Improved detection of Bartonella DNA in mammalian hosts and arthropod vectors by real-time PCR using the NADH dehydrogenase gamma subunit (nuoG)

We used a whole-genome scanning technique to identify the NADH dehydrogenase gamma subunit (nuoG) primer set that is sensitive and specific enough to detect a diverse number of Bartonella species in a wide range of environmental samples yet maintains minimal cross-reactivity to mammalian host and arthropod vector organisms.     

Preparation of Rocky Mountain spotted fever vaccine suitable for human immunization.

Rocky Mountain spotted fever vaccine was produced from rickettsiae grown in chicken embryo cells in roller bottle cultures. The rickettsiae were concentrated and purified by passage through a sucrose gradient and inactivated with formalin. This vaccine satisfactorily passed preinactivation and final container testing and is believed to be superior to the presently available yolk sac vaccine.     

Discrepancies in Weil-Felix and microimmunofluorescence test results for Rocky Mountain spotted fever.

Only 4.2% of 284 single specimens and 17.6% of 51 pairs of sera reactive in Weil-Felix agglutination tests for Rocky Mountain spotted fever were confirmed by a specific Rickettsia rickettsii microimmunofluorescence test.     

Kidney lesions in Rocky Mountain spotted fever: a light-, immunofluorescence-, and electron-microscopic study

The essential pathologic lesion in Rocky Mountain spotted fever (RMSF) is a vasculitis that may involve the kidneys as well as the heart, brain, skin, and subcutaneous tissues. Histopathologic information concerning the response of the kidneys in RMSF is rather limited, however. In this study renal tissue from 17 children who died of RMSF was examined by light, electron, and immunofluorescence microscopy. A lymphocytic or mixed inflammation, or both, involving vessels and interstitium of the kidney was found in all patients. In addition, 10 patients had histologic evidence of acute tubular necrosis, and another 3 had glomerular lesions consisting of focal segmental tuft necrosis or increased cellularity secondary to neutophilic infiltration, or both. Immunofluorescence- and electron-microscopic studies failed to demonstrate immune-complex deposition within glomeruli, a finding that suggests that immunoglobulin and classic immune complexes were not involved in the pathogenesis of the renal lesions at the time of death. These findings suggest the possibility that the pathogenesis of the renal lesion in RMSF may be due to a direct action of the organism (Rickettsia rickettsii) on the vessel wall.     

Detection and identification of spotted fever group rickettsiae in Dermacentor species from southern California

Dermacentor occidentalis Marx and Dermacentor variabilis (Say) commonly bite humans in California. These Dermacentor species may play a role in transmitting spotted fever group (SFG) rickettsiae to humans in many parts of the state where Dermacentor andersoni Stiles, a known vector for the etiologic agent of Rocky Mountain spotted fever, Rickettsia rickettsii, is absent. However, the specific rickettsial agents present in these ticks and their current prevalence are poorly understood. In total, 365 D. occidentalis and 10 D. variabilis were collected by flagging vegetation at 16 sites in five counties of southern California. The presence of SFG rickettsial DNA in these ticks was detected with rOmpA and GltA gene polymerase chain reaction (PCR) assays. The rickettsial species were identified by sequencing PCR amplicons. Of 365 D. occidentalis, 90 (24.7%) contained R. rhipicephali DNA, 28 (7.7%) contained DNA of unclassified genotype 364D, two (0.55%) contained R. bellii DNA, and one (0.3%) contained R. rickettsii DNA. Of 10 D. variabilis, four (40%) contained only R. rhipicephali. Four new genotypes of R. rhipicephali were discovered. For the first time, we detected R. rickettsii in D. occidentalis. Our study provides the first molecular data on the prevalence and species identification of SFG rickettsiae circulating in populations of these California ticks. Because neither D. variabilis nor R. rickettsii were abundant, 364D should be evaluated further as a potential cause of human SFG rickettsioses in southern California.     

Detection of the Rocky Mountain spotted fever agent, Rickettsia rickettsii, in dead ticks, Dermacentor andersoni.

Rickettsia rickettsii remained viable and retained chromophilic properties for not more than 24 h after Dermacentor andersoni were killed by freezing. Antigenic reactivity was detected for at least 71 days by direct immunofluorescence. However, rickettsiae in ticks suffocated with mineral oil remained pathogenic for at least 14 days. Accordingly, ticks removed from a host by mineral oil or dying from desiccation in transit are still suitable for rickettsial examination.     

Rickettsial infection of the pulmonary microcirculation: the basis for interstitial pneumonitis in Rocky Mountain spotted fever

Pulmonary tissue from 10 patients with fatal Rocky Mountain spotted fever was examined by brightfield microscopy for histopathologic lesions and by immunofluorescence for Rickettsia rickettsii. The distribution of rickettsiae and the vasculitis of the pulmonary microcirculation coincided. The lungs demonstrated the consequent interstitial pneumonia--alveolar septal congestion and interstitial edema; alveolar edema, fibrin, macrophages, and hemorrhage; and interlobular septal edema. The effects of the rickettsial damage to the pulmonary microcirculation are an important component of the pathophysiology of severe Rocky Mountain spotted fever. The distribution of rickettsial organisms within the lung indicates that person to person aerosol transmission is extremely unlikely.     

Prevalence of vectors of the spotted fever group Rickettsiae and murine typhus in a Bedouin town in Israel

A survey of the vectors of spotted fever group Rickettsiae and of murine typhus was carried out in Rahat, a Bedouin town in the Negev Desert, where the diseases are endemic. Houses with known cases of spotted fever group Rickettsiae or murine typhus were compared with those without reported clinical cases. A neighboring Jewish community, Lehavim, where no cases of spotted fever group Rickettsiae and murine typhus were reported in recent years, was used as a control. In the houses of patients with spotted fever group Rickettsiae in Rahat, an average of 7.4 times more ticks were found than in control houses. Out of 190 ticks isolated from sheep and goats or caught by flagging in Rahat, 90% were Rhipicephalus sanguineus (Latreille), 7.9% Rhipicephalus turanicus Pomerantzev, and 2.1% were Hyalomma sp. In the houses of patients with murine typhus, three times more rats were caught and, on the average, each rat was infested with 2.2 times more fleas than rats in the control houses. Out of 323 fleas collected from 35 Norwegian rats (Rattus norvegicus Berkenhout), 191 were Xenopsylla cheopis Rothschild and 132 Echidnophaga murina Tiraboschi. Thus, there was a six to seven times higher probability of encountering a tick or flea vector where infections had occurred than in control houses in Rahat. The percentage of rats seropositive to Rickettsia typhi was similar in study and control households (78.3 and 76.2, respectively). In the control settlement, Lehavim, only three Mus musculus L. were caught, which were not infested with ectoparasites and their sera were negative for murine typhus. Out of 10 dogs examined in this settlement, 15 R. sanguineus and eight specimens of the cat flea (Ctenocephalides felis felis Bouché) were isolated. No rats were caught in this settlement. These data indicate that there is a correlation among the density of domestic animals, their ectoparasites, and the incidence of spotted fever group Rickettsiae and murine typhus in Rahat.     

Computer simulation of management strategies for American dog ticks (Acari: Ixodidae) and Rocky Mountain spotted fever

Computer models were developed to simulate the effects of management technologies on populations of the American dog tick, Dermacentor variabilis (Say), principal vector of Rocky Mountain spotted fever (RMSF) in eastern North America. The technologies modeled were area-wide acaricide application, acaricide-food-baited tubes for self-treatment by small mammals, dipping of dogs in acaricides, acaricide-impregnated plastic dog collars, reduction of small mammal host populations (host management), and removal of vegetation that protects free-living tick stages (vegetative management). Submodels for each of these technologies were incorporated into a model (ADTSIM) for the population dynamics of the tick and RMSF transmission. Comparisons of simulated and observed data were used to verify reasonable accuracy of the submodels. Repetitive simulations were made to identify levels and timing of each control method (alone or combined) required to reduce tick populations below a RMSF transmission threshold of 252 unfed adults/ha. Eight to 30 acaricide applications, depending on acaricide and percentage of population treated, were needed during a 10-yr period to reduce densities of ticks below the threshold. The baited-tube method, host management, and vegetative management (depending on level and frequency of treatment) also were capable of reducing tick density below the threshold. However, acaricide-impregnated plastic dog collars did not reduce tick density below the threshold unless at least 50% of the hosts of adult ticks were domestic dogs. Integrated strategies were developed for management of ticks and RMSF in six selected states. These strategies reduced numbers of human cases of RMSF 90% or more by year 20 by maintaining tick densities between 100 and 252 unfed adults/ha.     

Analysis of Genetic Algorithm for Rule-Set Production (GARP) modeling approach for predicting distributions of fleas implicated as vectors of plague, Yersinia pestis, in California

More than 20 species of fleas in California are implicated as potential vectors of Yersinia pestis. Extremely limited spatial data exist for plague vectors-a key component to understanding where the greatest risks for human, domestic animal, and wildlife health exist. This study increases the spatial data available for 13 potential plague vectors by using the ecological niche modeling system Genetic Algorithm for Rule-Set Production (GARP) to predict their respective distributions. Because the available sample sizes in our data set varied greatly from one species to another, we also performed an analysis of the robustness of GARP by using the data available for flea Oropsylla montana (Baker) to quantify the effects that sample size and the chosen explanatory variables have on the final species distribution map. GARP effectively modeled the distributions of 13 vector species. Furthermore, our analyses show that all of these modeled ranges are robust, with a sample size of six fleas or greater not significantly impacting the percentage of the in-state area where the flea was predicted to be found, or the testing accuracy of the model. The results of this study will help guide the sampling efforts of future studies focusing on plague vectors.     

Potential for free radical-induced lipid peroxidation as a cause of endothelial cell injury in Rocky Mountain spotted fever.

Cells infected by Rickettsia rickettsii, the causative agent of Rocky Mountain spotted fever, display unusual intracellular morphological changes characterized by dilatation of the membranes of the endoplasmic reticulum and outer nuclear envelope. These changes are consistent with those that might be expected to occur following peroxidation of membrane lipids initiated by oxygen radical species, such as the hydroxyl radical or a variety of organic radicals. Using a fluorescent probe, we have found significantly increased levels of peroxides in human endothelial cells infected by R. rickettsii. Studies with desferrioxamine, an iron chelator effective in preventing formation of the hydroxyl radical from hydrogen peroxide and the superoxide free radical, reduced peroxide levels in infected cells to those found in uninfected cells. This observation suggests that the increased peroxides in infected cells may be lipid peroxides, degradation products of free radical attack on polyenoic fatty acids. The potential for lipid peroxidation as an important mechanism in endothelial cell injury caused by R. rickettsii is discussed.     

Computer simulation of Rocky Mountain spotted fever transmission by the American dog tick (Acari: Ixodidae)

A computer model was developed for simulation of the transmission of Rickettsia rickettsii, the causative agent of Rocky Mountain spotted fever (RMSF), by the American dog tick, Dermacentor variabilis (Say). The model of RMSF was combined with a model for population dynamics of the American dog tick and included simulation of infection and transmission of rickettsiae between ticks and host mammals and transmission of RMSF to humans. The model simulated the effects of biotic and environmental variables such as weather, host density, habitat, transovarial transmission, fecundity of infected ticks, and infectivity level of ticks and mammals. Some parameters in the model were fitted by iterative simulations to produce realistic rates of R. rickettsii infection in adult ticks and small and medium-sized mammal hosts. Parameters also were fitted to yield the historical average number of RMSF cases for Virginia. Comparisons of the simulated and actual number of cases for nine other states indicated a reasonable level of validity for the model. A theoretical tick density threshold of 252 unfed adult ticks/ha for transmission of RMSF was determined from a relationship between rate of transmission to humans and density of ticks. The transmission threshold can be used for additional modeling efforts to study the effects of management technologies on tick densities and RMSF human cases. The model can serve as a framework for modeling other tick-borne diseases such as Lyme disease, babesiosis, and heartwater.     

Comparison of serologic methods for the diagnosis of Mediterranean spotted fever.

During summer Mediterranean spotted fever (MSF) is prevalent in the subcostal part of Croatia (North Dalmatia) as well as in other areas of the Mediterranean coast. We compared the specificity and sensitivity of complement fixation (CF), latex agglutination (Latex-R. conorii), microimmunofluorescence (micro-IF) and enzyme-linked immuno assay (ELISA) for detection and measuring antibodies against Rickettsia conorii in 78 sera from 46 patients with/or suspected Mediterranean spotted fever. The seroreactivity with SFG antigens containing the Rickettsia conorii-antigen(s) as a common determinant, was positive in all four serological tests suggesting that Rickettsia conorii was the probable causative agent of infection in our patients.     

Distribution, seasonality, and hosts of the Rocky Mountain wood tick in the United States

Anaplasma marginale Theiler is a tick-borne pathogen that causes anaplasmosis in cattle. There are approximately 20 tick species worldwide that are implicated as vectors of this pathogen. In the United States, Dermacentor andersoni Stiles and Dermacentor variabilis (Say) are the principal vectors. The risk of transmission of anaplasmosis to cattle has been largely based on the distribution of D. andersoni in the United States. We developed a centralized geographic database that incorporates collection records for D. andersoni from two large national databases. We reviewed the geographic records in each database and postings from MEDLINE and AGRICOLA to produce a national county-level distribution map based on a total of 5,898 records. The records spanned the period from 1903 through 2001 with the majority between 1921 to 1940. Populations of D. andersoni were recorded from 267 counties in 14 states and were distinguished as either established or reported. We found 180 counties with established populations of D. andersoni and 87 counties with reported occurrences in 14 states with the majority of established populations reported from Montana, Idaho, and Oregon. D. andersoni populations in the United States currently extend from the western portions of Nebraska and the Dakotas westward to the Cascade Mountains and from the northern counties of Arizona and New Mexico northward to the Canadian border. The data will be useful for identifying regions at increased risk of acquiring anaplasmosis in the United States. Based upon the database collection records, we also present a summary of recorded hosts for D. andersoni and comments on its seasonal occurrence.     

Microimmunofluorescence test for the serological study of rocky mountain spotted fever and typhus.

A microimmunofluorescence test was used to study antibody responses to various spotted fever group and typhus group rickettsiae during Rocky Mountain spotted fever (RMSF) and epidemic typhus (ET). Patients with RMSF reacted most strongly to Rickettsia rickettsii; those with ET reacted predominantly to R. prowazekii. The degree of cross-reaction to other rickettsial strains varied from patient to patient, but a particular pattern of cross-reaction was consistently observed in serial sera from the same patient. Fresh isolates from three Montana RMSF cases were indistinguishable from each other and from strain R of R. rickettsii used as a standard antigen in all tests. Immunoglobulin M (IgM) antibodies were usually present in high titer in early-convalescent-phase sera from RMSF, as well as ET, patients. After RMSF, IgM antibodies persisted for a few months and, in one instance, for as long as 10 months. IgM responses to laboratory-acquired infections were infrequent in persons previously vaccinated with antigens related to the infecting strain. Previous antigenic conditioning from infection or vaccination may have accounted partly for the apparent lack of IgM response in a few study participants.