Large Subunit Ribosomal RNA Research Articles

Leaf Spots on bodhi tree (Ficus religiosa) caused by Diaporthe tulliensis.

Ficus religiosa L., known as bodhi tree, is an ornamental tree and widely planted as an avenue and roadside tree due to ovate-rounded leaves with narrow, elongated tips. During 2018-2021, circular to oval-shaped leaf spots with pale white centers and brown-black edges surrounded by a chlorotic halo were observed on the leaves of more than 200 bodhi trees all year round in a park in Zhanjiang, Guangdong (N 21°15'22.29''; E110°23'1.03''). The leaf disease incidences were usually 15-80%, in severe cases, up to 100% in autumn and winter every year, and some trees shed all leaves(Fig S1). Repeated annual defoliation may weaken the tree and decrease the aesthetic value in the landscape. Diseased tissues (5 × 5 mm) of five symptomatic infected leaves were surface sterilized in 3% hydrogen peroxide solution for 3 min, rinsed thrice with sterile water, plated on potato dextrose agar (PDA) amended with ampicillin (50mg/L), and incubated at 25-28 ℃ in the dark for 3-7 days. Five strains with similar morphology were obtained by transferring hyphal tips of the colonies to fresh PDA and further isolating by single spore method. Fungal colonies were flat and spreading, with sparse, white aerial mycelium, and black pycnidial conidiomata semi-immersed in PDA after 30-days incubation at 25-28 ℃ in dark. Conidiophores were hyaline and α-conidia were single-celled, oval to fusiform, guttulate, 5.3 × 2.5 μm (n>50), similar to Diaporthe sp. (Crous et al. 2015), but no β and γ -conidia were observed. The internal transcribed spacer(ITS), large subunit ribosomal RNA gene(LSU), calmodulin (CAL) and β-tubulin(TUB) gene regions of a representative strain were amplified using specific primers reported by White et al. (1990), Gao et al (2017) and Gomes et al (2013), and submitted to GenBank (ITS: OM200852, LSU: OM228732, CAL: OM244761, TUB: OM244760). NCBI Blastn showed above 99% identity to D. tulliensis (anamorph: Phomopsis heveicola ) isolates of ITS (MT974186, MN393590 and KX457967), LSU (KR936131), CAL (MW759801), and TUB (KR936132 and MN399886), respectively (Crous et al. 2015; Huang et al. 2021; Gong et al. 2020; Bai et al. 2017). Based on the concatenated ITS, CAL, and TUB, a Maximum Likelihood phylogenetic analyses using MEGA 10.1.8 clustered the fungus with D. tulliensis in a clade with a 93% bootstrap support(Fig S2). Therefore, the fungus was identified as D. tulliensis based on morpho- molecular characteristics. Healthy detached leaves were sanitized thrice with 70% alcohol, and rinsed with sterile water. PDA plugs with actively growing 10-days-old mycelium were placed on predetermined sites, put into a sealed box with above 80% relative humidity and incubated at room temperature (25-28℃). Each isolate was inoculated at 25 needle-wounded and unwounded sites, PDA plugs without mycelium served as controls. Symptomatic spots appeared on all wounded leaves by 7 days post-inoculation (dpi) and on all unwounded leaves by 12 dpi. No symptoms appeared on controlled leaves. Pure cultures were recovered from inoculated leaves and showed identical morpho-molecular criteria to the original isolates. More than 70 pathogenic fungi are known to cause diseases on F. religiosa while there is no record of D. tulliensis infecting bodhi according to the U.S. National Fungus Collection (Farr and Rossman 2022). This report could provide basic understanding and alerting role for horticulturist in daily management.

Fungal endophyte Epicoccum nigrum 38L1 inhibits in vitro and in vivo the pathogenic fungus Fusarium graminearum

Wheat (Triticum aestivum L.) and its products constitute a major part of the daily human and animal diet. However, Fusarium head blight caused by Fusarium graminearum is one of the most devastating fungal diseases in wheat. In this study, a fungal endophyte strain 38L1 was newly isolated from leaf of maize (Zea mays L.) and identified as Epicoccum nigrum using molecular phylogenetic analysis of the rDNA internal transcribed spacer (ITS), large subunit ribosomal RNA (LSU) and beta-tubulin (TUB) gene sequences. In dual cultures and double plate assays, strain 38L1 markedly inhibited the mycelial growth of F. graminearum strain PH-1 and four other pathogenic fungi including Botrytis cinerea strain P-1, Colletotrichum gloeosporioides strain Y-3, Botrytis cinerea strain F-2, and Sclerotinia sclerotiorum strain D-4, indicating a broad antifungal activity. Besides, culture filtrates of strain 38L1 indicated a high fungicidal effect resulting in a significant reduction of mycelial growth, mycelial biomass and spore germination of F. graminearum strain PH-1. Furthermore, the application of conidial suspensions and/or culture filtrates of E. nigrum strain 38L1 with spore suspensions of F. graminearum strain PH-1 on wheat spikes resulted in a significant biocontrol efficacy of Fusarium head blight in growth chamber and field conditions. Hence, these results suggest that E. nigrum strain 38L1 could be a potential biocontrol agent against F. graminearum strain PH-1 in vitro and in vivo.

Updated taxonomy of Chinese Clavaria subg. Syncoryne (Clavariaceae, Agaricales): description of two new species and one newly recorded species

Species of Clavaria (Clavariaceae, Agaricales) collected from China were studied using morphological and molecular methods. Two species, C. aspersa and C. hupingshanensis, are here described as new to science; C. aspersa possesses simple, white basidiomata in gregarious or scattered fascicles, whereas C. hupingshanensis possesses simple, rose-white to seashell-pink basidiomata in gregarious to caespitose clusters. In addition, C. amoenoides is described as a newly recorded species for China; this species is characterized by simple, very pale orange-yellow to picric-yellow basidiomata. A phylogenetic analysis was conducted based on a combined dataset of internal transcribed spacer, nuclear ribosomal RNA large subunit, and the RNA polymerase II second largest subunit sequences. The phylogenetic reconstruction resolved accessions of the three species into three independent lineages within the Clavaria. The morphology of the three species is described in detail and is illustrated with line drawings and photographs. Holotypes of the new species are deposited in the Mycological Herbarium of Hunan Normal University. The sequences newly generated in this study have been deposited in GenBank. An updated key to the known Clavaria species in China is provided.

Development of a nested PCR assay for specific detection of Metschnikowia bicuspidata infecting Eriocheir sinensis.

In recent years, the “milky disease” caused by Metschnikowia bicuspidata has seriously affected the Eriocheir sinensis culture industry. Discovering and blocking the transmission route has become the key to controlling this disease. The existing polymerase chain reaction (PCR) detection technology for M. bicuspidata uses the ribosomal DNA (rDNA) sequence, but low sensitivity and specificity lead to frequent false detections. We developed a highly specific and sensitive nested PCR method to detect M. bicuspidata, by targeting the hyphally regulated cell wall protein (HYR) gene. This nested HYR-PCR produced a single clear band, showed no cross-reaction with other pathogens, and was superior to rDNA-PCR in specificity and sensitivity. The sensitivity of nested HYR-PCR (6.10 × 101 copies/μL) was greater than those of the large subunit ribosomal RNA gene (LSU rRNA; 6.03 × 104 copies/μL) and internal transcribed spacer (ITS; 6.74 × 105 copies/μL) PCRs. The nested HYR-PCR also showed a higher positivity rate (71.1%) than those obtained with LSU rRNA (16.7%) and ITS rDNA (24.4%). In conclusion, we developed a new nested HYR-PCR method for the specific and sensitive detection of M. bicuspidata infection. This will help to elucidate the transmission route of M. bicuspidata and to design effective management and control measures for M. bicuspidata disease.

New records of three commercial fish hosts for two Unicapsula spp. and Kudoa megacapsula (Myxozoa: Myxosporea: Multivalvulida)

Infections caused by multivalvulid myxosporeans belonging to genera Unicapsula and Kudoa (Cnidaria: Myxozoa) occasionally affect commercial marine fish species. Postmortem myoliquefaction caused by a variety of Kudoa spp., including K. thyrsites, and unsightly cyst or pseudocyst formation, caused by K. amamiensis, U. muscularis, and other kudoid species, negatively affect commercial values of fillets. However, multivalvulid infections are often latent and imperceptible in the market. Biodiversity, host range, and epidemiology remain to be explored. Here, myxosporean infection was detected in four commercial fish species from southern China, using morphological and molecular analyses. Three Unicapsula spp. (U. pyramidata in Nemipterus japonicus; U. pflugfelderi in Dentex angolensis transported from the Eastern Central Atlantic Ocean, off West African coast; and U. aequilobata in Decapterus macarellus) and Kudoa megacapsula in Nemipterus virgatus were observed to form pseudocysts in the myofibers of the host trunk muscles. All fish hosts identified here, except for U. pyramidata, are new records. Kudoa megacapsula was morphologically characterized by gigantic, cruciform myxospores with four wing-like shell valves morphologically comparable to previous Japanese records of the same species in aquaculture facilities, acquiring fly from China or Korea (Sphyraena pinguis and Seriola quinqueradiata, respectively). Molecular analyses established the conspecificity of the present Chinese isolate with previously recorded Japanese isolates. To our knowledge, for the first time, a partial large subunit ribosomal RNA gene sequence of K. megacapsula was obtained, showing close phylogenetic relationships with Kudoa spp. harboring cruciform myxospores, such as K. thyrsites, K. gunterae, K. whippsi, and K. lateolabracis.

Boletus littoreus (Boletaceae, Boletales), a new species with a white basidioma from Hainan, China

Boletus littoreus (Boletaceae, Boletales) is described as a new species from tropical China based on morphological and molecular evidence from the nuclear ribosomal large subunit RNA (28S), the translation elongation factor 1-α gene (TEF1), and the RNA polymerase II second largest subunit gene (RPB2). It is morphologically characterized by white basidiomata, a stipe without reticulations, a pileipellis composed of elliptic-fusiform to subglobose, sometimes filamentous, chains of hyphae, and is associated with trees of Dipterocarpaceae. An illustrated morphological description is provided along with comparison to related and similar species.

Morphological and phylogenetic analyses reveal two new species and a new record of Phyllosticta (Botryosphaeriales, Phyllostictaceae) from Hainan, China.

The fungal genus Phyllosticta has been reported from all around the world and accommodates numerous pathogenic and endophytic species isolated from a wide range of plant hosts. Based on multilocus phylogenies from a combined dataset of genes encoding internal transcribed spacer (ITS), large subunit of ribosomal RNA (LSU rDNA), translation elongation factor 1 alpha (TEF1α), actin (ACT) and glycerol-3-phosphate dehydrogenase (GPDH), in conjunction with morphological characteristics, we describe two new species P.oblongifoliae sp. nov. and P.pterospermi sp. nov., as well as a new Chinese record P.capitalensis. Their similarity and dissimilarity to morphologically-allied and phylogenetically-related species are also annotated and discussed.

Morphological and molecular identification of Ellipsoidisporodochium gen. nov. (Tubakiaceae, Diaporthales) in Hainan Province, China

During a survey of microfungi on symptomatic leaves from Photinia serratifolia (Desfontaines) Kalkman, an interesting species of asexual ascomycetes were found. Based on multilocus phylogenies from a combined dataset of genes encoding internal transcribed spacer (ITS), large subunit of ribosomal RNA (LSU), translation elongation factor 1 alpha (tef1), part of the beta-tubulin gene region (tub2), and partial RNA polymerase II second largest subunit (rpb2), in conjunction with morphological characteristics, we describe a new genus of Tubakiaceae, Ellipsoidisporodochium gen. nov. (type species: Ellipsoidisporodochium photiniae sp. nov.). The genus has fusoid to ampulliform sporodochia and obovoid to ellipsoid conidia. Their relatedness to allied taxa are also annotated and discussed.

Parasites with zoonotic potential found in commercially important fish in Tamaulipas, Northeastern Mexico.

Human population is exposed to numerous parasitic ichthyozoonoses. Although Tamaulipas state (northeastern Mexico) is well known for its fishing and aquaculture industry, there are few reports of this type of zoonosis. Therefore, it is imperative to investigate whether the parasites that affect these fish may represent a zoonotic risk for the inhabitants of the area. The objective of this study was to identify molecular and/or morphologically muscle parasites of fish from coastal locations in Tamaulipas, Mexico, and assess the risk of infection for humans. Between 2017 and 2018, 764 individual fish belonging to 28 species were examined for parasites. Collected worms were processed for their identification using morphological characteristics. In addition, partial sequences of the large subunit (28S) ribosomal RNA gene were obtained from some species to corroborate their identity. Prevalence and mean intensity of all registered infections were calculated. A total of seven species of parasites were found: cestodes (Poecilancistrium caryophyllum), trematodes (Clinostomum tataxumui, Clinostomum cichlidorum), nematodes (Eustrongylides sp., Contracaecum sp.) and pentastomids (Sebekia purdieae, Sebekia sp.). Parasites infected 10 species belonging to different fish families (Ariidae, Centrarchidae, Centropomidae, Cichlidae, Eleotridae, Ictaluridae, Mugilidae and Sciaenidae). Congeneric species of parasites or related to those registered in this study have been identified as zoonotic agents in other regions of the world. Despite the low levels of infection (2.6-16.6% prevalence and 1-5.5 parasites per infected host), there is a latent risk of transmission to humans, so it is recommended to avoid eating raw or undercooked fish meat.

English

The genus Basidiobolus contains large groups of terrestrial fungi including the etiological agents of human gastrointestinal basidiobolomycosis (GIB). This study aimed to identify Basidiobolus species from the common house gecko and to compare them with human GIB isolates. Gecko and human GIB samples were collected from Muhayil Aseer area, south Saudi Arabia (2017-2019). Isolation of fungi from the gut contents of geckos was performed using Sabouraud dextrose agar incubated aerobically at 30°C for five days. Suspected Basidiobolus species were tentatively identified using routine bench tests and phenotypes were authenticated by phylogenetic analysis of the large subunit ribosomal RNA gene. Isolates (n = 10) were found to have zygomycete-like phenotypic characteristics. In the 28S ribosomal RNA gene phylogenetic tree, the strains assembled in the subclade encompass Basidiobolus spp. along with previously reported isolates from human’ GIB. The strains had a close resemblance with Basidiobolus haptosporus (99.97%) as well as with B. haptosporus var. minor (99.97%). One isolate (L3) falls within the subclade containing B. haptosporus strain NRRL28635. The recovery of similar isolates from both humans and gecko lizards in one geographic region is an important development toward knowing risk factors for GIB.   Key words: Zygomycetes, Entomophthoromycota, Basidiobolus, lizards.

The <i>COI</i> haplotype diversity of the pelagic polychaete <i>Tomopteris</i> (Annelida: Tomopteridae) collected from the Pacific coast off Kii Peninsula, central Japan

The polychaete genus Tomopteris is an important component of the zooplankton and is also known for its communication using bioluminescence. However, there have been few studies about the species diversity and ecology of Tomopteris because of taxonomic and handling difficulties resulting from the fragility of their bodies. In this study, we performed DNA barcoding and species delimitation analysis to better understand the species diversity of Tomopteris using 17 specimens collected on a research cruise conducted off the eastern Kii Peninsula, Japan. Due to damage to the detailed morphological characteristics necessary for identifications based on the current taxonomy, none of the specimens was morphologically identifiable at the species level. The analyses of the mitochondrial cytochrome c oxidase subunit I (COI) and nuclear large ribosomal subunit (28S) RNA gene sequences revealed 7–8 molecularly delimited species among them, which was greater than the number of species currently recognized in Japan. These results suggest that the traditional morphology-based taxonomy overlooks the taxonomic complexity of Tomopteris, thus a combinatory analysis of both morphology and DNA barcoding will be desirable for assessing the true species biodiversity of Tomopteris.

Penicillium vietnamense sp. nov., the First Novel Marine Fungi Species Described from Vietnam with a Unique Conidiophore Structure and Molecular Phylogeny of Penicillium Section Charlesia

Penicillium vietnamense sp. nov. was isolated from Nha Trang Bay, Vietnam in June 2017. It is phylogenetically distinct from the sister species of Penicillium section Charlesia series Indica based on multi-locus sequence typing results using internal transcribed spacer, large subunit ribosomal RNA, β-tubulin, calmodulin, and RNA polymerase II second largest subunit regions. It showed strong growth on Czapek yeast autolysate agar at 37 °C, a strong acid production on Creatine sucrose agar, and produced short stipes, small vesicles, and subglobose to globose conidia delicately roughened with very short ridges. As the first novel marine fungi species described from Vietnam and discovered in a unique environment, the data could be significant for understanding the taxonomy and geographical distribution of marine fungi in tropical coastal systems such as Vietnam.

Molecular and Morphological Characteristics of a New Species Collected from an Insect (Cicindela transbaicalica) in Korea

To exploit insect-derived fungi, insects were collected from seven different regions in Korea, including Gyeongbuk, Goryeong, and several fungi were isolated from them. A fungal strain designated 21-64-D was isolated from riparian tiger beetle (Cicindela transbaicalica) and morphologically identified as a species belonging to the genus Oidiodendron. Phylogenetic analysis using the nucleotide sequences of internal transcribed spacer (ITS) regions and the partial sequence of the large subunit of the nuclear ribosomal RNA (LSU) gene revealed the distinct phylogenetic position of the isolate among recognized Oidiodendron species including its closest neighbors O. chlamydosporicum, O. citrinum, O. maius, and O. pilicola. The hyphal and conidial morphology of the strain, particularly club-shaped hyphae, clearly differentiated it from its close relatives. Results indicated that 21-64-D is a novel species in the genus Oidiodendron, for which the name Oidiodendron clavatum sp. nov. is proposed.

Stigmatomyces aff. limnophorae on dipteran hosts in Peninsular Malaysia

Flies parasitized by Laboulbeniales, captured in Selangor state, were examined and identified as Boettcherisca javanica, Boettcherisca sp ., and Hypopygiopsis violacea. The fungus was identified as Stigmatomyces aff. limnophorae based on morphology and phylogenetic analysis of sequences of the partial nuclear small and large subunit ribosomal RNA genes. This fungus represents a new record for Malaysia; and these are the first reports of Boettcherisca and Hypopygiopsis as hosts for any laboulbenialean species.

First Report of Epicoccum layuense Causing Leaf Brown Spot on Camellia oleifera in Hefei, China.

Camellia oleifera, a major tree species for producing edible oil, is originated in China. Its oil is also called ''eastern olive oil'' with high economic value due to richness in a variety of healthy fatty acids (Lin et al. 218). However, leaves are susceptible to leaf spot disease (Zhu et al. 2014). In May 2021, we found circular to irregular reddish-brown lesions, 4-11 mm in diameter, near the leaf veins or leaf edges on 30%-50% leaves of 1/3 oil tea trees in a garden of Hefei City, Anhui Province, China (East longitude 117.27, North latitude 31.86) (Figure S1 A). To isolate the causal agents, symptomatic leaves were cut from the junction of diseased and healthy tissues (5X5 mm) and treated with 70 % alcohol for 30 secs and 1 % NaClO for 5 min, and subsequently inoculated onto PDA medium for culture. After 3 days, hyphal tips were transferred to PDA. Eventually, five isolates were obtained. Then the isolates were cultured on PDA at 25°C for 7 days and the mycelia appeared yellow with a white edge and secreted a large amount of orange-red material to the PDA (Figure S1 B and C). Twenty days later, the mycelium appeared reddish-brown, and sub-circular (3-10 mm) raised white or yellow mycelium was commonly seen on the Petri dish, and black particles were occasionally seen. Meanwhile, the colonies on the PDA produced abundant conidia. Microscopy revealed that conidia were globular to pyriform, dark, verrucose, and multicellular with 14.2 to 25.3 μm (=19.34 μm, n = 30) diameter (Figure S1 D). The morphological characteristics of mycelial and conidia from these isolates are similar to that of Epicoccum layuense (Chen et al.2020). To further determine the species classification of the isolates, DNA was extracted from 7-day-old mycelia cultures and the PCR-amplified fragments were sequenced for internal transcribed spacer (ITS), beta-tubulin and 28S large subunit ribosomal RNA (LSU) gene regions ITS1/ITS4, Bt2a/Bt2b and LR0R/LR5, followed by sequencing and molecular phylogenetic analysis of the sequences analysis (White et al. 1990; Glass and Donaldson 1995; Vilgalys and Hester 1990). Sequence analysis revealed that ITS, beta-tubulin, and LSU divided these isolates into two groups. The isolates AAU-NCY1 and AAU-NCY2, representing the first group (AAU-NCY1 and AAU-NCY5) and the second group (AAU-NCY2, AAU-NCY3 and AAU-NCY4), respectively, were used for further studies. Based on BLASTn analysis, the ITS sequences of AAU-NCY1 (MZ477250) and AAU-NCY2 (MZ477251) showed 100 and 99.6% identity with E. layuense accessions MN396393 and KY742108, respectively. And, the beta-tubulin sequences (MZ552310; MZ552311) showed 99.03 and 99.35% identity with E. layuense accessions MN397247 and MN397248, respectively. Consistently, their LSU (MZ477254; MZ477255) showed 99.88 and 99.77% identity with E. layuense accessions MN328724 and MN396395, respectively. Phylogenetic trees were built by maximum likelihood method (1,000 replicates) using MEGA v.6.0 based on the concatenated sequences of ITS, beta-tubulin and LSU (Figure S2). Phylogenetic tree analysis confirmed that AAU-NCY1 and AAU-NCY2 are closely clustered with E. layuense stains (Figure S2). To test the pathogenicity, conidial suspension of AAU-NCY2 (106 spores/mL) was prepared and sterile water was used as the control. Twelve healthy leaves (six for each treatment) on C. oleifera tree were punched with sterile needle (0.8-1mm), the sterile water or spore suspension was added dropwise at the pinhole respectively (Figure S1 E and F). The experiment was repeated three times. By ten-day post inoculation, the leaves infected by the conidia gradually developed reddish-brown necrotic spots that were similar to those observed in the garden, while the control leaves remained asymptomatic (Figure S1 G and H). DNA sequences derived from the strain re-isolated from the infected leaves was identical to that of the original strain. E. layuense has been reported to cause leaf spot on C. sinensis (Chen et al. 2020), and similar pathogenic phenotypes were reported on Weigela florida (Tian et al. 2021) and Prunus x yedoensis Matsumura in Korea ( Han et al. 2021). To our knowledge, this is the first report of E. layuense causing leaf spot on C. oleifera in Hefei, China.

Six New Species of Leucoagaricus (Agaricaceae) from Northeastern China

Six new species, Leucoagaricus albosquamosus, Leucoagaricus atroviridis, Leucoagaricus aurantioruber, Leucoagaricus candidus, Leucoagaricus centricastaneus and Leucoagaricus virens, collected from northeastern China are described based on morphological characters and molecular evidence. Illustrations of fresh basidiomata and line drawings of key anatomical characters are provided. A phylogenetic tree inferred from internal transcribed spacer (ITS) region and large subunit ribosomal RNA gene (LSU) sequences shows that three of the new taxa are nested within the section Leucoagaricus and two of the new taxa are in the subgenus Sericeomyces, whereas the other new taxus is clustered with Leucoagaricus viriditinctus and Leucoagaricus irinellus, forming a clade that does not fit in any known section.

A New Lichenized Fungus, Psoroglaena humidosilvae, from a Forested Wetland of Korea, with a Taxonomic Key to the Species of Psoroglaena.

Psoroglaena humidosilvae Lee is described as a new lichen species from a wetland forest in South Korea. The new species is distinct from P. stigonemoides (Orange) Henssen by little projections locally present on the thallus; smaller, paler, and globose perithecia; smaller asci; and smaller ascospores generally 3-septate. Molecular analyses employing internal transcribed spacer (ITS), mitochondrial small subunit (mtSSU), and nuclear large subunit ribosomal RNA (LSU) sequences strongly support P. humidosilvae as a nonidentical species in the genus Psoroglaena. A surrogate key is provided to assist in the identification of all 22 species of Psoroglaena.

First Report of a New Disease Caused by Fusarium tricinctum on Apple Tree in China.

First Report of a New Disease Caused by Fusarium tricinctum on Apple Tree in China.

Morphological and molecular phylogenetic analysis of Bivesicula claviformis Yamaguti, 1934 infecting the tomato hind Cephalopholis sonnerati (Serranidae) in Saudi Arabia

Groupers are large predators in the Serranidae family that feed on fish and small invertebrates. Identification of parasitic taxa of groupers was done concerning their morphological and biological characteristics. A total of 30 Cephalopholis sonnerati (Serranidae) specimens were investigated for digenetic trematodes in the Red Sea, Saudi Arabia. Only one bivesiculid species, belonging to the Bivesiculidae family, has been identified, with a prevalence rate of 60% (18/30) among infected fish. The Bivesicula species obtained from the fish host’s intestine is morphologically and morphometric compatible with Bivesicula claviformis, which was previously identified from Epinephelus fasciatus (Serranidae) in North Borneo. In addition, maximum parsimony based on Tamura-Nei model was utilized to determine the phylogeny of the recovered Bivesicula species using partial small subunit ribosomal RNA gene (18S rRNA) and large subunit ribosomal RNA gene (28S rRNA) sequences. The query sequences showed the identities of 99.39% and 94.02% for 18S (AJ287485.1) and 28S rRNA (AY222182.1) of the previously described Bivesicula claviformis, respectively. The present study demonstrated that Bivesicula claviformis is the first reported of this genus as endoparasites from C. sonnerati, as well as providing novel DNA data for this species.

Redescription of Ptychobothrium belones (Dujardin, 1845) (Cestoda: Bothriocephalidea) from needlefishes (Beloniformes: Belonidae) in the Mediterranean Sea.

The bothriocephalidean tapeworm Ptychobothrium belones (Dujardin, 1845) Lönnberg, 1889 is redescribed on the basis of new materials collected from the intestine of the Mediterranean needlefish Tylosurus imperialis (Rafinesque) (Beloniformes: Belonidae) off Tunisia in the central Mediterranean Sea. This species is characterised by a laterally compressed hexagonal to fan-shaped scolex with an apical disc and two well-developed bothria, an oval ovary and numerous testes arranged in lateral and median fields. The typical diagnostic feature of P. belones is the medullary yolk follicles located between the muscle fibres of the internal longitudinal muscles (paramuscular) and the uterine duct strongly sinuous, S-shaped in mature proglottides. New molecular data (partial sequence of nuclear large subunit ribosomal RNA gene) confirmed the close relationship with specimens previously collected by Brabec et al. (2006, 2015) from the banded needlefish Strongylura leiura (Bleeker) off the Maldives, Indian Ocean, but these specimens are most likely another species. Thus, Ptychobothrium belones as previously reported represents most likely more than one species and its cosmopolitan distribution and host specificity revised by Kuchta et al. (2008b) should be revisited. The type material of P. belones was reported as collected in Mediterranean Sea off Sète, France, from Belone belone (Linnaeus); however, this host was probably misidentified, being confused with morphologically similar small specimens of T. imperialis. This study confirms that P. belones is an exclusive parasite of needlefishes (Belonidae) but at least one further species infecting S. leiura may exist.