Detection of the etiologic agent of human ehrlichiosis by polymerase chain reaction

Anderson, Burt; Sumner, John W.; Dawson, Jacqueline E.; Tzianabos, Theodore; Greene, Cornelia R.; Olson, James G.; Fishbein, Daniel B.; Olsen-Rasmussen, Melissa; Holloway, Brian P.; George, E H

doi:10.1128/jcm.30.4.775-780.1992

Cited by 229 publications

(71 citation statements)

References 10 publications

(10 reference statements)

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“…The recently introduced technique of PCR has great potential for improving the ability to diagnose infectious diseases caused by obligate intracellular bacteria. It has been employed to detect various kinds of rickettsial agents of humans and animals (1,2,8,14). A disadvantage of PCR-based DNA detection is the need to confirm the authenticity of the amplified products either by Southern blot hybridization, by DNA sequencing, or by restriction endonuclease cleavage patterns.…”

Section: Discussionmentioning

confidence: 99%

“…Oligonucleotide primers and probes were obtained from a commercial source (DNAFax Co., Ltd., Taipei, Taiwan) and used without further purification. Primer EP1 was constructed to correspond to nucleotides 53 to 69 on the sense strand of E. platys, a region of the 16S rDNA near the 5Ј end that has been reported to be highly variable in other ehrlichiae (1). Table 1 shows the sequences of three primers and the corresponding sequences for all ehrlichiae that are currently known to infect dogs and some other ehrlichial species.…”

Section: Methodsmentioning

confidence: 99%

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Specific amplification of Ehrlichia platys DNA from blood specimens by two-step PCR

Chang

Pan

1996

J Clin Microbiol

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A two-step PCR method for diagnosis of canine infectious cyclic thrombocytopenia was established. Three primers derived from the 16S rRNA gene sequence were used to amplify genomic DNA specifically from Ehrlichia platys. Two-round amplification with DNA templates prepared from E. platys-infected blood specimens produced 742-and 385-bp fragments, but these products were not found when an Ehrlichia canis-infected blood sample and Escherichia coli were used. This method, for which the minimum detectable copy number in the blood specimen was estimated to be five ehrlichial inclusions within platelets, is more sensitive than single PCR amplification. These results demonstrate that this two-step PCR is highly sensitive and efficient for detecting the etiologic agent of canine infectious cyclic thrombocytopenia in blood. The same technique was applied to blood specimens collected from a dog inoculated with E. platys. Amplification of the target DNA fragments was observed with blood collected on the fifth day after inoculation, which indicates that this method is also feasible for early diagnosis of E. platys infection.

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Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Specific amplification of Ehrlichia platys DNA from blood specimens by two-step PCR

Chang

Pan

1996

J Clin Microbiol

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“…PCR was used to detect and differentiate Anaplasma and Ehrlichia using species‐specific primers (Anderson et al., ; Dawson et al., , ; Murphy et al., ; Kocan et al., ; Inokuma et al., ; Kawahara et al., ; Torina et al., ), the details of which are shown in Table . PCRs were performed in an automatic DNA thermocycler (Bio‐Rad, Hercules, CA, USA), and PCR products were separated by 1.5% agarose gel electrophoresis to assess the presence of specific bands indicative of Anaplasma and Ehrlichia spp.…”

Section: Methodsmentioning

confidence: 99%

“…For this reason, the diagnosis of Ehrlichia and Anaplasma infection in humans is commonly based on clinical manifestations and serological analyses. Several useful and more sensitive molecular tools, such as polymerase chain reaction (PCR) and nested‐PCR, have been used to detect and identify Ehrlichia and Anaplasma species in both hosts and vectors (Anderson et al., ; Dawson et al., , ; Murphy et al., ; Kocan et al., ; Inokuma et al., ; Kawahara et al., ; de la Fuente et al., ; Torina et al., ).…”

Section: Introductionmentioning

confidence: 99%

Molecular Survey ofAnaplasmaandEhrlichiaof Red Deer and Sika Deer in Gansu, China in 2013

Chen

Liu

et al. 2015

Transbound Emerg Dis

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Anaplasma and Ehrlichia are important emerging tick-borne pathogens in both humans and animals. Here, we conducted a molecular surveillance study in Gansu, China to assess the prevalence of Anaplasma and Ehrlichia spp. in red deer and sika deer based on polymerase chain reaction (PCR) analysis and sequencing of 16S rRNA or msp genes. PCR revealed that the prevalence of Anaplasma ovis, Anaplasma bovis and Anaplasma platys of the Qilian Mountain samples was 32%, 9% and 9%, respectively; the prevalence of Anaplasma ovis, Anaplasma bovis, Anaplasma platys was 20%, 15% and 15% among the Long Mountain samples, respectively. Of the Long Mountain samples, two (5%) of the 40 samples were positive for Ehrlichia canis, but all 44 of the Qilian Mountain samples were negative for E. canis, and no other Anaplasma or Ehrlichia spp. were found in the samples. The phylogenetic tree showed that the newly isolated Anaplasma and Ehrlichia spp. could be classified as belonging to four clades, including an A. bovis cluster, A. ovis cluster, A. platys cluster and E. canis cluster. In addition, Bartonella schoenbuchensis was firstly identified in blood samples from red deer in Gansu, China. Our results provide important data to increase the understanding of the epidemiology of anaplasmosis and ehrlichiosis of red deer and sika deer and will assist with the implementation of measures to control anaplasmosis and ehrlichiosis transmission to red deer, sika deer and other animals in Gansu, China.

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“…The first-round PCR used the primers ECC and ECB, while the nested PCR used primers ECC and HE3. 4 The PCRs were carried out using 2.5× HotMasterMix (Eppendorf, New York, NY) and PCR-grade water. All primers and probes were obtained from Sigma Genosys.…”

Section: Polymerase Chain Reaction (Pcr)mentioning

confidence: 99%

Overcoming Barriers to the Transformation of the Genus Ehrlichia

Long

Whitworth

Walker

et al. 2005

Annals of the New York Academy of Sciences

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While bacterial transformation has evolved since the early 20th century to allow for the genetic manipulation of a variety of microbial agents, rickettsial organisms have proved resistant to such advances until only recently. The Ehrlichia are small, gram-negative, obligately intracellular bacterial parasites, which belong to the family Anaplasmataceae and cause a variety of infections in human and animal hosts. E. chaffeensis is the causative agent of human monocytotropic ehrlichiosis and is transmitted by Amblyomma americanum, the Lone Star tick. In this work, we describe the first report of successful transformation of a closely related ehrlichial species, the murine monocytotropic species E. muris. Application of these techniques should allow for a wide variety of molecular studies to be performed that were previously impossible. This heralds the beginning of a new era in ehrlichial research.

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Detection of the etiologic agent of human ehrlichiosis by polymerase chain reaction

Cited by 229 publications

References 10 publications

Specific amplification of Ehrlichia platys DNA from blood specimens by two-step PCR

Specific amplification of Ehrlichia platys DNA from blood specimens by two-step PCR

Molecular Survey ofAnaplasmaandEhrlichiaof Red Deer and Sika Deer in Gansu, China in 2013

Overcoming Barriers to the Transformation of the Genus Ehrlichia

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