This report presents evidence that dogs recover from acute canine monocytic ehrlichiosis (CME) after 16 days of doxycycline treatment (10 mg/kg of body weight every 24 h). Blood PCR was as valuable as splenic aspirate PCR for early diagnosis of acute CME. Splenic aspirate PCR was, however, superior to blood PCR for the evaluation of ehrlichial elimination.Canine monocytic ehrlichiosis (CME) is a tick-borne disease with a global distribution (9). Diagnosis of CME is confirmed by demonstration of morulae in blood smears, serology, culturing of the rickettsiae, and PCR using Ehrlichia canis-specific primers. Tetracyclines are commonly used in the treatment of CME, with doxycycline in particular being the most acceptable and widely used (2, 3).Blood samples are presently used for PCR evaluation of ehrlichial infection and response to therapy (15). A previous study has demonstrated ehrlichial DNA in splenic aspirates of two subclinically experimentally infected dogs whose blood and bone marrow aspirates were found to be negative for E. canis DNA, suggesting the importance of splenic aspirates in the diagnosis of ehrlichial carriers (6). This finding questioned the validity of previous blood-based PCR studies for evaluating treatment regimens for CME, and the length of doxycycline treatment required for E. canis infection remained uncertain.The aims of this study were (a) to investigate whether splenic aspirates are superior to blood samples as a sample source for E. canis PCR; (b) to determine whether dogs with acute CME remain persistently infected despite concurrent treatment; and (c) to determine the duration of doxycycline treatment required to eliminate E. canis DNA, as measured by PCR in cases of acute CME.Five beagle dogs, 4 to 6 years old and negative serologically and by PCR for E. canis, were used in this study. The dogs were artificially infected by intravenous injection of 5 ml of E. canisinfected blood. This blood was drawn from an acutely ill, naturally infected dog. Infection of the donor dog with E. canis was confirmed by serology using the indirect immunofluorescence antibody (IFA) test (11), by blood culture of DH82 cells (4), and by p30-based nested PCR for E. canis (14).

SummaryThe tick Ixodes ricinus is responsible for the transmission of a number of bacterial, protozoan and viral diseases to humans and animals in Europe and š Č ȇ Northern Africa. Female I. ricinus from England, Switzerland and Italy have been found to harbour an intracellular a-proteobacterium, designated IricES1, within the cells of the ovary. IricES1 is the only prokaryote known to exist within the mitochondria of any animal or multicellular organism. To further examine the distribution, prevalence and mode of transmission of IricES1, we performed polymerase chain reaction screening of I. ricinus adults from 12 countries across its geographic distribution, including tick colonies that have been maintained in the laboratory for varying periods of time. IricES1 was detected in 100% of field-collected female ticks from all countries examined (n = 128), while 44% of males were found to be infected (n = 108). Those males that are infected appear to harbour fewer bacteria than females. Sequencing of fragments of the 16S rRNA and gyrB genes revealed very low nucleotide diversity among various populations of IricES1. Transmission of IricES1 from engorged adult females to eggs was found to be 100% (n = 31). In tick colonies that had been maintained in the laboratory for several years, a relatively low prevalence was found in females (32%; n = 25). To our knowledge, IricES1 is the most widespread and highly prevalent of any tick-associated symbiont.

GM1-ganglioside receptor binding by the B subunit of cholera toxin (CtxB) is widely accepted to initiate toxin action by triggering uptake and delivery of the toxin A subunit into cells. More recently, GM1 binding by isolated CtxB, or the related B subunit of Escherichia coli heat-labile enterotoxin (EtxB), has been found to modulate leukocyte function, resulting in the down-regulation of proinflammatory immune responses that cause autoimmune disorders such as rheumatoid arthritis and diabetes. Here, we demonstrate that GM1 binding, contrary to expectation, is not sufficient to initiate toxin action. We report the engineering and crystallographic structure of a mutant cholera toxin, with a His to Ala substitution in the B subunit at position 57. Whereas the mutant retained pentameric stability and high affinity binding to GM1-ganglioside, it had lost its immunomodulatory activity and, when part of the holotoxin complex, exhibited ablated toxicity. The implications of these findings on the mode of action of cholera toxin are discussed.