Fleas and flea-borne diseases of North Africa

Fleas and flea-borne diseases of North Africa. 1 Basma El Hamzaoui1,2; Antonio Zurita3*; Cristina Cutillas3, Philippe Parola1,2 2 1Aix Marseille Univ; IRD; AP-HM; SSA; VITROME; 3 2IHU Méditerranée Infection; Marseille; France 4 3Department of Microbiology and Parasitology. Faculty of Pharmacy. University of Seville. 5 Profesor García González 2, 41012 Seville, Spain. 6 Emails: BE: [email protected]; AZ: [email protected]; CC: cutill[email protected]; PP : 7 [email protected] 8 *Corresponding author: Dr. Antonio Zurita. 3Department of Microbiology and Parasitology. 9 Faculty of Pharmacy. University of Seville. Profesor García González 2, 41012 Seville, Spain. 10 Email address: [email protected] 11 12 13 14 15 16 17 18 19 20 Content 21 Abstract 22 Introduction 23 Fleas of medical-veterinary importance in North Africa 24  Pulex irritans 25  Echidnophaga gallinacea 26  Ctenocephalides felis 27  Ctenocephalides canis 28  Spilopsyllus cuniculi 29  Xenopsylla cheopis 30  Archaeopsylla erinacei 31 Flea-borne diseases of North Africa 32  Plague 33  Flea-borne rickettsial diseases 34  Murine typhus 35  Rickettsia felis infection and flea-borne spotted fever 36  Bartonelloses 37  Cat-scratch disease 38  Trench fever 39  Parasitic diseases 40  Dipylidium caninum infection or Dipylidiasis 41  Flea-borne viral diseases 42 Means of flea control and identification 43 44 Abstract 45 North Africa has an interesting and rich wildlife including hematophagous arthropods, and 46 specifically fleas, which constitute a large part of the North African fauna, and are recognised 47 vectors of several zoonotic bacteria. Flea-borne organisms are widely distributed throughout 48 the world in endemic disease foci, where components of the enzootic cycle are present. 49 Furthermore, flea-borne diseases could re-emerge in epidemic form because of changes in the 50 vector–host ecology due to environmental and human behaviour modifications. We need to 51 know the real incidences of flea-borne diseases in the world due to this incidence could be much 52 greater than are generally recognized by physicians and health authorities. As a result, diagnosis 53 and treatment are often delayed by health care professionals who are unaware of the presence 54 of these infections and thus do not take them into consideration when attempting to determine 55 the cause of a patient’s illness. In this context, this bibliographic review aims to summarise the 56 main species of fleas present in North Africa, their geographical distribution, flea-borne 57 diseases, and their possible re-emergence. 58 59 Keywords: Fleas, North Africa, flea-borne diseases, vectors 60 61 List of abbreviations: 62 MALDI–TOF MS: Matrix Assisted Laser Desorption Ionisation – Time Of Flight Mass 63 Spectrometry 64 Introduction 65 Fleas (Insecta, Siphonaptera) are obligate hematophagous ectoparasites. They are wingless 66 insects which are small (2-10 mm) and generally present a laterally flattened body. Fleas have 67 three thoracic segments, each with a pair of well-developed legs (Beaucournu and Launay, 68 1990; Bitam et al., 2010) enabling adults to jump long distances (Bitam et al., 2010). They are 69 ectoparasites that usually infest mammals and rodents but rarely birds. Flea species distribution 70 extends to the seven continents, including Antarctica (Whiting et al., 2008). Furthermore, fleas 71 are able to inhabit a very wide range of habitats and hosts (Whiting, 2002). The greatest 72 diversity of species can be observed in the temperate regions of the globe (Lewis, 1993). 73 Fleas are holometabolous insects that complete their cycle from egg to adult via three larval 74 stages and a pupal stage (Zakson-Aiken et al., 1996). The completion of the entire life cycle 75 varies among species but takes on average three to five weeks depending on the temperature 76 and humidity conditions. Flea larvae are vermiform and legless, with chewing mouthparts 77 (Dryden and Rust, 1994). 78 Flea species adapt to their hosts but do not do so exclusively. This fact could explain the role 79 of the so-called rat flea Xenopsylla cheopis (Rothschild, 1903) in the epidemiology of human 80 plague, or the existence of highly promiscuous fleas species, such as the so-called human flea, 81 Pulex irritans (Linnaeus, 1758) or the cat flea Ctenocephalides felis (Bouché, 1835) which 82 occur on a wide variety of carnivorous animal species (Gratz, 1999). 83 Fleas’ behaviour towards their hosts makes it possible to classify them in three categories, 84 including i) fleas that live permanently on their host, such as X. cheopis, P. irritans, 85 Ctenocephalides canis and C. felis, ii) fleas that permanently live in their hosts’ nest or burrows, 86 parasitizing them only during blood meals, such as Ceratophyllus gallinae; iii) and the so-called 87 penetrating fleas, such as females of the genera Tunga and Neotunga, which are able to burrow 88 into their hosts’ dermal tissue, where they increase dramatically in size (up to l000-fold) 89 accompanied by an extensive morphological degeneration (Durden and Traub, 2002). Females 90 of Echidnophaga gallinacea can also fix themselves around the eyes of poultry after 91 fertilization (Franc, 1994a). 92 Within Siphonaptera, the Pulicidae family exhibits an interesting diversity of host specificity 93 patterns and ecological habits (Beutel et al., 2008). Within this family, P. irritans, C. felis and 94 even X. cheopis have been the most studied species due to their cosmopolitan distribution 95 together with the fact that these species are closely related to humans (Durden and Traub, 2002). 96 The importance of fleas in human public health is mostly related to their ability to transmit 97 infectious disease agents during the blood meal. Some fleas are indeed vectors of human 98 infectious diseases, such as bubonic plague, caused by Yersinia pestis (Zeppelini et al., 2016), 99 murine typhus, caused by Rickettsia typhi (Peniche Lara et al., 2012) and flea-borne spotted 100 fever caused by Rickettsia felis (Angelakis et al., 2016) . They are also probably involved in 101 the transmission of Bartonella henselae the agent of cat-scratch disease (Bitam et al., 2010; 102 Chomel and Kasten, 2010). 103 “North Africa” is a collective term including Mediterranean countries and territories situated 104 in the northern-most region of the African continent, including Morocco, Algeria, Tunisia, 105 Libya and Egypt. It is a total area of around five million kilometres, more than 90% of which 106 is desert (Peel et al., 2007). North Africa is vulnerable to various climates, with the coast being 107 characterised by a Mediterranean climate of wet winters and dry summers, while non-coastal 108 areas are characterised by an arid desert climate with generally hot summers, cold winters and 109 little rainfall (Radhouane, 2013). This climatic diversification contributes to the richness of the 110 North African fauna, including fleas. 111 In this review, we focus on fleas of veterinary and clinical importance from North Africa, 112 particularly those species belonging to the Pulicidae family. We also discuss innovative 113 methods for the identification of fleas and their associated pathogens. 114 Fleas of medical-veterinary importance in North Africa 115 The Pulicidae family consists of four tribes, 21 genera and 167 species (Beutel et al., 2008). 116 Some authors (Lewis, 1998) considered Pulicidae as including Tungidae. However, Whiting et 117 al. (2008) placed this family as a monophyletic and phylogenetically distant group from 118 Tungidae (Beutel et al., 2008). Most fleas of veterinary importance are grouped in this family 119 since they may act as vectors of some infectious diseases, may play a role as intermediate hosts 120 for several parasites, and many cause allergic reactions in animals and humans, associated with 121 their bloodsucking habits (Dobler and Pfeffer, 2011). 122  Pulex irritans (Figure 1.1): 123 This flea has a cosmopolitan distribution and is often referred as the “human flea”. However, 124 this species often parasitizes a wide variety of hosts, including large wild mammals and rodents, 125 although rarely found on rats (Gratz, 1999). Under natural conditions, this flea is active 126 throughout the year, with a peak in summer (Beaucournu and Launay, 1990). Its legs are 127 sufficiently developed to jump up to 50 cm and it has the particularity of being able to survive 128 for a year without feeding, since this flea is capable of absorbing quantities of blood equivalent 129 to 20 times its weight (Belthoff et al., 2015). 130 The presence of this flea in North Africa has been reported several times. P. irritans was 131 collected on dogs from Tunisia (Tunis, Maktar, Siliana). P.irritans was also found in north132 western Libya (Tripoli) on eight farm dogs (Kaal et al., 2006), and in Morocco (Agadir, 133 Casablanca,Tiznit) (Boudebouch et al., 2011). G. Blanc and M. Baltazard (1945) long suspected 134 the role of P. irritans in the transmission of the plague in Morocco (Audouin-Rouzeau, 2003). 135 They conducted nine experiments with naturally infected P. irritans on plague victims. In the 136 first eight experiments, all guinea pigs were not infected, even when 240 fleas were used. For 137 their ninth experiment, G. Blanc and M. Baltazard (1945) collected 720 fleas from six plague 138 victims, and one guinea pig bitten by a flea was infected. This experiment enabled the authors 139 to state that infected human fleas could transmit diseases (Audouin-Rouzeau, 2003). 140  Echidnophaga gallinacea (Figure 1.2): 141 This species is commonly known as the “Stick tight flea” and is about 2 mm long. Females of 142 this species are able to remain attached for up to six weeks to a single host site, causing 143 ulceration at the attachment site. The eggs are then deposited in the ulcers that are formed on 144 the skin of the host. Later, the larvae fall to the ground and feed on any organic debris found 145 (Boughton et al., 2006). In many cases, a large number of fleas can congregate around the eyes 146 and on the bare skin of poultry, which makes it difficult to remove them (Elston, 2001). 147 E. gallinacea is more active in summer, causing serious trouble with livestock, especially in 148 rural areas. In North Africa, E. gallinacea has been reported in Morocco (Rabat, Agadir, Salé, 149 Safi, Essaouira, Casablanca), Algeria (Saïda), in Tunisian islands (Djerba and Zembra) 150 (Beaucournu and Launay, 1990) and has also been isolated from dogs in Libya (Kaal et al., 151 2006). 152 Loftis et al. (2006) conducted a surveillance study of fleas on mammals and associated 153 pathogens in Egypt, with E. gallinacea being the main species collected. In addition, these 154 authors were able to detect spotted fever by Rickettsia sp, similar to RF2125, in all collected 155 specimens (Loftis et al., 2006). 156  Ctenocephalides felis (Figure 1.3): 157 C. felis is commonly known as the “cat flea”. C. felis originates from Africa and now parasites 158 pets and livestock very easily and regularly in warm and hot climates (Beaucournu and Ménier, 159 1998). It is considered to be the main flea species infecting domestic carnivores in many 160 countries around the world (Rust, 2016). 161 This species is cosmopolitan and sedentary on its host and is mainly active in summer. All 162 mammals living in the same biotope are susceptible to be infested by C. felis (Beaucournu and 163 Launay, 1990). Its host specificity is low, thus, it can be found on other animals, including small 164 ruminants, cattle, primates, rodents, poultry and even opossums. The direct transmission 165 between individuals is frequent, although it appears to be rarer in dogs than in cats (Dobler and 166 Pfeffer, 2011). 167 C. felis has been found in Tunisia, and has been collected from cats, dogs, sheep and goats. 168 Several microorganisms have been detected in C. felis in Tunisia, including Bartonella spp., R. 169 felis (Zouari et al., 2017). It has also been collected from hedgehogs in Algeria. These authors 170 reported the presence of R. felis DNA in all C. felis collected. The results of this study can 171 therefore help human and veterinary clinicians to focus on a broader spectrum of pathogens and 172 take them into consideration during diagnosis (Leulmi et al., 2016). 173 The presence of this flea has also been reported in Morocco (Casablanca, Tiznit) on domestic 174 animals, and molecular biology demonstrated the presence of Bartonella clarridgeiae, B. 175 henselae and R. felis in these fleas (Boudebouch et al., 2011). It has also been collected from 176 cats and dogs from a farm in Libya (Kaal et al., 2006). At this point, it should be highlighted 177 the ausence of Rickettsia asembonensis in C. felis collected from North Africa. This pathogen 178 has been widely detected in C. felis and C. canis collected from several regions of sub‐Saharan 179 Africa such as Rwanda, Zambia or Kenya (Nziza et al., 2018; Moonga et al., 2019), however ; 180 currently, there is no studies which confirm the presence of this bacteriim in the cat flea from 181 North Africa. 182 This flea can quickly change hosts, which can play a role in the transmission of pathogens. The 183 saliva of C. felis has irritating properties, which generally leads to dermatosis due to the bites. 184 It can also cause anemia during a massive infestation (Gaguere and Prelaud, 2006). 185  Ctenocephalides canis (Figure 1.4): 186 C. canis is also known as the “dog flea”. Despite its name, it has been demonstrated that the 187 prevalence of C. felis in dogs is higher than that of C. canis (Linardi and Santos, 2012). C. canis 188 has a very similar morphology to C. felis subspecies. Thus, we can easily discriminate between 189 C. canis and C. felis felis but it used to be difficult to differentiate between C. canis and C. felis 190 strongylus or C. felis orientis. These differences are mainly based on the shape of the frons of 191 the cephalic capsule, the presence and shape of the dorsal incrassation and the number of setae 192 on the occiput (Linardi and Santos, 2012). They also differentiate in the average jump height, 193 which is 15.5 cm in C. canis and 13.2 cm in C. felis (Beaucournu and Launay, 1990). Although 194 both species are recognised vectors of R. felis and several Bartonella spp. pathogens, it is 195 important to discriminate between C. felis and C. canis, since many authors have reported a 196 much lower prevalence of these bacteria in C. canis than in C. felis (Kumsa et al., 2014; 197 Lawrence et al., 2015). 198 This is a species that is sedentary on its host and infests mainly domestic and wild canids. The 199 red fox is its primary host, but epidemiological studies have shown that C. canis can also be 200 found on cats, albeit with a lower prevalence than C. felis (Beaucournu and Ménier, 1998; 201 Linardi and Santos, 2012; Marrugal et al., 2013). 202 This species of flea is frequent in North Africa, and has been collected from dogs from Tunisia, 203 where molecular analysis showed the presence of Bartonella spp. in 23.5% of collected C. canis 204 (Zouari et al., 2017). A preliminary study conducted in Egypt on domestic rodents revealed the 205 presence of C. canis in the Dakahlia governorate (Soliman and Mikhail, 2011). In northern 206 family Rickettsiaceae (Maina et al., 2016). R. felis, R. asembonensis, and “Candidatus 349 Rickettsia senegalensis” belong to the spotted fever group rickettsiae (SFGR) that genetically 350 clusters within the transitional group of rickettsiae (Gillespie et al., 2007). We have just 351 mentioned that R. felis is associated with flea-borne spotted fever, however, the pathogenicity 352 of R. asembonensis and “Ca. R. senegalensis” is currently unknown. These three agents have a 353 worldwide distribution and they have been detected in humans and non-human primates (Tay 354 et al., 2015; Kho et al., 2016). R. asembonensis DNA has been detected in fleas from three 355 families of fleas (Pulicidae, Ceratophyllidae and Coptopsyllidae) with highest prevalence rates 356 reported in C. felis and C.canis (Roucher et al., 2012; Jiang et al., 2013). In spite of that, in 357 North Africa this bacterium only has been detected in E. gallinacea collected from Egypt 358 (Loftis et al., 2006). 359 Bartonelloses 360 Bartonelloses are zoonosis caused by Gram-negative aerobic bacteria of the genus Bartonella 361 (Iannino et al., 2018). Many species have been described. They include recognized pathogens 362 as well as bacteria of unknown pathogenicity. (Stuckey et al., 2017). 363  Cat-scratch disease 364 This disease was first described in 1889 and it is caused by B. henselae (Wong et al., 1995). 365 The domestic cat is the main reservoir of this bacterium (Chomel, 1996). 366 Transmission to humans takes place through a cat scratch or bite, but contamination by C. felis 367 flea faeces and transmission by regurgitation have also been demonstrated (Bouhsira et al., 368 2013b). Infection with B. henselae can cause fever, hepatitis, endocarditis, bacillary 369 angiomatosis and bacillary peliosis (Durden and Traub, 2002). 370 B. henselae has been detected by qPCR in dog blood from Algiers, Algeria (Azzag et al., 2012; 371 Bessas et al., 2016). B. henselae has also been detected in C. canis and C. felis, collected from 372 domestic animals in Sfax, Jendouba and Manouba in Tunisia, (Belkhiria et al., 2017; Zouari et 373 al., 2017). This bacteria was detected using qPCR in C. felis collected from sheep, cats and dogs 374 in Casablanca, Morocco (Boudebouch et al., 2011). 375  Trench fever: 376 Trench fever is a zoonosis caused by infection by the bacterium B. quintana, a gram-negative 377 bacterium which is considered as a re-emerging human pathogen (Anderson and Neuman, 378 1997; Faccini-Martínez et al., 2017). This disease was described during the Second World War 379 following several cases of soldiers who suffered from the disease (Kostrzewski, 1949). B. 380 quintana may also be responsible for bacillary angiomatosis (Relman et al., 1990), endocarditis 381 (Drancourt et al., 1995) and chronic lymphadenopathy (Raoult et al., 1994). 382 It is transmitted by body lice (P. h. humanus) (Coulaud et al., 2014), but some authors have 383 recently shown the ability of C. felis to transmit B. quintana, which was found in flea faeces 11 384 days after an infectious meal (Bouhsira et al., 2013a; Kernif et al., 2014). This bacterium has 385 also been detected in P. irritans in Gabon (Rolain et al., 2005). 386 The presence of this bacterium has often been detected by molecular biology in body lice 387 collected from homeless people in northern Algeria (Louni et al., 2018) and also in patients 388 during an endocarditis study conducted in Casablanca and Marrakech (Morocco) (Boudebouch 389 et al., 2017), and, finally, in 12 endocarditis patients in Sfax (Tunisia) (Znazen et al., 2005). 390 Parasitic diseases 391  Dipylidium caninum infection or Dipylidiasis 392 This is a medium-sized tapeworm which frequently parasitizes the small intestine of dogs and 393 cats. Nevertheless, they can occasionally parasitize humans (Moskvina and Ermolenko, 2016), 394 in which case it causes Dipylidiasis. Its intermediate hosts are C. felis and C. canis and its final 395 hosts are dogs and cats. These parasites deplete the hosts’ nutrients during their presence in the 396 digestive tract and the spoliation of all the nutrients explains the clinical signs (Neira O et al., 397 2008). The parasite is transmitted to the host by ingestion of contaminated fleas or faeces. 398 Furthermore, lice can, exceptionally, transmit the worm (Franc, 1994b). 399 This parasite has been reported in three countries of northern Africa. I It was mentioned in a 400 study that targeted wild canids in Tunisia, when the parasite was detected in 55% of foxes and 401 jackals (Lahmar et al., 2014). It was also detected in Egypt, in a study of intestinal parasites 402 from 113 samples of stray cat faeces taken north of the Nile delta (Khalafalla, 2011). Even in 403 Morocco, D. caninum has been found in stray dogs in urban and rural areas of Rabat with a 404 percentage of 40.4% of all samples tested (57 dogs) (Pandey et al., 1987). 405 Flea-borne viral diseases 406 The biological transmission of viruses by fleas has not been widely studied, the only known 407 virus transmitted by fleas is Myxomavirus (myxomatosis virus) (Sobey et al., 1977)(Kerr et al., 408 2015). This virus is transmitted by the rabbit flea, S. cuniculi. This involves mechanical 409 contamination by mouth parts and the virus is released when a bite occurs (Shepherd and 410 Edmonds, 1980). Clinical signs are usually severe and death occurs within 10 to 12 days. 411 However, rabbits with milder signs, including those suffering from the amyxomatous form or 412 those that have been previously vaccinated, may survive with nursing care (Meredith, 413 2013).The epidemiology of this virus in North Africa has never been studied, but the presence 414 of the flea vector has been reported in several Moroccan cities (Beaucournu and Launay, 1990). 415 Although several human viral pathogens have been isolated or detected in fleas, the role of fleas 416 in their transmission is either unknown or considered to be incidental. These viruses include 417 those causing lymphocytic choriomeningitis, tick-borne encephalitis and Russian spring418 summer encephalitis (Durden and Traub, 2002). 419 The feline leukaemia virus (FeLV) is a frequent virus in domestic cats and it was still suspected 420 to be transmitted by an arthropod vector. Some authors (Vobis et al. 2003) developed an 421 experimental model to study the vector capacity of C. felis to transmit the FeLV virus (Vobis 422 et al., 2003). The fleas were fed for 24 hours with blood from a FeLV-infected cat, and FeLV 423 was finally detected in fleas and their faeces. The fleas could even transmit the FeLV virus from 424 one blood sample to another. The results indicate that cat fleas are potential vectors for FeLV 425 RNA in vitro and probably also in vivo (Vobis et al., 2003) 426 Identification and laboratory rearing 427 The correct identification of flea species is essential in any research or control project. Current 428 identification methods are mainly based on morphological identification. However, to perform 429 detailed morphological identification, it is necessary to clear flea samples with 10% KOH or 430 NaOH (Lewis, 1993), examine under a stereomicroscope, and then mount and photograph them. 431 Molecular biology has been used over the last 20 years for flea identification and detection of 432 their associated pathogens (Zurita et al., 2015). These approaches are limited by the length of 433 time, the availability of reference sequences in the GenBank database, the cost associated with 434 molecular biology approaches, as well as the small number of entomologists specialised in flea 435 taxonomy (Yssouf et al., 2016). Recently, mass spectrometry has emerged as an innovative 436 identification tool for arthropods, especially fleas (Yssouf et al., 2014). The use of MALDI TOF 437 MS requires the development of an adequate protocol in order to standardise sample preparation 438 methods and to allow for subsequent exchanges using the database between several research 439 laboratories (Nebbak et al., 2017). In addition, MALDI TOF MS has proved its effectiveness 440 in identifying fleas which may or may not be infected by a pathogen and even in distinguishing 441 between fleas infected with two pathogens of the same family (El Hamzaoui et al., 2018). 442 Nevertheless, since MALDI-TOF MS techniques have demonstrated some differences in the 443 MS spectra of specimens preserved in different storage conditions (Nebbak et al., 2017; Zurita 444 et al., 2018), it is still necessary to combine morphological, molecular and proteomics methods 445 in order to carry out an efficient specific identification within the Order Siphonaptera. 446 Flea rearing is also a key step in the study of the biology of Siphonaptera, their morphology and 447 their vectorial capacity. The cat flea, C. felis, is found worldwide and has been reported to 448 parasitize many species of wild and domestic animals (Rust and Dryden, 1997). In addition, C. 449 felis has been described as having low host specificity. Indeed, it is a flea that feeds on a variety 450 of animals and rodents, and is therefore the right choice for developing a laboratory breeding 451 programme (Dryden and Rust, 1994). The artificial rearing system described by Wade and 452 Georgi (1988) includes a heated Plexiglas box from which flea chambers are suspended and in 453 which a source of human blood is heated (previously stored at 4°C) (Wade and Georgi, 1988). 454 To maintain the required temperature difference of 10°C, the entire system is housed in a 455 temperature-controlled chamber. The temperature should be maintained between 25°C and 456 35°C, with a relative humidity of 75 to 80% using a tray filled with water. Fleas are raised in 457 the dark 24 hours a day (Kernif et al., 2015). 458 Rearing fleas makes it possible to study their vectorial capacity. An experimental model of 459 artificial infection of fleas with a strain of B. quintana has shown the ability of C. felis to acquire 460 the bacterium and transmit it, alive, in faeces. Several flea groups were fed with blood mixed 461 with the bacterial inoculum at different concentrations. qPCR showed the presence of B. 462 quintana in faeces and immunohistochemistry localised the bacterium in the digestive tract of 463 C. felis (Kernif et al., 2014). 464 Control 465 Insecticides are the most widely-used means of flea control, including powder sprays in nests, 466 burrows, or house walls in infested areas (Rust, 2016). Molt inhibitors are also used to control 467 fleas in their larval stages. It is strongly recommended to control their host in combination with 468 flea control using insecticides, in order to avoid what happened in the case of plague (Franc, 469 1994a). 470 Conclusion: 471 For several decades, we have witnessed the re-emergence of several vector-borne zoonotic 472 pathologies. Metagenomics and molecular biology have revolutionized the epidemiology of 473 these diseases; however, some areas remain poorly explored as North Africa. 474 Fleas are hematophagous, wingless insects that have the ability to jump. Their ability to transmit 475 pathogens explains their importance in human and animal health. They are spread all over the 476 world and some species do not require the presence of a specific host. This review summarizes 477 the latest data on flea vectors and flea-borne diseases in North Africa (Morocco, Algeria, 478 Tunisia, Egypt and Libya). 479 For this reason, we have selected disease’s vector species described in North Africa in recent 480 years, we have also reported the cases of vector-borne diseases by fleas diagnosed to update the 481 epidemiological situation in this region. 482 483 Declarations: 484 All authors consent to the publication 485 Ethics approval and consent to participate: not applicable 486 No competing interests exist 487 Funding: not applicable 488 Availability of data and materials: not applicable 489 Author Contributions: 490 Wrote the paper: BE AZ PP CC 491 Acknowledgements: Our thanks to Jean-Michel Bérenger (IHU Méditerranée Infectio) and Dr. 492 Phillip Kaufman (University of Florida) for facilitating access to some pictures of fleas. 493 494 495 496 497 498 Figures 499 Figure 1: 1. Pulex irritans female, 2. 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