Detection of rickettsial DNA in ixodid ticks recovered from dogs and cats in Japan

DNA from ticks recovered from 1137 dogs and 133 cats from all over Japan were examined for Rickettsia infection by citrate synthase gene (gltA)-based PCR and partial nucleotide sequencing. A total of 91 dog tick samples and 18 cat tick samples showed a single band of the appropriate size in the nested PCR. Sequence analysis was successfully performed on 102 samples. DNA of Rickettsia japonica or closely related Rickettsia spp. strains were detected from 38 ticks in 16 prefectures mainly in western Japan. The other 33, detected from 13 prefectures including Hokkaido and Okinawa, were found to be Rickettsia helvetica or closely related strains. A total of 29 DNA that showed highest homology with Rickettsia akari or closely related strains were detected in 19 prefectures, widespread throughout Japan. Rickettsia canada-like DNA was detected from Haemaphysalis sp. removed from a dog in Fukuoka, and ;Candidatus Rickettsia tarasevichiae'-like DNA was from Ixodes sp. removed from a dog in Hokkaido.     

Prevalence of Rickettsia species antibodies and Rickettsia species DNA in the blood of cats with and without fever

Rickettsia species antibodies have been detected in some cats but it is unknown whether infected cats develop clinical signs. The prevalence of Rickettsia species deoxyribonucleic acid (DNA) in blood from clinically ill cats has not been determined. The objective of this study was to determine if cats with fever (body temperature >or=102.5 degrees F [39.2 degrees C]) were more likely to have evidence of rickettsial infection than healthy, age-matched, control cats with a body temperature<102.5 degrees F. Rickettsia species polymerase chain reaction (PCR) assays were performed to detect rickettsial DNA extracted from blood (71 paired samples), indirect immunofluorescence assays (IFA) were performed to detect serum antibodies against Rickettsia felis (90 paired samples) and Rickettsia rickettsii (91 paired samples), and the results between pairs were compared. All samples were negative for Rickettsia species DNA. More cats with fever were seropositive for R felis or R rickettsii than control cats, but results were not statistically significant. Results of this pilot study failed to show an association between Rickettsia species DNA or Rickettsia species antibodies and fever.     

Prevalence of tick-borne Rickettsia and Ehrlichia in Ixodes persulcatus and Ixodes ovatus in Tokachi district, Eastern Hokkaido, Japan

DNA from 111 ticks collected by flagging in Tokachi district, Eastern Hokkaido, Japan were examined for infection with Rickettsia and Ehrlichia, by PCR and sequencing methodology. For Rickettsia, analysis of the partial sequence of the citrate synthase gene was successfully performed on 11 DNA samples from I. persulcatus, and 7 of them showed 99.8% identical with Rickettsia helvetica while the other 4 showed 99.8% identical with ;Candidatus Rickettsia tarasevichiae'. For Ehrlichia, a partial sequence of the 16S rRNA gene detected from I. persulcatus was 100% identical with that from Ehrlichia muris, and another DNA sample from I. ovatus showed 99.8% identical with Ehrlichia species detected from I. ovatus. The results suggest that the pathogens detected here might be distributed in this area.     

Molecular survey of Rickettsia spp. in sick dogs in Italy

The aim of this retrospective study was to investigate the prevalence of Rickettsia spp. DNA in the blood of sick dogs from Italy. Canine blood samples (n=650) submitted for molecular testing of Rickettsia spp. to a diagnostic laboratory from February 2003 to March 2006 were studied. The Rickettsia spp. DNA detection was performed by Light Cycler real-time PCR using hybridization probes separately conducted with specific primers and probes. The total percentage of Rickettsia spp.-positive dog samples was 1.5% (10 out of 650). The percentage of Rickettsia spp.-positive dog samples submitted from north, central and southern Italy were 0.4% (1/248), 1.4% (3/219) and 3.3% (6/183), respectively. Five out of 138 dogs (3.6%) from Sicily were positive on Rickettsia PCR testing. A statistical difference was found between the percentages of positive samples from the Yorkshire terrier group (10.7%) compared with the mixed breed group (0.7%). No statistical differences were found between seasonal period, region and gender. Based on molecular data, there is infrequent rickettsiemia in dogs.     

Prevalence of Bartonella species, hemoplasmas, and Rickettsia felis DNA in blood and fleas of cats in Bangkok, Thailand

Flea infestations are common in Thailand, but little is known about the flea-borne infections. Fifty flea pools and 153 blood samples were collected from client-owned cats between June and August 2009 from veterinary hospitals in Bangkok, Thailand. Total DNA was extracted from all samples, and then assessed by conventional PCR assays. The prevalence rates of Bartonella spp. in blood and flea samples were 17% and 32%, respectively, with DNA of Bartonella henselae and Bartonella clarridgeiae being amplified most commonly. Bartonella koehlerae DNA was amplified for the first time in Thailand. Hemoplasma DNA was amplified from 23% and 34% of blood samples and flea pools, respectively, with 'Candidatus Mycoplasma haemominutum' and Mycoplasma haemofelis being detected most frequently. All samples were negative for Rickettsia felis. Prevalence rate of B. henselae DNA was increased 6.9 times in cats with flea infestation. Cats administered flea control products were 4.2 times less likely to be Bartonella-infected.     

Infection by spotted fever rickettsiae in people, dogs, horses and ticks in Londrina, Parana State, Brazil

Spotted fever is a disease caused by bacteria from the genus Rickettsia of the spotted fever group (SFG). Rickettsia rickettsii is likely the main agent of Brazilian spotted fever (BSF). With the objective of gathering information on the circulation of SFG rickettsiae in Londrina, Parana state, ticks from dogs and horses and also blood from dogs, horses and humans were collected in a neighbourhood of the city which presented potential for circulation of rickettsiae between hosts and vectors. Amblyomma cajennense, Dermacentor nitens, and Rhipicephalus sanguineus ticks were subjected to Polymerase Chain Reaction targeting a fragment of the Rickettsia gltA gene. This specific gene encodes the enzyme citrate synthase of Rickettsia spp., and results on all ticks were negative. Human and animal sera were tested by Indirect Immunofluorescence Assay in which R. rickettsii and R. parkeri were used as antigens. Sera from 4.7% human, 2.7% canine and 38.5% equine were positive for R. rickettsii. For R. parkeri, 0.9% human, 2.7% canine and 11.5% equine samples were positive. All samples reactive to R. parkeri also reacted to R. rickettsii. An epidemiological questionnaire was applied, but there were no statistically significant results. Comparison of our serological results with previous studies in Brazil, among BSF endemic and non-endemic areas, indicates that there is no established rickettsial infection in the study area, a statement corroborated with our molecular analysis. Nonetheless, as humans of the present study are highly exposed to tick infestations, health education within the population is needed to obtain efficient tick control.     

Rickettsiae in arthropods collected from the North African Hedgehog (Atelerix algirus) and the desert hedgehog (Paraechinus aethiopicus) in Algeria

Hedgehogs have become a popular pet despite their potential role in zoonotic disease transmission. We conducted an entomological study in a mountainous region of northeast Algeria in which we collected 387 fleas (Archeopsylla erinacei) and 342 ticks (Rhipicephalus sanguineus and Haemaphysalis erinacei) from Paraechinus aethiopicus and Atelerix algirus hedgehogs. Of the hedgehogs sampled, 77.7% and 91% were infested with fleas and ticks, respectively. Significantly more ticks and fleas were collected from A. algirus than from P. aethiopicus. Rickettsia felis was detected in 95.5% of fleas and R. massiliae was detected in 6.25% of Rh. sanguineus ticks by molecular tools. A new Rickettsia species of the spotted fever group was detected in 11.25% of Rh. sanguineus and in 77% of H. erinacei ticks. Overall, we show that hedgehogs can act as hosts for ectoparasites infected with several rickettsial agents. These data justify a more detailed investigation of animal reservoirs for Rickettsiae.     

Molecular detection of rickettsial agents in ticks and fleas collected from a European hedgehog (Erinaceus europaeus) in Marseilles, France

The European hedgehog (Erinaceus europaeus) is a synanthropic nocturnal insectivore commonly found in the countryside and in the parks and gardens. Because hedgehogs are already involved in the transmission of a number of zoonoses, including salmonellosis and ringworm, we decided to study their possible role in the epidemiology of the spotted fever group of Rickettsia. We collected ticks and fleas from a hedgehog that was captured in the city of Marseilles in France. Using a genus-specific quantitative PCR (qPCR) analysis followed by a species-specific qPCR analysis for positive samples, we observed that 91.7% (11/12) of the Rhipicephalus sanguineus ticks were positive for Rickettsia massiliae and 99.2% (128/129) of the Archaeopsylla erinacei fleas were positive for Rickettsia felis. Hedgehogs carry infected ectoparasites and then likely ensure the dissemination of spotted fever group Rickettsiae, and their epidemiological role requires further investigation.     

唐山地区蜱传斑点热群立克次体分子流行病学

根据已发表的长角血蜱16SrRNA序列及斑点热群立克次体外膜蛋白A（OmpA）基因序列设计2对特异性引物,对唐山地区采集的长角血蜱进行PCR检测,并对阳性样本进行测序和序列分析,抽检样本建立分子系统进化树。结果表明,在315份蜱DNA样本中检测出25份阳性样本,阳性率为7.94%;序列分析结果显示唐山地区长角血蜱携带立克次体同处于一个分支,与日本株立克次体同源性最高（93.30%）,其次是福建株立克次体（92.11%）,黑龙江立克次体绥芬株（90.45%）、虎林株（90.42%）。结论得出唐山地区蜱传斑点热感染较严重,分子进化分析结果显示唐山地区蜱传斑点热群立克次体可能为一新种。     

Rickettsial infection in domestic mammals and their ectoparasites in El Valle de Antón, Coclé, Panamá

The present research evaluated the presence of Rickettsia spp. on ectoparasites of horses and dogs (using PCR techniques), and their sera (using immunofluorescence assay) in El Valle de Antón town in Panama. A total of 20 horses and 20 dogs were sampled, finding four species of ectoparasites on dogs (the ticks Rhipicephalus sanguineus, Amblyomma ovale, Amblyomma oblongoguttatum, and the flea Ctenocephalides felis), and two tick species on horses (Amblyomma cajennense and Dermacentor nitens). DNA of Rickettsia amblyommii was found in pools of A. cajennense, D. nitens, and R. sanguineus, while Rickettsia felis was detected in C. felis pools. Overall, 70% (14/20) and 65% (13/20) of the horses and dogs, respectively, were seroreactive (titer ≥ 64) to spotted fever group rickettsiae. Sera from six dogs and five horses reacted to R. amblyommii antigens with titers at least four-fold higher than those for the other antigens tested (Rickettsia bellii, Rickettsia parkeri, Rickettsia rhipicephali, R. felis, and R. rickettsii). These serological results, coupled with our molecular findings, suggest that these dogs and horses were infected by Rickettsia amblyommii. More studies need to be realized afford to identify the Rickettsia species responsible for other serological and molecular positive results, and their ecological importance.     

Prevalence of Bartonella species,hemoplasmas, and Rickettsia felis DNA in blood and fleas of cats in Bangkok,Thailand

Flea infestations are common in Thailand, but little is known about the flea-borne infections. Fifty flea pools and 153 blood samples were collected from client-owned cats between June and August 2009 from veterinary hospitals in Bangkok, Thailand. Total DNA was extracted from all samples, and then assessed by conventional PCR assays. The prevalence rates of Bartonella spp. in blood and flea samples were 17% and 32%, respectively, with DNA of Bartonella henselae and Bartonella clarridgeiae being amplified most commonly. Bartonella koehlerae DNA was amplified for the first time in Thailand. Hemoplasma DNA was amplified from 23% and 34% of blood samples and flea pools, respectively, with ‘Candidatus Mycoplasma haemominutum’ and Mycoplasma haemofelis being detected most frequently. All samples were negative for Rickettsia felis. Prevalence rate of B. henselae DNA was increased 6.9 times in cats with flea infestation. Cats administered flea control products were 4.2 times less likely to be Bartonella-infected.     

Antibodies to Ehrlichia canis, Ehrlichia platys, and Spotted Fever Group Rickettsiae in Louisiana Dogs

Antibodies to Ehrlichia canis, Ehrlichia platys, and spotted fever group (SFG) rickettsiae were detected by indirect immunofluorescence in sera from 27 ill individually owned thrombocytopenic dogs (platelet concentrations less than 200,000 platelets/microliters) and 59 healthy kenneled dogs located in southern Louisiana. Platelet concentrations less than 100,000 platelets/microliters were detected in 63% of ill thrombocytopenic dogs and 6.8% of healthy kennel dogs. One ill thrombocytopenic dog had intracytoplasmic E platys morulae detected within platelets. The prevalence of increased serum antibody titers to E canis and E platys was 25.9% and 40.7% for the ill thrombocytopenic dogs and 20.3% and 54.2% for the healthy kennel dogs, respectively. All dogs with seropositivity to E canis had increased antibody titers of greater than or equal to 1:100 to E platys. Simultaneous examination of increased serum antibody titers (greater than or equal to 1:64) to four SFG rickettsiae indicate that Rickettsia rhipicephali and Rickettsia montana accounted for the majority of the antibodies detected in these dogs. Of 86 dogs tested, 44.2% were seronegative to E canis, E platys, and SFG rickettsiae.     

Prevalence of Rickettsia felis DNA in the blood of cats and their fleas in the United States

Rickettsia felis is associated with fever, headache, myalgia, and macular rash in some infected humans and has been detected in the cat flea (Ctenocephalides felis) in many countries around the world. While some naturally exposed cats have been assessed for antibodies against R felis, to our knowledge, no one has reported use of polymerase chain reaction (PCR) to attempt to amplify R felis DNA from client-owned cats and the fleas collected from them. In this study, we assayed 92 pairs of cat blood and flea extracts from Alabama, Maryland and Texas, using PCR assays that amplify a region of the citrate synthase gene (gltA) and the outer membrane protein B gene (ompB). Of the 92 pairs, 62 of 92 (67.4%) flea extracts and none of the cat blood samples were positive for R felis DNA.     

Molecular evidence of vector-borne pathogens in dogs and cats and their ectoparasites in Algiers,Algeria

In Algeria, only limited information is currently available on the prevalence of emergent canine and feline vector-borne diseases. The aim of the present work was to detect by qPCR vector-associated bacteria in stray dogs and cats and their ectoparasites from Algiers.18/117 (15.38%) dogs and 2/107 (1.87%) cats were positive for at least one vector-borne agent. Coxiella burnetii and Bartonella henselae were identified in 1/117 (0.85%) dog individually. Ehrlichia canis DNA was detected in 17/117 (14.52%) dogs. 1/107 (0.93%) cat was positive to C. burnetii and another 1/107 (0.93%) to B. henselae.DNA of Rickettsia massiliae, Rickettsia conorii and E. canis was detected in Rhipicephalus sanguineus. Cat fleas were infected with Rickettsia felis, B. henselae and Bartonella clarridgeiae. B. vinsonii subsp. berkhoffii was identified in Xenopsylla cheopis collected from dogs.The findings of this study indicate that dogs and cats from Algeria are exposed to multiple tick and flea-borne pathogens.     

Serological and molecular evidence of exposure to arthropod-borne organisms in cats from northeastern Spain

One hundred sixty-eight cat sera from Spain were tested for IgG antibodies to Rickettsia conorii (Rc), Ehrlichia canis (Ec), Anaplasma phagocytophilum (Ap) and Bartonella henselae (Bh) antigens using IFA and for FeLV antigen and FIV antibody by ELISA. For 47 whole blood samples, PCR testing was performed for Rickettsia, Ehrlichia and Bartonella. Seroprevalences were: Bh (71.4%), Rc (44%), Ec (11.3%), FeLV (8.5%), FIV (7.4%) and Ap (1.8%). Bh antibodies were associated with seroreactivity to both Ec and Rc antigens. FIV antibodies were associated with illness and cats older than 2 years. Bartonella henselae and B. clarridgeiae (Bcl) DNA was amplified from seven and one sample, respectively.     

Serologic and molecular evidence of coinfection with multiple vector-borne pathogens in dogs from Thailand

Forty-nine dogs from Thailand were evaluated for serologic evidence of exposure or polymerase chain reaction (PCR) evidence of infection with vectorborne pathogens, including Ehrlichia sp. (Ehrlichia canis, Ehrlichia chaffeensis, Ehrlichia equi, and Ehrlichia risticii), Bartonella vinsonii subsp. berkhoffi (Bvb), spotted fever group (SFG) rickettsiae (Rickettsia rickettsii), Typhus group (TG) rickettsiae (Rickettsia canada, Rickettsia prowazekii, and Rickettsia typhi), and Babesia sp. (Babesia canis and Babesia gibsonii). All study dogs had at least 1 of 3 entry criteria: fever, anemia, or thrombocytopenia. By immunofluorescence antibody (IFA) testing, seroreactivity was most prevalent to E chaffeensis (74%) and E canis (71%) antigens, followed by E equi (58%), Bvb (38%), E risticii (38%), R prowazekii (24%), B canis (20%), R rickettsii (12%), R canada (4%), and B gibsonii (4%) antigens. There was 100% concordance between E canis IFA and Western blot immunoassay (WI) for 35 of 35 samples; 2 samples were IFA and WI reactive only to E equi antigens. By PCR amplification, 10 dogs were found to be infected with E canis, 5 with Ehrlichia platys, and 3 with B canis. Sequencing of PCR products was undertaken to compare Ehrlichia strains from Thailand to strains originating from the United States. Partial DNA sequence analysis confirmed infection with E canis and E platys, with identical 16S rRNA sequence alignment to E canis (U26740) and to E platys (M83801), as reported in GenBank. Partial E canis P28.1 and P28.2 amino acid sequences from Thai dogs were divergent from analogous sequences derived from North American E canis (AF082744) strains, suggesting that the Thai dogs were infected with a geographically distinct strain of E canis compared to North American strains. The results of this study indicate that dogs in Thailand have substantial exposure to vectorborne diseases and that coinfection with these pathogens may be common.     

Rickettsiae in arthropods collected from red foxes (Vulpes vulpes) in France

The aim of our study was to detect the presence of Rickettsia spp. and Bartonella spp. in ticks and fleas collected from red foxes (Vulpes vulpes) in southeastern France during 2008. Using a genus-specific quantitative PCR (qPCR) assay, which was followed by a species-specific qPCR assay for the positive samples, 45.2% (33/73) of ticks (Rhipicephalus turanicus) were found to be infected with Rickettsia massiliae. 10.5% (2/19) of the fleas (Archaeopsylla erinacei) collected in the study tested positive for Rickettsia felis. A genus-specific qPCR assay did not reveal any Bartonella species in any of the ticks or fleas collected. The role of red foxes in the epidemiology of spotted fever caused by Rickettsiae species requires further investigation.     

Flea-borne pathogens in the cat flea Ctenocephalides felis and their association with mtDNA diversity of the flea host

Flea-borne pathogens were screened from 100 individual cat fleas using a PCR approach, of which 38 % were infected with at least one bacterium. Overall, 28 % of the flea samples were positive for Bartonella as inferred from ITS DNA region. Of these, 25 % (7/28) were identified as Bartonella clarridgeiae, 42.9 % (12/28) as Bartonella henselae consisted of two different strains, and 32.1 % (9/28) as Bartonella koehlerae, which was detected for the first time in Malaysia. Sequencing of gltA amplicons detected Rickettsia DNA in 14 % of cat flea samples, all of them identified as Rickettsia asembonensis (100 %). None of the flea samples were positive for Mycoplasma DNA in 16S rRNA gene detection. Four fleas were co-infected with Bartonella and Rickettsia DNAs. Statistical analyses reveal no significant association between bacterial infection and mtDNA diversity of the cat flea. Nevertheless, in all types of pathogen infections, infected populations demonstrated lower nucleotide and haplotype diversities compared to uninfected populations. Moreover, lower haplotype numbers were observed in infected populations.     

Rocky Mountain spotted fever in an urban canine population

A survey for the prevalence of Rocky Mountain spotted fever antibodies in dogs from a well-defined endemic area in Columbus, Ohio (Franklin County), was conducted during the summer of 1981. Seventy-three blood samples from dogs in this area were tested by microimmunofluorescence for antibodies to Rickettsia rickettsii and other rickettsial antigens. Thirty-three (45.2%) of these samples were positive for R rickettsii, with titers ranging from 1:8 to 1:2,048. For comparison, 137 blood samples from dogs at the Franklin County Dog Pound also were tested. One dog (0.7%) from the comparison group was seropositive to R rickettsii, with a 1:64 titer. The results indicated a nidus of this disease within the city of Columbus, Ohio.     

High prevalence of “Candidatus Rickettsia amblyommii” in Amblyomma ticks from a Spotted Fever Endemic Region in North Argentina

Ticks from an endemic Spotted Fever region in Argentina were analysed by PCR for Spotted Fever Group Rickettsiae. DNA of “Candidatus Rickettsia amblyommii” was found in 21.3% of Amblyomma hadanii and in 44.0% of A. neumanni. Amblyomma sculptum (formerly A. cajennense) and Haemaphysalis juxtakochi were negative for rickettsial DNA. DNA of Rickettsia rickettsii, the etiological agent of the clinical cases reported within the studied region was not detected in the analysed sample.