Partial 16S rDNA Sequencing Research Articles

Exopolysaccharides from an Ophiopogon japonicus endophyte inhibit proliferation and migration in MC-4 human gastric cancer cells

Background: Recently, natural products have gained increasing attention. Endophytic bacteria represent a source of natural anticancer products, and are well-known for producing diverse bioactive anti-cancer compounds. Herein, we investigated the antitumor activity of exopolysaccharides (EPSs) extracted from endophytes in Ophiopogon japonicus . Methods: The endophyte bacterium MD4 was isolated from O. japonicus . To identify MD4, a partial sequence of 16S rDNA was analyzed. The phenol-sulfuric acid method was used to measure the EPSs concentration. Cell viability was tested by the MTT assay to quantify the EPS cytotoxicity effect. Wounding healing, gelatin zymography, and ELISA were used to measure cell migration ability. Results: MD4 was identified as a Staphylococcus sp . with 99% similarity. The results of the phenol sulfuric acid method showed that the purified EPS concentration from MD4 was 28.068 µg/µL. There was a concentration-dependent cytotoxic effect of MD4-EPS on MC-4 cells, with IC50 values of 0.891 µg/µL. The results of the MTT assay showed that MD4-EPS effectively inhibited the migration of MC-4 cells as the concentration increased. The gelatin zymography analysis showed that the enzymatic activities of the migration-related MMP2 and MMP9 proteins decreased after EPS treatment. Furthermore, the protein secretion levels of MMP2 and MMP9 after MD4-EPS treatment were significantly lower than those in the control and PBS groups. Conclusions: Our results demonstrate that the EPSs derived from the MD4 endophyte of O. japonicus acted as natural antitumor agents to inhibit the migration of gastric cancer cells.

Biogreen remediation of chromium-contaminated soil using Pseudomonas sp. (RPT) and neem (Azadirachta indica) oil cake

The prospective of indigenous Pseudomonas sp. (RPT) and neem oil cake for enhanced removal of chromium (Cr) from contaminated soil microcosm was explored in this study. The bacteria were isolated from Cr-contaminated soil and identified as Pseudomonas sp. based on partial 16S rDNA sequencing. The isolate RPT showed high Cr(VI) tolerance (1000 mg/l) and removal rate (64.4%) in batch experiments. Transmission electron microscopy observation showed that the isolate effectively precipitated the Cr both intra- and extracellularly. Microcosm studies revealed that neem oil cake amendment (7.5% w/v) enhanced Cr(VI) removal (82%) from contaminated soil. Furthermore, soil enzyme activities were increased in the biostimulated soil. The obtained results indicated that the application of neem oil cake along with indigenous Cr(VI)-resistant bacteria could inspire the bioremediation of Cr(VI)-contaminated soil field scale.

Identification of predominant epiphytic and endophytic bacterial isolates in rice seeds effective for enhancement of seed germination and plant growth

ABSTRACT-During rice growing season, the symptomless rice seeds from different paddy fields in Guilan province, Iran, were collected. After isolation of epiphytic and endophytic bacteria, 39 isolates including 19 epiphytes and 20 endophytes were selected based on the predominant characteristics. Five Operational Taxonomic Units (OTUs) were obtained based on PCR-RFLP of 16S r-DNA in these isolates. According to biochemical tests and partial sequencing of 16S-rDNA, in both epiphyte and endophyte bacteria of rice seeds, the most populated OTUs (V and II) were identified as Pantoea ananatis and Pseudomonas oryzihabitans, respectively. Six representative isolates from these two OTUs were selected to be evaluated for their abilities for rice seed germination and growth enhancement. Among them, P. oryzihabitans was not beneficially effective. However, JpB1 isolate of P. ananatis was considered to be the most effective plant growth promoting isolate, since it showed stable beneficial effects on most surveyed characteristics in both rice seed germination and growth enhancement experiments. Although, OpB3 isolate of P. ananatis produced IAA in higher amount and solubilized phosphate more than the other isolates followed by JpB1 and P. ananatis L3pB3, it was not beneficially effective on rice seed germination.

Hidden mycota of pine needles: Molecular signatures from PCR-DGGE and Ribosomal DNA phylogenetic characterization of novel phylotypes

Previous studies for enumerating fungal communities on pine needles relied entirely on phenotypic diversity (microscopy) or identification based on DNA sequence data from those taxa recovered via cultural studies. To bypass limitations of the culturing methods and provide a more realistic diversity estimate, we employed and assessed a PCR-DGGE based method coupled with rDNA phylogenetic sequence analyses to characterize fungal taxa associated with pine needles. Fresh (living) and decayed needles from three hosts of the Pinaceae (Keteleeria fortunei, Pinus elliottii and P. massoniana) were examined. Morphological studies reveal that the most abundant species associated with decayed needles were Cladosporium cladosporioides and an unidentified Trichoderma species followed by Gliocephalotrichum sp., Gliocladium sp., Lophodermium pinastri, Paecilomyces varioti, Phaeostalagmus cyclosporus and a Phoma sp, which are commonly occurring fungi. Community genomic data from freshly collected and decayed pine needles recovered 40 operational taxonomic units, which appear to be mostly undetected members of the natural fungal consortium. Sequence analyses revealed a number of phylotypes or “species” that were not recovered using traditional morphological and cultural approaches previously used. Phylogenetic data from partial 18S rDNA sequence data reveal that most phylotypes represent potential novel phylogenetic fungal lineages with affinities to the Dothideomycetes, Leotiomycetes, Lecanoromycetes and Sordariomycetes and were not identical to previously known endophytes or saprobes. Although the major ecological roles of these phylotypes in pine needles are still enigmatic, this study provides new insights in hidden fungal diversity that mycologists are possibly ignoring given the discrepancies associated with available methods. To what extent do previously recovered identified species (either as saprobes or endophytes) from morphological or culturing studies act as pioneer decomposers or constitute an integral part of endophytic community warrants further investigation.

Molecular and morphological characterization of Paurodontoides siddiqii n. sp. (Nematoda: Hexatylina, Sphaerulariidae) associated with bark samples of Pinus eldarica from western Iran.

A new species of Paurodontoides, P. siddiqii n. sp., is described and illustrated based on its morphological, morphometric, and molecular characters. The new species is characterized by a female 550-729 μm long, lip region continuous with body contour, stylet length 7.0-8.0 μm long or c. 1.0-1.2 times the lip region diameter, lateral fields with four smooth incisures, excretory pore at 85-125 μm from anterior end located at the base of the pharyngeal bulb or posterior to it, basal pharyngeal bulb with a short posterior extension projecting into the intestine, monodelphic-prodelphic reproductive system with prominent 19-22 μm long post-uterine sac, and elongate conoid tail with a filiform terminus. The new species is compared with two known species of the genus. It differs from the type species of the genus, P. linfordi, by having slightly shorter stylet, lateral field with smooth incisures, different position of the excretory pore, and absence of male. Compared to P. latus, the new species has a shorter body, shorter stylet, different position of the excretory pore, female tail shape and absence of male. The new species was also compared with close species of the genus Paurodontus because of lateral field marked with four lines, asymmetrical stylet knobs and absence of male. Molecular phylogenetic studies of the new species using partial sequences of 18S rDNA revealed that it forms a clade with a species of the genus Ficotylus. In phylogenetic analyses using partial sequences of the 28S rDNA D2-D3 domain, the new species formed a monophyletic group with a species of the genus Veleshkinema and Sphaerularia spp. (Sphaerulariinae).

Intranuclear coccidiosis in tortoises — discovery of its causative agent and transmission

Intranuclear coccidiosis of testudines (known as TINC) is an emerging disease in chelonians. Although endogenous stages were repeatedly detected in various tissues, attempts to find the oocysts in faeces failed, leaving the question of the transmission and classification of the causative agent of TINC unresolved. We recorded small spherical oocysts (∼6–7 μm in diameter) of an eimeriid coccidium in faeces of a leopard tortoise (Stigmochelys pardalis). Sporulated oocysts were used for the experimental oral inoculation of juvenile coccidia-free tortoises representing 5 species (S. pardalis, Testudo graeca, T. hermanni, T. horsfieldii, and Geochelone sulcata). The oocysts’ association with TINC was confirmed based on clinical signs, histopathological findings of intranuclear endogenous stages of the coccidium in many organs (including intestine), and by the partial 18S rDNA sequence analysis of the DNA isolated from organs of the experimentally infected animals and from a single naturally infected as well as from all experimentally infected tortoises. Breeding colonies of chelonians should be screened for this pathogen in order to prevent its further spread and unwanted introduction into endangered free-ranging chelonian populations.

Three, two, one! Revision of the long-bodied sphaerodorids (Sphaerodoridae, Annelida) and synonymization of Ephesiella, Ephesiopsis and Sphaerodorum.

BackgroundLong-bodied sphaerodorids (Annelida, Sphaerodoridae) is the common name for members of the three closely and morphologically homogenous currently accepted genera of benthic marine bristle worms: Ephesiella, Ephesiopsis and Sphaerodorum. Members of this group share the presence of two dorsal and longitudinal rows of macrotubercles with terminal papillae, and two longitudinal rows of microtubercles, features that are unique among sphaerodorids. Genera are distinguished by the chaetae morphology. Members of Ephesiella are characterised by having compound chaetae (except, sometimes, simple chaetae in the first chaetigers), Sphaerodorum bear only simple chaetae, and Ephesiopsis have both compound and simple chaetae in all parapodia.MethodsMitochondrial (partial COI and 16S rDNA) and nuclear (partial 18S rDNA and 28S rDNA) sequence data of long-bodied sphaerodorids with compound and simple chaetae, and an outgroup of additional seven sphaerodorid species were analysed separately and in combination using Bayesian inference (BA), and Maximum Likelihood (ML) methods. Long-bodied sphaerodorids from around the world (including type specimens) were examined under a range of optical equipment in order to evaluate putative generic and specific diagnostic features, in addition to intraspecific variability.ResultsPhylogenetic analyses of mitochondrial and nuclear DNA sequences of specimens identified as Ephesiella and Sphaerodorum, based on chaeta morphology, were performed. Sphaerodorum and Ephesiella were recovered as paraphyletic and nested within each other. Revision of current nominal species diagnostic features are performed and discussed.DiscussionResults contradict current generic definitions. Recovery of paraphyletic compound and simple chaetae clades urge the synonymization of these two genera of long-bodied sphaerodorids. Morphological data also suggest the synonymization of Ephesiopsis.

A new species of Haliotrema (Monogenea: Ancyrocephalidae (sensu lato) Bychowsky & Nagibina, 1968) from holocentrids off Langkawi Island, Malaysia with notes on the phylogeny of related Haliotrema species

Haliotrema susanae sp. nov. is described from the gills of the pinecone soldierfish, Myripristis murdjan off Langkawi Island, Malaysia. This species is differentiated from other Haliotrema species especially those from holocentrids in having a male copulatory organ with bract-like extensions at the initial of the copulatory tube, grooved dorsal anchors and ventral anchors with longer shafts. The maximum likelihood (ML) analysis based on partial 28S rDNA sequences of H. susanae sp. nov. and 47 closely related monogeneans showed that H. susanae sp. nov. is recovered within a monophyletic clade consisting of only species from the genus Haliotrema. It is also observed that H. susanae sp. nov. forms a clade with H. cromileptis and H. epinepheli which coincides with a similar grouping by Young based on solely morphological characteristics. The morphological and molecular results validate the identity of H. susanae sp. nov. as belonging to the genus Haliotrema.

Psychropotid holothurians (Echinodermata: Holothuroidea: Elasipodida) of the tropical Western Pacific collected by the KEXUE expedition with description of one new species

ABSTRACTTwo holothurian species of the family Psychropotidae were collected by RV KEXUE during the cruise to tropical Western Pacific. One of them, Benthodytes manusensis is described as new to science. The new species has a broad brim and reddish light violet body, with three pairs of well-developed dorsal papillae, arranged in two single rows, evenly distributed. Body wall ossicles were comprised of rods and large crosses with a bipartite central apophysis. We also provide the Genbank accession numbers of partial sequences of 16S rDNA and mitochondrial cytochrome oxidase subunit I gene. In addition, notes are presented on the morphology and distribution of another species Psychropotes depressa (Théel).http://zoobank.org/urn:lsid:zoobank.org:pub:22B43A54-1B05-4812-A098-448A0044B49C

Phylogenetic relationships within Dicrocoeliidae (Platyhelminthes: Digenea) from birds from the Czech Republic using partial 28S rDNA sequences

Partial (D1-D3) 28S rRNA gene sequences from 16 isolates of digenean parasites of the family Dicrocoeliidae recovered from 16 bird species from the Czech Republic were used for phylogenetic reconstruction. Comparison with sequences available from GenBank suggests that the genus Brachylecithum is paraphyletic, requiring further validation and possible systematic revision. Although partial 28S rDNA is relatively conserved, analyses suggest that the following taxa are synonymous: Lutztrema attenuatum = L. monenteron = L. microstomum, Brachylecithum lobatum = B. glareoli. Zonorchis petiolatus is reassigned back to the genus Lyperosomum with L. collurionis as a junior synonym. The study revealed how complicated the systematics of the family Dicrocoeliidae is currently. The morphology of the group is variable, and the current distinguishing characters at species and even generic level are not sufficiently distinctive; it is difficult to identify the specimens correctly and identification of GenBank isolates is not reliable. Extensive sampling of isolates for both molecular and morphological studies is necessary to resolve the relationships within the family.

On the systematics of some marine haploporids (Trematoda) with the description of a new species of Megasolena Linton, 1910

Megasolena mikra sp. nov. is described from the queen angelfish, Holacanthus ciliaris (Linnaeus), off Florida, USA. The new species can be differentiated from all other species of Megasolena Linton, 1910 except Megasolena littoralis Muñoz, George-Nascimento, and Bray, 2017 in possessing testes that are smaller in diameter than the ovary. The new species can be differentiated from M. littoralis in lacking tegumental spines and possessing oral sucker papillae. Molecular data are provided for two species each of Cadenatella Dollfus, 1946, Hapladena Linton, 1910, and Megasolena Linton, 1910. Bayesian inference analysis of concatenated internal transcribed spacer region-2 (ITS2) and partial 28S rDNA sequences of 50 haploporoids revealed 1) a monophyletic Atractotrematidae Yamaguti, 1939 sister to the rest of the haploporoids tested; 2) a paraphyletic Megasoleninae Manter, 1935 – if Hapladena is included; and 3) a monophyletic Cadenatellinae Gibson and Bray, 1982 sister to the ‘mugilid’ haploporids. The ‘mugilid’ haploporids formed a monophyletic clade consisting of the subfamilies Chalcinotrematinae Overstreet and Curran, 2005, Forticulcitinae Blasco-Costa, Balbuena, Kostadinova, and Olson, 2009, Haploporinae Nicoll, 1914, and Waretrematinae Srivastava, 1937. Based on our analysis we restrict the Megasoleninae to include Megasolena, Vitellibaculum Montgomery, 1957, and Metamegasolena Yamaguti, 1970, all of which have species with two testes. To accommodate the former megasolenine taxa with a single testis, we erect the Hapladeninae subf. nov. for species in Hapladena and tentatively, Myodera Montgomery, 1957. Our results further support that haploporoids had a common marine ancestor with two testes, and that members of the Haploporoidea Nicoll, 1914 underwent diversification following a shift from a primarily marine life history with eupercarian hosts to a more euryhaline one with diadromous hosts (namely mullet).

Molecular survey of piroplasm species from selected areas of China and Pakistan

BackgroundPiroplasmosis is an important animal disease that is a major constraint to the development of the livestock industry, often resulting in significant economic losses. Therefore, there is an urgent need to further understand the etiology of this and other tick-borne infections.MethodsBlood samples were collected from tick-infested animals from the Chakwal, Jhang, and Faisalabad districts of Punjab, Pakistan and from peri-urban areas around Hohhot, Inner Mongolia, China to investigate the presence of Babesia and Theileria species. In total, 450 blood samples were collected with FTA cards from cattle of the study areas of Pakistan; the genomic (g)DNA of one hundred and twenty samples from cattle in Inner Mongolia were provided by the Lanzhou Veterinary Research Institute, China. Following the extraction of gDNA, the 18S rRNA gene (V4 hypervariable region) of piroplasms was amplified in all samples using semi-nested PCR. Positively identified samples were sequenced for the identification of Theileria and Babesia species. The partial full-length sequence of 18S rDNA was amplified for species confirmation of Theileria-positive samples, whereas the RAP-1c gene was amplified for Babesia bigemina-positive samples.ResultsSemi-nested PCR results revealed that 144 (25.26%) samples were positive for piroplasms. Theileria annulata was the most prevalent species (115/144; 20.17%), followed by Theileria orientalis (16/144; 2.80%). Among Babesia, the only species recorded was Babesia bigemina (13/144; 2.28%).ConclusionThe present study reveals new data on the prevalence of piroplasm species in bovine populations of selected areas of China and Pakistan and their phylogenetic relationships. It is also the first detailed report of T. orientalis from native animals in Pakistan.

Records of opecoeline species Pseudopecoelus cf. vulgaris and Anomalotrema koiae Gibson & Bray, 1984 (Trematoda, Opecoelidae, Opecoelinae) from fish of the North Pacific, with notes on the phylogeny of the family Opecoelidae.

Opecoelid species Pseudopecoelus cf. vulgaris and Anomalotrema koiae Gibson & Bray, 1984 were found in fish collected in the boreal waters of the North Pacific. Pseudopecoelus cf. vulgaris differs from Pseudopecoelus vulgaris (Manter, 1934) in terms of the egg size. This is the first record of A. koiae in the North Pacific, and the second of Pseudopecoelus cf. vulgaris. Previously, A. koiae was recorded only in North Atlantic fish. Partial sequences of 28S rDNA obtained for these two species and six other previously unsequenced representatives of the family Opecoelidae were analysed together with data from GenBank. Phylogenetic analysis supports the allocation of the six clades of opecoelids - Helicometrinae, Opecoelinae, Opistholebetinae, 'freshwater Plagioporinae', 'marine Plagioporinae B' and 'marine Plagioporinae C', and confirms the paraphyly of the group 'deep-sea Plagioporinae'. Our phylogeny does not support previous hypotheses about the monophyly of opecoelines with a uroproct.

Pseudomonas fluorescens group bacterial strains are responsible for repeat and sporadic postpasteurization contamination and reduced fluid milk shelf life

Postpasteurization contamination (PPC) of high temperature, short time-pasteurized fluid milk by gram-negative (GN) bacteria continues to be an issue for processors. To improve PPC control, a better understanding of PPC patterns in dairy processing facilities over time and across equipment is needed. We thus collected samples from 10 fluid milk processing facilities to (1) detect and characterize PPC patterns over time, (2) determine the efficacy of different media to detect PPC, and (3) characterize sensory defects associated with PPC. Specifically, we collected 280 samples of high temperature, short time-pasteurized milk representing different products (2%, skim, and chocolate) and different fillers over 4 samplings performed over 11 mo at each of the 10 facilities. Standard plate count (SPC) as well as total GN, coliform, and Enterobacteriaceae (EB) counts were performed upon receipt and after 7, 10, 14, 17, and 21 d of storage at 6°C. We used 16S rDNA sequencing to characterize representative bacterial isolates from (1) test days with SPC >20,000 cfu/mL and (2) all samples with presumptive GN, coliforms, or EB. Day-21 samples were also evaluated by a trained defect judging panel. By d 21, 226 samples had SPC >20,000 cfu/mL on at least 1 d of shelf life; GN bacteria were found in 132 of these 226 samples, indicating PPC. Crystal violet tetrazolium agar detected PPC with the greatest sensitivity. Spoilage due to PPC was predominantly associated with Pseudomonas (isolated from 101 of the 132 samples with PPC); coliforms and EB were found in 27 and 37 samples with spoilage due to PPC, respectively. Detection of Pseudomonas and Acinetobacter was associated with lower flavor scores; coagulated, fruity fermented, and unclean defects were more prevalent in d-21 samples with PPC. Repeat isolation of Pseudomonas fluorescens group strains with identical partial 16S rDNA sequence types was observed in 8 facilities. In several facilities, specific lines, products, or processing days were linked to repeat product contamination with Pseudomonas with identical sequence types. Our data show that PPC due to Pseudomonas remains a major challenge for fluid milk processors; the inability of coliform and EB tests to detect Pseudomonas may contribute to this. Our data also provide important initial insights into PPC patterns (e.g., line-specific contamination), supporting the importance of molecular subtyping methods for identification of PPC sources.

Description of a new species of the genus Anguillonema Fuchs, 1938 (Nematoda: Sphaerularioidea) with an identification key to the species.

Anguillonema iranicum n. sp. is described and illustrated as the second species of this genus from Iran, based on morphological, morphometric and molecular characteristics. It is identified by a short, thin body, a continuous lip region, six lines on the lateral field, a short, thin stylet, a posteriorly located pharyngo-intestinal junction to excretory pore, the presence of a post-vulval uterine sac, vulval position at 89% (87.4-89.9%) of body length, an elongate conoid tail with a rounded to pointed tip and not dorsally bent, and common functional males with short spicules and lacking a bursa. Morphological differences between the new species and the three known species of the genus, namely A. amolensis, A. crenati and A. poligraphi, are discussed. Molecular phylogenetic studies of the new species using partial 18S rDNA sequence revealed that it formed a sister clade with three species of Howardula, one species of Anguillonema and one unidentified isolate. In phylogenetic analyses using partial sequences of 28S rDNA D2-D3 segment, the new species formed a clade with two isolates of Parasitylenchus. A key to identification of Anguillonema species is also presented.

Myxidium shedkoae Sokolov, 2013 (Myxozoa: Myxidiidae), a parasite of the gallbladder of Perccottus glenii Dybowski, 1877 (Actinoptrygii: Odontobutidae): Supplementary data on morphology and phylogenetic position based on 18s rDNA sequence analysis.

This paper is the first report on the molecular characterisation of myxozoan parasites from the odontobutid fish Chinese (Amur) sleeper (Perccottus glenii Dybowski, 1877). The authors determined the partial 18S rDNA sequence of Myxidium shedkoae Sokolov, 2013 from the gallbladder of the fish. Phylogenies reconstructed using maximum likelihood and Bayesian inference analysis revealed that M. shedkoae belongs to the hepatic biliary group of myxozoans (after Kristmundsson and Freeman, 2013) as a member of the clade consisting of Zschokkella sp. KLT-2014, Myxidium truttae and Zschokkella nova. Some new morphological features of the parasite are also presented.

Potential anti-biofilm producing marine actinomycetes isolated from sea sediments in Thailand

Marine actinomycetes are well known as novel resources of bioactive compounds including anti-biofilm agents with various therapeutic applications. In this study, one hundred and one marine actinomycetes were isolated from sea sediments collected from Andaman sea and the Gulf of Thailand. Morphological study and partial 16S rDNA sequence analysis revealed that most of isolates belong to genus Streptomyces (n = 90, 89.11%) and the rest are rare actinomycetes (n = 11, 10.89%) including Actinopolymorpha, Actinomycetospora, Dietzia Nocardiopsis, Micromonospora and Mycobacterium. Among them, ten and thirteen strains significantly reduced more than 60% of biofilm formation of Escherichia coli and Staphylococcus aureus, respectively. But no such activity was found against Pseudomonas aeruginosa. Further analysis of their supernatants indicated that most of marine actinomycetes secreted non-toxic anti-biofilm agents and some accompanied with protease activity. These are preferred characteristics of anti-biofilm substances that potentially restricted biofilm forming of E. coli and S. aureus and simultaneously prevented bacteria to develop resistance.

Disseminated nontuberculous mycobacterial disease in a cat caused by Mycobacterium sp. strain MFM001

Mycobacterium sp. strain MFM001 (MFM001), a new strain of slow-growing nontuberculous mycobacteria not associated with the Mycobacterium avium complex, led to disseminated mycobacterial disease characterized by transmural granulomatous gastroenterocolitis of lepromatous type in a cat treated with immunosuppressive medication. MFM001 was found to be closely related to M. kyorinense or M. celatum by partial sequence of 16S rDNA, and identical with an unspecified M. sp. strain B10-07.09.0206 isolated from a human pulmonary infection in Germany. MFM001 should be considered as a pathogenic organism in cats, especially in immunocompromised animals.

Two new species and two new records of myxomycetes from subtropical forests in China

Two new myxomycetes from subtropical forests of China, Arcyria aeruginosa sp. nov. and Diderma subochraceum sp. nov., are described and illustrated. Arcyria aeruginosa occurred on rotten stem of bamboo and differs from all known species of the genus in having dense spines on inner surface of cups. The diagnostic feature of D. subochraceum is single-layered peridium which dehisces by longitudinal or stellate slit. Preliminary phylogenetic analyses based on partial 18S rDNA sequences support the establishment of the two new species. Additionally, Didymium saturnus H.W. Keller and Fuligo luteonitens L.G. Krieglst. & Nowotny are reported from China for the first time.

A rare and unusual new bittiine genus with two new species from the South Pacific (Cerithiidae, Gastropoda).

A new genus, Limatium gen. n., and two new species, L. pagodula sp. n. and L. aureum sp. n. are described, found on outer slopes of barrier reefs and fringing reefs in the South Pacific. They are rare for cerithiids, which typically occur in large populations. The two new species are represented by 108 specimens sampled over a period of 30 years, only 16 of which were collected alive. Three subadults from the Philippines and Vanuatu likely represent a third species. In addition to their rarity, Limatium species are atypical for cerithiids in their smooth, polished, honey to golden brown shells with distinctive white fascioles extending suture to suture. The radula presents a unique morphology that does not readily suggest an affinity to any of the cerithiid subfamilies. Two live-collected specimens, one of each species and designated as holotypes, were preserved in 95% ethanol and sequenced. Bayesian analysis of partial COI and 16S rDNA sequences demonstrates a placement in the Bittiinae, further extending our morphological concept of the subfamily.