Immunity of fleas (Order Siphonaptera).

The immune response of arthropod vectors plays a key role in the spread and transmission of vector-borne diseases. Although fleas transmit several human pathogens (e.g., Bartonella henselae, Rickettsia felis, R. typhi, and Yersinia pestis), few studies have examined how these vectors respond to infection. In hematophagous arthropods, imbibed pathogens must survive the hostile environment of blood meal digestion, which includes proteolytic digestive enzymes, protease inhibitors and expression of genes associated with protection of epithelial linings. Additionally, insect epithelial cells exhibit local immune defense against ingested pathogens by producing antimicrobial peptides and reactive oxygen species. This review details these and other aspects of insect immunity as it relates to fleas, with an emphasis on the gut immune response to two blood-borne pathogens, R. typhi and Y. pestis.

A 2015 outbreak of flea-borne rickettsiosis in San Gabriel Valley, Los Angeles County, California.

Although flea-borne rickettsiosis is endemic in Los Angeles County, outbreaks are rare. In the spring of 2015 three human cases of flea-borne rickettsiosis among residents of a mobile home community (MHC) prompted an investigation. Fleas were ubiquitous in common areas due to presence of flea-infested opossums and overabundant outdoor cats and dogs. The MHC was summarily abated in June 2015, and within five months, flea control and removal of animals significantly reduced the flea population. Two additional epidemiologically-linked human cases of flea-borne rickettsiosis detected at the MHC were suspected to have occurred before control efforts began. Molecular testing of 106 individual and 85 pooled cat fleas, blood and ear tissue samples from three opossums and thirteen feral cats using PCR amplification and DNA sequencing detected rickettsial DNA in 18.8% of the fleas. Seventeen percent of these cat fleas tested positive for R. felis-specific DNA compared to under two (<2) percent for Candidatus R. senegalensis-specific DNA. In addition, serological testing of 13 cats using a group-specific IgG-ELISA detected antibodies against typhus group rickettsiae and spotted fever group rickettsiae in six (46.2%) and one (7.7%) cat, respectively. These results indicate that cats and their fleas may have played an active role in the epidemiology of the typhus group and/or spotted fever group rickettsial disease(s) in this outbreak.

Rickettsial seropositivity in the indigenous community and animal farm workers, and vector surveillance in Peninsular Malaysia.

Rickettsioses are emerging zoonotic diseases that are often neglected in many countries in Southeast Asia. Rickettsial agents are transmitted to humans through exposure to infected arthropods. Limited data are available on the exposure of indigenous community and animal farm workers to the aetiological agents and arthropod vectors of rickettsioses in Peninsular Malaysia. Serological analysis of Rickettsia conorii and Rickettsia felis was performed for 102 individuals from the indigenous community at six rural villages and 87 workers from eight animal farms in Peninsular Malaysia in a cross-sectional study. The indigenous community had significantly higher seropositivity rates for R. conorii (P<0.001) and R. felis (P<0.001), as compared to blood donors from urban (n=61). Similarly, higher seropositivity rates for R. conorii (P=0.046) and R. felis (P<0.001) were noted for animal farm workers, as compared to urban blood donors. On the basis of the sequence analysis of gltA, ompA and ompB, various spotted fever group rickettsiae closely related to R. raoultii, R. heilongjiangensis, R. felis-like organisms, R. tamurae, Rickettsia sp. TCM1, R. felis, Rickettsia sp. LON13 and R. hulinensis were identified from tick/flea samples in animal farms, indigenous villages and urban areas. This study describes rickettsial seropositivity of the Malaysian indigenous community and animal farm workers, and provides molecular evidence regarding the presence of rickettsial agents in ticks/fleas infesting domestic animals in Peninsular Malaysia.

Serological evidence of exposure to Rickettsia felis and Rickettsia typhi in Australian veterinarians.

BACKGROUND: Rickettsia felis and Rickettsia typhi are emerging arthropod-borne zoonoses causing fever and flu-like symptoms. Seroprevalence and risk factors associated with exposure to these organisms was explored in Australian veterinarians. METHODS: One hundred and thirty-one veterinarians from across Australia were recruited to participate in a cross-sectional survey. Veterinarians provided a single blood sample and answered a questionnaire on potential risk factors influencing their exposure to R. felis and R. typhi. Indirect microimmunofluorescence antibody testing (IFAT) was used to identify evidence of serological exposure of the participants to R. felis and R. typhi. Results were analyzed and a logistical regression model performed to predict risk factors associated with seropositivity. RESULTS: In total, 16.0% of participants were seropositive to R. felis, 4.6% to R. typhi and 35.1% seropositive to both, where cross-reactivity of the IFAT between R. felis and R. typhi precluded a definitive diagnosis. Veterinarians residing within the south-eastern states of Victoria and Tasmania were at a higher risk of exposure to R. felis or generalised R. felis or R. typhi exposure. Older veterinarians and those that recommended flea treatment to their clients were found to be significantly protected from exposure. CONCLUSIONS: The high exposure to R. felis amongst veterinary professionals suggests that flea-borne spotted fever is an important cause of undifferentiated fever conditions that may not be adequately recognized in Australia.

Caso probable de fiebre manchada ( Rickettsia felis) transmitida por pulgas / Probable case of flea-borne spotted fever ( Rickettsia felis )

Rickettsia felis es el agente etiológico de la fiebre manchada transmitida por pulgas, cuyo principal vector y reservorio es Ctenocephalides felis . Típicamente, la enfermedad se presenta como fiebre aguda asociada a cefalea, astenia, exantema máculo-papular generalizado y, en algunos casos, con escara de inoculación. En los últimos años, R. felis ha venido adquiriendo un papel importante en la etiología del síndrome febril agudo, calificándola como una enfermedad emergente y subdiagnosticada. La inmunofluorescencia indirecta es actualmente el método diagnóstico de referencia. Sin embargo, esta técnica presenta limitaciones relacionadas con la reacción cruzada que existe entre las diferentes especies del género Rickettsia . En el presente reporte se describe el caso de un paciente de 16 años con síndrome febril agudo secundario a infección probable por R. felis .

Rickettsia felis is the etiologic agent of flea-borne spotted fever, with Ctenocephalides felis as its main vector and reservoir. Typically, the disease presents as acute fever associated with headache, asthenia, generalized maculo-papular rash, and in some cases, an inoculation eschar. In recent years, R. felis has acquired an important role in the etiology of the acute febrile syndrome; it is indeed an emerging infectious disease, albeit underdiagnosed. Indirect immunofluorescence assay (IFA) is currently the reference diagnostic method. However, this technique has limitations related to the cross reactivity among different species of rickettsiae. Herein, we describe a case of a 16 year-old patient with an acute febrile syndrome secondary to probable infection with R. felis.

Seroprevalence and risk factors for Rickettsia felis exposure in dogs from Southeast Queensland and the Northern Territory, Australia.

BACKGROUND: The recent detection of Rickettsia felis DNA in dogs in Australia suggests that dogs are potential mammalian reservoir hosts for this emerging rickettsia. To date, there is no published report addressing the seroprevalence of R. felis in dogs in Australia. METHODS: Antigens for R. felis were produced by inoculating confluent XTC-2 monolayer cell cultures with three pools of cat flea (Ctenocephalides felis) homogenates. Infection was confirmed by real-time (qPCR), conventional or nested PCRs targeting the ompB, gltA, 17 kDa and ompA genes. Two hundred and ninety-two dogs from Southeast Queensland and the Northern Territory were tested for the presence of R. felis antibodies using a microimmunofluorescence (IF) test and the seroprevalence and associated risk factors for exposure were determined using both uni- and multi-variate analyses. RESULTS: Rickettsia felis was successfully isolated in cell culture from all three cat-flea pools. One hundred and forty-eight dogs (50.7%) showed seropositivity with titres ≥64 and 54 (18.5%) with titres ≥128. At antibody titres ≥64, dogs with active ectoparasite control were less likely to be seropositive to R. felis (OR: 2.60; 95% CI: 1.20 - 5.56). CONCLUSIONS: This first reported isolation of R. felis in cell culture in Australia allowed for the production of antigen for serological testing of dogs. Results of this serological testing reflects the ubiquitous exposure of dogs to R. felis and advocate for owner vigilance with regards to ectoparasite control on domestic pets.

[Probable case of flea-borne spotted fever (Rickettsia felis)]. / Caso probable de fiebre manchada (Rickettsia felis) transmitida por pulgas.

Rickettsia felis is the etiologic agent of flea-borne spotted fever, with Ctenocephalides felis as its main vector and reservoir. Typically, the disease presents as acute fever associated with headache, asthenia, generalized maculo-papular rash, and in some cases, an inoculation eschar. In recent years, R. felis has acquired an important role in the etiology of the acute febrile syndrome; it is indeed an emerging infectious disease, albeit underdiagnosed. Indirect immunofluorescence assay (IFA) is currently the reference diagnostic method. However, this technique has limitations related to the cross reactivity among different species of rickettsiae. Herein, we describe a case of a 16 year-old patient with an acute febrile syndrome secondary to probable infection with R. felis.

Seroprevalence of Rickettsia bellii and Rickettsia felis in dogs, São José dos Pinhais, State of Paraná, Brazil.

Brazilian spotted fever (BSF) is a vector-borne zoonosis caused by Rickettsia rickettsii bacteria. Dogs can be host sentinels for this bacterium. The aim of the study was to determine the presence of antibodies against Rickettsia spp. in dogs from the city of São José dos Pinhais, State of Paraná, Southern Brazil, where a human case of BSF was first reported in the state. Between February 2006 and July 2007, serum samples from 364 dogs were collected and tested at 1:64 dilutions by indirect immunofluorescence assay (IFA) against R. rickettsii and R. parkeri. All sera that reacted at least to one of Rickettsia species were tested against the six main Rickettsia species identified in Brazil: R. rickettsii, R. parkeri, R. bellii, R. rhipicephali, R. amblyommii and R. felis. Sixteen samples (4.4%) reacted to at least one Rickettsia species. Among positive animals, two dogs (15.5%) showed suggestive titers for R. bellii exposure. One sample had a homologous reaction to R. felis, a confirmed human pathogen. Although Rickettsia spp. circulation in dogs in the area studied may be considered at low prevalence, suggesting low risk of human infection, the present data demonstrate for the first time the exposure of dogs to R. bellii and R. felis in Southern Brazil.

Seroprevalence of Rickettsia bellii and Rickettsia felis in dogs, São José dos Pinhais, State of Paraná, Brazil / Soroprevalência de Rickettsia bellii e Rickettsia felis em cães, São José dos Pinhais, Paraná, Brasil

Brazilian spotted fever (BSF) is a vector-borne zoonosis caused by Rickettsia rickettsii bacteria. Dogs can be host sentinels for this bacterium. The aim of the study was to determine the presence of antibodies against Rickettsia spp. in dogs from the city of São José dos Pinhais, State of Paraná, Southern Brazil, where a human case of BSF was first reported in the state. Between February 2006 and July 2007, serum samples from 364 dogs were collected and tested at 1:64 dilutions by indirect immunofluorescence assay (IFA) against R. rickettsii and R. parkeri. All sera that reacted at least to one of Rickettsia species were tested against the six main Rickettsia species identified in Brazil: R. rickettsii, R. parkeri, R. bellii, R. rhipicephali, R. amblyommii and R. felis. Sixteen samples (4.4 percent) reacted to at least one Rickettsia species. Among positive animals, two dogs (15.5 percent) showed suggestive titers for R. bellii exposure. One sample had a homologous reaction to R. felis, a confirmed human pathogen. Although Rickettsia spp. circulation in dogs in the area studied may be considered at low prevalence, suggesting low risk of human infection, the present data demonstrate for the first time the exposure of dogs to R. bellii and R. felis in Southern Brazil.

A febre maculosa brasileira (FMB) é uma zoonose veiculada por carrapatos e causada pela bactéria Rickettsia rickettsii, podendo os cães ser hospedeiros sentinelas para essa bactéria. O objetivo do estudo foi determinar a presença de anticorpos contra Rickettsia spp. em cães de São José dos Pinhais, estado do Paraná, Sul do Brasil. Entre fevereiro de 2006 e julho de 2007, amostras séricas de 364 cães foram coletadas e testadas na diluição de 1:64 por Reação de Imunofluorescência Indireta (RIFI) contra R. rickettsii e R. parkeri. Todos os soros reagentes para pelo menos uma espécie de Rickettsia foram testados contra as seis principais espécies de Rickettsia identificadas no Brasil: R. rickettsii, R. parkeri, R. bellii, R. rhipicephali, R. amblyommii e R. felis. Dezesseis amostras (4,4 por cento) reagiram para pelo menos uma espécie de Rickettsia. Dos animais positivos, dois cães (15,5 por cento) apresentaram títulos sugestivos de exposição a R. bellii. Uma amostra apresentou reação homóloga frente à R. felis, um agente patogênico confirmado para seres humanos. Muito embora os resultados demonstrem uma baixa prevalência de Rickettsia spp. em cães, sugerindo um baixo risco de infecção humana, este estudo relatou pela primeira vez a evidência de exposição a R. bellii e R. felis em cães no Sul do Brasil.

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.

Rickettsia felis infection, Tunisia.

We report, for the first time, serologic evidence of Rickettsia felis and R. aeschlimannii infections acquired in Tunisia from 1998 to 2003. We found that most patients with antibodies against both R. conorii and R. typhi had serologic evidence of R. felis infection.

Serological evidence of infection with Rickettsia typhi and Rickettsia felis among the human population of Catalonia, in the northeast of Spain.

Murine typhus (MT) is a cause of fever of intermediate duration in the south of Spain. Rickettsia typhi has been described as the MT etiological agent. Rickettsia felis produces an infection similar to MT. The aim of the study is to determine their seroprevalence in humans in Catalonia. Antibodies to Rickettsia typhi and Rickettsia felis from 217 serum samples were examined by indirect immunofluorescence assay (IFA). Age, gender, residence area, contact with animals, and occupation were surveyed. Rickettsia typhi and Rickettsia felis seroprevalences were 8.8% and 3.2%, respectively. Rickettsia typhi was present in 7.6% of the samples in urban, 8.5% in semirural, and in 21.4% in rural areas, whereas Rickettsia felis was present in 3.5% in urban, 1.7% in semirural, and 7.1% in rural area. The only statistically significant association observed was that between Rickettsia felis seropositivity and age. Our data seem to indicate the presence of Rickettsia typhi and Rickettsia felis in humans in Catalonia.

Human Rickettsia felis infection, Canary Islands, Spain.

We report the first cases of human infection by Rickettsia felis in the Canary Islands. Antibodies against R. felis were found in 5 adsorbed serum samples from 44 patients with clinically suspected rickettsiosis by Western blot serology. Fleas from 1 patient's dog were positive for R. felis by polymerase chain reaction.

Detection of R. felis and R. typhi in fleas using monoclonal antibodies.

Rickettsia typhi and R. felis are flea-transmitted human pathogenic rickettsial species. To investigate the distributional dynamics of these rickettsiae we designed a micro-immunofluorescence assay (MIF) using species-specific monoclonal antibodies (MAbs) applied to flea cryosections. Our assay was performed in less than 3 h and its applicability was demonstrated by the detection of R. typhi in 50 artificially infected human body lice but in none of 50 uninfected lice. With MIF, we identified 31 positive among 32 fleas proven with PCR to be naturally infected with R. felis; and 7 positive among 32 fleas proven with PCR to be naturally infected with R. typhi. No cross-detection was observed with both MAbs. Fresh R. felis-infected fleas were significantly more MIF-positive than long conserved R. typhi-infected fleas (31/32 vs. 7/32, P < 0.01). This discrepancy may be linked to degradation of antigens by long-term freezing. For R. typhi-infected fleas, our assay was significantly more efficient when applied to fleas in early stages of infection (less than 15 days) by comparison with fleas frozen more than 20 days after infection (7/15 vs. 0/17, P = 0.01). This difference may be related to an antigenic modification caused by selection pressure in the vector and host process. The sensitivity of the described method did not exceed 47% (7/15) for R. typhi but, in contrast, was 97% for R. felis. Thus, our method appears to be useful for surveillance in R. felis infections, but requires further studies for the detection of R. typhi.

Antigenic classification of Rickettsia felis by using monoclonal and polyclonal antibodies.

Rickettsia felis is a flea-transmitted rickettsia. There is a discrepancy between its reported phylogenic and phenotypic identifications. Following the first report of R. felis, it was considered by tests with serologic reagents to be closely related to another recognized flea-transmitted rickettia, R. typhi. Subsequently, it appeared to be more closely related to spotted fever group (SFG) rickettsiae by genetic analysis. In the present work, R. felis was studied by microimmunofluorescence (MIF) serologic typing and with monoclonal antibodies (MAbs). Mouse polyclonal antisera to R. felis cross-reacted only with SFG rickettsiae. A neighbor-joining analysis based on MIF indicated that R. felis is actually related to SFG rickettsiae antigenically, clustering with R. australis, R. akari, and R. montanensis. A panel of 21 MAbs was raised against a 120-kDa protein antigen or a 17-kDa polypeptide of R. felis. They cross-reacted with most members of the SFG rickettsiae but not with R. prowazekii, R. typhi, or R. canadensis of the typhus group (TG) rickettsiae. Sixty-four MAbs previously generated to seven other ricketttsial species were tested with R. felis. Three MAbs reacted with the 120-kDa antigen and were generated by R. africae, R. conorii, and R. akari, respectively. They exhibited cross-reactivities with R. felis. All our data show that R. felis harbors the antigenic profile of an SFG rickettsia.