Difference in sulfur regulation mechanism between tube-dwelling and free-moving polychaetes sympatrically inhabiting deep-sea hydrothermal chimneys.

The environment around deep sea hydrothermal vents is characterized by an abundance of sulfur compounds, including toxic hydrogen sulfide. However, numerous communities of various invertebrates are found in it. It is suggested that invertebrates in the vicinity of hydrothermal vents detoxify sulfur compounds by biosynthesis of taurine-related compounds in the body. On the other hand, the vent endemic polychaete Alvinella pompejana has spherocrystals composed of sulfur and other metals in its digestive tract. It was considered that the spherocrystals contribute to the regulation of sulfur in body fluids. Paralvinella spp. and Polynoidae. gen. sp. live sympatrically and in areas most affected by vent fluid. In this study, we focused on the digestive tract of Paralvinella spp. and Polynoidae. gen. sp. to examine whether they have spherocrystals. We also investigated the possible involvement of bacteria in the digestive tract in spherulization. Examination with a scanning electron microscope (SEM) equipped with Energy Disperse X-ray Spectroscopy (EDS) detected spherocrystals containing sulfur and iron in the digestive tract of Paralvinella spp. In contrast, such spherocrystals were not observed in that of Polynoidae. gen. sp. although sulfur is detected there by inductively coupled plasma-optical emission spectrometry (ICP-OES). Meta-16S rRNA analysis indicated that the floras of the digestive tracts of the two species were very similar, suggesting that enteric bacteria are not responsible for spherocrystal formation. Analysis of taurine-related compounds indicated that the digestive tissues of Polynoidae. gen. sp. contain a higher amount of hypotaurine and thiotaurine than those of Paralvinella spp. Therefore, the two sympatric polychaetes use different strategies for controlling sulfur, i.e., Paralvinella spp. forms spherocrystals containing elemental sulfur and iron in the digestive tract, but Polynoidae. gen. sp. accumulates taurine-related compounds instead of spherocrystals. Such differences may be related to differences in their lifestyles, i.e., burrow-dweller or free-moving, or may have been acquired phylogenetically in the evolutionary process.

Development of an immunochromatographic test for the detection of Mycoplasma pneumoniae GroES antigen.

Mycoplasma pneumoniae frequently causes community-acquired pneumonia in children; ß-lactam antibiotics are ineffective against this bacterium because of its lack of a cell wall. Hence, a rapid and simple detection method is required to ensure appropriate treatment. In this study, we developed a rapid and simple immunochromatography-based detection method using monoclonal antibodies that react with the co-chaperone GroES of M. pneumoniae. Mice were immunized with recombinant GroES, and hybridoma cells producing anti-GroES monoclonal antibodies were established. For the development of the immunochromatographic test, antibody pairs with superior reactivity and specificity were selected. The developed immunochromatographic test could detect 0.1 ng/mL of recombinant GroES within 20 min. Moreover, no cross-reaction was observed with other microorganisms, including six Mycoplasma species, 20 other bacterial species, and one yeast species. Macrolide-resistant and -susceptible M. pneumoniae clinical isolates were detected at approximately 104 to 105 colony-forming units/mL. The study indicates that immunochromatographic tests targeting GroES are useful for rapid and simple detection of M. pneumoniae.

Identification of cells expressing two peptidoglycan recognition proteins in the gill of the vent mussel, Bathymodiolus septemdierum.

In symbiotic systems in which symbionts are transmitted horizontally, hosts must accept symbionts from the environment while defending themselves against invading pathogenic microorganisms. How they distinguish pathogens from symbionts and how the latter evade host immune defences are not clearly understood. Recognition of foreign materials is one of the most critical steps in stimulating immune responses, and pattern recognition receptors (PRRs) play vital roles in this process. In this study, we focused on a group of highly conserved PRRs, peptidoglycan recognition proteins (PGRPs), in the deep-sea mussel, Bathymodiolus septemdierum, which harbours chemosynthetic bacteria in their gill epithelial cells. We isolated B. septemdierum PGRP genes BsPGRP-S and BsPGRP-L, which encode a short- and a long-type PGRP, respectively. The short-type PGRP has a signal peptide and was expressed in the asymbiotic goblet mucous cells in the gill epithelium, whereas the long-type PGRP was predicted to include a transmembrane domain and was expressed in gill bacteriocytes. Based on these findings, we hypothesize that the secreted and transmembrane PGRPs are engaged in host defence against pathogenic bacteria and/or in the regulation of symbiosis via different cellular localizations and mechanisms.

Long-term Cultivation of the Deep-Sea Clam Calyptogena okutanii: Changes in the Abundance of Chemoautotrophic Symbiont, Elemental Sulfur, and Mucus.

Survival of deep-sea Calyptogena clams depends on organic carbon produced by symbiotic, sulfur-oxidizing, autotrophic bacteria present in the epithelial cells of the gill. To understand the mechanism underlying this symbiosis, the development of a long-term cultivation system is essential. We cultivated specimens of Calyptogena okutanii in an artificial chemosynthetic aquarium with a hydrogen sulfide (H2S) supply system provided by the sulfate reduction of dog food buried in the sediment. We studied morphological and histochemical changes in the clams' gills by immunohistochemical and energy-dispersive X-ray analyses. The freshly collected clams contained a high amount of elemental sulfur in the gill epithelial cells, as well as densely packed symbiotic bacteria. Neither elemental sulfur nor symbiotic bacteria was detected in any other organs except the ovaries, where symbiotic bacteria, but not sulfur, was detected. The longest survival of an individual clam in this aquarium was 151 days. In the 3 clams dissected on Days 57 and 91 of the experiment, no elemental sulfur was detected in the gills. The symbiotic bacteria content had significantly decreased by Day 57, and was absent by Day 91. For comparison, we also studied the deep-sea mussel Bathymodiolus septemdierum, which harbors a phylogenetically close, sulfur-oxidizing, symbiotic bacterium with similar sulfur oxidation pathways. Sulfur particles were not detected, even in the gills of the freshly collected mussels. We discuss the importance of the proportion of available H2S and oxygen to the bivalves for elemental sulfur accumulation. Storage of nontoxic elemental sulfur, an energy source, seems to be an adaptive strategy of C. okutanii.

Effects of Hemagglutination Activity in the Serum of a Deep-Sea Vent Endemic Crab, Shinkaia Crosnieri, on Non-Symbiotic and Symbiotic Bacteria.

In deep-sea hydrothermal environments, most invertebrates associate with dense populations of symbiotic microorganisms in order to obtain nutrition. The molecular interactions between deep-sea animals and environmental microbes, including their symbionts, have not yet been elucidated in detail. Hemagglutinins/lectins, which are carbohydrate-binding proteins, have recently been reported to play important roles in a wide array of biological processes, including the recognition and control of non-self materials. We herein assessed hemagglutination activity in the serum of a deep-sea vent endemic crab, Shinkaia crosnieri, which harbors chemosynthetic epibionts on its plumose setae. Horse and rabbit erythrocytes were agglutinated using this serum (opt. pH 7.5 and opt. temperature 15°C). Agglutinating activity was inhibited by eight kinds of sugars and several divalent cations, did not require any divalent metal ions, and remained detectable even after heating the serum at 100°C for 30 min. By using fluorescently labeled serum, we demonstrated that deep-sea crab serum components bound to the epibionts even in the presence of sugars. This study represents the first immunological assessment of a deep-sea vent endemic crab and demonstrated the possibility of a non-lectin-mediated symbiont-host interaction.

Macrolide antibiotic-mediated downregulation of MexAB-OprM efflux pump expression in Pseudomonas aeruginosa.

Macrolide antibiotics modulate the quorum-sensing system of Pseudomonas aeruginosa. We tested the effect of macrolide antibiotics on the cell density-dependent expression of the MexAB-OprM efflux pump and found that 1.0 mug/ml (MIC/6.25) of azithromycin suppressed the expression of MexAB-OprM by about 70%, with the result that the cells became two- to fourfold more susceptible to antibiotics such as aztreonam, tetracycline, carbenicillin, chloramphenicol, and novobiocin.