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As an intestinal organism settled long-term within the gut of marine fish, Vibrio scophthalmi is a potential object for the bacterium genetic variation and adaptation research. The genetic diversity, antimicrobial resistance phenotype, and genotype of 33 V. scophthalmi isolated from diseased marine fish intestines between 2002 and 2020 were evaluated. The results showed that all isolates were frequently resistant to penicillins, cephalosporins, aminoglycosides, and macrolides and displayed multidrug-resistant (MDR) phenotype in vitro. Thirty percent of isolates were resistant to more than 20 different drugs. The average insensitive (resistant and intermediate) rate of V. scophthalmi isolates was 49.5%~81.8% between 2002 and 2020, but the t-test revealed that there was no significant difference in the drug-resistance rate of V. scophthalmi isolates with typical interannual variability. Eleven antimicrobial resistance genes (strB, strA, ant(3 ˝)-I, mphA, blaPSE, qnrS, tetC, tetE, tetM, tetS, and int1) were detected in these isolates, but the antimicrobial resistance phenotypes and genotypes of these isolates were not consistent. Enterobacterial repetitive intergenic consensus polymerase chain reaction (ERIC-PCR) analysis indicated that 33 isolates could be divided into two clusters (G1 and G3) and two single isolates (G2 and G4), and the G2 cluster was isolated from South Sea C. undulates with typical geographical species differences. There was no significant correlation between the drug susceptibility and the genetic types of V. scophthalmi isolates. The results reveal the mismatch phenomenon between antimicrobial resistance and genotype of inherent V. scophthalmi in the marine fish intestines, and the antimicrobial susceptibility isolates might be a potential risk source for storage and transmission of resistance genes.

Melanochlamys aquilina sp. nov. (Figure 2) http://zoobank.org/ urn:lsid:zoobank.org:act: 482066FA-C7B3-4855-A945-57989C50DC14 Type specimens. Holotype: MBM285089 (body length 13.5 mm), from the type locality, December 2019; Paratypes: MBM285090, two specimens, collected with the holotype at the type locality, December 2019. Type locality. A sea cucumber culture pond on the coast of Laizhou Bay, Shandong Province, 37°36′N, 118°57′E. Distribution and habitat. Only known from the type locality. Intestinal tracts with shells of Cylichnatys angusta (Gould, 1859). Etymology. The name is derived from the Latin aquilinus, meaning ‘of the eagle’, referring to the eagle-like distal end of the penis. Description. External morphology (Fig. 2A). Living animals up to 14.5 mm in length. Body elongate, nearly cylindrical. Cephalic shield occupying about half of the length of the body; anterior end typically rounded and narrow, posterior end widening and truncated. Posterior shield elongate, ending in two short and wide caudal lobes of similar size, separated by a U-shaped notch. Parapodia developed, running along almost the entire length of the animal, and folded up to the edge of the dorsum. No eyes are visible externally. The genital opening located near the posterior end of the parapodia on the right side of the body. Animal dark blue to black in color. Dorsal surface of the cephalic shield with a central, longitudinal line with darker pigment. Shell (Figs 2B, C) brownish in color, well-calcified but thin, fragile (only upper part could be dissected). Shell surface sculptured with weak irregular growth lines. Spire small, with a rounded apex. Inner lip with a thin, straight posterior projection which is separated from subsequent whorl by a large indentation (see Fig. 2B, arrow). Body whorl expanded, the area near apical margin with prominent external carina (Fig. 2C). Posterior process elongate and curved, directed backward and separated from the spire by a deep notch. Male reproductive anatomy (Figs 2D, E). Prostate elongate, simple, of equal width along entire length. Width of prostate variable in the specimens dissected. Prostate attached to a tissue mass, where the penis, the spermatic bulb and the excurrent sperm groove connected. Penis elongated, with a cuticularized penial spine on the tip. Penial spine slightly curved, with a broad proximal base that abruptly narrows into a pointed distal tip. Spermatic bulb consisting of a convoluted duct ending into a wider vesicle at its distal end and having a retractor muscle attached to it. Size of spermatic bulb variable between studied individuals. Remarks. Melanochlamys aquilina sp. nov. is characterized by having a uniform, dark external coloration with a dark longitudinal line along the central part of the cephalic shield. Externally, the new species is most similar to Melanochlamys fukudai from Japan. That latter, however, anatomically differs from the new species in having an expanded, straight distal end of the prostate, and a much less developed excurrent sperm groove. In addition, Melanochlamys fukudai differs from Melanochlamys aquilina sp. nov. in having the inner lip of shell equipped with a thick, involuted posterior projection, and a shorter posterior process that is separated from the spire by a shallower notch. Phylogenetic analyses. The two consensus trees inferred using BI and ML criteria were generally congruent (Fig. 3). The analyses based on concatenated dataset (COI, 16S and H3) showed that Melanochlamys aquilina sp. nov. clustered together with the other Melanochlamys spp. forming a well-supported clade (PP=0.96 BS=82), sister to Melanochlamys fukudai (PP=1, BS=99). The phylogenetic tree inferred using BI criteria based on single gene COI (Suppl. Fig. 1) shows similar overall topology. Melanochlamys aquilina sp. nov. sequences were recovered as a well-supported lineage (PP =1) clearly distinct from other congeners. The genus Melanochlamys, including the new species, was recovered as monophyletic with full support (PP=1). These results support the systematic placement of the new species in the genus Melanochlamys and its separation from other congeners. Species delimitation analyses. The ABGD analysis of the COI sequences resulted in the delimitation of six species from the North Pacific (M. aquilina, M. diomedea, M. ezoensis, M. fukudai, M. kohi, M. chabanae), with values of prior intraspecific divergence (P) being ≥ 0.0129. These groups correspond to clades recovered by BI and ML analyses (Fig. 3). With available molecular data, the analysis of a 579-bp fragment of the COI gene yielded a 7.8%–8.7% pairwise distance between Melanochlamys aquilina sp. nov. and M. fukudai, a divergence much higher than the known intraspecific variation of Melanochlamys spp. (0–2.5%). The pairwise distance between Melanochlamys aquilina sp. nov. and other congeners ranged from 17.0% –24.0%. These results provide additional support for the separation of Melanochlamys aquilina sp. nov. from its congeners. ......continued on the next page

Genus Melanochlamys Cheeseman, 1881 Type species: Melanochlamys cylindrica Cheeseman, 1881, by original designation. Auckland harbour, New Zealand., Published as part of Zhang, Shuqian, Liao, Meijie, Wang, Yingeng, Kong, Miao & Li, Bin, 2020, Morphological and molecular evidence of a new species of Melanochlamys (Gastropoda: Heterobranchia) from the Bohai Sea, China, pp. 399-410 in Zootaxa 4861 (3) on page 401, DOI: 10.11646/zootaxa.4861.3.6, http://zenodo.org/record/4428611, {"references":["Cheeseman, T. F. (1881) On a new genus of opisthobranchiate Mollusca. Transactions of the New Zealand Institute, 13, 224."]}