Evaluation of a Yersinia pestis mutant impaired in a thermoregulated type VI-like secretion system in flea, macrophage and murine models.

Type VI secretion systems (T6SSs) have been identified recently in several Gram-negative organisms and have been shown to be associated with virulence in some bacterial pathogens. A T6SS of Yersinia pestis CO92 (locus YPO0499-YPO0516) was deleted followed by investigation of the phenotype of this mutation. We observed that this T6SS locus of Y. pestis was preferentially expressed at 26 degrees C in comparison to 37 degrees C suggesting a possible role in the flea cycle. However, we found that the deletion of T6SS locus YPO0499-YPO0516 in Y. pestis CO92 had no effect on the ability of this strain to infect the oriental rat flea, Xenopsylla cheopis. Nevertheless, this mutant displayed increased intracellular numbers in macrophage-like J774.A1 cells after 20 h post-infection for bacterial cells pre-grown at 26 degrees C indicating that expression of this T6SS locus limited intracellular replication in macrophages. In addition, deletion of the YPO0499-YPO0516 locus reduced the uptake by macrophages of the Y. pestis mutant pre-grown at 37 degrees C, suggesting that this T6SS locus has phagocytosis-promoting activity. Further study of the virulence of the T6SS mutant in murine bubonic and inhalation plague models revealed no attenuation in comparison with the parental CO92 strain.

New approaches to detection and identification of Rickettsia africae and Ehrlichia ruminantium in Amblyomma variegatum (Acari: Ixodidae) ticks from the Caribbean.

Imported from Africa in the 1700s and despite frequent modern eradication efforts, Amblyomma variegatum (F.) spread through the Caribbean by cattle transport, small ruminants, and migrating birds. A. variegatum is a vector for Rickettsia africae, the causative agent of African tick bite fever, and Ehrlichia ruminantium, the causative agent of heartwater. We examined 95 A. variegatum and six Rhipicephalus (Boophilus) microplus (Canestrini) collected from cattle at an abattoir in Antigua. Engorged tick extracts adsorbed on Nobotu filter paper strips and new nested polymerase chain reaction (PCR) assays for E. ruminantium and Dermatophilus congolensis were used to evaluate these ticks for the presence of these pathogenic bacteria. Amblyomma ticks (62.4%) contained R. africae DNA by PCR/restriction fragment length polymorphism analysis and DNA sequencing of the OmpA and 17-kDa antigen genes. Twenty Amblyomma and two Rh. microplus contained E. ruminantium DNA. No E. chaffeensis, Anaplasma phagocytophilum, Coxiella burnetii, or D. congolensis DNA was detected in these ticks. The continued presence of Am. variegatum in the Caribbean poses a significant risk of infection in cattle with E. ruminantium and in humans by R. africae. Eradication efforts are essential to prevent the further spread of Am. variegatum.

Detection and identification of bacterial agents in Ixodes persulcatus Schulze ticks from the north western region of Russia.

Ixodes persulcatus Schultze ticks are traditionally associated with transmission of Lyme disease, babesiosis, and tick-borne encephalitis. Here we compared the prevalence of infection with Borrelia burgdorferi, and rickettsial and ehrlichial agents in I. persulcatus ticks collected in different locations of the North Western administrative region of Russia. Altogether, 27.7% of ticks were infected with at least one organism, while the DNA of two or more bacteria was found in 11.8% of ticks tested. The highest average prevalence of Anaplasmataceae (20.8%) was detected in ticks from Arkhangel'sk province, while the prevalence in ticks from Novgorod province and St. Petersburg, respectively, was 7.3% and 12.2%. Only Ehrlichia muris DNA was identified by DNA sequencing. In comparison, the prevalence of B. burdorferi DNA was 16.6%, 5.8%, and 24.5% in the respective locations. The 382-bp amplicon of gltA from Candidatus Rickettsia tarasevichiae was detected in 2.75% and 1.6%, respectively, of ticks from Arkhangel'sk and Novgorod provinces, extending further west and north the area where this rickettsia is known to be present. DNA of the rickettsia-like endosymbiont Montezuma was primarily associated with female ticks, 8-28% of which were infected. Since I. persulcatus is so commonly infected with multiple agents that may cause human diseases, exposure to these ticks poses significant risk to human health in this region.

Prevalence of bacterial agents in Ixodes persulcatus ticks from the Vologda Province of Russia.

The prevalence of rickettsiae, ehrlichiae, and the rickettsia-like endosymbiont called Montezuma relative to that of Borrelia was determined in questing Ixodes persulcatus (I. persulcatus) ticks collected in 2002-2003 from Vologda Province, Russia. Ehrlichia muris, Anaplasma phagocytophilum, Montezuma, and new spotted fever group rickettsiae were detected by polymerase chain reaction (PCR) for the first time in this area. The rickettsiae were all Candidatus Rickettsia tarasevichiae, the furthest west this organism has been detected. After Borrelia, Montezuma was the agent most frequently detected; it may be present throughout the distribution of I. persulcatus in Russia. Ehrlichiae and rickettsiae frequently share the same tick host with Borrelia burgdorferi sensu lato so cotransmission and mixed infections in vertebrate hosts, including humans, may occur.