Prevalence of Lyme Disease and Relapsing Fever Borrelia spp. in Vectors, Animals, and Humans within a One Health Approach in Mediterranean Countries

Abstract

The genus Borrelia has been divided into Borreliella spp., which can cause Lyme Disease (LD), and Borrelia spp., which can cause Relapsing Fever (RF). The distribution of genus Borrelia has broadened due to factors such as climate change, alterations in land use, and enhanced human and animal mobility. Consequently, there is an increasing necessity for a One Health strategy to identify the key components in the Borrelia transmission cycle by monitoring the human-animal-environment interactions. The aim of this study is to summarize all accessible data to increase our understanding and provide a comprehensive overview of Borrelia distribution in the Mediterranean region. Databases including PubMed, Google Scholar, and Google were searched to determine the presence of Borreliella and Borrelia spp. in vectors, animals, and humans in countries around the Mediterranean Sea. A total of 3026 were identified and screened and after exclusion of papers that did not fulfill the including criteria, 429 were used. After examination of the available literature, it was revealed that various species associated with LD and RF are prevalent in vectors, animals, and humans in Mediterranean countries and should be monitored in order to effectively manage and prevent potential infections.

1. Introduction

One Health approach is a holistic perspective that deals with complicated issues involving human, animal, and environmental health [1]. Such approaches have become increasingly important in recent years due to critical changes that have altered the interactions among humans, animals, and their environment [2]. Human population growth as well as their expansion into new geographical areas results in closer proximity between people and animals, facilitating the transmission of diseases among them [3,4]. Additionally, climate change and alterations in land use, along with increased movement of people, animals, and animal products through international travel and trade, further exacerbates the spread of endemic diseases [5,6].

Borrelia spp. are microaerophilic, slow-growing spirochetal bacteria that have been associated with human diseases [7]. Depending on the infecting species, it can cause Lyme Disease (LD) [8], also known as Lyme borreliosis, or Relapsing Fever (RF) [9].

LD is the most prevalent vector-borne disease in moderate climates of the northern hemisphere with an increasing incidence [10]. It is estimated that approximately 476,000 cases are diagnosed and treated annually in the United States [11], and over 200,000 cases per year in Europe [12].

RF can be either louse-borne (LBRF) or tick-borne (TBRF) [13] and is predominantly found in Africa, Spain, Saudi Arabia, Asia, and western United States, with epidemics still occurring in developing countries due to poor living conditions, famine, and war [14]. Currently, LBRF is mostly observed in the Horn of Africa and among immigrants originating from endemic areas [15]. TBRF has recently been associated with travel-related disease after travelling in endemic areas [16].

Borrelia spp. can be transmitted by hard and soft ticks, as well as human body lice. The distribution of these arthropod vectors is closely linked to the presence of the bacteria in both animals and humans [17]. Animals serve as hosts for both ticks and bacteria, playing a crucial role in the circulation of Borrelia spp. [18]. Therefore, studying the distribution of ticks in relation to the surrounding environment and the prevalence of suitable hosts for ticks and Borrelia spp. is essential for better understanding the factors contributing to human diseases [19].

The aim of this article is to review the current data on the presence of Borrelia spp. in vectors, animals, and humans around the Mediterranean Sea in order to monitor their human-animal-environment interactions within a One Health approach. Such information provides insight for the better surveillance and management of Borrelia infections.

2. Methods

A review of the literature was conducted on the available publications from 1936 to February of 2024 by searching PubMed, Google Scholar, and Google. We included studies related to the presence of LD Borreliella spp. and RF Borrelia spp. in countries around the Mediterranean Sea in the vectors of transmission, the animal hosts, as well as the humans that can become infected. Search terms included “Lyme Disease”, “Lyme borreliosis”, “Relapsing Fever”, “Borreliella”, “Borrelia”, “bacteria”, “human-animal-environment interaction”, “One Health”, “humans”, “animals”, “ticks”, “lice”. Articles that were not written in English were excluded from this study (Figure 1).

Figure 1.

Flow diagram of study selection.

3. Bacteriology

Species of the Borrelia genus are members of the Borreliaceae family of the Spirochaetales phylum and are thus characterized by the typical spirochaete (spiral) shape [20]. They are microaerophilic and host-associated since they rely on their hosts for most of their nutrients in an obligate parasitic lifestyle [21]. They possess an outer membrane around their protoplasmic cylinder analogous to that of Gram-negative bacteria, but without lipopolysaccharides [22]. They have several axial filaments attached to each pole, called endoflagella [13], between the outer membrane and the peptidoglycan layer, inside the periplasmic space, which help the bacteria move forward in a corkscrew-like motion [8]. Their genome consists of one highly conserved linear chromosome and several plasmids that can be either linear or circular and their number varies highly among strains [23]. Borrelia genus is divided into two genera related to human disease, Borreliella for the Lyme group and Borrelia for the Relapsing Fever group (Figure 2). Even though they belong to the same genus, they are known to have significant differences regarding their biological characteristics, their epidemiology, as well as their pathogenicity [24]. According to the phylogenetic data, although they share a genetic similarity, they are different, and they belong to independent clades under a common ancestor. For this reason, it has been proposed and recently validated [25], to include the Lyme group Borrelia into a new genus taxon named Borreliella [26].

Figure 2.

Phylogenetic tree of the genus Borrelia from LD and RF groups constructed with the neighbor joining method with 1000 Bootstrap replications in MEGA 11.0 software based on the flaB gene.

3.1. LD

LD is caused by the Borreliella genospecies. Currently, it contains twenty-six species, twenty-three validated, and three proposed (Table 1) [27,28]. It consists of bacteria 10–20 μm long, approximately 0.3 μm in diameter and the number of their endoflagella ranges from seven to twelve [24]. The species that most commonly cause human disease are Borreliella burgdorferi, Borreliella afzelii, Borreliella garinii, Borreliella mayonii, Borreliella spielmanii, and Borreliella bavariensis. Bl. burgdorferi is responsible for the majority of the LD cases in North America, Bl. garinii in Asia, and Bl. afzelii along with Bl. garinii in Europe [29,30].

Table 1.

Borreliella spp. of the LD Group.

Borreliella Species	Human Infection	Geographical Distribution	Hosts	Vectors	References
Borreliella afzelii	Yes	Europe, Asia	Rodents	I. ricinus, I. persulcatus, I. pavlovsky	[31,32,33,34]
Borreliella americana	Unknown	North America, Poland	Rodents, Birds	I. pacificus, I. minor, I. ricinus	[35,36,37]
Borreliella andersonii	Unknown	USA	Cottontail rabbits	I. scapularis, I. dentatus	[38,39,40]
Borreliella bavariensis	Yes	Europe, Asia	Rodents	I. ricinus, I. persulcatus	[41,42,43]
Borreliella bissettiae	Yes	USA, Europe	Rodents	I. spinipalpis, I. ricinus, I. pacificus	[34,44,45,46,47]
Borreliella burgdorferi	Yes	America, Europe, Asia	Rodents, Mammals, Birds	I. scapularis, I. pacificus, I. ricinus, I. kaiseri	[34,48,49,50]
Borreliella californiensis	Unknown	USA, Europe	Kangaroo Rats	I. jellisoni, I. spinipalipis, I. pacificus, I. ricinus	[34,46,51,52,53]
Borreliella carolinensis	Unknown	USA, Europe	Rodents	I. minor, I. ricinus, I. affinis	[37,54,55,56,57,58]
Borreliella chilensis	Unknown	Chile	Rodents	I. stilesi	[59]
Borreliella finlandensis	Unknown	Finland	Unknown	I. ricinus	[60]
Borreliella garinii	Yes	Europe, Asia	Rodents, Birds	I. ricinus, I. persulcatus	[34,61]
Candidatus Borreliella ibitipoquensis	Unknown	Brazil	Birds	I. paranaensis	[62]
Borreliella japonica	Unknown	Japan	Rodents	I. ovatus	[63,64]
Borreliella kurtenbachii	Unknown	North America	Rodents	I. scapularis	[58,65,66]
Borreliella lanei	Unknown	USA, Europe	Lagomorph	I. spinipalpis, I. pacificus, I. ricinus	[34,44,52,53,67,68]
Borreliella lusitaniae	Yes	Europe, North Africa	Lizards, Rodents	I. ricinus	[69,70,71]
Borrelia maritima	Unknown	North America	Unknown	Ixodes spinipalpis	[72]
Borreliella mayonii	Yes	USA	Rodents	I. scapularis	[73,74,75]
Candidatus Borrelia paulista	Unknown	Brazil	Rodents	Unknown	[76]
Candidatus Borrelia sibirica	Unknown	Siberia	Rodents	I. apronophorus, I. persulcatus, I. trianguliceps	[77]
Borreliella sinica	Unknown	China	Rodents	I. ovatus	[78]
Borreliella spielmanii	Yes	Europe	Rodents	I. ricinus	[79,80,81,82]
Borreliella tanukii	Unknown	Japan	Unknown	I. tanuki	[83,84,85]
Borreliella turdi	Unknown	Japan, Europe	Birds	I. turdus, I. frontalis, I. ricinus	[84,85,86,87]
Borreliella valaisiana	No	Europe, Asia	Birds	I. ricinus	[88,89,90,91,92]
Borreliella yangtzensis	Unknown	Asia	Rodents	I. granulatus	[93,94,95,96,97]

3.2. RF

RF is caused by the species of the relapsing fever complex, which consists of 22 validated and several proposed species (Table 2) [98,99]. Bacteria from the RF complex tend to have shorter bodies than the Lyme group and the number of their endoflagella ranges from 15 to 20 [98]. Depending on the vector of transmission, the species of the RF group are categorized in the soft-tick-borne relapsing fever (STBRF), the hard-tick-borne relapsing fever (HTBRF), and the louse-borne relapsing fever (LBRF) group [99]. Additionally, STBRF are further classified according to the geographical area into the Old World strains and the New World strains [100]. Another category is the avian RF which consists of Borrelia anserina and is transmitted by the soft tick Argas spp. [101].

Table 2.

Borrelia spp. of the RF Group.

Borrelia Species	Human Infection	Geographical Distribution	Hosts	Vectors	References
Louse-Borne Relapsing Fever
Borrelia recurrentis	LBRF	Virtually worldwide, currently Ethiopia, Sudan	Human	Pediculus humanus	[14,102]
Old World Soft-Tick Borne Relapsing Fever
Candidatus Borrelia algerica	TBRF	Algeria	Human	Unknown	[103]
Borrelia caucasica	STBRF	Azerbaijan, Georgia, Armenia, Ukraine	Rodents, Human	O. verrucosus	[104,105,106,107]
Borrelia crocidurae	STBRF	Western and Northern Africa	Insectivores, Rodents, Camels, Human	O. sonrai, O. erraticus	[105,107,108]
Borrelia duttonii	STBRF neurological signs, neonatal infection	Central, Eastern, and Southern Africa	Human	O. moubata	[105,107,109,110]
Borrelia graingeri	Unknown	East Africa	Rodents	O. graingeri	[105,107,111,112]
Borrelia harveyi	Unknown	Kenya	Monkey	Unknown	[105,113,114]
Borrelia hispanica	STBRF Ocular and neurological symptoms (rare)	Iberian Peninsula and Northwestern Africa	Insectivores, Rodents, Foxes, Jackals, Dogs, Human	O. erraticus, O. marocanus, O. occidentalis	[105,107,115]
Candidatus Borrelia kalaharica	STBRF	Kalahari Desert (Botswana and Namibia)	Human	O. savignyi	[116]
Borrelia latyschewii	Unknown	Central Asia, Middle East	Unknown	O. tartakovsky	[105,114,117]
Borrelia merionesi	No	Morocco and Atlantic coastal areas of the Sahara Desert	Rodents	O. merionesi, O. costalis, O. erraticus	[118]
Borrelia microti	STBRF	Iran, Afghanistan	Human	O. erraticus	[107,119,120,121]
Borrelia persica	STBRF, neurological symptoms (rare), respiratory distress (rare)	Central Asia, Middle East, Egypt, Greece	Rodents, Carnivores, Dogs, Cats, Human	O. tholozani	[102,105,122]
Borrelia tillae	No	Southern Africa	Rodents	O. zumpti	[123]
New World Soft-Tick Borne Relapsing Fever
Borrelia brasiliensis	Unknown	Brazil	Unknown	O. brasiliensis	[124,125]
Borrelia coriaceae	No	USA	Rodents, Deer	O. coriaceus	[125,126,127,128]
Borrelia dugesii	Unknown	Mexico	Unknown	O. dugesii	[105,129]
Borrelia hermsii	STBRF, neurological symptoms (rare), neonatal infections (rare)	British Columbia, Western USA, France	Rodents, Deer, Dog, Human	O. hermsi	[102,105,126,128,130]
Borrelia johnsonii	TBRF	USA	Bats	O talaje	[131,132,133]
Borrelia mazzottii	No	Mexico	Unknown	O. talaje	[134]
Borrelia parkeri	Unknown	Western USA	Horses	O. parkeri	[130,135]
Borrelia turicatae	STBRF, Ocular and neurological symptoms	British Columbia, USA, Mexico, Africa, Europe, Asia	Rodents, Dog, Human	O. turicata, O. maritimus, C. capensis	[102,105,136]
Borrelia venezualensis	STBRF	Central America and northern South America	Human	O. rudis	[105,137]
Hard-Tick Borne Relapsing Fever
Borrelia lonestari	STARI?	USA	Rodents, Birds, Deer, Human	Amblyomma americanum (lone star tick)	[133,138,139,140,141,142]
Borrelia miyamotoi	Flu-like syndrome, HTBRF, neurological symptoms	Asia, Europe, USA	Rodents, Birds, Human	I. ricinus, I. persulcatus, I. scapularis	[143,144,145,146]
Candidatus Borrelia texasensis	Unknown	USA (Texas)	Unknown	Dermacentor variabilis	[147]
Borrelia theileri	No	Africa, Australia, North and South America	Cattle, Sheep, Goats, Camels	Rhipicephalus spp.	[105,110,148,149]
Avian Relapsing Fever
Borrelia anserina	No	Worldwide	Birds	Argas spp.	[105,110,150,151,152]

4. One Health Approach

Global health challenges are escalating, and hence, One Health approaches emerge as a pivotal framework for addressing such complex health matters [153]. This holistic perspective acknowledges the interconnection of these domains and emphasizes the need for collaboration among medical, veterinary, environmental, and other relevant fields [154]. In recent years, the multifaceted nature of Borrelia infections has underlined the necessity for a comprehensive One Health approach. By integrating insights from diverse scientific areas, a One Health perspective offers synergistic efforts to address the complexities of Borrelia infections, enhancing the ability to protect both human and animal populations while promoting environmental sustainability (Figure 3) [155].

Figure 3.

Interactions among humans, animals, and the environment within a One Health concept.

5. Lyme Disease

5.1. Clinical Manifestations

The clinical manifestations of LD can vary widely, as well as the severity and duration of the symptoms, and can affect multiple body systems [156]. The organs most frequently involved are skin, joints, nervous system, heart, and eyes [30,157]. In general, Bl. burgdorferi is more often associated with Lyme arthritis, Bl. garinii with neuroborreliosis, and Bl. afzelii with skin manifestations [30]. The early infection is localized, but the spirochete can spread if left untreated [158]. The most common manifestation of LD that occurs in 80–90% of the cases is a rash known as erythema migrans (EM) [159], which can have divergent characteristics [160,161]. It typically appears as an annular, erythematous rash at the site of the tick bite, which may or may not present with a central clearing [162]. It is a skin redness that appears 1–36 days (with an average of 7–14 days) after the tick bite with a tendency to expand, eventually reaching a diameter of even up to 70 cm (with a median of 10–16 cm) [163,164]. Multiple EM lesions can also occur, with a primary lesion at the site of the tick bite and secondary skin lesions caused by hematogenous dissemination [165]. Additionally, many patients also experience systemic symptoms like fatigue, fever, chills, arthralgia, myalgia, and regional lymphadenopathy [166,167]. Non-cutaneous organ involvement is also possible. Lyme neuroborreliosis occurs in approximately 12% of untreated patients who may present with cranial neuropathy (unilateral or bilateral facial nerve palsy), lymphocytic meningitis, radiculitis, or encephalitis [168,169,170]. Lyme arthritis is the most common late manifestation in the USA, presenting in approximately 60% of untreated patients, but only in a small percentage in Europe [8]. It usually appears months or even up to 2 years after initial infection and typically affects only one or a few large joints, especially the knee [171]. In rare but potentially fatal cases, Lyme carditis appears in 5% of untreated patients usually causing atrioventricular conduction block [172]. The development of chronic congestive cardiomyopathy has also been linked to LD [173,174,175].

5.2. Epidemiology

LD is endemic in temperate regions of the Northern Hemisphere, such as North America, Europe, and Asia [30]. The documented incidence for countries in Europe and Asia varies from negligible in the United Kingdom, Turkey, and Japan to significant in the Netherlands, Belgium, Austria, Slovenia, Lithuania, and Estonia [176,177]. Although incidents have been recorded in countries like China and Mongolia, information on the prevalence of LD in Asia is limited [178,179]. More than 90% of cases in North America are documented from only two regions of the country: the northeast and mid-Atlantic region and the north central region [180]. Both regions have grown significantly over the past 20 years and have extended up to south Canada [181,182]. LD rates are increasing worldwide [181,183] due to increased awareness [184] along with the expanding distribution of the tick vectors due to environmental factors [156,185].

5.2.1. Transmission Vectors Infected with Borreliella spp. in Countries around the Mediterranean Sea

Infectious agents causing LD are transmitted by slow-feeding hard-bodied ticks (ixodid vectors) of the Ixodes ricinus complex and are the only natural agents known through which humans can become infected (Figure 4) [186]. In northeastern, mid-Atlantic, and north central United States, the main vector of transmission is Ixodes scapularis (black-legged tick) and in the Pacific Coast states, Ixodes pacificus [187]. In Asia and in Eastern Europe, the main vector tick is Ixodes persulcatus, while in Western Europe it is Ixodes ricinus [8,188]. Nymphal ticks, rather than the adult ticks, are primarily responsible for transmission of the disease [189]. Spirochaetes are located mainly in the midgut of the tick before attachment and only in small numbers in the salivary glands of the ticks, however, the number in the salivary glands increases rapidly post attachment [190]. Thus, the risk of transmission increases with increased duration of tick attachment and the nymphal ticks are more likely to remain undetected for longer due to their smaller size [191]. The ticks feed on a single vertebrate host when in their larval, nymphal, and adult female phases. Adult male ticks may take sporadic small meals. When a tick, usually a larva or a nymph, feeds on an infected host, the bacteria are acquired by the tick and are subsequently transferred to its midgut [192]. The spirochetes migrate to new hosts, including humans, when the next developing stage of the tick feeds on them [193]. Nevertheless, transovarial transmission which occurs from female to offspring, is rare, resulting in a dead end for the bacteria [194]. A study concerning the human transplacental transmission has been performed, and although Bl. burgdorferi was not directly linked to any of the unfavorable outcomes, the frequency of adverse outcomes emphasizes the need for further studies [195].

Figure 4.

Transmission of Borreliella spirochetes.

Prevalence of the Borreliella complex bacteria have been identified in questing as well as feeding ticks across the Mediterranean. Nonetheless, there are no data on the following Mediterranean countries: Gibraltar, Monaco, Malta, Bosnia and Herzegovina, Montenegro, Albania, Cyprus, Syria, Lebanon, Palestine, Israel, and Libya.

Western Mediterranean (Spain, France)

In Spain, questing I. ricinus ticks were tested for the presence of Borreliella and Bl. garinii was found to be the most prevalent species followed by Bl. valaisiana, Bl. lusitaniae, Bl. afzelii, and Bl. burgdorferi [196]. Additionally, Bl. turdi was found in I. frontalis ticks collected from vegetation of urban and sub-urban areas [197] as well as in I. frontalis, I. ricinus, and Haemaphysalis punctata ticks collected from passerine birds [87]. Moreover, Hyalomma lusitanicum ticks from wild ungulates revealed that they were positive for Bl. afzelii [198]. Borreliella species have also been found in non-Ixodes ticks, as reported in studies with Haemaphysalis spp. ticks, Dermacentor spp., and Rhipicephalus bursa from the vegetation [199,200]. Finally, Borreliella DNA was reported in a male Dermacentor reticulatus removed from a patient [201]. In France, questing I. ricinus ticks from peri-urban forests demonstrated the presence of Bl. afzelii, Bl. burgdorferi, and Bl. garinii followed by Bl. valaisiana, Bl. lusitaniae, and Bl. spielmanii [202]. Moreover, Borreliella carolinensis was reported in one questing I. ricinus tick from a French forest [203]. Furthermore, ticks feeding on humans revealed the presence of Bl. burgdorferi in I. ricinus, and Bl. garinii in I. ricinus and in unidentified Ixodes spp. [204]. Larva ticks engorged on birds were also tested and it was found that Borreliella DNA was present in I. ricinus, I. frontalis, Ixodes spp., and Haemaphysalis spp. [205]. Additionally, I. ricinus ticks from rats and Hyalomma marginatum from cattle from the island of Corsica [206], as well as a questing Dermacentor reticulatus tick from central France were also positive for Borreliella DNA [207].

North Central Mediterranean (Italy, Slovenia)

In Italy, questing I. ricinus ticks, as well as I. ricinus ticks feeding on wild ungulates and domestic animals, were found to be infected with Bl. afzelii, Bl. burgdorferi, Bl. garinii, Bl. valaisiana, and Bl. lusitaniae [208,209]. Moreover, I. ricinus ticks feeding on humans revealed the presence of Borreliella DNA with Bl. afzelii being the most prevalent, followed by Bl. lusitaniae, Bl. garinii, Bl. valaisiana, and Bl. burgdorferi [210]. In addition, Bl. spielmanii and Bl. valaisiana were identified in Ixodes ventalloi and Ixodes acuminatus ticks feeding on feral cats and birds from Pianosa island in the Tuscany archipelago [211], and in Hyalomma rufipes ticks feeding on birds from Ponza island [212]. Likewise, in Slovenia, Bl. garinii was the most predominant species in questing I. ricinus ticks, followed by Bl. afzelii, Bl. valaisiana, Bl. burgdorferi, and Bl. lusitaniae [213]. Finally, Bl. garinii and Bl. valaisiana were found on Ixodes spp. ticks from passerine birds feeding in Slovenia [214].

Balkans (Croatia, Greece)

In Croatia, I. ricinus ticks tested positive for Bl. afzelii, Bl. garinii, Bl. valaisiana, Bl. burgdorferi [215], as well as Bl. lusitaniae [216]. In Greece, Ixodes spp. feeding on birds tested positive for Bl. afzelii and Bl. garinii [214].

Eastern Mediterranean (Turkey)

Questing I. ricinus ticks from Turkey were cultivated leading to the identification of Bl. lusitaniae, Bl. afzelii, Bl. garinii, Bl. burgdorferi, and Bl. valaisiana [217]. Additionally, Bl. burgdorferi was found in Rhipicephalus turanicus collected from wild boar [218], as well as in Hyalomma marginatum, Hyalomma excavatum, Hyalomma spp., and Haemaphysalis parva ticks collected from humans [219]. Borreliella were also prevalent in Rhipicephalus annulatus, in Hyalomma anatolicum, and I. ricinus questing or feeding on humans, pets, or other animals [220].

Southern Mediterranean (Egypt, Tunisia, Algeria, Morocco)

Several tick species feeding on one-humped camels from Cairo, Egypt, demonstrated the presence of Bl. afzelii in Hyalomma dromedarii and H. marginatum, as well as Bl. burgdorferi in H. dromedarii and Amblyomma testudinarium [221]. Moreover, Borreliella were also identified in Rhipicephalus sanguineus from dogs and in Hyalomma anatolicum excavatum from cattle [222]. In Tunisia, questing I. ricinus ticks were positive for Borreliella DNA [223] and after cultivation, Bl. lusitaniae and Bl. garinii were isolated [223,224]. In Algeria, questing I. ricinus ticks were infected with Bl. garinii [225] and I. ricinus ticks feeding on cattle with Bl. lusitaniae [216]. In Morocco, questing I. ricinus ticks were found to be infected with Bl. lusitaniae, Bl. garinii, and Bl. burgdorferi [226].

In conclusion, the characterized species of the Borreliella complex that have been identified among ticks throughout the Mediterranean basin are Bl. burgdorferi, Bl. afzelii, Bl. garinii, Bl. valaisiana, Bl. spielmanii, Bl. lusitaniae, Bl. turdi, and Bl. carolinensis. Specifically, Bl. burgdorferi has been identified in Spain, France, Italy, Slovenia, Croatia, Turkey, Egypt, and Morocco, Bl. afzelii in Spain, France, Italy, Slovenia, Croatia, Greece, Turkey, and Egypt, Bl. garinii in Spain, France, Italy, Slovenia, Croatia, Greece, Turkey, Tunisia, Algeria, and Morocco, Bl. valaisiana in Spain, France, Italy, Slovenia, Croatia, and Turkey. Additionally, Bl. spielmanii in France and Italy, Bl. lusitaniae in Spain, France, Italy, Slovenia, Croatia, Turkey, Tunisia, Algeria, and Morocco, Bl. turdi in Spain and France and finally, Bl. carolinensis in France (Figure 5 and Table 3).

Figure 5.

Map of Borreliella spp. in vectors in countries around the Mediterranean Sea.

Table 3.

Borreliella spp. in vectors around the Mediterranean Sea.

Country	Borreliella Species
	Borreliella spp.	Bl. burgdorferi	Bl. afzelii	Bl. garinii	Bl. valaisiana	Bl. spielmanii	Bl. lusitaniae	Bl. turdi	Bl. carolinensis
Spain	✓	✓	✓	✓	✓		✓	✓
France	✓	✓	✓	✓	✓	✓	✓	✓	✓
Italy	✓	✓	✓	✓	✓	✓	✓
Slovenia	✓	✓	✓	✓	✓		✓
Croatia	✓	✓	✓	✓	✓		✓
Greece			✓	✓
Turkey	✓	✓	✓	✓	✓		✓
Egypt	✓	✓	✓
Tunisia	✓			✓			✓
Algeria	✓			✓			✓
Morocco	✓	✓		✓			✓

5.2.2. Animal Hosts Infected with Borreliella spp. in Countries around the Mediterranean Sea

Reservoir hosts are important for the transmission of the bacterium, as they can become heavily infested with ticks and transmit the bacterium to them when they feed on their blood [227]. More than 100 animal species have been identified as Borreliella hosts, including small mammals, birds, and lizards [70,228,229]. The main hosts in North America are small mammals, such as mice, chipmunks, and squirrels, with the white-footed mice being the most common [230,231,232]. In Europe and Asia, the primary hosts are rodents like voles, shrews, and mice like the yellow-necked mice, especially for Bl. afzelii [27,192]. Birds can also become infected with Borreliella and serve as important hosts mainly for Bl. garinii as they can make long-range displacements [233]. Although deer develop antibodies when exposed to Borreliella via tick bites [234], they are incompetent reservoir hosts and do not transmit the infection to feeding ticks [235,236,237].

Infections of the Borreliella complex have been described in several animal species across the Mediterranean. However, there are no data on animal infections and the species involved in the following Mediterranean countries: Gibraltar, Monaco, Malta, Montenegro, Cyprus, Syria, Lebanon, Palestine, Israel, Libya, and Morocco.

Western Mediterranean (Spain, France)

In Spain, Borreliella have been identified in several host species. Wood mice tested positive for Borreliella and voles for Bl. afzelii [199]. Additionally, spleen samples from wild ungulates revealed the prevalence of Bl. afzelii [198]. Antibodies against Borreliella were identified in dogs [238], in red foxes [239], and in wolves [240]. Finally, chamois [241] and roe deer [198] were also presenting antibodies. In France, rodents have been identified as important host species of Borreliella [242,243]. Indeed, Siberian chipmunks were infected with Bl. burgdorferi, Bl. afzelii, Bl. garinii [244], and Bl. spielmanii [245], bank voles with Bl. afzelii, Bl. burgdorferi, and Bl. garinii [245] and wood mice with Bl. afzelii [245]. Additionally, red squirrels have also tested positive for Bl. burgdorferi, Bl. afzelii, and Bl. garinii [246]. Moreover, antibodies against Borreliella have also been identified in dogs [247], horses, and ponies [248]. Last but not least, skin biopsy specimens from deer and roe deer were DNA-positive for Bl. burgdorferi, Bl. garinii or Bl. afzelii, and one Bl. burgdorferi specimen was also positive in culture, indicating the presence of alive spirochetes in deer skin [249].

North Central Mediterranean (Italy, Slovenia)

In Italy, blood samples from rodents were positive for Borreliella DNA [250], and ear biopsies from bank voles and wood or yellow-necked mice revealed the presence of Bl. afzelii, Bl. valaisiana, or Bl. lusitaniae [251]. Foxes were also found to host several Borreliella genospecies, specifically Bl. burgdorferi, Bl. afzelii, Bl. garinii, Bl. valaisiana, Bl. lusitaniae, or Bl. bissettiae, and one fox was co-infected with both Bl. afzelii and Bl. bissettiae [252]. Antibodies against Borreliella were identified in dogs from rural as well as from urban areas [253,254], as well as in horses [255]. Furthermore, Bl. lusitaniae DNA has also been identified in horses [256] and in wall lizards from southern Italy [257]. Besides the animals mentioned above, wild brown hares captured in central Italy were found to carry anti-Borreliella antibodies [258], and one blood sample from a wild boar was positive for Borreliella DNA [259]. Additionally, farm-reared pheasants were seropositive [260] and feral pigeons were positive for the presence of Borreliella DNA [261]. Finally, it was demonstrated that Borreliella DNA was found in blood samples collected from the heart of hunted red deer [262], as well as antibodies against Borreliella in blood samples from fallow deer from farms [263]. In Slovenia, Borreliella DNA was found in heart and lung biopsies from rodents including yellow-necked mice, bank voles, wood mice, Mediterranean water shrews, and a white-toothed shrew [264], and after cultivation, Bl. afzelii was isolated [264]. In addition, antibodies against Borreliella were found in deer [265] and in dogs [266].

Balkans (Croatia, Bosnia and Herzegovina, Albania, Greece)

In Croatia, rodents from eight different collection sites were PCR-positive for Bl. afzelii [267] and a small seropositivity against Borreliella was identified in dogs [266,268,269]. Dogs were also seropositive in Bosnia and Herzegovina against Borreliella [266] and in Albania, for Bl. garinii [270]. In Greece, dogs [271,272], sheep [273], and horses [274] were also found to present antibodies against Borreliella.

Eastern Mediterranean (Turkey)

In Turkey, antibodies against Borreliella were identified in dogs and horses [275] and against Bl. afzelii in mice from North Turkey [276]. Finally, sick house cats tested positive for Borreliella DNA [277].

Southern Mediterranean (Egypt, Tunisia, Algeria)

In Egypt, blood samples from cattle were analyzed and Borreliella DNA was detected [222]. Additionally, dogs were found to have anti-Borreliella antibodies [278], as well as Borreliella DNA [279]. Moreover, Borreliella DNA was also identified in blood samples from healthy one-humped camels [221]. In Tunisia, Borreliella DNA was found in blood samples from goats, sheep, cattle, and camels [280]. Furthermore, lizards were also positive for Bl. lusitaniae as demonstrated by DNA identification from skin, liver, and/or kidney tissue samples, as well as by cultivation of a skin sample [281]. Finally, in Algeria, anti-Borreliella antibodies were found in dogs [282], as well as in horse blood samples [283].

In conclusion, the circulating Borreliella spp. that have been characterized from animals around the Mediterranean are Bl. burgdorferi, Bl. afzelii, Bl. garinii, Bl. valaisiana, Bl. spielmanii, Bl. bissettiae, and Bl. lusitaniae. Specifically, Bl. burgdorferi has been identified in Spain, France, Italy, Slovenia, and Egypt, Bl. afzelii in Spain, France, Italy, Slovenia, Croatia, and Turkey, Bl. garinii in France, Italy, Slovenia, and Albania, Bl. valaisiana in Italy, Bl. spielmanii in France, Bl. bissettiae in Italy, and Bl. lusitaniae in Italy and Tunisia (Figure 6 and Table 4).

Figure 6.

Map of Borreliella spp. in animals in countries around the Mediterranean Sea.

Table 4.

Borreliella spp. in animals around the Mediterranean Sea.

Country	Borreliella Species
	Borreliella spp.	Bl. burgdorferi	Bl. afzelii	Bl. garinii	Bl. valaisiana	Bl. spielmanii	Bl. bissettiae	Bl. lusitaniae
Spain
France
Italy
Slovenia
Croatia
Bosnia and Herzegovina
Albania
Greece
Turkey
Egypt
Tunisia
Algeria

Rodents Dogs Deer Foxes/Wolves Goats/Chamois Horses Squirrels/Chipmunks Hares Birds Lizards Sheep Cats Cattle Camels Wild boar.

5.2.3. Borreliella spp. in Humans in Countries around the Mediterranean Sea

Many Borreliella infections in humans are reported using molecular and/or serological assays around the Mediterranean and are associated with different genospecies of the Borreliella complex. However, up to date, there are no reports of human infection or very little is known on its prevalence and the species involved in the following Mediterranean countries: Gibraltar, Monaco, Malta, Montenegro, Cyprus, Syria, Lebanon, Palestine, Libya, Tunisia, Algeria, and Morocco.

Western Mediterranean (Spain, France)

In Spain, serum samples from the general population [284] as well as from patients bitten by ticks [201] were found to have antibodies against Bl. burgdorferi using the Bl. burgdorferi B31 strain. Additionally, Bl. garinii was isolated from skin biopsy specimens from patients with EM [285]. In France, synovial fluids were tested for the presence of Borreliella DNA, revealing that the most prevalent species was Bl. burgdorferi, followed by Bl. afzelii and Bl. garinii [286]. Furthermore, skin samples from patients with borrelial lymphocytoma resulted in the molecular identification of Bl. afzelii, Bl. burgdorferi, and Bl. garinii, which was also positive in cultivation [287]. Recently, a case of LD caused by Bl. spielmanii was also reported manifesting as a large EM [288].

North Central Mediterranean (Italy, Slovenia)

Skin biopsies from patients in Italy with clinically diagnosed LD resulted in the isolation of Bl. afzelii, Bl. garinii, and Bl. burgdorferi [289]. Additionally, patients with early or late LD symptoms were found to be infected with Bl. afzelii and Bl. garinii. One co-infection of Bl. garinii with Bl. afzelii and another with Bl. valaisiana were also reported [290]. On the other hand, serum samples from blood donors in South Tyrol revealed the presence of anti-Bl. afzelii and anti-Bl. bavariensis antibodies using Western Blot analysis [291]. Several Borrelia species have also been responsible for human infections in Slovenia. Culture-positive skin samples from patients with only EM as the clinical manifestation of LD indicated that the majority of the infections were due to Bl. afzelii, followed by Bl. garinii and Bl. burgdorferi [292]. Except from the species reported above, in Slovenia, Bl. bavariensis has also been isolated from humans with LD [41,293]. In addition, Bl. spielmanii was also isolated from skin biopsy specimens of patients with EM [294,295]. Moreover, patients with borrelial lymphocytoma were found to be infected by Bl. afzelii, Bl. garinii, or Bl. bissettiae [296].

Balkans (Croatia, Bosnia and Herzegovina, Albania, Greece)

Human infections have also been reported in the Balkan region. In Croatia, antibodies against Bl. burgdorferi (strain B31) were found among various population groups [297]. On the other hand, molecular identification of Bl. afzelii DNA was reported in serum samples from patients with EM from northwest Croatia [298]. Additionally, a woman with EM was found to be infected by Bl. garinii [299]. In Bosnia and Herzegovina, a case of Lyme neuroborreliosis was reported in a male patient two months after a tick bite in a forest from the southern part of the country [300] and, in Albania, the Institute of Public Health reported the diagnosis of LD cases [270] without determination of the specific Borreliella species responsible for the disease. Finally, in Greece, serum samples from patients with suspected zoonotic infection were tested and the presence of anti-Borreliella antibodies was reported [301]. Positive and equivocal samples were further analyzed by Western Blot indicating Bl. afzelii infection [301].

Eastern Mediterranean (Turkey, Israel)

In Turkey, LD cases have been documented [302], as well as seropositivity against Bl. burgdorferi from randomly selected subjects [303] and farmers and forestry workers [304]. A case of LD was also reported in Israel in 1991 [305].

Southern Mediterranean (Egypt)

Finally, in Egypt, people with close contact to feverish cattle and dogs with loss of body weight have been found positive for Borreliella with serological and molecular assays [222].

To summarize, the circulating Borreliella spp. from the Borreliella complex that have been characterized from human infections around the Mediterranean Sea are Bl. burgdorferi, Bl. afzelii, Bl. garinii, Bl. valaisiana, Bl. bavariensis, Bl. spielmanii, and Bl. bissettiae. Specifically, Bl. burgdorferi has been reported in Spain, France, Italy, Slovenia, and Croatia, Bl. afzelii in Spain, France, Italy, Slovenia, Croatia, and Greece, Bl. garinii in Spain, France, Italy, Slovenia, and Croatia, Bl. valaisiana in Italy, Bl. bavariensis in Italy and Slovenia, Bl. spielmanii in France and Slovenia, and Bl. bissettiae in Slovenia (Figure 7 and Table 5, Box 1).

Figure 7.

Map of Borreliella spp. in humans in countries around the Mediterranean Sea.

Table 5.

Borreliella spp. in humans around the Mediterranean Sea.

Country	Borreliella Species
	Borreliella spp.	Bl. burgdorferi	Bl. afzelii	Bl. garinii	Bl. valaisiana	Bl. bavariensis	Bl. spielmanii	Bl. bissettiae
Spain	✓	✓	✓	✓
France	✓	✓	✓	✓			✓
Italy	✓	✓	✓	✓	✓	✓
Slovenia	✓	✓	✓	✓		✓	✓	✓
Croatia	✓	✓	✓	✓
Albania	✓
Greece	✓		✓
Turkey	✓
Israel	✓
Egypt	✓

Box 1. Prevention of LD.

Spending time in tick infested areas always carries the risk of contracting LD. However, several measures can be implemented to minimize this risk. First, tick repellent should be applied to both clothing and skin before venturing outdoors. It is also important to consistently wear long-sleeved shirts and long trousers, ensuring that they remain tucked into socks throughout the entire duration spent outside. In the case of accompanying pets, acaricides should also be used. During outdoor activi-ties, it is recommended to avoid areas with dense grass and leaf litter. After outdoor activities, a thor-ough examination should be conducted on clothing, gear, and pets for the presence of ticks. Addition-ally, inspection of the human body, including the scalp, is of great importance. If any ticks are found, appropriate removal using tweezers is essential as soon as possible. Finally, modifying the surround-ings of residential areas by clearing leaf litter, placing wood chips in areas where lawns meet forests, applying acaricides, and constructing fences to prevent the entry of tick infested animals is advisable to reduce tick populations.

6. Relapsing Fever

6.1. Clinical Manifestations

RF can be transmitted to humans through the bites of infected soft ticks, hard ticks, or body lice causing STBRF, HTBRF, and LBRF respectively. The clinical manifestations can vary, but RF hallmark is recurring episodes of high fever which can reach as high as 41 °C, spaced by afebrile periods [13]. RF patients usually experience headache, myalgia, arthralgia, and chills. Other symptoms such as fatigue, nausea and vomiting, abdominal pain and diarrhea, skin rash, neurological symptoms such as confusion and disorientation, and cardiac symptoms such as palpitations and chest pain may also occur. The severity of symptoms can vary from almost asymptomatic to lethal, depending on the species of Borrelia involved, the host immune response, and the stage of the infection. Untreated RF can lead to serious complications such as meningitis, hepatitis, and multi-organ failure in all three types of RF.

6.1.1. TBRF

The symptoms of STBRF usually begin approximately one week after the bite of an infected tick, however the incubation period ranges from 4 to 18 days [9]. Patients typically develop an abrupt fever onset of 38.7–40 °C but can reach even up to 41 °C [99]. The first febrile episode is usually the longest and lasts for 12 h to 17 days with a median of 3 days, is frequently accompanied by nonspecific symptoms including headache, arthralgia, myalgia, and nausea, and ends with a crisis of shivering chills or rigors [306,307]. The average time between the first episode and the first relapse is 7 days and the number of recurrences of fever is up to 13 if left untreated [308]. Due to their increased neurotropism, B. duttonii and B. turicatae infections have neurological symptoms that are more severe than those of other RF pathogens, with meningitis and facial palsy among the most prevalent symptoms [102].

Untreated B. duttonii infections have fatality rates that range from 4 to 10%; however, if appropriate antibiotics are given right away, the death rate is less than 2% [307]. Infection during pregnancy is associated with an increased fatality rate in pregnant women, with 10–15% cases of neonatal deaths and a 43.6% perinatal lethality worldwide [309,310].

6.1.2. LBRF

The incubation period is 4–18 days with a median of 7 days [311]. The symptoms initiate suddenly with a fever of almost 40 °C accompanied by stiffness, often followed by headache, anorexia, nausea, vomiting, diarrhea, and generalized pain (muscle, joint, back pain) [311,312]. The length of LBRF episodes ranges from 4 to 10 days, with a median of 5 days, and it takes 5–9 days between the episodes [307,312]. In general, patients with LBRF may have 3 to 5 febrile episodes [308]. Neurological symptoms are less frequent than in TBRF [312]. Fatality rates for untreated disease range from 30 to 80% [313], however, quick administration of the proper antibiotics reduces the fatality rate to 2–5% [306,307]. Infection during pregnancy is associated with adverse outcomes in three out of four pregnancies [313].

6.2. Epidemiology

6.2.1. TBRF

TBRF has a worldwide distribution, with endemic regions in all continents except Australia and Antarctica [16,99]. Over the past few decades, there have been changes in its epidemiology due to the emergence of new TBRF species, the finding of hard tick RF vectors, the extension of the vectors’ geographic range, the improvements in diagnostic techniques, and the increased awareness among healthcare professionals [143,314]. Some areas such as Europe and Japan have developed into endemic areas and in contrast, others like South America and central Africa were once thought to be more endemic than they actually are [9,307]. Additionally, with instances being recorded in people who have visited endemic regions, TBRF has also become recognized as a disease connected to travel [315].

6.2.2. LBRF

The epidemiology of the once-universal LBRF has radically declined during the past few decades [316]. The disappearance of this infection has been directly linked to the decline in the Pediculus humanus humanus lice infestation of clothing, due to improved living standards along with the use of insecticides [317]. LBRF is still endemic almost exclusively in the Horn of Africa [14], especially in Ethiopia [318,319], Somalia [320] and Eritrea [321], and it occasionally spreads to nearby countries like Sudan [308]. Remaining endemic foci of LBRF are also reported to be small parts of Asia [322] and Latin America [323]. Also, in large Mediterranean cities, homeless populations are at risk of re-emerging LBRF following reported huge lice infestation [324,325,326].

6.2.3. Transmission Vectors of Borrelia spp. Identified in Countries around the Mediterranean Sea

Initially, it was considered that TBRF was transmitted solely by fast-feeding soft-bodied ticks (argasid vectors) of the Ornithodoros genus [107], however, it is now known that Borrelia spp. of the RF group can be transmitted from hard-ticks [143] and human body lice too [327]. In North America, Ornithodoros hermsii and Ornithoodoros turicata are the main tick species to transmit STBRF [306], along with Ornithodoros rudis in South America [328], Ornithodoros erraticus and Ornithodoros tholozani in Eurasia [16,329], and Ornithodoros moubata and Ornithodoros sonrai in Africa [330]. Soft tick bites are rarely noticed since they feed rapidly (15–90 min) and afterwards they return to their habitat. The STBRF-causing spirochetes enter the midgut during the blood meal of Ornithodoros ticks on an infected host, and in the weeks that follow, they colonize the salivary glands where they are maintained transstadially and transovarially [98]. As a result, spirochetes that colonize the salivary gland are more likely to cause an early mammalian infection, leading to rapid transmission of STBRF to the next mammalian host shortly after a bite (Figure 8) [331]. Infection can possibly occur after just 15 s of tick attachment as a tick’s salivary glands are persistently infected [332]. All tick stages are obligate blood feeders and thus potential vectors of Borrelia transmission [306]. Infected ticks remain infectious for their entire lifespan, and they can live up to 10 years infecting several hosts [9,333]. Hard ticks that are known to cause HTBRF are Ixodes spp., Amblyomma spp., Dermacentor spp., and Rhipicephalus spp. [138,334]. The human body louse Pediculus humanus is the only vector of LBRF [316]. Agents of the Borrelia RF group are known to infect tick species throughout the Mediterranean. However, there are insufficient data about Gibraltar, Monaco, Malta, Croatia, Bosnia and Herzegovina, Montenegro, Albania, Greece, Cyprus, Syria, Lebanon, and Libya.

Figure 8.

Transmission of RF Borrelia spirochetes.

Western Mediterranean (Spain, France)

In Spain, DNA from B. miyamotoi was detected in questing I. ricinus nymph and adult ticks [196]. This presence was confirmed across various ecological regions, including coastal, plateau, and mountain areas [335]. Nymph and male I. ricinus ticks feeding on roe deer were also carrying B. miyamotoi bacteria [336]. Additionally, on Espartar Island, a small island in the western Mediterranean Sea, Ornithodoros maritimus ticks from a cave occupied by small seabirds tested positive for Borrelia spp., showing a 99% sequence identity to Borrelia turicatae [337]. In France, DNA samples from free-living I. ricinus nymph ticks collected in the Sénart forest [338] and from the Alpes-Maritimes region [339] revealed the presence of B. miyamotoi. Moreover, B. miyamotoi was prevalent in engorged I. ricinus larva feeding on breeding birds [205] and in I. ricinus and Haemaphysalis punctata ticks feeding on domestic and wild animals from the island of Corsica [340]. Furthermore, Argas vespertilionis ticks collected from a bat-infested attic were also analyzed and a RF Group Borrelia sp. with a 100% sequence identity to strain CPB1 was found [341]. In addition, ticks removed from humans were also tested for the presence of RF Borrelia spp. and it was found that I. ricinus were positive [204,342].

North Central Mediterranean (Italy, Slovenia)

Questing I. ricinus ticks collected by dragging from different altitudes from the mountain areas of northwestern Italy [343], and from both the western and eastern Alps [344], tested positive for B. miyamotoi DNA. B. miyamotoi was also detected in questing I. ricinus ticks from two regions in Slovenia, Littoral Inner Carniola, and the Coastal-Karst area [213].

Eastern Mediterranean (Turkey, Palestine, Israel)

B. miyamotoi DNA was detected in adult I. ricinus ticks collected from the countryside of the European site of Turkey as well as from parks of the metropolitan area of Istanbul [345]. Moreover, Hyalomma aegyptium ticks feeding on Testudo graeca from four different collection sites of Turkey demonstrated the presence of RF Borrelia spp. [346]. Additionally, ticks collected from caves from several locations of the West Bank, Palestine, demonstrated the presence of Borrelia persica in Ornithodoros tholozani [347]. B. persica was also prevalent in nymph female and male O. tholozani ticks from different parts of Israel [348,349].

Southern Mediterranean (Egypt, Tunisia, Algeria, Morocco)

In Egypt, B. miyamotoi was identified in various tick species gathered from one-humped camels in Cairo and Giza. Specifically, these species include Hyalomma dromedarii, Rhipicephalus annulatus, Amblyomma hebraeum, Amblyomma lepidum, Amblyomma cohaerens, and Amblyomma variegatum [221]. Additionally, Borrelia spirochetes were identified in Ornithodoros savignyi from a village of Giza governorate [350]. Furthermore, R. annulatus ticks collected from donkeys tested positive for Borrelia theileri [351], and Ornithodoros erraticus ticks collected from Nile Grass Rat burrows with Borrelia crocidurae [352]. In Tunisia, Rhipicephalus sanguineus ticks feeding on hedgehogs were infected with a RF Group Borrelia sp. [353] and O. erraticus ticks collected from rodent burrows with B. crocidurae [354]. Additionally, B. hispanica was also identified in Ornithodoros sonrai, O. erraticus, and Ornithodoros normandi ticks from small mammal burrows collected from nine different sites [330]. In Algeria, B. crocidurae was also demonstrated in O. sonrai, O. erraticus, and O. normandi ticks from small mammal burrows collected from 18 different sites [330]. In addition, identification of B. hispanica DNA was found in O. occidentalis ticks and B. turicatae in Carios capensis ticks from seabird nests [355]. Furthermore, Argas persicus ticks collected from infested hen farms were also tested and it was revealed that Borrelia anserina was prevalent [356]. Hyalomma aegyptium ticks feeding on Testudo graeca from seven different collection sites of north Algeria as well as from 15 collection sites of central and mostly north Morocco were found to be positive for RF Borrelia spp. [346]. O. sonrai and O. erraticus ticks from small mammal burrows in Morocco were also tested, revealing the presence of B. hispanica, B. crocidurae, and Borrelia merionesi [330,357]. Finally, Ornithodoros marocanus complex ticks from nine sites in Morocco were found infected with B. hispanica and B. crocidurae [358].

To sum up, the circulating Borrelia spp. that have been identified in vectors around the Mediterranean are B. miyamotoi, B. hispanica, B. crocidurae, B. merionesi, B. turicatae, B. anserina, B. persica, and B. theileri. B. miyamotoi has been reported in Spain, France, Italy, Slovenia, Turkey, and Egypt, B. hispanica in Tunisia, Algeria, and Morocco, and B. crocidurae in Egypt, Tunisia, Algeria, and Morocco. In addition, B. merionesi has been found in ticks in Morocco, B. turicatae in Spain and Algeria, B. anserina in Algeria, B. persica in Palestine and Israel, and B. theileri in Egypt (Figure 9 and Table 6).

Figure 9.

Map of Borrelia spp. in vectors in countries around the Mediterranean Sea.

Table 6.

Borrelia spp. in vectors around the Mediterranean Sea.

Country	Borrelia Species
	Borrelia spp.	B. miyamotoi	B. hispanica	B. crocidurae	B. merionesi	B. turicatae	B. anserina	B. persica	B. theileri
Spain		✓				✓
France	✓	✓
Italy		✓
Slovenia		✓
Turkey	✓	✓
Palestine								✓
Israel								✓
Egypt		✓		✓					✓
Tunisia	✓		✓	✓
Algeria	✓		✓	✓		✓	✓
Morocco	✓		✓	✓	✓

6.2.4. Borrelia spp. in Animals in Countries around the Mediterranean Sea

The hosts of RF bacteria vary depending on the specific species. The majority of species can infect humans and small mammals, and a few species are also able to infect birds, as well as domestic or wild mammals [99]. The main hosts of Borrelia hermsii are small mammals such as chipmunks, squirrels, and mice [359] and of Borrelia miyamotoi are a wide range of mammals, including rodents and birds [145,360]. On the other hand, Borrelia anserina infects exclusively birds and is not associated with human disease [152]. Humans are the primary hosts of Borrelia recurrentis and Borrelia duttonii [102,361,362,363]. Several animal species across the Mediterranean have been found to be infected by species of the Borrelia RF group. Limited data are available, though, for the following Mediterranean countries: Gibraltar, Monaco, Malta, Bosnia and Herzegovina, Montenegro, Albania, Cyprus, Syria, Lebanon, Palestine, and Libya.

Western Mediterranean (Spain, France)

In Spain, B. hispanica has been identified in dogs and a street cat through positive blood smear tests and molecular analysis of 16S rRNA which revealed a 100% similarity with B. hispanica for all samples [364]. In France, research on the prevalence of RF spirochetes in bank voles confirmed the presence of B. miyamotoi [365].

North Central Mediterranean (Italy, Slovenia)

In Italy, pigeons were discovered to have antibodies against B. anserina, after analysis with surface immunofluorescence assay (SIFA) and Western blotting [151]. However, in Slovenia, molecular analysis of rodent biopsy specimens indicated the presence of B. miyamotoi [264].

Balkans (Croatia)

In Croatia, B. miyamotoi has been identified in various rodents, including common shrews, yellow-necked mice, and striped field mice [267].

Eastern Mediterranean (Turkey, Palestine, Israel)

B. miyamotoi has been identified in blood samples from wild rodents from Turkey, also with the highest infection rate in Ural field mice, followed by yellow-necked mice, and bank voles [366]. In addition, in West Bank, Palestine, rodents [349] and rock hyraxes [367] were found to be infected with B. persica. In Israel, several species have been identified as potential hosts for RF Borrelia spp.. Natural infection of B. persica in cats and dogs has been confirmed with positive blood smears and molecular assays [368], as well as with serological assays [369]. Moreover, molecular assays with blood samples from rodents and canids from all over Israel demonstrated that the species most commonly infected with B. persica was the social vole, followed by the red fox, the fat sand rat, the golden jackal, and the Cairo spiny mouse [349]. Additionally, besides golden jackals and red foxes, striped hyenas and European badgers also tested positive for B. persica DNA [370]. Finally, B. persica was also identified in blood and spleen samples from rock hyraxes [367].

Southern Mediterranean (Egypt, Tunisia, Algeria, Morocco)

In Egypt, serum samples from various animals were tested for anti-Borrelia spp. antibodies, with the highest infection rate found in camels, followed by sheep, goats, cows, and buffaloes [350]. Furthermore, blood samples from healthy one-humped camels from Cairo and Giza indicated infections with B. crocidurae and B. miyamotoi [221]. In addition, B. theileri has been identified in cattle [371,372] and in sheep blood [372,373]. In Tunisia, blood samples from fat sand rats were examined microscopically and found to be infected with a Borrelia spp. which could not be further characterized [374]. In Algeria, blood samples from cattle were examined, revealing an infection with RF Borrelia species [375]. Furthermore, B. theileri has also been identified in blood samples from sheep and goats by molecular assays [376]. In Morocco, rodents underwent testing for the presence of Borrelia spp. Thick blood film analysis was conducted in blood samples and molecular assays on brain tissue samples, revealing the presence of spirochetes in both blood and brain tissue samples [330,357]. Positive samples were not further determined to species level, except from one area, where B. merionesi was identified [357]. B. crocidurae has also been reported in rodent and insectivore samples [377].

Concluding, the circulating Borrelia spp. that have been identified in animals across the Mediterranean are B. miyamotoi, B. hispanica, B. persica, B. crocidurae, B. merionesi, B. anserina, and B. theileri. B. miyamotoi has been reported in France, Slovenia, Croatia, Turkey, and Egypt, B. hispanica in Spain, B. persica in Palestine and Israel, B. crocidurae in Egypt and Morocco, B. anserina in Italy, B. merionesi in Morocco, and B. theileri in Egypt and Algeria (Figure 10 and Table 7).

Figure 10.

Map of Borrelia spp. in animals in countries around the Mediterranean Sea.

Table 7.

Borrelia spp. in animals around the Mediterranean Sea.

Country	Borrelia Species
	Borrelia spp.	B. hispanica	B. persica	B. crocidurae	B. miyamotoi	B. anserina	B. merionesi	B. theileri
Spain
France
Italy
Slovenia
Croatia
Turkey
Palestine
Israel
Egypt
Tunisia
Algeria
Morocco

Dogs Cats Rodents Birds Hyraxes Foxes/Jackals/Hyenas Sheep Goats Cattle/Buffalos Camels Badger.

6.2.5. Borrelia spp. in Humans in Countries around the Mediterranean Sea

Infections due to the RF Group Borrelia are also present around the Mediterranean. There are no reports of human infection or very little is known on its prevalence and the species involved in the following Mediterranean countries: Gibraltar, Monaco, Malta, Slovenia, Croatia, Bosnia and Herzegovina, Montenegro, Albania, Turkey, Syria, Lebanon, Palestine, and Tunisia.

Western Mediterranean (Spain, France)

In Spain, RF cases have been documented in individuals within one week after exposure to ticks in mountainous regions [378] or following visits to holiday farms [314], working on a pig farm [314], or among intravenous drug users [379]. Moreover, between 1994 and 2016, numerous cases of RF were diagnosed through direct visualization of spirochetes in blood samples. Some of these samples were forwarded for identification, all of which were identified as B. hispanica [380]. Furthermore, from 2004 to 2015 numerous patients were diagnosed with TBRF based on microscopic analysis of blood or cerebrospinal fluid samples and had clinical data similar to those of a B. hispanica infection [381]. In France, B. miyamotoi and B. hermsii have been identified among patients with symptoms related to a tick bite [382]. In addition, a B. miyamotoi infection was confirmed by serological assays after a B. miyamotoi-infected tick bite [204]. Finally, B. recurrentis infections have been identified in homeless individuals by microimmunofluorescence [324,325] and Western Blotting [324].

North Central Mediterranean (Italy)

In Italy, the first autochthonous case of B. crocidurae infection was documented in a 51-year-old woman from Macedonia who was living in Italy from 15 years old and was diagnosed with meningitis due to B. crocidurae as shown by PCR from CSF specimen after a ten-day history of headache, vertical diplopia, and low-grade fever [383].

Balkans (Greece)

Following a two-week camping excursion on the Greek island of Tilos, during which he explored several caves, a traveler from Belgium reported experiencing recurrent febrile episodes. Upon admission to the hospital, spirochetes were detected on his blood smear, and molecular assays to identify the responsible Borrelia sp. confirmed a B. persica infection [384].

Eastern Mediterranean (Cyprus, Israel)

In Cyprus, a number of cases involving RF among civilians and military personnel, all of whom had somehow spent time in caves or near tick habitats, have been documented over the past century, and in some cases spirochetes were also observed in the blood smears of these individuals as well [385,386]. Also, a case of RF contracted in Cyprus and imported in the United Kingdom was published regarding a soldier who had spent three weeks on exercise in Cyprus and spent the night inside a cave where he observed ticks on his feet when he woke up [387]. In Israel, several cases of RF have been reported during the past years. Outbreaks in 2009 and 2010 have been reported among soldiers during cave entry and prolonged close contact with the ground in the context of a military exercise [388]. Additional cases have been documented, including those diagnosed by a hematological laboratory [389], neonatal cases [390] with severe Jarisch–Herxheimer reaction [391], cases who received placebos in a double-blind trial for post-exposure treatment to prevent TBRF [392], and cases confirmed by molecular assays for B. persica infection [348,393]. B. persica was also identified in spirochetemic patients from Israel as well as in patients with similar clinical manifestations, but with negative blood smears [394], and in a TBRF patient with adult respiratory distress syndrome (ARDS) [395].

Southern Mediterranean (Egypt, Libya, Algeria, Morocco)

A case of RF in a patient with nephritis and subarachnoid hemorrhage, confirmed by blood film, was report in Egypt after spending the night on the ground before the onset of the symptoms [396]. Additionally, humans have been found to carry antibodies against Borrelia spp. antigen [350]. In Libya, cases of RF with confirmed diagnosis by blood film, have been reported in one case of a young Palestinian driver who had not been anywhere else other than Benghazi for at least one month before the onset of his symptoms, an Italian P.O.W. whose specific route before the illness onset is unknown, but he came from Western Cyrenaica, an Indian pioneer who had spent his time wholly in Cyrenaica and in Benghazi, and a British lance-corporal who spent two nights in Tobruk and otherwise was totally in Benghazi [397]. In Algeria, Borrelia spp. DNA has been identified in feverish patients from Oran, and one sample was indicative of a new Borrelia spp. called Candidatus Borrelia algerica [103]. Finally, in Morocco, cases of RF have been confirmed in patients with unexplained fever by positive blood film [357], as well as by molecular assays demonstrating B. hispanica infections [398,399].

To sum up, the circulating Borrelia spp. that have been reported to cause human infections in countries around the Mediterranean Sea are B. hispanica, B. persica, B. crocidurae, B. miyamotoi, B. hermsii, Candidatus B. algerica, and B. recurrentis. B. hispanica has been reported in Spain and Morocco, B. persica in Greece and Israel, B. crocidurae in Italy, B. miyamotoi in France, B. hermsii in France, Candidatus B. algerica in Algeria, and B. recurrentis in France (Figure 11 and Table 8, Box 2).

Figure 11.

Map of Borrelia spp. in humans in countries around the Mediterranean Sea.

Table 8.

Borrelia spp. in humans around the Mediterranean Sea.

Country	Borrelia Species
	Borrelia spp.	B. hispanica	B. persica	B. crocidurae	B. miyamotoi	B. hermsii	Candidatus B. algerica	B. recurrentis
Spain	✓	✓
France					✓	✓		✓
Italy				✓
Greece			✓
Cyprus	✓
Palestine	✓
Israel	✓		✓
Egypt	✓
Libya	✓
Algeria							✓
Morocco	✓	✓

Box 2. Prevention of RF.

Preventing TBRF relies on minimizing exposure to soft ticks. This can be achieved by avoiding habitats where soft ticks are prevalent, such as caves, animal burrows, huts, and cabins. Furthermore, it is recom-mended to use tick repellents on clothing and skin, along with wearing long-sleeved shirts and trousers tucked into socks when engaging in outdoor activities. On the other hand, preventing LBRF primarily de-pends on maintaining personal and clothing hygiene to prevent infestation by body lice.

6.2.6. Imported Human Cases of RF in Mediterranean Countries

Cases of RF are not only contracted in Mediterranean countries but can also be transferred there due to travelling or immigration in other countries, especially from central Africa and Asia. In France, several cases of B. crocidurae infection have been identified, most of which were in immigrants from Somalia [400,401,402,403] passing through Mauritania [315] or immigrants from Mali [315]. Additionally, a case of B. hispanica was identified after travel in Morocco and Spain [315] and another of B. persica after a trip to Uzbekistan and Tajikistan [404]. In Italy, the vast majority of imported RF cases are due to B. recurrentis in immigrants from the Horn of Africa, e.g., Somalia, Eritrea, and Ethiopia [15,405,406]. The immigration route usually includes more or less prolonged stays in Sudan and Libya [326,407,408], and in some cases the immigration route includes Kenya and South Sudan, before crossing Sudan and Libya [409,410]. However, a case of B. recurrentis has also been reported in an immigrant from Mali, who passed by Algeria and Libya before arriving to Italy [411]. Also, a B. crocidurae infection was identified in Italy after travel to Senegal [412]. Finally, B. recurrentis infection has been demonstrated in Israel in Ethiopian Pilgrims [413] and in Libya and Italy in immigrants from Somalia with Germany as their final destination, who were passing through Libya and Italy as intermediate stations [14] (Table 9).

Table 9.

Imported RF cases in Mediterranean countries.

Country of Manifestation	Country of Origin/Immigration Route	Borrelia Species	References
France	Senegal	B. crocidurae	[400,401,402,403]
	Senegal and Mauritania	B. crocidurae	[315]
	Mali	B. crocidurae	[315]
	Morocco and Spain	B. hispanica	[315]
	Uzbekistan and Tajikistan	B. persica	[404]
Italy	Eritrea/Somalia/Ethiopia and Libya	B. recurrentis	[15]
	Somalia and Libya	B. recurrentis	[326,408]
	Somalia, Kenya, South Sudan, Sudan, and Libya	B. recurrentis	[409]
	Somalia/Sudan and Libya	B. recurrentis	[407]
	East Africa	B. recurrentis	[405]
	Mali, Algeria, and Libya	B. recurrentis	[411]
	Somalia, Kenya, Uganda, Sudan, and Libya	B. recurrentis	[410]
	Somalia	B. recurrentis	[406]
	Senegal	B. crocidurae	[412]
Israel	Ethiopia	B. recurrentis	[413]
Libya and Italy as intermediate destinations	Somalia, Ethiopia, Kenya, South Sudan, and Sudan	B. recurrentis	[14]

7. Conclusions

In order to successfully monitor and limit the spread of Borrelia infections, it is vital that the medical, veterinary, and environmental sectors collaborate. For this reason, we provide insight for every country around the Mediterranean Sea with available information regarding the prevalence of all Borrelia spp. that have been implicated in tick, animal, and human infection. In summary, LD agents were mostly found in the European part of the Mediterranean. Several Borrelia spp. were identified in hard ticks and in numerous animals, such as rodents, wild ungulates, farm animals, and pets. RF agents were identified all around the Mediterranean in both soft ticks as well as hard ticks. Many animal species were reported to be infected with RF agents, such as rodents, pets, and farm animals. In conclusion, infections caused by Borrelia spp. are still a serious issue that afflict thousands of people every year and hence demand our undivided attention.

Supplementary Materials

The tables with all the references of the Borrelia spp. in humans, animals and vectors can be downloaded at https://www.mdpi.com/article/10.3390/pathogens13060512/s1, Table S1: Lyme Group Borreliella spp. in humans [41,201,222,265,270,284,285,286,287,288,289,290,291,292,293,294,295,296,297,298,299,300,301,302,303,304,305,382,414,415,416,417,418,419,420,421,422,423,424,425,426,427,428,429,430,431,432,433,434,435,436,437,438,439,440,441,442,443,444,445,446,447,448,449,450,451,452,453,454,455,456,457,458,459,460,461,462,463,464,465,466,467,468,469,470,471,472,473,474,475,476,477,478,479,480,481,482,483,484,485,486,487,488,489,490,491,492,493,494,495,496,497,498,499,500,501,502,503,504,505,506,507,508,509,510,511,512,513,514,515,516,517]; Table S2: Lyme Group Borreliella spp. in animals [198,199,221,222,238,239,240,241,242,243,244,245,246,247,248,249,250,251,252,253,254,255,256,257,258,259,260,261,262,263,264,265,266,267,268,269,270,271,272,273,274,275,276,277,278,279,280,281,282,283,365,373,447,496,502,518,519,520,521,522,523,524,525,526,527,528,529,530,531,532,533,534,535,536,537,538]; Table S3: Lyme Group Borreliella spp. in ticks [61,87,196,197,198,199,200,201,202,203,204,205,206,207,208,209,210,211,212,213,214,215,216,217,218,219,220,221,222,223,224,225,226,233,244,250,251,257,259,279,281,285,291,295,335,336,338,339,340,342,343,346,365,447,473,495,501,502,525,535,539,540,541,542,543,544,545,546,547,548,549,550,551,552,553,554,555,556,557,558,559,560,561,562,563,564,565,566,567,568,569,570,571,572,573,574,575,576,577,578,579,580,581,582,583,584,585,586,587,588,589,590,591,592,593,594,595,596,597,598,599,600,601,602,603,604,605,606,607,608,609,610,611,612,613,614,615,616,617,618,619,620,621,622,623,624,625,626,627,628,629,630,631,632,633,634,635,636,637,638,639,640,641]; Table S4: Relapsing Fever Group Borrelia spp. in humans [103,204,314,324,325,347,348,350,357,378,379,380,381,382,383,384,385,386,387,388,389,390,391,392,393,394,395,396,397,398,399,642,643,644,645,646,647,648,649,650,651]; Table S5: Relapsing Fever Group Borrelia spp. in animals [151,221,264,267,330,349,350,357,364,365,366,367,368,369,370,371,372,373,374,375,376,377,652,653]; Table S6: Relapsing Fever Group Borrelia spp. in ticks [196,204,205,213,221,330,335,336,337,338,339,340,341,342,343,344,345,346,347,348,349,351,352,353,354,355,356,357,358,365,447,540,551,559,562,563,566,568,570,575,576,582,643,653,654,655].

Author Contributions

Data curation, M.K.; writing—original draft preparation, M.K.; review and editing, M.D.; supervision, E.A. All authors have read and agreed to the published version of the manuscript.

Data Availability Statement

The original contributions presented in the study are included in the article/Supplementary Material, further inquiries can be directed to the corresponding author.

Conflicts of Interest

The authors declare no conflicts of interest.

Funding Statement

This research received no external funding.