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Human infection with Candidatus Neoehrlichia mikurensis, China.
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MedLine Citation:
PMID:  23017728     Owner:  NLM     Status:  MEDLINE    
To identify Candidatus Neoehrlichia mikurensis infection in northeastern China, we tested blood samples from 622 febrile patients. We identified in 7 infected patients and natural foci for this bacterium. Field surveys showed that 1.6% of ticks and 3.8% of rodents collected from residences of patients were also infected.
Hao Li; Jia-Fu Jiang; Wei Liu; Yuan-Chun Zheng; Qiu-Bo Huo; Kun Tang; Shuang-Yan Zuo; Kun Liu; Bao-Gui Jiang; Hong Yang; Wu-Chun Cao
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't    
Journal Detail:
Title:  Emerging infectious diseases     Volume:  18     ISSN:  1080-6059     ISO Abbreviation:  Emerging Infect. Dis.     Publication Date:  2012 Oct 
Date Detail:
Created Date:  2012-09-28     Completed Date:  2013-02-19     Revised Date:  2013-07-11    
Medline Journal Info:
Nlm Unique ID:  9508155     Medline TA:  Emerg Infect Dis     Country:  United States    
Other Details:
Languages:  eng     Pagination:  1636-9     Citation Subset:  IM    
Beijing Institute of Microbiology and Epidemiology, Beijing, People’s Republic of China.
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MeSH Terms
Anaplasmataceae / classification,  genetics,  isolation & purification*
Anaplasmataceae Infections / epidemiology*,  microbiology,  veterinary
China / epidemiology
Communicable Diseases, Emerging / epidemiology*,  microbiology,  veterinary
DNA, Bacterial / blood,  genetics
DNA, Ribosomal / genetics
Ixodes / microbiology*
Middle Aged
RNA, Ribosomal, 16S / genetics
Rodent Diseases / epidemiology*,  microbiology
Rodentia / microbiology
Sequence Analysis, DNA
Reg. No./Substance:
0/DNA, Bacterial; 0/DNA, Ribosomal; 0/RNA, Ribosomal, 16S

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Journal Information
Journal ID (nlm-ta): Emerg Infect Dis
Journal ID (iso-abbrev): Emerging Infect. Dis
Journal ID (publisher-id): EID
ISSN: 1080-6040
ISSN: 1080-6059
Publisher: Centers for Disease Control and Prevention
Article Information
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Print publication date: Month: 10 Year: 2012
Volume: 18 Issue: 10
First Page: 1636 Last Page: 1639
PubMed Id: 23017728
ID: 3471638
Publisher Id: 12-0594
DOI: 10.3201/eid1810.120594

Human Infection with Candidatus Neoehrlichia mikurensis, China Alternate Title:Human Infection with Candidatus N. mikurensis
Hao Li1
Jia-Fu Jiang1
Wei Liu1
Yuan-Chun Zheng
Qiu-Bo Huo
Kun Tang
Shuang-Yan Zuo
Kun Liu
Bao-Gui Jiang
Hong Yang
Wu-Chun Cao
Beijing Institute of Microbiology and Epidemiology, Beijing, People’s Republic of China (H. Li, J.-F. Jiang, W. Liu, K. Tang, S.-Y. Zuo, K. Liu, B.-G. Jiang, H. Yang, W.-C. Cao);
and Mudanjiang Forestry Central Hospital, Mudanjiang, People’s Republic of China (Y.-C. Zheng, Q.-B. Huo)
Correspondence: Address for correspondence: Wu-Chun Cao, State Key Laboratory of Pathogen and Biosecurity, Beijing Institute of Microbiology and Epidemiology, 20 Dong-Da St, Fengtai District, Beijing 100071, People’s Republic of China; email:

Candidatus Neoehrlichia mikurensis was detected in 1999 in Ixodes ricinus ticks in the Netherlands and referred to as an Ehrlichia spp.–like agent (1). It was then classified as a new member of family Anaplasmataceae on the basis of ultrastructure and phylogenetic analysis (2). The agent was detected in ticks and small wild mammals in Europe and Asia (16) and has recently been reported to infect humans, especially immunocompromised patients in Europe (710). However, no cases of infection have been identified outside Europe. Moreover, the agent has not yet been isolated in pure culture, and its antigens are not available.

To investigate human infections with tick-borne agents in China, we initiated a surveillance study at Mudanjiang Forestry Central Hospital (Mudanjiang, China). This hospital is one of the largest hospitals treating patients with tick-borne infectious diseases in northeastern China, where various tick-borne agents have been detected in ticks and animal hosts (1115).

The Study

During May 2–July 30, 2010, a total of 622 febrile patients, who had histories of recent tick bites and sought treatment at Mudanjiang Forestry Central Hospital (Figure 1) were screened for the infections of tick-borne agents. When patients were admitted, peripheral blood samples were collected and treated with EDTA. DNA as extracted by using the QIAmp DNA Blood Mini Kit (QIAGEN, Germantown, MD, USA).

For a broad-range assay, a nested PCR specific for the 16S rRNA gene (rrs) was used to detect organisms in the family Anaplasmataceae. For positive samples, 2 heminested PCRs were used to amplify the entire rrs gene. For further confirmation, a nested PCR specific for the 60-kDa heat shock protein gene (groEL) was performed. Detailed cycling conditions for all amplifications are described in the Technical Appendix.

Seven patients were found to be infected with Candidatus N. mikurensis by amplifications of the rrs and groEL genes. Amplified rrs gene (1,501 bp) and partial groEL gene (1,230 bp) sequences from these patients were identical. These sequences were also identical to genes of Candidatus N. mikurensis detected in ticks and rodents in the Asian region of Russia (5).

Serum samples were collected from patients during the acute (2–12 days after onset of illness) or convalescent (34–42 days after onset of illness) phases of illness. All samples were negative for IgG against Anaplasma phagocytophilum, Ehrlichia chaffeensis, Borrelia burgdorferi, Rickettsia heilongjiangensis, and tick-borne encephalitis virus when tested by indirect immunofluorescence assay.

All 7 patients were farmers residing in the villages in Mudanjiang. Their median age was 41 years (range 29–67 years) and 5 were men. None had been vaccinated against tick-borne encephalitis. The patients had onset of illness during May 20–July 13, 2010. The median time from the tick bite to the onset of illness and from the onset of illness to the physician visit was 8 days (range 2–35 days) and 7 days (range 1–12 days), respectively.

All patients were otherwise healthy, and none had a history of underlying immunocompromised conditions. Fever, headache, and malaise were reported for all 7 patients. Other major manifestations included nausea (5/7), vomiting (5/7), myalgia (4/7), and stiff neck (4/7). Less common symptoms were arthralgias (2/7), cough (2/7), diarrhea (1/7), confusion (1/7), and erythema (1/7). Skin erythema (multiple and oval) was seen on the neck of 1 patient.

Laboratory test results showed leukopenia in 1 patient, leukocytosis in 1 patient, thrombocytopenia in 2 patients, and anemia in 2 patients. Serum levels of alanine aminotransferase and aspartate aminotransferase were within reference ranges for all patients. Wright–Giemsa stained peripheral blood smears did not show morulae or other blood parasites.

To identify local natural foci, we performed a field investigation on infections of Candidatus N. mikurensis in ticks and rodents from areas of residences of the patients. During May–July 2010, a total of 516 host-seeking ticks, including 316 I. persulcatus, 187 Haemaphysalis concinna, and 13 Dermacentor silvarum, were collected on vegetation and individually examined. Candidatus N. mikurensis DNA was detected in 6 (1.9%) I. persulcatus and 2 (0.8%) H. concinna ticks, but no DNA was detected in D. silvarum ticks (Table).

A total of 211 rodents of various species were captured by using snap traps. After rodent species was identified, spleen specimens were collected for DNA extraction and PCR. Eight rodents of 3 species, 5 (4.6%) Clethrionomys rufocanus, 2 (5.7%) Rattus norvegicus, and 1 (33.3%) Tamias sibiricus, were positive for Candidatus N. mikurensis (Table).

Nucleotide sequences of rrs and groEL genes of 8 ticks and 8 rodents were identical to each other and to sequences obtained from the 7 patients. Phylogenetic analysis of rrs genes showed that nucleotide sequences identified were identical to those of Candidatus N. mikurensis from Japan and the Asian region of Russia but different from sequences from Europe (99.6%–99.8% similarity) (Figure 2, panel A). Similar phylogenetic relationships were observed in a neighbor-joining tree based on groEL gene nucleotide sequences. In comparison with sequences from humans and ticks in Europe, the groEL gene sequences identified in the study showed 97.6%–98.4% similarity (Figure 2, panel B).


We have detected Candidatus N. mikurensis DNA in blood samples from 7 patients collected during the period of acute illness, which suggests that this bacterium was the etiologic agent of the infections. Our findings demonstrated human infections with Candidatus N. mikurensis in China. The rrs and groEL gene nucleotide sequences of this Candidatus N. mikurensis variant were identical to those obtained from ticks and rodents in the Asian region of Russia, which have not been reported to cause human infection.

Unlike reported cases in elderly or immunocompromised patients in whom disease developed (710), all 7 patients in our study had relatively mild disease. Major clinical manifestations and laboratory findings of the cases in our report, such as leukocytosis, were not similar to those of previously reported cases. It is noteworthy that the patients reported in this study were previously healthy. Thus, their clinical manifestations might be typical of Candidatus N. mikurensis infection in an otherwise healthy population. However, the number of cases in our study was limited, and clinical data were not inclusive. Clinical characteristics of Candidatus N. mikurensis infection should include detailed descriptions of additional cases.

Our finding of a Candidatus N. mikurensis variant in 1.6% of ticks and 3.8% of rodents tested suggested natural foci of the bacterium in Mudanjiang. Therefore, clinical diagnosis of Candidatus N. mikurensis infection should be considered in patients who have been exposed to areas with high rates of tick activity. It is noteworthy that Candidatus N. mikurensis was originally detected in R. norvegicus from Guangzhou Province in southeastern China (4), thereby indicating the potential threat to humans in areas other than northeastern China.

In summary, we identified Candidatus N. mikurensis as an emerging human pathogen in China. Further studies should be conducted to isolate this bacterium and investigate its epidemiologic, genetic, and pathogenic features. To guide diagnostic testing and treatment, physicians should be aware that human Candidatus N. mikurensis infections are in Heilongjiang Province and that PCR can be used as a diagnostic technique for identifying suspected infections.

Technical Appendix

PCR, morphologic, and serologic procedures used for detection of Candidatus Neoehrlichia mikurensis, Mudanjiang, China.


Suggested citation for this article: Li H, Jiang J-F, Liu W, Zheng Y-C, Huo Q-B, Tang K, et al. Human infection with Candidatus Neoehrlichia mikurensis, China. Emerg Infect Dis [Internet]. 2012 Oct [date cited].

1These authors contributed equally to this article.

This study was supported by the Natural Science Foundation of China (81130086, 81072250, and 81172729), the Chinese Basic Research Project (2010CB530201), and the Special Fund for Health Research in the Public Interest (201202019).

Mr Li is a medical student in the State Key Laboratory of Pathogen and Biosecurity, Beijing Institute of Microbiology and Epidemiology. His research interests include the microbiology, epidemiology, and ecology of vector-borne diseases.

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[Figure ID: F1]
Figure 1 

Location of Mudanjiang, Heilongjiang Province, China, where Candidatus Neoehrlichia mikurensis was detected.

[Figure ID: F2]
Figure 2 

A) Neighbor-joining trees based on the 16S rRNA gene (rrs) and B) the 60-kDa heat shock protein gene (groEL) of Candidatus Neoehrlichia mikurensis, China, generated by using Molecular Evolutionary Genetics Analysis software version 4.0, ( the maximum composite-likelihood method, and bootstrap analysis of 1,000 replicates. Asterisks indicate nucleotide sequences of Candidatus N. mikurensis determined in this study. Numbers on branches indicate percentage of replicates that reproduced the topology for each clade. Scale bars indicate estimated evolutionary distance. A total of 1,303 positions for rrs and 953 positions for groEL were analyzed. Sources of Candidatus N. mikurensis sequences are shown between species names and GenBank accession numbers.

[TableWrap ID: T1] Table  Prevalence of Candidatus Neoehrlichia mikurensis in ticks and rodents, Mudanjiang, China
Species No. positive/no. tested (%)
Ixodes persulcatus 6/316 (1.9)
Haemaphysalis concinna 2/187 (0.8)
Dermacentor silvarum 0/13 (0)
Total 8/516 (1.6)
Clethrionomys rufocanus 5/109 (4.6)
Rattus norvegicus 2/35 (5.7)
Apodemus peninsulae 0/30 (0)
Apodemus agrarius 0/25 (0)
Mus musculus 0/9 (0)
Tamias sibiricus 1/3 (33.3)
Total 8/211 (3.8)

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Keywords: Keywords: Candidatus Neoehrlichia mikurensis, bacteria, human infection, ticks, rodents, vector-borne infections, China.

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