BACKGROUND: Scrub typhus, severe fever with thrombocytopenia syndrome (SFTS) and human granulocytic anaplasmosis (HGA) are important arthropod-borne infectious diseases in Korea and share a common point that they are transmitted by arthropod bites mostly during outdoor activities and there are considerable overlaps of epidemiologic and clinical features at presentation. We investigated the co-infection of these infections. METHODS: The study subjects were patients with laboratory-confirmed scrub typhus who were enrolled retrospectively in 2006. SFTS virus (SFTSV) infection was confirmed by a reverse transcriptase polymerase chain reaction (PCR) to amplify partial L segment of SFTSV for molecular diagnosis. HGA was confirmed by a nested PCR to amplify 16S rRNA gene of Anaplasma phagocytophilum. Direct sequencing of the positive PCR products was performed. Clinical features of co-infected subjects were described. RESULTS: One-hundred sixty-seven patients with scrub typhus were included in the analysis. Co-infection of A. phagocytophilum was identified in 4.2% of scrub typhus patients (7/167). The route of co-infection was uncertain. The co-infected patients had not different clinical manifestations compared to the patients with scrub typhus only. All the study subjects were negative for SFTSV. CONCLUSION: We found retrospective molecular evidence of the co-infection of scrub typhus and HGA in Korea. HGA may be more prevalent than expected and need to be considered as an important differential diagnosis of febrile patients in Korea.
Anaplasma phagocytophilum ; Anaplasmosis ; Animals ; Arthropods ; Coinfection ; Communicable Diseases ; Diagnosis ; Diagnosis, Differential ; Fever ; Genes, rRNA ; Humans ; Korea ; Polymerase Chain Reaction ; Retrospective Studies ; Reverse Transcriptase Polymerase Chain Reaction ; Scrub Typhus ; Thrombocytopenia

Rodents are well-known reservoirs and vectors of many emerging and re-emerging infectious diseases, but little is known about their role in zoonotic disease transmission in Bhutan. In this study, a cross-sectional investigation of zoonotic disease pathogens in rodents was performed in Chukha district, Bhutan, where a high incidence of scrub typhus and cases of acute undifferentiated febrile illness had been reported in people during the preceding 4–6 months. Twelve rodents were trapped alive using wire-mesh traps. Following euthanasia, liver and kidney tissues were removed and tested using PCR for Orientia tsutsugamushi and other bacterial and rickettsial pathogens causing bartonellosis, borreliosis, human monocytic ehrlichiosis, human granulocytic anaplasmosis, leptospirosis, and rickettsiosis. A phylogenetic analysis was performed on all rodent species captured and pathogens detected. Four out of the 12 rodents (33.3%) tested positive by PCR for zoonotic pathogens. Anaplasma phagocytophilum, Bartonella grahamii, and B. queenslandensis were identified for the first time in Bhutan. Leptospira interrogans was also detected for the first time from rodents in Bhutan. The findings demonstrate the presence of these zoonotic pathogens in rodents in Bhutan, which may pose a risk of disease transmission to humans.
Anaplasma ; Anaplasma phagocytophilum ; Anaplasmosis ; Animals ; Bartonella ; Bartonella Infections ; Bhutan ; Communicable Diseases, Emerging ; Ehrlichiosis ; Euthanasia ; Humans ; Incidence ; Kidney ; Leptospira ; Leptospira interrogans ; Leptospirosis ; Liver ; Orientia tsutsugamushi ; Polymerase Chain Reaction ; Rodentia ; Scrub Typhus ; Zoonoses

The identification and characterization of pathogenic and zoonotic tick-borne diseases like granulocytic anaplasmosis are essential for developing effective control programs. The differential diagnosis of pathogenic Anaplasma phagocytophilum and non-pathogenic A. phagocytophilum-like Anaplasma spp. is important for implementing effective treatment from control programs. The objective of the present study was to investigate the prevalence of Anaplasma spp. in horses in Korea by nucleotide sequencing and restriction enzyme fragment length polymorphism assay. Of the 627 horses included in the study, only 1 (0.2%) was infected with A. phagocytophilum. Co-infection with A. phagocytophilumlike Anaplasma spp. was not detected in the study. The 16S rRNA sequence of A. phagocytophilum was similar (99.5–100%) to A. phagocytophilum 16S rRNA isolated from horses in other countries. PCR adapted to amplify A. phagocytophilum groEL and msp2 genes failed to generate amplicons, suggesting genetic diversity in these genes. This study is the first molecular detection of A. phagocytophilum in horses in Korea. Human granulocytic anaplasmosis and animal infection of A. phagocytophilum have been reported in Korea recently. Because of vector tick distribution, global warming, and the increase of the horse industry, horses should be considered as a potential reservoir for A. phagocytophilum, and cross infectivity should be evaluated even though a low prevalence of infection was detected in this study. Furthermore, continuous surveillance and effective control measures for A. phagocytophilum should be established to prevent disease distribution and possible transmission to humans.
Anaplasma phagocytophilum ; Anaplasma ; Anaplasmosis ; Animals ; Coinfection ; Diagnosis, Differential ; Genetic Variation ; Global Warming ; Granulocytes ; Horses ; Humans ; Korea ; Phylogeny ; Polymerase Chain Reaction ; Polymorphism, Restriction Fragment Length ; Prevalence ; Tick-Borne Diseases ; Ticks

Members of the genus Anaplasma are important emerging tick-borne pathogens in both humans and animals in tropical and subtropical areas. Here, we investigated the presence of Anaplasma spp. in 621 sheep and 710 goats from six provinces of China. Polymerase chain reaction (PCR) and DNA sequencing were conducted to determine the prevalence of Anaplasma (A.) phagocytophilum, A. ovis and A. bovis targeting the 16S ribosomal RNA or the major surface protein 4 gene. PCR revealed Anaplasma in 39.0% (240/621) of sheep and 45.5% (323/710) of goats. The most frequently detected species was A. ovis (88/621, 14.2% for sheep; 129/710, 18.2% for goats), followed by A. bovis (60/621, 9.7% for sheep; 74/710, 10.4% for goats) and A. phagocytophilum (33/621, 5.3% for sheep; 15/710, 2.1% for goats). Additionally, eight sheep and 20 goats were found to be infected with three pathogens simultaneously. DNA sequencing confirmed the presence of these three Anaplasma species in the investigated areas, and phylogenetic analysis indicated that there was geographic segregation to a certain extent, as well as a relationship between the host and cluster of A. ovis. The results of the present study provide valuable data that helps understand the epidemiology of anaplasmosis in ruminants from China.
Anaplasma ovis ; Anaplasma phagocytophilum ; Anaplasma* ; Anaplasmosis ; Animals ; China* ; Epidemiology ; Goats* ; Humans ; Polymerase Chain Reaction ; Prevalence ; RNA, Ribosomal, 16S ; Ruminants ; Sequence Analysis, DNA ; Sheep*

Tick-borne rickettsial diseases (TBRD) are commonly encountered in medical and veterinary clinical settings. The control of these diseases is difficult, requiring disruption of a complex transmission chain involving a vertebrate host and ticks. The geographical distribution of the diseases is related to distribution of the vector, which is an indicator of risk for the population. A total of 1,107 ticks were collected by tick dragging from forests, ecotourism parks and hosts at 101 sites in 22 of the 32 states of Mexico. Collected ticks were placed in 1.5 mL cryovials containing 70% ethanol and were identified to species. Ticks were pooled according to location/host of collection, date of collection, sex, and stage of development. A total of 51 ticks were assayed by polymerase chain reaction (PCR) to confirm species identification using morphological methods. A total of 477 pools of ticks were assayed using PCR techniques for selected tick-borne pathogens. Anaplasma phagocytophilum was the most commonly detected pathogen (45 pools), followed by, Ehrlichia (E.) canis (42), Rickettsia (R.) rickettsii (11), E. chaffeensis (8), and R. amblyommii (1). Rhipicephalus sanguineus was the tick most frequently positive for selected pathogens. Overall, our results indicate that potential tick vectors positive for rickettsial pathogens are distributed throughout the area surveyed in Mexico.
Anaplasma phagocytophilum ; Animals* ; Ehrlichia ; Ehrlichia canis ; Ehrlichia chaffeensis ; Ethanol ; Forests ; Humans* ; Mexico* ; Polymerase Chain Reaction ; Rhipicephalus sanguineus ; Rickettsia ; Ticks* ; Vertebrates

North Korea is located on the northern part of the Korean Peninsula in East Asia. While tick-borne pathogens of medical and veterinary importance have been reported from China and South Korea, they have not been reported from North Korea. To screen for zoonotic tick-borne pathogens in North Korea, ticks were collected from domestic goats. A total of 292 (27 nymph, 26 male, 239 female) Haemaphysalis (H.) longicornis were collected and assayed individually for selected tick-borne pathogens. A total of 77 (26.4%) were positive for Anaplasma bovis, followed by Bartonella (B.) grahamii (15, 5.1%), Anaplasma phagocytophilum (12, 4.1%), Bartonella henselae (10, 3.4%), and Borrelia spp. (3, 1.0%) based on 16S ribosomal RNA and ITS species-specific nested polymerase chain reaction. Using the groEL-based nested PCR, a total of 6 and 1 H. longicornis were positive for B. grahamii and B. henselae, respectively. All products were sequenced and demonstrated 100% identity and homology with previously reported sequences from other countries in GenBank. This is the first report of the detection of tick-borne pathogens in the North Korea and suggests that farm animals may act as reservoirs for zoonotic tick-borne pathogens.
Anaplasma phagocytophilum ; Anaplasma* ; Animals, Domestic ; Bartonella henselae ; Bartonella* ; Borrelia* ; China ; Databases, Nucleic Acid ; Democratic People's Republic of Korea* ; Far East ; Goats* ; Humans ; Korea ; Male ; Nymph ; Polymerase Chain Reaction ; Prevalence* ; RNA, Ribosomal, 16S ; Ticks

OBJECTIVETo type the Chinese Anaplasma phagocytophilum isolates by Multispacer typing (MST).

METHODSBased on the genomes of the 4 published Anaplasma strains, 4 genomic sequences were analyzed by Mauve 2.3.1 software and variable spacer sequences were selected for designing primers with the bio-software Primer Premier 5.0. A total of 11 Chinese A. phagocytophilum isolates, obtained from different areas of China during 2009-2012 were assayed by the MST. Twenty two intergenic sequences for each isolate tested and the reference A. phagocytophilum strain Webster and A. phagocytophilum strain HZ were concatenated in the order of HGA-mst 1F/1R-mst 2F/2R, HGA-mst 22F/22R.

RESULTSTwenty two pairs of primers were successfully used for typing the Human granulocytic anaplasmosis (HGA) strains in the study. Those 22 intergenic sequences exhibited a great diversity among the strains tested and each of the strain tested was identified as unique genotype, according to the alignment analysis of the 22 concatenated intergenic sequences. Of these single nucleotide polymorphism (SNPs) identified in the study, the nucleotide transitions shared the highest percentage (60.2%, 251/417) and then the nucleotide transversion, accounted for 23.0% (96/417) and the indel events (insertion/deletion) were observed of 16.7% (70/417)SNPs. Phylogenetic analysis indicated that the 5 strains from patients (LZ-H1, LZ-H2, LZ-H3, LZ-H4, LZ-H5) from Laizhou areas, Shandong province and 1 tick strain (LZ-T1) from Haemaphysalis longicornis collected from the same areas where the patients lived were grouped in the same clan with the reference A. phagocytophilum strain Webster and strain HZ. Beijing isolates (BJ-H1) grouped with Xinjiang isolates (XJ-H1 and XJ-H3) while another tick isolates from Laizhou areas (LZ-T2) and another Xinjiang human isolate(XJ-H2)were in the same clan, which was closely related to the isolates from severe patients in Laizhou.

CONCLUSIONChinese HGA isolates exhibited a great diversity of intergenic regions. MST seemed a valuable tool for the detection and tracing for any endemic strains of Anaplasma during the outbreak investigations in the public health events.

Anaplasma phagocytophilum ; classification ; genetics ; Bacterial Typing Techniques ; methods ; China ; DNA, Ribosomal Spacer ; genetics ; Polymorphism, Single Nucleotide

OBJECTIVELyme disease and Human granulocytic anaplasmosis are tick-borne diseases caused by Borrelia burgdorferi and Anaplasma phagocytophilum respectively. We have investigated infection and co-infection of the two diseases in the population of forest areas of eight provinces in China by measuring seroprevalence of antibodies against B. burgdorferi and A. phagocytophilum.

METHODSForest areas in 8 provinces were chosen for investigation using whole sampling and questionnaire survey methods. 3 669 serum samples from people in the forest areas were tested for the presence of antibodies by indirect immunofluorescent assay (IFA).

RESULTSSeroprevalence against B. burgdorferi was 3% to 15% and against A. phagocytophilum was 2% to 18% in the study sites in the 8 provinces in China. We also found co-infection of B. burgdorferi and A. phagocytophilum in 7 of the 8 provinces (the exception being the Miyun area in Beijing). The seroprevalence for both B. burgdorferi and A. phagocytophilum was significantly higher among people exposed to ticks than among people who were not exposed to ticks.

CONCLUSIONWe conclude that both pathogens are endemic in the forest areas in the eight provinces, but the prevalence of B. burgdorferi and A. phagocytophilum differs between the provinces.

Adolescent ; Adult ; Anaplasma phagocytophilum ; pathogenicity ; Anaplasmosis ; blood ; epidemiology ; Animals ; Borrelia burgdorferi ; pathogenicity ; Child ; China ; Coinfection ; Female ; Humans ; Lyme Disease ; blood ; epidemiology ; Male ; Middle Aged ; Seroepidemiologic Studies ; Tick-Borne Diseases ; blood ; epidemiology ; Trees ; Young Adult

Anaplasma phagocytophilum ; Anaplasmosis ; epidemiology ; Animals ; China ; epidemiology ; Dermacentor ; microbiology ; Humans ; Molecular Epidemiology

OBJECTIVETo evaluate the Anaplasma phagocytophilum (A. phagocytophilum), Ehrlichia canis (E. canis), Dirofilaria immitis (D. immitis) (canine heartworm), Borrelia burgdorferi (B. burgdorferi) infections in countryside dogs from Yunnan, Hainan and Anhui provinces.

METHODSSerum samples were collected from 26 dogs in Yunnan, Hainan and Anhui provinces. The samples were tested using a commercial ELISA rapid diagnostic assay kit (SNAP(®) 4Dx(®); IDEXX Laboratories, Inc. U.S.A.). Meanwhile, indirect immunofluorescence assay (IFA) recommended by WHO was conducted to detect IgG to A. phagocytophilum. Two methods were analyzed and compared.

RESULTSThe number of serologically positive dogs for IgG to A. phagocytophilum was only 2 which was from Hainan province and none of the 26 dogs responded positive for E. canis, D. immitis (canine heartworm), and B. burgdorferi by ELISA rapid diagnostic method. The number of serologically positive dogs for IgG to A. phagocytophilum was 13 (50%) by IFA method. Data of the two methods were analyzed by statistical software and the difference was statistically significant (P=0.002).

CONCLUSIONSIt can be concluded that IFA method was more sensitive than ELISA rapid diagnostic method. However, we need conduct further and intensive epidemiology survey on tick-born diseases pathogens including A. phagocytophilum, E. canis, D. immitis (canine heartworm), and B. burgdorferi which have public health significance.

Anaplasma phagocytophilum ; immunology ; Animals ; Borrelia burgdorferi ; immunology ; China ; epidemiology ; Dirofilaria immitis ; immunology ; Dirofilariasis ; blood ; epidemiology ; immunology ; Disease Vectors ; Dog Diseases ; epidemiology ; Dogs ; Ehrlichia canis ; immunology ; Ehrlichiosis ; blood ; epidemiology ; immunology ; Enzyme-Linked Immunosorbent Assay ; methods ; Fluorescent Antibody Technique, Indirect ; methods ; Immunoglobulin G ; blood ; Lyme Disease ; blood ; epidemiology ; immunology ; Tick-Borne Diseases ; epidemiology