Aerobic ammonia-oxidizing bacteria (AAOB) are known to have an important function in the marine nitrogen cycle. Anaerobic ammonium oxidation (anammox) carried out by some members of Planctomycetales is also an important process in marine ecosystems. Ammonia-monooxygenase gene (amoA) fragments were amplified to investigate the potential for nitrification and the diversity of the AAOB in two marine sponges Ircinia strobilina and Mycale laxissima. All of the AmoA sequences obtained from the two sponges clustered with the AmoA sequences of the Betaproteobacteria Nitrosospira spp. To investigate the anaerobic ammonia-oxidizing bacteria (AnAOB) in sponges, 16S rRNA gene fragments of Planctomycetales and anammox bacteria were also amplified with specific primers, and clone libraries were constructed. The Planctomycetales diversity detected in the two sponges was different. The Planctomycetales community in M. laxissima was affiliated with Pirellula, Planctomyces and anammox bacteria, while all of the I. strobilina Planctomycetales clones were solely affiliated with the candidate phylum 'Poribacteria'. Interestingly, sequences related to anammox genera were recovered only from M. laxissima. This is the first report of anammox bacteria in marine sponges. It is intriguing to find AAOB and AnAOB in M. laxissima, but the nature of their interaction with the sponge host and with each other remains unclear. This work further supports the potential of sponge-associated microorganisms for nitrification and sheds light on anammox as a new aspect of the nitrogen cycle in marine sponges.

Actinobacteria exclusively within the sub-class Acidimicrobidae were shown by 16S rDNA community analysis to be major components of the bacterial community associated with two sponge species in the genus Xestospongia. Four groups of Actinobacteria were identified in Xestospongia spp., with three of these four groups being found in both Xestospongia muta from Key Largo, Florida and Xestospongia testudinaria from Manado, Indonesia. This suggests that these groups are true symbionts in these sponges and may play a common role in both the Pacific and Atlantic sponge species. The fourth group was found only in X. testudinaria and was a novel assemblage distantly related to any previously sequenced actinobacterial clones. The only actinobacteria that were obtained in initial culturing attempts were Gordonia, Micrococcus and Brachybacterium spp., none of which were represented in the clone libraries. The closest cultured actinobacteria to all the Acidimicrobidae clones from Xestospongia spp. are 'Microthrix parvicella' and Acidimicrobium spp. Xestospongia spp. can now be targeted as source material from which to culture novel Acidimicrobidae to investigate their potential as producers of bioactive compounds. Isolation of sponge-associated Acidimicrobidae will also make it possible to elucidate their role as sponge symbionts.