Publications

4474

ABSTRACTThree cohorts of farmed yellowtail kingfish (Seriola lalandi) from South Australia were examined forChlamydia-like organisms associated with epitheliocystis. To characterize the bacteria, 38 gill samples were processed for histopathology, electron microscopy, and 16S rRNA amplification, sequencing, and phylogenetic analysis. Microscopically, the presence of membrane-enclosed cysts was observed within the gill lamellae. Also observed was hyperplasia of the epithelial cells with cytoplasmic vacuolization and fusion of the gill lamellae. Transmission electron microscopy revealed morphological features of the reticulate and intermediate bodies typical of members of the orderChlamydiales. A novel 1,393-bp 16S chlamydial rRNA sequence was amplified from gill DNA extracted from fish in all cohorts over a 3-year period that corresponded to the 16S rRNA sequence amplified directly from laser-dissected cysts. This sequence was only 87% similar to the reported “CandidatusPiscichlamydia salmonis” (AY462244) from Atlantic salmon and Arctic charr. Phylogenetic analysis of this sequence against 35ChlamydiaandChlamydia-like bacteria revealed that this novel bacterium belongs to an undescribed family lineage in the orderChlamydiales. Based on these observations, we propose this bacterium of yellowtail kingfish be known as “CandidatusParilichlamydia carangidicola” and that the new family be known as “CandidatusParilichlamydiaceae.”

Abstract Chemoheterotrophic marine bacteria of the SAR11 clade are Earth’s most abundant organisms. Following the first cultivation of a SAR11 bacterium, ‘Candidatus Pelagibacter ubique’ strain HTCC1062 (Ca. P. ubique) in 2002, unusual nutritional requirements were identified for reduced sulfur compounds and glycine or serine. These requirements were linked to genome streamlining resulting from selection for efficient resource utilization in nutrient-limited ocean habitats. Here we report the first successful cultivation of Ca. P. ubique on a defined artificial seawater medium (AMS1), and an additional requirement for pyruvate or pyruvate precursors. Optimal growth was observed with the collective addition of inorganic macro- and micronutrients, vitamins, methionine, glycine and pyruvate. Methionine served as the sole sulfur source but methionine and glycine were not sufficient to support growth. Optimal cell yields were obtained when the stoichiometry between glycine and pyruvate was 1:4, and incomplete cell division was observed in cultures starved for pyruvate. Glucose and oxaloacetate could fully replace pyruvate, but not acetate, taurine or a variety of tricarboxylic acid cycle intermediates. Moreover, both glycine betaine and serine could substitute for glycine. Interestingly, glycolate partially restored growth in the absence of glycine. We propose that this is the result of the use of glycolate, a product of phytoplankton metabolism, as both a carbon source for respiration and as a precursor to glycine. These findings are important because they provide support for the hypothesis that some micro-organisms are challenging to cultivate because of unusual nutrient requirements caused by streamlining selection and gene loss. Our findings also illustrate unusual metabolic rearrangements that adapt these cells to extreme oligotrophy, and underscore the challenge of reconstructing metabolism from genome sequences in organisms that have non-canonical metabolic pathways.

ABSTRACT The bacteria that cause necrotizing hepatopancreatitis in Penaeus vannamei adversely affect penaeid shrimp cultured in the western hemisphere. 16S rRNA and gyrase B gene analyses determined the taxonomic position of these bacteria. The name “ Candidatus Hepatobacter penaei” is proposed for these pathogenic bacteria, which are members of the Rickettsiales order.

Abstract Bacteria from the uncultured SUP05/Arctic96BD-19 clade of gamma proteobacterial sulfur oxidizers (GSOs) have the genetic potential to oxidize reduced sulfur and fix carbon in the tissues of clams and mussels, in oxygen minimum zones and throughout the deep ocean (>200 m). Here, we report isolation of the first cultured representative from this GSO clade. Closely related cultures were obtained from surface waters in Puget Sound and from the deep chlorophyll maximum in the North Pacific gyre. Pure cultures grow aerobically on natural seawater media, oxidize sulfur, and reach higher final cell densities when glucose and thiosulfate are added to the media. This suggests that aerobic sulfur oxidation enhances organic carbon utilization in the oceans. The first isolate from the SUP05/Arctic96BD-19 clade was given the provisional taxonomic assignment ‘Candidatus: Thioglobus singularis’, alluding to the clade’s known role in sulfur oxidation and the isolate’s planktonic lifestyle.

This study addressed the taxonomic position and group classification of a phytoplasma responsible for virescence and phyllody symptoms in naturally diseased Madagascar periwinkle plants in western Malaysia. Unique regions in the 16S rRNA gene from the Malaysian periwinkle virescence (MaPV) phytoplasma distinguished the phytoplasma from all previously described ‘ Candidatus Phytoplasma ’ species. Pairwise sequence similarity scores, calculated through alignment of full-length 16S rRNA gene sequences, revealed that the MaPV phytoplasma 16S rRNA gene shared 96.5 % or less sequence similarity with that of previously described ‘ Ca. Phytoplasma ’ species, justifying the recognition of the MaPV phytoplasma as a reference strain of a novel taxon, ‘Candidatus Phytoplasma malaysianum’. The 16S rRNA gene F2nR2 fragment from the MaPV phytoplasma exhibited a distinct restriction fragment length polymorphism (RFLP) profile and the pattern similarity coefficient values were lower than 0.85 with representative phytoplasmas classified in any of the 31 previously delineated 16Sr groups; therefore, the MaPV phytoplasma was designated a member of a new 16Sr group, 16SrXXXII. Phytoplasmas affiliated with this novel taxon and the new group included diverse strains infecting periwinkle, coconut palm and oil palm in Malaysia. Three phytoplasmas were characterized as representatives of three distinct subgroups, 16SrXXXII-A, 16SrXXXII-B and 16SrXXXII-C, respectively.

Two strains (JC85T and JC108) of Gram-stain-negative, motile bacteria were isolated from endolithic beach sand samples on an oligotrophic medium. Based on the 16S rRNA gene sequence analysis, both strains were identified as belonging to the genus Rhizobium . Strain JC108 had 16S rRNA gene sequence similarity of 100 % with Rhizobium pusense NRCPB10T and formed a cluster with this strain. Strain JC85T had 96.9 % 16S rRNA gene sequence similarity and was 18 % related (based on DNA–DNA hybridization) to Rhizobium borbori DN316T. With other strains of the genus Rhizobium , the 16S rRNA gene sequence similarity was less than 96.3 %. Strain JC85T could tolerate up to 3 % salinity, fix N2, was resistant to ampicillin (10 µg) and was positive for catalase and oxidase. The major fatty acid was C18 : 1ω7c (69 %) with minor amounts of C19 : 0 cyclo ω8c (8.9 %), C16 : 0 (6.9 %), C12 : 0 (5.7 %) and C19 : 1ω7c/C19 : 1ω6c (2.2 %). Polar lipids of strain JC85T include two unidentified aminophospholipids (APL1,2), two unidentified phospholipids (PL1,2), phosphatidylcholine and four unidentified lipids (L1–4). Q-10 is the major quinone of strain JC85T. Based on polyphasic taxonomic analysis, strain JC85T represents a novel species for which, the name Rhizobium subbaraonis JC85T is proposed. The type strain is JC85T ( = DSM 24765T = KCTC 23614T).