SUMMARY: Tick-borne rickettsioses are caused by obligate intracellular bacteria belonging to the spotted fever group of the genusRickettsia. These zoonoses are among the oldest known vector-borne diseases. However, in the past 25 years, the scope and importance of the recognized tick-associated rickettsial pathogens have increased dramatically, making this complex of diseases an ideal paradigm for the understanding of emerging and reemerging infections. Several species of tick-borne rickettsiae that were considered nonpathogenic for decades are now associated with human infections, and novelRickettsiaspecies of undetermined pathogenicity continue to be detected in or isolated from ticks around the world. This remarkable expansion of information has been driven largely by the use of molecular techniques that have facilitated the identification of novel and previously recognized rickettsiae in ticks. New approaches, such as swabbing of eschars to obtain material to be tested by PCR, have emerged in recent years and have played a role in describing emerging tick-borne rickettsioses. Here, we present the current knowledge on tick-borne rickettsiae and rickettsioses using a geographic approach toward the epidemiology of these diseases.

The rapid and reliable identification of arthropod vector species is an essential component of the fight against vector-borne diseases. However, owing to the lack of entomological expertise required for the morphological identification method, development of alternative and complementary tools is needed. This review describes the main methods used for arthropod identification, focusing on the emergence of protein profiling using MALDI-TOF MS technology. Sample preparation, analysis of reproducibility, database creation and blind tests for controlling accuracy of this tool for arthropod identification are described. The advantages and limitations of the MALDI-TOF MS method are illustrated by emphasizing different hematophagous arthropods, including mosquitoes and ticks, the top two main vectors of infectious diseases.

Ticks are considered the second vector of human and animal diseases after mosquitoes. Therefore, identification of ticks and associated pathogens is an important step in the management of these vectors. In recent years, Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) has been reported as a promising method for the identification of arthropods including ticks. The objective of this study was to improve the conditions for the preparation of tick samples for their identification by MALDI-TOF MS from field-collected ethanol-stored Malian samples and to evaluate the capacity of this technology to distinguish infected and uninfected ticks. A total of 1,333 ticks were collected from mammals in three distinct sites from Mali. Morphological identification allowed classification of ticks into 6 species including Amblyomma variegatum, Hyalomma truncatum, Hyalomma marginatum rufipes, Rhipicephalus (Boophilus) microplus, Rhipicephalus evertsi evertsi and Rhipicephalus sanguineus sl. Among those, 471 ticks were randomly selected for molecular and proteomic analyses. Tick legs submitted to MALDI-TOF MS revealed a concordant morpho/molecular identification of 99.6%. The inclusion in our MALDI-TOF MS arthropod database of MS reference spectra from ethanol-preserved tick leg specimens was required to obtain reliable identification. When tested by molecular tools, 76.6%, 37.6%, 20.8% and 1.1% of the specimens tested were positive for Rickettsia spp., Coxiella burnetii, Anaplasmataceae and Borrelia spp., respectively. These results support the fact that MALDI-TOF is a reliable tool for the identification of ticks conserved in alcohol and enhances knowledge about the diversity of tick species and pathogens transmitted by ticks circulating in Mali. Ticks are among the most important vectors and reservoirs of several animal and human pathogens such as viruses, bacteria and protozoa. However, very few studies have been done on ticks in Mali. At present, little information is available about tick species infesting livestock or human tick-borne diseases transmitted in Mali. The identification of tick species and the determination of pathogens associated are essential to evaluate epidemiology and risks of human and animal diseases: the One Health approach. Current identification methods are time consuming, expensive and laborious. Previous studies have shown that MALDI-TOF mass spectrometry analyses may allow accurate tick species identification. A recent study suggested that it was possible to identify ticks preserved in alcohol by MALDI-TOF MS. The aim of the present study was to improve tick leg sample preparation conditions for their identification by MALDI-TOF MS from Malian ethanol-preserved specimens collected in the field. This study provided 99.4% concordance between morphological and MALDI-TOF identification. The detection of microorganisms was also performed by molecular biology revealing the presence of the presence of Rickettsia spp., Coxiella burnetii, Borrelia spp. and Anaplasmataceae. These results support the use of MALDI-TOF MS in entomology, tick diseases epidemiology and improve the knowledge of tick species-diversity and tick-borne pathogens circulating in Mali.

A method for rapid species identification of ticks may help clinicians predict the disease outcomes of patients with tick bites and may inform the decision as to whether to administer postexposure prophylactic antibiotic treatment. We aimed to establish a matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) spectrum database based on the analysis of the legs of six tick vectors: Amblyomma variegatum , Rhipicephalus sanguineus , Hyalomma marginatum rufipes , Ixodes ricinus , Dermacentor marginatus , and Dermacentor reticulatus . A blind test was performed on a trial set of ticks to identify specimens of each species. Subsequently, we used MALDI-TOF MS to identify ticks obtained from the wild or removed from patients. The latter tick samples were also identified by 12S ribosomal DNA (rDNA) sequencing and were tested for bacterial infections. Ticks obtained from the wild or removed from patients ( R. sanguineus , I. ricinus , and D. marginatus ) were accurately identified using MALDI-TOF MS, with the exception of those ticks for which no spectra were available in the database. Furthermore, one damaged specimen was correctly identified as I. ricinus , a vector of Lyme disease, using MALDI-TOF MS only. Six of the 14 ticks removed from patients were found to be infected by pathogens that included Rickettsia , Anaplasma , and Borrelia spp. MALDI-TOF MS appears to be an effective tool for the rapid identification of tick vectors that requires no previous expertise in tick identification. The benefits for clinicians include the more targeted surveillance of patients for symptoms of potentially transmitted diseases and the ability to make more informed decisions as to whether to administer postexposure prophylactic treatment.

In recent years, the scope and importance of emergent vector-borne diseases has increased dramatically. In Algeria, only limited information is currently available concerning the presence and prevalence of these zoonotic diseases. For this reason, we conducted a survey of hematophagous ectoparasites of domestic mammals and/or spleens of wild animals in El Tarf and Souk Ahras, Algeria. Using real-time PCR, standard PCR and sequencing, the presence of Bartonella spp., Rickettsia spp., Borrelia spp. and Coxiella burnetii was evaluated in 268/1626 ticks, 136 fleas, 11 Nycteribiidae flies and 16 spleens of domestic and/or wild animals from the El Tarf and Souk Ahras areas. For the first time in Algeria, Bartonella tamiae was detected in 12/19 (63.2 %) Ixodes vespertilionis ticks, 8/11 (72.7 %) Nycteribiidae spp. flies and in 6/10 (60 %) bat spleens (Chiroptera spp.). DNA from Coxiella burnetii, the agent of Q fever, was also identified in 3/19 (15.8 %) I. vespertilionis from bats. Rickettsia slovaca, the agent of tick-borne lymphadenopathy, was detected in 1/1 (100 %) Haemaphysalis punctata and 2/3 (66.7 %) Dermacentor marginatus ticks collected from two boars (Sus scrofa algira) respectively. Ri. massiliae, an agent of spotted fever, was detected in 38/94 (40.4 %) Rhipicephalus sanguineus sensu lato collected from cattle, sheep, dogs, boars and jackals. DNA of Ri. aeschlimannii was detected in 6/20 (30 %) Hyalomma anatolicum excavatum and 6/20 (30 %) Hy. scupense from cattle. Finally, Ri. felis, an emerging rickettsial pathogen, was detected in 80/110 (72.7 %) Archaeopsylla erinacei and 2/2 (100 %) Ctenocephalides felis of hedgehogs (Atelerix algirus). In this study, we expanded knowledge about the repertoire of ticks and flea-borne bacteria present in ectoparasites and/or tissues of domestic and wild animals in Algeria.

The identification of mosquito vectors is typically based on morphological characteristics using morphological keys of determination, which requires entomological expertise and training. The use of protein profiling by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS), which is increasingly being used for the routine identification of bacteria, has recently emerged for arthropod identification. To investigate the usefulness of MALDI-TOF-MS as a mosquito identification tool, we tested protein extracts made from mosquito legs to create a database of reference spectra. The database included a total of 129 laboratory-reared and field-caught mosquito specimens consisting of 20 species, including 4 Aedes spp., 9 Anopheles spp., 4 Culex spp., Lutzia tigripes, Orthopodomyia reunionensis and Mansonia uniformis. For the validation study, blind tests were performed with 76 specimens consisting of 1 to 4 individuals per species. A cluster analysis was carried out using the MALDI-Biotyper and some spectra from all mosquito species tested. Biomarker mass sets containing 22 and 43 masses have been detected from 100 specimens of the Anopheles, Aedes and Culex species. By carrying out 3 blind tests, we achieved the identification of mosquito vectors at the species level, including the differentiation of An. gambiae complex, which is possible using MALDI-TOF-MS with 1.8 as the cut-off identification score. A cluster analysis performed with all available mosquito species showed that MALDI-Biotyper can distinguish between specimens at the subspecies level, as demonstrated for An gambiae M and S, but this method cannot yet be considered a reliable tool for the phylogenetic study of mosquito species. We confirmed that even without any specific expertise, MALDI-TOF-MS profiling of mosquito leg protein extracts can be used for the rapid identification of mosquito vectors. Therefore, MALDI-TOF-MS is an alternative, efficient and inexpensive tool that can accurately identify mosquitoes collected in the field during entomological surveys.