Isolation and characterization of low nucleic acid (LNA)-content bacteria

Most planktonic bacteria are 'uncultivable' with conventional methods. Flow cytometry (FCM) is one approach that has been taken to study these bacteria. In natural aquatic environments, bacteria with high nucleic acid (HNA) and low nucleic acid (LNA) content are commonly observed with FCM after staining with fluorescent dyes. Although several studies have focused on the relative abundance and in situ activities of these two groups, knowledge on the growth of particularly LNA bacteria is largely limited. In this study, typical LNA bacteria were enriched from three different freshwater sources using extinction dilution (ED) and fluorescence-activated cell sorting (FACS). We have shown for the first time that LNA bacteria can be isolated and cultivated by using sterile freshwater as a growth medium. During growth, the typical LNA characteristics (that is, low-fluorescence intensity and sideward scatter (SSC)) remained distinct from those of typical HNA bacteria. Three LNA pure cultures that are closely affiliated to the Polynucleobacter cluster according to 16S rRNA sequencing results were isolated. Owing to their small size, cells of the isolates remained intact during cryo-transmission electronic microscopy examination and showed a Gram-negative cell-wall structure. The extremely small cell volume (0.05 mu m(3)) observed for all three isolates indicates that they are among the smallest free-living heterotrophic organisms known in culture. Their isolation and cultivation allow further detailed investigation of this group of organisms under defined laboratory conditions. The ISME Journal (2009) 3, 889-902; doi: 10.1038/ismej.2009.46; published online 7 May 200

Echinodermata: The Complex Immune System in Echinoderms

View references (418) The Echinodermata are an ancient phylum of benthic marine invertebrates with a dispersal-stage planktonic larva. These animals have innate immune systems characterized initially by clearance of foreign particles, including microbes, from the body cavity of both larvae and adults, and allograft tissue rejection in adults. Immune responsiveness is mediated by a variety of adult coelomocytes and larval mesenchyme cells. Echinoderm diseases from a range of pathogens can lead to mass die-offs and impact aquaculture, but some individuals can recover. Genome sequences of several echinoderms have identified genes with immune function, including expanded families of Toll-like receptors, NOD-like receptors, and scavenger receptors with cysteine-rich domains, plus signaling pathways and cytokines. The set of transcription factors that regulate proliferation and differentiation of the cellular immune system are conserved and indicate the ancestral origins of hematopoiesis. Both larval and adult echinoderms are in constant contact with potential pathogens in seawater, and they respond to infection by phagocytosis and encapsulation, and employ proteins that function in immune detection and response. Antipathogen responses include activation of the SpTransformer genes, a complement system, and the production of many types of antimicrobial peptides. Echinoderms have homologues of the recombinase activating genes plus all associated genes that function in vertebrates for immunoglobulin gene family rearrangement, although their gene targets are unknown. The echinoderm immune system has been characterized as unexpectedly complex, robust, and flexible. Many echinoderms have very long life-spans that correlate with an excellent capacity for cell damage repair. In many marine ecosystems, echinoderms are keystone predators and herbivores, and therefore are species that can serve as optimal sentinels of environmental health. Coelomocytes can be employed in sensor systems to test for the presence of marine pollutants. When Elie Metchnikoff inserted a rose prickle into a larval sea star and observed chemotaxis, phagocytosis, and encapsulation by the mesenchyme cells, he initiated not only the field of immunology but also that of comparative immunology, of which the echinoderms have been an important part