Aureispira anguillae sp. nov., isolated from Japanese eel Anguilla japonica leptocephali.

A novel filamentous eel-leptocephalus pathogenic marine bacterium, designated strain EL160426T, was isolated from Japanese eel, Anguilla japonica, leptocephali reared at a laboratory in Mie, Japan. In experimental infection studies on eel larvae, the strain EL160426T caused massive larval mortality and was reisolated from moribund leptocephali. Characteristically, observations of infected larvae found that EL160426T forms columnar colonies on the cranial surface of larvae. The novel isolate exhibited growth at 15-30 °C, pH 7-9, and seawater concentrations of 60-150% (W/V). Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain EL160426T was most closely related to Aureispira maritima 59SAT with 97.7% sequence similarity. The whole genome sequence analysis of the strain EL160426T showed that the strain maintained a circular chromosome with a size of approximately 7.58 Mbp and the DNA G + C content was 36.2%. The major respiratory quinone was MK-7 and the predominant cellular fatty acids were 16:0, 20:4 w6c (arachidonic acid), 17:0 iso and 16:0 N alcohol. DNA relatedness between the closest phylogenetic neighbor strain EL160426T and A. maritima (JCM23207T) was less than 13%. On the basis of the polyphasic taxonomic data, the strain represents a novel species of the genus Aureispira, for which the name Aureispira anguillae sp. nov. is proposed. The type strain is EL160426T (= JCM 35024 T = TSD-286 T).

Gilvimarinus japonicus sp. nov., a cellulolytic and agarolytic marine bacterium isolated from coastal debris.

A cellulolytic and agarolytic bacterial strain, designated 12-2T, was isolated from a piece of cotton rope fragment washed ashore on a beach and was studied phenotypically, genotypically and phylogenetically. Analyses of 16S rRNA and gyrB gene sequences and DNA base composition suggested that the strain is a member of the genus Gilvimarinus. However, levels of 16S rRNA and gyrB gene sequence similarity between it and the type strains of Gilvimarinus species were no higher than 97.9 and 78.7 %, respectively, suggesting that the strain is distinct. Moreover, the results of DNA-DNA hybridization experiments and physiological characterization clearly differentiated the strain from its closest neighbours. The strain is therefore considered to represent a novel species of the genus Gilvimarinus, for which the name Gilvimarinus japonicus sp. nov. is proposed. The type strain is 12-2T (=NBRC 111987T=KCTC 52141T).

Sulfitobacter pacificus sp. nov., isolated from the red alga Pyropia yezoensis.

Three Gram-negative, aerobic, halophilic bacterial strains, SCM2-10(T), SCM-4, and 14C-6, were isolated from the algal medium of the red alga Pyropia yezoensis (previously classified as Porphyra yezoensis) grown in laboratory experiments. Phylogenetic analysis based on 16S rRNA gene sequences revealed that the organisms with high similarities to the three isolates are Sulfitobacter geojensis MM-124(T) (98.7%), Sulfitobacter noctilucicola NB-77(T) (98.7%), Sulfitobacter noctilucae NB-68(T) (98.6%), Sulfitobacter mediterraneus CH-B427(T) (97.6%), and Sulfitobacter porphyrae SCM-1(T) (97.6%), and that the three isolates belong to the genus Sulfitobacter, within the class Alphaproteobacteria. The DNA G+C contents of the three isolates were found to be in the range of 56.5-57.1 mol%. DNA-DNA hybridization experiments demonstrated that the genomic relatedness between strain SCM2-10(T) and type strains of other Sulfitobacter species was in the range of 6.2-27.1%. The predominant respiratory quinone of the three isolates was identified as ubiquinone-10. The dominant polar lipids in the three isolates were found to be phosphatidylcholine, phosphatidylethanolamine, phosphatidylglycerol, and an unidentified amino lipid. The major fatty acid in the three isolates is C(18:1)ω7c. Strain SCM2-10(T) demonstrated unique phenotypic characteristics, which differed from those of type strains of other Sulfitobacter species. Based on the phylogenetic, genetic, phenotypic, and chemotaxonomic data, we propose a novel species of the genus Sulfitobacter, which we named as Sulfitobacter pacificus sp. nov. The type strain of this species is strain SCM2-10(T) (=LMG 27113(T) = NBRC 109915(T)).

Occurrence, molecular characterization, and antimicrobial susceptibility of Aeromonas spp. in marine species of shrimps cultured at inland low salinity ponds.

We aimed to document the risk of Aeromonas spp. in marine shrimp species cultured in inland low salinity ponds in Thailand. In 14 of 18 shrimp samples retrieved from inland grow-up ponds, Aeromonas spp. were detected at ranges from 4667 to 1,500,000 CFU/g body weight. The phylogenetic tree constructed with the gyrB and cpn60 concatenated sequences indicated that the 87 isolates consisted of Aeromonas veronii (70%), Aeromonas aquariorum (18%), Aeromonas caviae (7%), Aeromonas jandaei (2%), and Aeromonas schubertii (2%). The potential virulence of the isolates was examined by phenotypic and PCR assays. Hemolytic activity and the extracellular activity of lipase, DNase, and gelatinase were observed in most isolates (94-99%). PCR revealed the presence of 9 genes related to virulence in the 87 isolates: act (75%), aer (74%), alt (30%), ast (1%), ascV (34%), aexT (24%), fla (92%), ela (34%), and lip (24%). The susceptibility profiles to 14 antimicrobial agents of isolates were typical for the genus, but resistance to cefotaxime, a third-generation cephalosporin, and imipenem were found in two A. aquariorum and in three A. veronii isolates, respectively. These resistances were confirmed by determining minimum inhibitory concentrations. Our results indicate that the microbiological risk posed by Aeromonas should be considered for marine shrimp species that are cultured in low-salinity ponds. These shrimps may also be a vehicle for the transfer of different genotypes of Aeromonas and antibiotic-resistant determinants to regions worldwide through trade.

Sulfitobacter porphyrae sp. nov., isolated from the red alga Porphyra yezoensis.

Gram-stain-negative, aerobic, halophilic bacteria, designated SCM-1(T), LCM10-1 and CTBL-B-147, were isolated from modified half-strength SWM-III medium, PES medium and thalli after laboratory cultivation of a red alga, Porphyra yezoensis. Phylogenetic analysis of 16S rRNA gene sequences indicated that the new isolates were affiliated to the genus Sulfitobacter of the class Alphaproteobacteria, and the 16S rRNA gene sequence similarity of the new isolates with the closest related species, Sulfitobacter mediterraneus CH-B427(T), was 98.8%. The DNA G+C contents of the new isolates were in the range of 61.4-62.3 mol%. DNA-DNA relatedness values of strain SCM-1(T) with other type strains of the genus Sulfitobacter were less than 15.9%. The new isolates contained Q-10 as the predominant ubiquinone, phosphatidylcholine, phosphatidylglycerol, an unidentified amino lipid and an unidentified lipid as the main polar lipids, and C(18 : 1)ω7c, C(19 : 1)ω7c and C(16 : 0) as the major fatty acids (>10% of the total). Strain SCM-1(T) could be differentiated from Sulfitobacter mediterraneus JCM 21792(T) by 35 morphological and phenotypic characteristics. On the basis of the phylogenetic, genetic and phenotypic properties of the new isolates, the name Sulfitobacter porphyrae sp. nov. is proposed, with strain SCM-1(T) ( = LMG 27110(T) = NBRC 109054(T)) as the type strain.

Isolation of hyphomonas strains that induce normal morphogenesis in protoplasts of the marine red alga Pyropia yezoensis.

Marine macroalgae cannot develop normal morphology under axenic conditions although normal morphogenesis can be sustained when certain bacteria are present. In this study, bacteria that induced normal morphogenesis in the red alga Pyropia yezoensis (Nori) were identified. The bacteria were isolated from algal media, thalli, tissue debris, and purified protoplasts during protoplast isolation from P. yezoensis laboratory cultures. 16S rRNA gene sequence analysis showed these bacterial isolates belonged to α-Proteobacteria (12 groups), γ-Proteobacteria (3 groups), and Flavobacteria (2 groups). Axenic protoplasts of P. yezoensis generated by removing epiphytic bacteria were co-cultured along with the bacterial isolates. Most axenic protoplasts showed irregular morphogenetic and anaplastic cells; cells with normal morphology were scarce. However, inoculation with 11 strains of Hyphomonas (α-Proteobacteria) led to significantly higher normal morphogenetic rates (4.5-7.3 %, P < 0.01 or 0.05) compared to axenic protoplasts (0.06 %). These Hyphomonas strains were recovered from all experiments; thus, certain Hyphomonas strains can induce normal morphogenesis in P. yezoensis protoplasts. Direct inoculation of the Hyphomonas strain exhibited higher morphogenetic activity than inoculation of its extracellular and intracellular products. This is the first study demonstrating the influence of specific bacteria on protoplast morphology in marine macroalgae.

Algimonas ampicilliniresistens sp. nov., isolated from the red alga Porphyra yezoensis, and emended description of the genus Algimonas.

Three strains (14A-2-7(T), 14A-3-1 and 14A-3) of Gram-stain-negative, prosthecate, motile bacteria were isolated from an algal medium supplemented with 10 mg ampicillin l(-1) (w/v), in which the red alga Porphyra yezoensis had been cultured. Based on the 16S rRNA gene sequence analysis, the three isolates formed a cluster with the genus Algimonas of the family Hyphomonadaceae. The sequences of the three isolates had high similarity with those of Algimonas porphyrae 0C-2-2(T) (97.6 % similarity) and Litorimonas taeanensis G5(T) (95.6 % similarity). The DNA G+C contents of the three isolates ranged from 54.3 to 55.0 mol%, which were more similar to that of A. porphyrae 0C-2-2(T) (58.5 mol%) than to that of L. taeanensis G5(T) (47.1 mol%). The DNA-DNA relatedness showed that the three isolates were representatives of the same species (88.1-94.0 % relatedness) and that strain 14A-2-7(T) was a representative of a different species from A. porphyrae 0C-2-2(T) and L. taeanensis G5(T) (1.2-8.6 % relatedness). The phenotypic characteristics of strain 14A-2-7(T) differed by 20 results and 30 results from A. porphyrae 0C-2-2(T) and L. taeanensis G5(T), respectively. The three isolates contained ubiquinone-10 as the predominant quinone and C18 : 1ω7c as the major fatty acid. Based on the polyphasic taxonomic analysis, the three isolates represent a novel species of the genus Algimonas, for which the name Algimonas ampicilliniresistens sp. nov. is proposed. The type strain is 14A-2-7(T) ( = LMG 26421(T) = NBRC 108219(T)). An emended description of the genus Algimonas is also proposed.

Streptomyces abietis sp. nov., a cellulolytic bacterium isolated from soil of a pine forest.

Cellulolytic bacteria A191(T), A192 and A193 isolated from the soil of Sakhalin fir forest in Hokkaido, Japan were studied phenotypically, genotypically and phylogenetically. Analysis of their 16S rRNA gene and gyrB sequences and DNA base composition suggested that these isolates were conspecific and members of the genus Streptomyces. However, levels of 16S rRNA gene and gyrB sequence similarity between the isolates and the type strains of their closest relatives in the genus Streptomyces were no higher than 97.9 and 95.0 %, respectively, implying that these isolates were distinctive. Moreover, the results of DNA-DNA hybridization experiments and physiological characterization clearly differentiated these isolates from their closest neighbours. It is therefore concluded that these isolates represent a novel species of the genus Streptomyces, for which the name Streptomyces abietis is proposed. The type strain is A191(T) ( = NBRC 109094(T) = DSM 42080(T)).

Algimonas porphyrae gen. nov., sp. nov., a member of the family Hyphomonadaceae, isolated from the red alga Porphyra yezoensis.

Three Gram-negative, stalked, motile bacteria, designated 0C-2-2(T), 0C-17 and LNM-3, were isolated from the red alga Porphyra yezoensis. 16S rRNA gene sequence analysis revealed that the three novel strains belonged to the family Hyphomonadaceae, and were closely related to Litorimonas taeanensis G5(T) (96.5 % 16S rRNA gene sequence similarity) and Hellea balneolensis 26III/A02/215(T) (94.3 %). The DNA G+C contents of the novel isolates (58.5-60.2 mol%) were clearly distinguished from those of L. taeanensis G5(T) (47.1 mol%) and H. balneolensis DSM 19091(T) (47.9 mol%). The G+C content of L. taeanensis G5(T) obtained in this study was quite different from a previous report (63.6 mol%). DNA-DNA hybridization experiments showed that the novel strains constituted a single species. Eleven phenotypic features of the three isolates differed from those of both related genera. The predominant respiratory quinone was ubiquinone-10 and the major fatty acid was C(18 : 1)ω7c. On the basis of this polyphasic taxonomic analysis, the novel strains represent a novel genus and species, for which the name Algimonas porphyrae gen. nov., sp. nov. is proposed. The type strain of Algimonas porphyrae is 0C-2-2(T) (= LMG 26424(T) = NBRC 108216(T)).

Polaribacter porphyrae sp. nov., isolated from the red alga Porphyra yezoensis, and emended descriptions of the genus Polaribacter and two Polaribacter species.

Three Gram-negative, non-motile, strictly aerobic strains, designated LNM-20(T), LCM-1 and LAM-13, were isolated from thalli of the marine red alga Porphyra yezoensis. Phylogenetic analysis based on 16S rRNA gene sequences revealed that the isolates were associated with the genus Polaribacter in the family Flavobacteriaceae and were most closely related to Polaribacter dokdonensis DSW-5(T) (96.2 % 16S rRNA gene sequence similarity) and Polaribacter gangjinensis K17-16(T) (95.0 %). The DNA G+C content of the isolates was 28.6-29.2 mol%. DNA-DNA hybridization analysis showed that the isolates belonged to a single species distinct from both of their closest relatives. The only isoprenoid quinone detected was menaquinone-6. The main polar lipids were phosphatidylethanolamine, two unidentified aminolipids and two unidentified lipids. The major fatty acids were iso-C15 : 0, iso-C15 : 1ω10c and iso-C15 : 0 3-OH. The phenotypic features of strain LNM-20(T) differed from those of their closest relatives in several regards (colony colour, growth with 1 % NaCl and on TSA plus 2.5 % NaCl, hydrolysis of Tweens 40 and 80, and oxidization of five carbon compounds). On the basis of phylogenetic, chemotaxonomic and phenotypic analysis, the isolates represent a novel species in the genus Polaribacter, for which the name Polaribacter porphyrae sp. nov. is proposed. The type strain is LNM-20(T) ( = LMG 26671(T)  = NBRC 108759(T)). Emended descriptions of the genus Polaribacter and P. dokdonensis and P. gangjinensis are also proposed.

Maritalea porphyrae sp. nov., isolated from a red alga (Porphyra yezoensis), and transfer of Zhangella mobilis to Maritalea mobilis comb. nov.

Three Gram-negative, motile, aerobic bacteria were isolated from cultures of the marine red alga Porphyra yezoensis. Phylogenetic analysis based on 16S rRNA gene sequences revealed that the novel strains were closely related to Maritalea myrionectae CL-SK30(T) (97.9% 16S rRNA gene sequence similarity) and Zhangella mobilis E6(T) (96.2 %). 16S rRNA gene sequence similarity between Z. mobilis E6(T) and M. myrionectae CL-SK30(T) was 97.9%. The DNA G+C contents of the isolates (49.4-50.0 mol%) were similar to those of M. myrionectae DSM 19524(T) (52.3 mol%) and Z. mobilis JCM 15144(T) (50.3 mol%). From these results, it was difficult to differentiate the genus Zhangella from the genus Maritalea. DNA-DNA hybridization demonstrated that the isolates belonged to a single species. The isolates could also be distinguished from M. myrionectae and Z. mobilis on the basis of chemotaxonomic and phenotypic features, including fatty acid composition (particularly C(16:1)ω7c), growth with 6-9% (w/v) NaCl, carbon utilization, oxidation patterns and so on. A novel species of the genus Maritalea is proposed to accommodate the three isolates, with the name Maritalea porphyrae sp. nov. The type strain is LCM-3(T) (=LMG 25872(T)=NBRC 107169(T)). Furthermore, it is proposed that Zhangella mobilis should be transferred from the genus Zhangella to the genus Maritalea, with the name Maritalea mobilis comb. nov. (type strain E6(T)=CGMCC 1.7002(T)=JCM 15144(T)).

Reduction of lipids in fish meal prepared from fish waste by a yeast Yarrowia lipolytica.

The high lipid content in fish waste is one of the reasons why fish meal made from fish waste is commercially rated as low grade meal. Microorganisms which have the ability to reduce crude lipids from samples of minced fish in solid-state fermentation were screened and a strain of Yarrowia lipolytica showed the highest efficiency for reducing the lipids by 29%. The lipid reduction by this strain was especially affected by the ratio of surface area to the weight in the fermented mince samples and by the water content, suggesting the importance of the oxygen supply. In the fermentation with intermittent mixing during 96 h incubation, reduction efficiency for crude lipids came to 46% but that for protein was less than 1%. With the fermentation, 41.5 g of crude lipids in 1 kg of the minces were estimated to be reduced to 22.4 g, indicating increase of protein content in the final product. Furthermore, the carbonyl value which is an indicator of lipid oxidation was relatively suppressed by the fermentation. These results suggest that the fermentation can improve the quality of fish meal from fish waste which is rich of lipids.

Analysis of a 30 kbp plasmid encoding histidine decarboxylase gene in Tetragenococcus halophilus isolated from fish sauce.

In order to analyze the genes related to the histamine production, a strain of histamine producing halophilic bacteria, referred to as strain H, was isolated using enrichment culture and dilution-to-extinction methods with histidine broth inoculated from the fish sauce mashes. The two Japanese fish sauce mashes used, accumulate over 1000 mg/l of histamine. Phenotypic and 16 S rRNA gene sequence analyses identified strain H as Tetragenococcus halophilus, the predominant histamine producing bacteria present during fish sauce fermentation. Genetic analyses (PCR and Southern blot) of the histamine producing strain confirmed that the strain harbored a 30 kbp plasmid (pHDC) encoding a single copy of the pyruvoyl dependent histidine decarboxylase gene (hdc). A comparison of hdcA that is a structural gene of histidine decarboxylase among strain H, Lactobacillus hilgardii 0006, L. sakei LTH2076, Oenococcus oeni 9204, T. halophilus and T. muriaticus JCM10006 (T) indicated >99% sequence similarity. The hdc gene cluster consisted of 4 ORFs, hdcP, hdcA, hdcB, and hdcRS, and were almost identical to that of L. hilgardii 0006 with 99% sequence similarity including the structural hdc spacer region. However, the approximately 500 bp regions upstream and downstream of the hdc gene were different between that of strain H and L. hilgardii 0006. The complete sequence of pHDC revealed 29,924 nucleotides including 28 ORFs, two pairs of IR (inverted repeat), similar sequence of plasmid conjugative elements, and a theta-type replicon. These results suggested that hdc could be encoded on transformable elements among lactic acid bacteria.

Non-Toxin-Producing Bacillus cereus Strains Belonging to the B. anthracis Clade Isolated from the International Space Station.

In an ongoing Microbial Observatory investigation of the International Space Station (ISS), 11 Bacillus strains (2 from the Kibo Japanese experimental module, 4 from the U.S. segment, and 5 from the Russian module) were isolated and their whole genomes were sequenced. A comparative analysis of the 16S rRNA gene sequences of these isolates showed the highest similarity (>99%) to the Bacillus anthracis-B. cereus-B. thuringiensis group. The fatty acid composition, polar lipid profile, peptidoglycan type, and matrix-assisted laser desorption ionization-time of flight profiles were consistent with the B. cereus sensu lato group. The phenotypic traits such as motile rods, enterotoxin production, lack of capsule, and resistance to gamma phage/penicillin observed in ISS isolates were not characteristics of B. anthracis. Whole-genome sequence characterizations showed that ISS strains had the plcR non-B. anthracis ancestral "C" allele and lacked anthrax toxin-encoding plasmids pXO1 and pXO2, excluding their identification as B. anthracis. The genetic identities of all 11 ISS isolates characterized via gyrB analyses arbitrarily identified them as members of the B. cereus group, but traditional DNA-DNA hybridization (DDH) showed that the ISS isolates are similar to B. anthracis (88% to 90%) but distant from the B. cereus (42%) and B. thuringiensis (48%) type strains. The DDH results were supported by average nucleotide identity (>98.5%) and digital DDH (>86%) analyses. However, the collective phenotypic traits and genomic evidence were the reasons to exclude the ISS isolates from B. anthracis. Nevertheless, multilocus sequence typing and whole-genome single nucleotide polymorphism analyses placed these isolates in a clade that is distinct from previously described members of the B. cereus sensu lato group but closely related to B. anthracis. IMPORTANCE The International Space Station Microbial Observatory (Microbial Tracking-1) study is generating a microbial census of the space station's surfaces and atmosphere by using advanced molecular microbial community analysis techniques supported by traditional culture-based methods and modern bioinformatic computational modeling. This approach will lead to long-term, multigenerational studies of microbial population dynamics in a closed environment and address key questions, including whether microgravity influences the evolution and genetic modification of microorganisms. The spore-forming Bacillus cereus sensu lato group consists of pathogenic (B. anthracis), food poisoning (B. cereus), and biotechnologically useful (B. thuringiensis) microorganisms; their presence in a closed system such as the ISS might be a concern for the health of crew members. A detailed characterization of these potential pathogens would lead to the development of suitable countermeasures that are needed for long-term future missions and a better understanding of microorganisms associated with space missions.

Antimicrobial effect of spices and herbs on Vibrio parahaemolyticus.

The antimicrobial effects of spices and herbs from 18 plant species were examined on a foodborne pathogen, Vibrio parahaemolyticus, with the use of combinations of temperatures and nutrient levels. Basil, clove, garlic, horseradish, marjoram, oregano, rosemary, and thyme exhibited antibacterial activities at incubation of 30 degrees C, while with the exception of horseradish, the same spices and additional 7 species exhibited the activities at 5 degrees C. The lowest MIC (minimum inhibitory concentration) was 0.125% observed in clove and marjoram at 30 degrees C in a nutrient rich medium. Lowering of incubation temperature produced little effect on the MICs except for turmeric. The decreasing of the MIC in turmeric appeared to be basically attributed to the sensitivity of the bacterium to coldness. In nutrient poor medium, the lowest was 0.001 and 0.00025% in marjoram at 30 degrees C and at 5 degrees C, respectively. The sensitivity to several spices and herbs was similar among different clinical serotypes including the emerging strain O3:K6. These results suggest that the spices and herbs can be practical for protecting seafood from the risk of contamination by V. parahaemolyticus and used in hurdle technology with low temperature.

Occurrence and density of vibrio parahaemolyticus in live edible crustaceans from markets in China.

Vibrio parahaemolyticus is a marine bacterium causing foodborne disease. Occurrence of the bacterium was investigated in six species of edible crustaceans available from markets in mainland China. The bacterium was detected in 22 of 45 whole-body, shell, and feces samples, including mitten crabs, which are supposed to be produced in freshwater ponds. The mean densities ranged from 2.8 log CFU/g in mitten crabs (Eriocheir sinensis) to 5.1 CFU/g in giant tiger prawns (Penaeus monodon). In hemolymph and muscle samples collected axenically, V. parahaemolyticus was detected in all of the prawns at a mean density of 2.6 log most probable number (MPN)/g, in two of five striped stone crabs (Charybdis feriatus) at a mean density of 1.1 log MPN/ml, and two of five mangrove mud crabs (Scylla serrata) at a mean density of 1.3 log MPN/ml. When six mitten crabs were collected from two freshwater ponds in China and were examined, V. parahaemolyticus was not detected. It seemed that cross-contamination occurred among live crustaceans at the markets. The results suggest that proper handling, storage, and cooking of these crustaceans will be necessary to lessen the risk of foodborne illness from V. parahaemolyticus.

Accumulation and depuration rates of paralytic shellfish poisoning toxins in the shore crab Telmessus acutidens by feeding toxic mussels under laboratory controlled conditions.

Accumulation and depuration rates of paralytic shellfish poisoning toxins (PSP) in the crab Telmessus acutidens were investigated by feeding toxic and non-toxic mussels under laboratory controlled conditions. The crab accumulated toxins in the hepatopancreas in proportion to the amount of toxic mussels they ingested, and the toxicity in the crab hepatopancreas became 3.2 fold of that in the prey mussels after 20 days of feeding. During depuration, a fast reduction of the total toxicity was observed in the crab, and the retention rate of the toxicity after 5 days depuration with feeding of non-toxic mussels was 45.8+/-18.7%. The reduction of the toxicity was moderated in the later period of depuration, and the retention rates of the total toxicity after 10 and 20 days were 54.1+/-29.8% and 14.5+/-9.0%, respectively. The toxin profiles in the crab and mussel were investigated by high performance liquid chromatography, and reductive conversions of the toxins were observed when the toxins were transferred from the mussel to the crab. Consequently, high concentrations of GTX2 and GTX3, and STX that were not detected in the prey mussels, were found in the crab.

Accumulation of paralytic shellfish poisoning toxins in the edible shore crab Telmessus acutidens.

Several shellfishes including the crab Telmessus acutidens and its prey bivalve Mytilus galloprovincialis were collected at Onahama in Japan to investigate the accumulation of paralytic shellfish poisoning (PSP) toxins during the blooming season of toxic dinoflagellates. The toxicity of the viscera of T. acutidens collected in 1999 was 30.0 and 80.0 MU/g, and that of M. galloprovincialis was 9.6 MU/g by mouse bioassay. PSP toxins in the crab viscera were identified by HPLC-FLD and ESI-MS, so this is the first observation of PSP toxins in T. acutidens. Carbamate toxins (GTX1-4, and STX) were the major component in the crab as well as in the mussels, and accounted for over 60% on a molar basis. However, the ratio of the N1-OH toxin to N1-H toxin of the crabs were largely different from that of the mussels, and a reductive conversion of the toxins in T. acutidens is concluded as the probable cause. In 2000, PSP toxins were also detected in both crabs and mussels, though the contents were very low compared with the levels observed in 1999. The difference in the toxin abundance suggests that the toxin content in the crab was affected by those of the prey.

Comparison of paralytic shellfish poisoning toxin between carnivorous crabs (Telmessus acutidens and Charybdis japonica) and their prey mussel (Mytilus galloprovincialis) in an inshore food chain.

Paralytic shellfish poisoning toxin in two shore crab species, Telmessus acutidens and Charybdis japonica, were compared with the toxin in the prey mussel Mytilus galloprovincialis and causative dinoflagellates Alexandrium tamarense, all having been collected at Onahama, Fukushima Prefecture, in the northern part of Japan. When the toxicities were detected in mussels by mouse bioassays, 73.7% of the sampled T. acutidens were toxic in the hepatopancreas. T. acutidens has been found to become toxic for three years, therefore, it can be concluded that the crab commonly and repeatedly accumulate the toxins via the food chain at Onahama. C. japonica was also expected to be a possible vector species, because small quantities of the toxins were detected in eight specimens of the crab by HPLC analysis. By the comparison of the toxin profiles in the dinoflagellates, mussels and the crab T. acutidens, reductive conversions of GTX1 and GTX4 were observed when the toxins passed through the three species in the food chain. But increases of STX and neoSTX by further reductive process were not observed in the crab. The absence of the STX group toxins in the crab suggests that the crab eliminates the toxin before such reductive process occur.

gyrB as a phylogenetic discriminator for members of the Bacillus anthracis-cereus-thuringiensis group.

Bacillus anthracis, the causative agent of the human disease anthrax, Bacillus cereus, a food-borne pathogen capable of causing human illness, and Bacillus thuringiensis, a well-characterized insecticidal toxin producer, all cluster together within a very tight clade (B. cereus group) phylogenetically and are indistinguishable from one another via 16S rDNA sequence analysis. As new pathogens are continually emerging, it is imperative to devise a system capable of rapidly and accurately differentiating closely related, yet phenotypically distinct species. Although the gyrB gene has proven useful in discriminating closely related species, its sequence analysis has not yet been validated by DNA:DNA hybridization, the taxonomically accepted "gold standard". We phylogenetically characterized the gyrB sequences of various species and serotypes encompassed in the "B. cereus group," including lab strains and environmental isolates. Results were compared to those obtained from analyses of phenotypic characteristics, 16S rDNA sequence, DNA:DNA hybridization, and virulence factors. The gyrB gene proved more highly differential than 16S, while, at the same time, as analytical as costly and laborious DNA:DNA hybridization techniques in differentiating species within the B. cereus group.