Canadian Journal of Microbiology

A large (47.75 ± 3.56 µm in diameter) Thiovulum bacterial strain forming white veils is described from a marine mangrove ecosystem. High sulfide concentrations (up to 8 mM of H2S) were measured on sunken organic matter (wood/bone debris) under laboratory conditions. This sulfur-oxidizing bacterium colonized the organic matter, forming a white veil. According to conventional scanning electron microscope (SEM) observations, bacterial cells are ovoid and slightly motile by numerous small flagella present on the cell surface. Large intracytoplasmic internal sulfur granules were observed, suggesting a sulfidic-based metabolism. Observations were confirmed by elemental sulfur distribution detected by energy-dispersive X-ray spectroscopy (EDXS) analysis using an environmental scanning electron microscope (ESEM) on non-dehydrated samples. Phylogenetic analysis of the partial sequence of 16S rDNA obtained from purified fractions of this Epsilonproteobacteraeota strain indicates that this bacterium belongs to the Thiovulaceae cluster and could be one of the largest Thiovulum ever described. We propose to name this species Candidatus Thiovulum sp. strain imperiosus.

Intracellular bacterial symbionts are known from various insect groups, particularly from those feeding on unbalanced diets, where the bacteria provide essential nutrients to the host. In the case of reed beetles (Coleoptera: Chrysomelidae, Donaciinae), however, the endosymbionts appear to be associated with specialized “glands” that secrete a material used for the beetles’ unusual water-tight cocoon. These glands were discovered over a century ago, but the bacteria they contain have yet to be characterized and placed in a phylogenetic context. Here, we describe the ultrastructure of two endosymbiotic species (“ Candidatus Macropleicola appendiculatae” and “ Candidatus Macropleicola muticae”) that reside in cells of the Malpighian tubules of the reed beetle species Macroplea appendiculata and Macroplea mutica , respectively. Fluorescent in situ hybridization using oligonucleotides targeting the 16S rRNA gene specific to Macroplea symbionts verified the localization of the symbionts in these organs. Phylogenetic analysis of 16S rRNA placed “Candidatus Macropleicola” in a clade of typically endosymbiotic Enterobacteriaceae (γ-proteobacteria). Finally, we discuss the evidence available for the hypothesis that the beetle larvae use a secretion produced by the bacteria for the formation of an underwater cocoon.

To characterize potentially important surface-exposed proteins of the phytoplasma causing chrysanthemum yellows (CY), new primers were designed based on the conserved regions of 3 membrane protein genes of the completely sequenced onion yellows and aster yellows witches’ broom phytoplasmas and were used to amplify CY DNA. The CY genes secY, amp, and artI, encoding the protein translocase subunit SecY, the antigenic membrane protein Amp and the arginine transporter ArtI, respectively, were cloned and completely sequenced. Alignment of CY-specific secY sequences with the corresponding genes of other phytoplasmas confirmed the 16S rDNA-based classification, while amp sequences were highly variable within the ‘Candidatus Phytoplasma asteris’. Five CY partial sequences were cloned into the pRSetC expression vector, and 3 of the encoded protein fragments (Amp 64/651, Amp 64/224, ArtI 131/512) were expressed as fusion antigens for the production of CY-specific polyclonal antibodies (A416 against Amp 64/224; A407 against ArtI 131/512). A416 recognized, in Western blots, the full-length Amp from CY-infected plants (periwinkle, daisy) and insect vectors ( Euscelidius variegatus , Macrosteles quadripunctulatus ). A416 also reacted to European aster yellows, to primula yellows phytoplasmas, to northern Italian strains of ‘Ca. Phytoplasma asteris’ from lettuce and gladiolus, but it did not react to American aster yellows phytoplasma.