New findings from an old pathogen: intraerythrocytic bacteria (family Anaplasmatacea) in red-backed salamanders Plethodon cinereus.

During a recent study of red-backed salamanders (Plethodon cinereus), we discovered an intraerythrocytic organism typified by violet-staining, intracellular inclusions, consistent with descriptions of Cytamoeba or Aegyptianella (bacteria). Here we characterize its taxonomic status using molecular techniques and ask basic questions about its nature. Blood smears from 102 salamanders were examined from Pennsylvania, New York, and Virginia to determine prevalence, and whole blood from several infected animals was tested using a PCR which targets the 16S rRNA gene of bacteria. Phylogenetic analysis of partial 16S rRNA gene sequence (1201 bp) indicated this organism was in the order Rickettsiales and is likely a member of the family Anaplasmatacea. The organism differed from currently described taxa and was clearly differentiated from Aegyptianella pullorum of birds and "Candidatus Hemobacterium ranarum" (formally A. ranarum) of frogs. Of all salamanders, 17 (16.7%) were infected and these were significantly larger (snout-vent length) and had higher body condition scores than uninfected ones, and males were more likely to be infected than females. Erythrocytes affected by the pathogen were 5% larger than unaffected ones, but otherwise similar in morphology. Infected animals tended to have a greater number of circulating white blood cells, based on estimates from smears, indicating a nonspecific response to the pathogen by the innate immune system. Given its phylogenetic position, this pathogen is likely transmitted by an arthropod vector, and the male-biased prevalence strongly implicates trombiculid mites, which also live in leaf litter and affect male salamanders more so than females.