posterior to the cloaca, respectively. The middle papilla of the three sessile papillae, directly anterior to cloaca was different in shape and size.

A distinct genetic population of Gongylonema pulchrum from water buffaloes in Nepal.

Whole-length esophagi of 111 Murrah cross water buffaloes (Bubalus bubalis) were collected in the Kathmandu and Chitwan districts of Nepal from December 2009 to February 2010. Gullet worms showing a typical epithelium-dwelling character were detected in 13 of 53 (24.5%) buffaloes in Kathmandu and in 5 of 58 (8.6%) buffaloes in Chitwan. The worms' morphology and measurements were identical to those of Gongylonema pulchrum Molin, 1857, except for the length of the left spicules relative to the body length. Scanning electron microscopy did not detect any further morphological differences regarding the collected specimen from Nepal compared with G. pulchrum . The ribosomal RNA gene (rDNA), including internal transcribed spacer (ITS) 1 and 2, and a partial region of the cytochrome c oxidase subunit I (COI) of mitochondrial DNA of the worms were characterized and compared with those of G. pulchrum collected from cattle, deer, wild boars, and monkeys in Japan and from cattle in Iran. The 18S, 5.8S, and 28S rDNA nucleotide sequences of the buffalo-collected worms had 99.8% (1,779/1,782), 100% (158/158), and 98.3-98.8% (3,494-3,507/3,551) identities, respectively, with those of G. pulchrum from the other host mammals. The ITS regions exhibited higher variations between the buffalo-collected worms and G. pulchrum from the other host mammals (85-88% identity for ITS1 and 56-80% identity for ITS2). The COI also showed lower identities (89.2-90.2%), although only a single amino acid substitution was noted compared with the majority of G. pulchrum samples collected in Japan. Based on these molecular genetic characters in the rDNA and COI mitochondrial DNA, together with a shorter left spicule length relative to body length, the gullet worms isolated from buffaloes in Nepal might belong to a distinct local or buffalo-preferring population of G. pulchrum, although its geographical distribution on the continent and host specificity remain to be clarified.

Rhabdochona (Rhabdochona) hypsibarbi n. sp. (Nematoda: Rhabdochonidae) from the freshwater cyprinid fish Hypsibarbus wetmorei (Smith) in northeast Thailand.

A new nematode species, Rhabdochona (Rhabdochona) hypsibarbi n. sp. (Rhabdochonidae), is described from the intestine of the freshwater cyprinid fish Hypsibarbus wetmorei (Smith) in the Mekong River, Nakhon Phanom Province, northeast Thailand. It is mainly characterized by medium-sized, bifurcate deirids, the presence of 14 anterior prostomal teeth and distinct basal teeth, length ratio of the muscular and glandular portions of esophagus (1:6-9), length of the left spicule (669-774 µm), absence of a dorsal barb on the right spicule, length ratio of spicules (1:4.9-6.0), arrangement of genital papillae, and smooth eggs without filaments or swellings. In addition, specifically unidentified fourth-stage larvae of Rhabdochona (Rhabdochona) sp., morphologically similar to R. hypsibarbi, were recorded from the red-tailed tinfoil Barbonymus altus (Günther) (Cyprinidae) in the Mekong River, Nakhon Phanom Province, northeast Thailand. Rhabdochona hypsibarbi is the fourth nominal species of Rhabdochona Railliet, 1916 reported from fishes in Thailand and the second species of the nominotypical subgenus found in this country.

A new species of Neoascarophis (Nematoda: Cystidicolidae) parasitic in Macrourus carinatus (Macrouridae) from Argentinean waters.

Nematodes of the cystidicolid Neoascarophis Machida, 1976 , are all parasites of macrourid fishes, making up at present 5 species. Several other unidentified species have also been reported in several fish species from the northern and southern Atlantic Ocean, including 1 from Macrourus carinatus (Günther) (Macrouridae) in the southwest Atlantic Ocean. During a parasitological survey carried out on samples of M. carinatus from Patagonian waters, nematodes referable to Neascarophis were found in ulcers in the gastric mucosa. These nematodes Neascarophis sphaerocaudata n. sp. closely resemble N. macrouri by the posterior position of the vulva and the dilated posterior extremity in females. However, the new species differs from N. macrouri mainly by its larger size, a larger muscular esophagus, and a widely globose posterior extremity in females. SEM study of cephalic structures also showed morphological differences between both species, especially in the morphology of the submedian labia and lateral pseudolabia. In view of these differences a new species is proposed.