A laboratory animal surveillance program intended to screen for the presence of adventitious pathogens was performed at the Laboratory Animal Center of the Academy of Military Medical Sciences, Beijing, China. Four blood samples from 10 captive-bred rhesus macaques (Macaca mulatta
) were presumed to be infected with Bartonella
spp. according to Giemsa-stained smears and transmission electron microscopy (Technical Appendix
). Further PCR and sequence analysis of 3 gene targets (internal transcribed spacer [ITS], gltA
, and rnp
B) confirmed the existence of B. quintana
in the 4 parasite-positive macaques (Technical Appendix Table 1
). In addition, B. quintana
was successfully isolated from the 4 monkeys by blood plating.
Figure 1 Monitoring surveillance of Bartonella quintana infection in macaques and identification of lice. A) Macaques were housed in linked cages (squares); dashed lines indicate wired net enabling direct contact between macaques, and solid line indicates wall (more ...)
During the 36-day period of observation, 3 screening tests of the 10 macaques showed an increasing prevalence of B. quintana: 4 were found positive at day 1, 7 positive at day 15, and all 10 positive at day 35 (, panel A). Close examination of the monkeys revealed no skin scratch or wound indicative of direct contact between them.
Examination for ectoparasites at the last day of observation (day 36) revealed that all 10 monkeys were infested with lice (mean 10.3 lice/monkey, range 4–28 lice). Lice from each infested monkey were combined in 2 pools. B. quintana
was identified in all pools of lice by PCR selective for ITS, glt
A, and rnp
B. Partial Cytb sequence (660-bp) of the louse was obtained (GenBank accession no. JX070558) (Technical Appendix Table 1
); phylogenetic analysis of Cytb identified the louse as a relative of lice of the genus Pedicinus
(, panel A). By means of stereomicroscopy, the louse was then identified as Pedicinus obtusus
(, panel B), a macaque-specific ectoparasite, according to morphologic criteria (9
Figure 2 Phylogenetic analyses of louse species and Bartonella spp. A) Phylogenetic tree of louse species based on the partial Cytb sequence (364-bp), obtained by using the neighbor-joining method with maximum composite likelihood analysis and bootstrap analysis (more ...)
Of the 60 rhesus macaques (27 male, 33 female) housed in 5 other rooms in individual cages in the same facility, an additional 30 were found to be positive for B. quintana by PCR. B. quintana prevalence among sexually immature macaques was higher than that among sexually mature macaques, but this difference was not significant (29/54 [53.7%] vs. 1/6 [16.7%], respectively; p = 0.195). B. quintana prevalence among male macaques was similar to that among females (15/27 [55.6%] vs. 15/33 [45.5%], respectively).
Nucleotide sequences of ITS (123-bp), gltA
(539-bp), and rnpB
(336-bp) from all macaques and pools of lice were identical; they differed from from those of B. quintana
strain Toulouse by1–3 bp . Phylogenetic markers rnpB
, 16S rRNA, and 23S rRNA (11
) were amplified and sequenced from the strain identified in this study (RM-11) (Technical Appendix Table 1
). Phylogenetic analysis of their combined sequence alignment placed the RM-11 strain on a separate branch along with the strain of B. quintana
from a cynomolgus macaque and in the same clade as strains from patients in Europe who had trench fever (strains Toulouse and Fuller) (, panel B).
To evaluate the ability of the isolate to cause disease, we intravenously inoculated 4 Bartonella
spp.–negative rhesus macaques with B. quintana
isolated from a blood sample of a macaque from this study and twice passaged on agar (detailed methods described in the Technical Appendix
). Bacteremia reached a peak in 1 monkey on day 7 postinoculation (160 CFU/mL), in 2 monkeys on day 14 postinoculation (290 and 240 CFU/mL), and in 1 monkey on day 42 postinoculation (240 CFU/mL). Bacteremia then dropped to below a detectable level after 15 weeks postinoculation for all monkeys (Technical Appendix
). A relapsing pattern of bacteremia was observed during the experiment. Rectal temperature, hemogram, and blood biochemistry results for the 4 monkeys remained within normal limits.
The animal facility employees who had direct contact with monkeys during cage cleaning and feeding activities were tested for B. quintana
infection. Paired serum samples collected at 2 time points 3 months apart were tested for IgG against B. quintana
by indirect immunofluorescence assay, as described (12
). The baseline serum samples were all negative at a dilution of 1:64. Among the serum samples collected 3 months later, 3 had IgG titers of 256, 1 had a titer of 512, and 4 were negative. For all blood samples collected at the 2 time points, PCR detection, blood-smear staining, and blood culture for Bartonella
spp. were negative. Analysis of questionnaires revealed that all 4 of the workers with evidence of seroconversion reported lice exposure; 2 of them were scratched or bitten by monkeys.