Bacterial communities of Bartonella-positive fleas: diversity and community assembly patterns

Abstract

We investigated the bacterial communities of nine Bartonella-positive fleas (n = 6 Oropsylla hirsuta fleas and n = 3 Oropsylla montana fleas), using universal primers, clone libraries, and DNA sequencing. DNA sequences were used to classify bacteria detected in a phylogenetic context, to explore community assembly patterns within individual fleas, and to survey diversity patterns in dominant lineages.

FIG. 1.

The proportions of 10 proteobacterial “families” within individual fleas are depicted. Samples are arranged based on Bartonella abundance data. Individual fleas tend to be dominated by one bacterial lineage. Numbers in parentheses indicate the total number of proteobacterial DNA sequences obtained from each flea. Fleas were collected from three different prairie dogs (prairie dog 1 harbored O. hirsuta fleas 1 to 3 [Oh_1 to Oh_3]; prairie dog 2, Oh_4; prairie dog 3, Oh_5 to Oh_6).

FIG. 2.

Maximum likelihood (GTR plus G correction model) tree of Bartonella from O. hirsuta (Oh) and O. montana (Om), with a number of sequences from GenBank. The clade with both O. hirsuta and O. montana is indicated by Om/h. Bartonella strains obtained from GenBank in the tree include B. washoensis (w; accession no. AF070463), two Bartonella sequences from Spermophilus sp. in China (C; accession no. DQ641913 and DQ641912), B. quintana (accession no. M73228), B. koehlerae (accession no. AF076237), B. grahamii (accession no. Z31349), B. bacilliformis (accession no. M65249), B. clarridgeiae (accession no. X97822), and a Bartonella sequence from another O. hirsuta (accession no. DQ473482). The tree was constructed using all of the Bartonella sequences detected, but some were removed to minimize the figure's size.

FIG. 3.

Maximum likelihood (GTR plus G correction model) tree of Rickettsiales detected in O. hirsuta (Oh) and O. montana (Om). Except for R. typhi, R. prowazekii, Rickettsiales bacterium “Montezuma” (R. ‘montezuma’), Orientalis tsutsugamushi, Anaplasma sp., Ehrlichia sp., and C. caryophilus, the taxon names indicate the genus of the host in which the bacteria were found (e.g., Wolbachia bacteria were found in a Nasonia parasitoid wasp host). Sequences from GenBank in the tree include Caedibacter caryophilus (accession no. AY753195), O. tsutsugamushi (accession no. U17258), Rickettsia typhi (accession no. AE017197), R. prowazekii (accession no. M21789), Rickettsiales bacterium “Montezuma” (accession no. AF493952), Anaplasma sp. (accession no. AY527214), Ehrlichia sp. (accession no. DQ324367), Hyalomma (accession no. AM181356), Acanthamoeba (accession no. AF132137), Anopheles (accession no. AY837738), Oh_GenBank (accession no. AY335925), Phtiropsylla (accession no. AY335932), Pulirritans (accession no. AY335926), Nephila (accession no. AF232234), Drosophila (accession no. AY833061), Nasonia (acces sion no. M84688), Myrmeleon (accession no. DQ068803), Paramecium (accession no. X58198), Louse 1 (accession no. AF467369), and Louse 2 (accession no. AF467370). The tree was constructed using all of the Rickettsiales sequences detected, but some were removed to minimize the figure's size.

FIG. 4.

Cumulative frequency distributions of pairwise genetic distances (GTR plus G corrected) for Bartonella and Rickettsiales strains from each flea species show that Rickettsiales lineages harbor more genetic diversity than Bartonella lineages. As genetic diversity within a lineage increases, the frequency distribution shifts to the right. Distributions of Bartonella genetic distances are presented as black lines, and Rickettsiales distances are presented as gray lines. Distributions for the two Bartonella lineages almost completely overlap and fall to the left of those for the Rickettsiales lineages. As an example of how to interpret this figure, the data point marked by the arrow indicates that 33.33% of the O. montana (Om) Rickettsiales (Rick) group 1 (G1) shares 99.65% or more sequence similarity.