Molecular characterization of Echinococcus granulosus isolates from Bulgarian human cystic echinococcosis patients

Although cystic echinococcosis (CE) is highly endemic in Bulgaria, there is still scarce information about species and/or genotypes of the Echinococcus granulosus complex that infect humans. Our study tackled the genetic diversity of E. granulosus complex in a cohort of 30 Bulgarian CE patients. Ten animal E. granulosus isolates from neighboring Greece were additionally included. Specimens were comparatively analyzed for partial sequences of five mitochondrial (mt) (cox I, nad I, rrnS, rrnL, and atp6) and three nuclear (nc) genes (act II, hbx 2, and ef-1α) using a PCR-sequencing approach. All 30 Bulgarian isolates were identified as E. granulosus sensu stricto (s.s.) and were showing identical sequences for each of the three examined partial nc gene markers. Based upon concatenated sequences from partial mtDNA markers, we detected 10 haplotypes: 6 haplotypes (H1-H6) clustering with E. granulosus s.s. (G1) and 4 haplotypes (H9-H13) grouping with E. granulosus s.s. (G3), with H1 and H10 being the most frequent in Bulgarian patients. The haplotypes H1, H4, and H11 were also present in Greek hydatid cyst samples of animal origin. In conclusion, E. granulosus s.s. (G1 and G3 genotypes) is the only causative agent found so far to cause human CE in Bulgaria. However, further studies including larger sample sizes and other additional geographic regions in Bulgaria will have to be performed to confirm our results

Coxiella burnetii Shedding in Milk and Molecular Typing of Strains Infecting Dairy Cows in Greece

Ruminants are considered the commonest animal reservoir for human infection of Coxiella burnetii, the Q fever causative agent. Considering the recently described importance of human Q fever in Greece, we aimed at providing the first comprehensive direct evidence of C. burnetii in dairy cows in Greece, including the genetic characterization of strains. The 462 examined dairy farms represented all geographical areas of Greece. One bulk tank milk sample was collected from every farm and tested for the presence of C. burnetii. Molecular genotyping of strains, performed directly on samples, revealed the existence of two separate clades characterized by single nucleotide polymorphism (SNP) genotypes of type 1 and type 2. The two clades were clearly distinguished in multiple locus variable-number tandem repeat analysis (MLVA) by two discriminative loci: MS30 and MS28. Whereas MLVA profiles of SNP-type 2 clade were closely related to strains described in other European cattle populations, the MLVA profile observed within the SNP type 1 clade highlighted a peculiar genetic signature for Greece, related to genotypes found in sheep and goats in Europe. The shedding of C. burnetii bearing this genotype might have yet undefined human epidemiological consequences. Surveillance of the genetic distribution of C. burnetii from different sources is needed to fully understand the epidemiology of Q fever in Greece

The geographical distribution of hard ticks (Acari: Ixodidae) in Greece and their role in the epidemiology of Crimean-Congo hemorrhagic fever

Crimean-Congo Hemorrhagic Fever (CCHF) is a severe acute febrile illness often accompanied by hemorrhagic manifestations; the fatality rate ranges between 5% - 30%. The causative agent is the Crimean-Congo hemorrhagic fever virus (CCHFV, genus Orthonairovirus, family Nairoaviridae). It is transmitted to humans by bites of infected ticks, mainly of the Hyalomma genus, or by direct contact with blood or tissues of patients or livestock at viremic phase. CCHFV is the most widespread tick-borne viral disease and has been reported in more than 30 countries in Africa, Asia, Eastern and Southern Europe and the Middle East. The disease has a wide distribution that correlates with the global distribution of Hyalomma ticks (50o north latitude limit). Nine genetic clades are currently recognized (four from Africa, three from Europe and two from Asia), which present a geographic correlation: “Africa 1”, “Africa 2”, “Africa 3” and “Africa 4” with strains from Central and South Africa; “Asia 1” from Middle East; “Asia 2” from China, Tajikistan, Kazakhstan and Uzbekistan; “Europe 1” from Balkan peninsula, Russia and Turkey; “Europe 2” consisting of the AP92 strain from Greece and “Europe 3” with strains from Russia. The strains that are circulating in Greece belong to the lineages “Europe 1” and “Europe 2” which differ genetically and in virulence level. The only human (fatal) case was reported in June 2008 and the strain belonged to lineage “Europe 1”, while the AP92 strain which belongs to lineage “Europe 2”, was isolated in 1975 from R. bursa ticks collected from goats in Vergina village in Central Macedonia. The purpose of the present study was to provide an overview on the prevalence and distribution of tick species in various regions of Greece (especially in areas where seroprevalence studies in humans have been previously performed), to identify the geographical parameters of the tick collection sites through the use of geographic and spatial analysis systems (GIS-SA), to estimate the CCHFV infection rate in ticks and to identify the genetic lineage of the circulating CCHFV strains. Therefore, a cross sectional field study on the distribution of Ixodidae ticks was carried out over three consecutive tick seasons (2012 - 2014); ticks were collected from small ruminants in 309 farms located in the mainland and five islands of Greece. In total, 2108 ticks were collected from sheep and goats. The identification of ticks to species level was carried out using taxonomic keys; two species of Rhipicephalus, two of Ixodes, five of Hyalomma, three of Haemaphysalis, and one of Dermacentor were identified. The species with the widest distribution were Rhipicephalus sanguineus sensu lato (s.l.) (64.8 %), R. bursa (25.9 %), and Dermacentor marginatus (4.1 %), whereas the least frequently collected species were Ixodes ricinus, I. gibbosus, Haemaphysalis parva, H. sulcata, H. punctata, Hyalomma marginatum, H. excavatum, H. dromedarii, H. rufipes, and H. impeltatum representing together less than 5.3 % of the collection. Grouping of ticks by collection date, location, animal host, tick species, and sex resulted in 1290 pools. Following tick homogenization and RNA extraction, an RT-nested PCR was applied for the detection of CCHFV. CCHFV was detected in 2.8% (36 of 1290) of the tick pools. Specifically, CCHFV was detected in 3.2% (27/843) of the R. sanguineus s.l. tick pools and in 2.4% (9 of 377) of the R. bursa tick pools. The regions with the highest CCHFV detection rate were Central and Western Macedonia (8.8%), Crete (6.2%) and Peloponnese (5.7%), while in Thessaly, Thrace and Epirus the percentage of positive pools were 3.1%, 1.4% and 1.2%, respectively. All H. marginatum ticks (the principal vector of CCHFV) were CCHFV negative. Phylogenetic analyses of the virus showed that sequences clustered into two genetic lineages, “Europe 1” (19/27) and “Europe 2” (8/27). Sequences of CCHFV lineage “Europe 1” were obtained mainly from R. sanguineus s.l. ticks (16/19), while sequences of lineage “Europe 2” were taken from R. bursa ticks (6/8). The sequences of the CCHFV lineage “Europe 2” of the present study differed from the prototype AP92 strain by 9.7% at nucleotide level. The present study gave a first insight into the tick fauna from mainland and islands in Greece. To date, there are no reports of CCHF cases in Greece (except the one in 2008 which was caused by a CCHFV strain belonging to the “Europe 1” lineage). The circulation of CCHFV strain(s) of lineage “Europe 2” in Greece, especially in regions where the seroprevalence is relatively high, together with the lack of human CCHF cases, suggests these strains are antigenic, with non- or low-pathogenic to humans. Further studies on these strains will increase our knowledge about the role of AP92-like strains in the CCHF epidemiology, which might be useful for drug and vaccine design. Regarding lineage “Europe 1”, it is probable that ticks of the Rhipicephalus genus are not competent virus vectors as are the ticks of Hyalomma genus. Further laboratory studies are needed to explore the role of Rhipicephalus spp. ticks in CCHFV maintenance and transmission.Ο αιμορραγικός πυρετός Κριμαίας-Κογκό (Crimean - Congo Hemorrhagic Fever– CCHF), είναι σοβαρή οξεία εμπύρετη νόσος που συνοδεύεται συχνά από αιμορραγικές εκδηλώσεις και παρουσιάζει θνητότητα που κυμαίνεται από 5%-30%. Αιτιολογικός παράγοντας της νόσου είναι ο ιός του αιμορραγικού πυρετού Κριμαίας-Κογκό (CCHFV, genus Orthonairovirus, family Nairoaviridae). Η μετάδοση της νόσου στον άνθρωπο γίνεται με νύγμα κροτώνων, κυρίως του γένους Hyalomma, ή μετά από άμεση επαφή των βλεννογόνων ή του δέρματος με αίμα, εκκρίματα ή ιστούς μολυσμένων ζώων ή ανθρώπων. Η γεωγραφική διασπορά του CCHFV είναι η πιο εκτεταμένη από τους ιούς που μεταδίδονται με κρότωνες, καθ΄ όσον η παρουσία του αναφέρθηκε σε περισσότερες από 30 χώρες στην Αφρική, Ασία, Ν.Α Ευρώπη και στη Μέση Ανατολή. Η κατανομή του ιού ακολουθεί τη διασπορά των κροτώνων του γένους Hyalomma, οι οποίοι εξαπλώνονται έως το 50° βόρειο γεωγραφικό πλάτος. Τα στελέχη του CCHF διαχωρίζονται σε 9 γονότυπους: οι γονότυποι “Africa 1”, “Africa 2” , “Africa 3” και “Africa 4” περιλαμβάνουν στελέχη διαφόρων χωρών της Αφρικής, ο γονότυπος “Asia 1” με στελέχη της Μέσης Ανατολής, ο “Asia 2” με στελέχη από την Κίνα, το Τατζικιστάν, το Καζακστάν, το Ουζμπεκιστάν, ο “Europe 1” με στελέχη από τα Βαλκάνια, τη Ρωσία και την Τουρκία, ο “Europe 2” που περιλαμβάνει ένα μόνο στέλεχος, το ελληνικό στέλεχος AP92 και ο γονότυπος “Europe 3” με στελέχη από τη Ρωσία. Στην Ελλάδα ανιχνεύθηκαν στελέχη που ανήκουν στους γονοτύπους “Europe 1” και “Europe 2”, τα οποία διαφέρουν σημαντικά στο γονιδίωμα και στη λοιμογόνο δύναμή τους. Η μόνη (και θανατηφόρα) περίπτωση CCHF στην Ελλάδα παρατηρήθηκε τον Ιούνιο του 2008 και το στέλεχος ανήκε στον γονότυπο “Europe 1” ενώ το στέλεχος AP92 που ανήκει στον γονότυπο “Europe 2” απομονώθηκε το 1975 από κρότωνες Rhipicephalus bursa στην Βεργίνα της Κεντρικής Μακεδονίας. Σκοπός της παρούσας μελέτης ήταν η καταγραφή της διασποράς και συχνότητας των κροτώνων σε διάφορες περιοχές της Ελλάδας (ιδιαίτερα σε αυτές όπου είχαν πραγματοποιηθεί προηγουμένως οροεπιδημιολογικές μελέτες στον γενικό πληθυσμό), η χαρτογράφησή τους μέσω χρήσης συστημάτων γεωγραφικής και χωρικής ανάλυσης (GIS- SA), ο έλεγχος των κροτώνων για μόλυνση με τον CCHFV, καθώς και η γενετική και η φυλογενετική ανάλυση των στελεχών της μελέτης. Συγκεκριμένα, διεξήχθη μια εκτεταμένη μελέτη του πληθυσμού και της διασποράς των σκληρών κρότωνων (Ixodidae) σε τρεις διαδοχικές περιόδους (2012 - 2014) σε 309 κτηνοτροφικές μονάδες αιγών και προβάτων που βρίσκονται στην ηπειρωτική χώρα και σε πέντε νησιά της Ελλάδας. Συνολικά συλλέχθηκαν 2108 κρότωνες. Η ταυτοποίησή τους με ειδικές για αρθρόποδα διχοτομικές κλείδες ανέδειξε δύο είδη Rhipicephalus, δύο Ixodes, πέντε Hyalomma, τρία Haemaphysalis και ένα Dermacentor. Τα είδη με την ευρύτερη κατανομή ήταν τα Rhipicephalus sanguineus sensu lato (s.l.) (64,8%), R. bursa (25,9%) και Dermacentor marginatus (4,1%), και ακολουθούσαν τα Ixodes ricinus, Ι. gibbosus, Haemaphysalis parva, Η. sulcata, Η. punctata, Hyalomma marginatum, Η. excavatum, Η. dromedarii, Η. rufipes και Η. impeltatum που όλα μαζί αντιπροσωπεύουν περίπου το 5,3% της συλλογής. Οι κρότωνες διαχωρίστηκαν σε 1290 ομάδες ανάλογα με το είδος, το φύλο, το ξενιστή, την ημερομηνία και την περιοχή συλλογής τους. Μετά από ομογενοποίηση, απομονώθηκε το ιικό RNA και εφαρμόστηκε αλυσιδωτή αντίδραση πολυμεράσης (RT-nested PCR) για την ανίχνευση του CCHFV. Ο ιός ανιχνεύθηκε σε 36 από τις 1290 (2,8%) ομάδες κροτώνων. Ειδικότερα, ανιχνεύθηκε σε 27 από τις 843 (3,2%) ομάδες κροτώνων του είδους R. sanguineus s.l. και σε 9 από τις 377 (2,4%) ομάδες του είδους R. bursa. Οι περιφέρειες με τα υψηλότερα ποσοστά ανίχνευσης του CCHFV ήταν η Κεντρική και Δυτική Μακεδονία (8,8%), η Κρήτη (6,2%) και η Πελοπόννησος (5,7%), ενώ στις Θεσσαλία, Θράκη και Ήπειρο τα ποσοστά ήταν 3,1%, 1,4% και 1,2% αντίστοιχα. Όλοι οι κρότωνες του είδους H. marginatum (που αποτελεί τον κύριο φορέα του CCHFV) ήταν αρνητικοί. Από την φυλογενετική ανάλυση των αλληλουχιών του ιού που ανιχνεύθηκαν σε 27 ομάδες κροτώνων φάνηκε η παρουσία των γονοτύπων “Europe 1” (19/27) και “Europe 2” (8/27). Ο γονότυπος “Europe 1” βρέθηκε κυρίως σε κρότωνες του είδους R. sanguineus s.l. (16/19), ενώ ο “Europe 2” σε κρότωνες του είδους R. bursa (6/8). Το γονιδίωμα του στελέχους CCHFV γονοτύπου “Europe 2” που ανιχνεύθηκε σε κρότωνες R. bursa διέφερε κατά 9,7% σε επίπεδο νουκλεοτιδίων από το πρότυπο στέλεχος AP92, το οποίο είχε απομονωθεί το 1975.Συμπερασματικά, η παρούσα μελέτη έδωσε μία πρώτη αποτύπωση σχετικά με τον πληθυσμό των κροτώνων στην ηπειρωτική χώρα και τα νησιά της Ελλάδας. Μέχρι σήμερα, δεν έχουν αναφερθεί κλινικές περιπτώσεις CCHF σε ανθρώπους (εκτός από μία περίπτωση το 2008 που οφειλόταν σε στέλεχος του γονοτύπου “Europe 1”). Η κυκλοφορία του CCHFV του γονοτύπου “Europe 2” σε περιοχές όπου ο οροεπιπολασμός είναι σχετικά υψηλός, σε συνδυασμό με την απουσία κλινικών περιστατικών, υποδηλώνει ότι αυτά τα στελέχη παρουσιάζουν αντιγονικότητα, αλλά έχουν μικρή η μηδαμινή λοιμογόνο δύναμη. Περαιτέρω μελέτες σε στελέχη του γονοτύπου “Europe 2” θα αυξήσουν τις γνώσεις μας σχετικά με το ρόλο αυτών των στελεχών τύπου ΑΡ92 στην επιδημιολογία του CCHF, οι οποίες θα μπορούσαν να είναι χρήσιμες για το σχεδιασμό φαρμάκων και εμβολίων. Όσον αφορά τον γονότυπο “Europe 1”, είναι πιθανό, οι κρότωνες του γένους Rhipicephalus να μην είναι ικανοί φορείς του ιού όπως είναι οι κρότωνες του γένους Hyalomma, οπότε θα απαιτηθούν περαιτέρω εργαστηριακές μελέτες για να διερευνηθεί ο ακριβής ρόλος τους στη συντήρηση και μετάδοση του CCHFV

Species diversity and spatial distribution of ixodid ticks on small ruminants in Greece

A cross sectional field study on the distribution of Ixodidae ticks was carried out over two consecutive tick seasons (2012 and 2013) in small ruminants (309 farms) located in mainland and five islands of Greece. A total of 2108 ticks (1199 females, 908 males, and 1 nymph) were collected from sheep and goats. Two species of Rhipicephalus, two of Ixodes, five of Hyalomma, three of Haemaphysalis, and one of Dermacentor were detected. The species with the widest distribution were Rhipicephalus sanguineus s.l. (64.8 %), R. bursa (25.9 %), and Dermacentor marginatus (4.1 %), whereas the least frequently collected species were Ixodes ricinus, I. gibbosus, Haemaphysalis parva, H. sulcata, H. punctata, Hyalomma marginatum, H. excavatum, H. dromedarii, H. rufipes, and H. impeltatum representing together less than 5.3 % of the collections. R. sanguineus s.l. was the predominant species, while D. marginatus and I. gibbosus were restricted to the northeastern Aegean Sea islands. This is the first survey on tick fauna from mainland and islands in Greece.The work is part of the FP7 project “ANTIcipating the Global Onset of Novel Epidemics” (Grant agreement no. 278976) supported by the European Commission.Peer Reviewe

First Identification and Molecular Characterization of Avian metapneumovirus Subtype B from Chickens in Greece

Avian metapneumovirus (aMPV) is considered a major pathogen for turkeys but its impact on chicken production is still partially neglected, even though it is fully acknowledged as a primary pathogen in chickens as well. The lack of structured diagnostic surveys does not allow a pervasive understanding of aMPV epidemiology. Being that aMPV is almost an everyday challenge for farmers and veterinarians, a more accurate report of its presence should be detailed, posing the basis for a deep and global epidemiologic analysis. With these premises, the present work aims to report the first detection and molecular characterization of aMPV subtype B field strains from unvaccinated chickens in Greece. The Greek strains appear to be phylogenetically related among each other and with other recent Mediterranean strains while being distant from the currently applied vaccines, thus stressing once more the necessity to evaluate aMPV diffusion and evolution

Molecular identification of tick-borne pathogens in ticks collected from dogs and small ruminants from Greece

Ticks are vectors for a variety of human and animal pathogens (bacteria, protozoa and viruses). In order to investigate the pathogens carried by ticks in Greece, a total of 179 adult ticks (114 female and 65 male) were collected from domestic animals (sheep, goats and dogs) from 14 prefectures of six regions of Greece. Among them, 40 were Dermacentor marginatus, 25 Haemaphysalis parva, 22 H. sulcata, one H. punctata, 13 Ixodes gibbosus, 77 Rhipicephalus sanguineus s.l. and one R. bursa. All ticks were tested for the presence of DNA of Anaplasma spp., Babesia spp., Coxiella burnetii, Rickettsia spp. and Theileria spp. The collected ticks were examined by PCR and reverse line blot (RLB) assay. A prevalence of 20.1% for Anaplasma spp., 15.6% for Babesia spp. (identifying B. bigemina, B. divergens, B. ovis and B. crassa), 17.9% for C. burnetii, 15.1% for Rickettsia spp., and 21.2% for Theileria spp. (identifying T. annulata, T. buffeli/orientalis, T. ovis and T. lestoquardi) was found. The results of this study demonstrate the variety of tick-borne pathogens of animal and human importance circulating in Greece, and that awareness is needed to minimize the risk of infection, especially among farmers and pet owners.This study was financially supported by the State Scholarship Foundation IKY, NSRF 2007–2013. IGFM is funded from the Own Research Program by University of Castilla-La Mancha.Peer Reviewe

How Safe to Eat Are Raw Bivalves? Host Pathogenic and Public Health Concern Microbes within Mussels, Oysters, and Clams in Greek Markets

Raw-bivalves consumption is a wide trend in Mediterranean countries. Despite the unambiguous nutritional value of seafood, raw consumption of bivalves may involve risks that could pose a significant threat to consumers’ health. Their filter-feeding behavior is responsible for the potential hosting of a wide variety of microorganisms, either pathogenic for the bivalves or public health threats. Under this prism, the current study was conducted in an effort to evaluate the risk of eating raw bivalves originating from the two biggest seafood markets in Thessaloniki, the largest production area of bivalves in Greece. Both microbiological and molecular methodologies were applied in order to assess the presence of various harmful microbes, including noroviruses, Bonamia, Marteilia, Esherichia coli, Salmonella, and Vibrio. Results indicated the presence of several Vibrio strains in the analyzed samples, of which the halophilic Vibrio harveyi was verified by 16S rRNA sequencing; other than this, no enteropathogenic Vibrio spp. was detected. Furthermore, although Esherichia coli was detected in several samples, it was mostly below the European Union (EU) legislation thresholds. Interestingly, the non-target Photobacterium damselae was also detected, which is associated with both wound infections in human and aquatic animals. Regarding host pathogenic microorganisms, apart from Vibrio harveyi, the protozoan parasite Marteilia refrigens was identified in oysters, highlighting the continuous infection of this bivalve in Greece. In conclusion, bivalves can be generally characterized as a safe-to-eat raw food, hosting more bivalve pathogenic microbes than those of public health concern

Coxiella burnetii Shedding in Milk and Molecular Typing of Strains Infecting Dairy Cows in Greece

Ruminants are considered the commonest animal reservoir for human infection of Coxiella burnetii, the Q fever causative agent. Considering the recently described importance of human Q fever in Greece, we aimed at providing the first comprehensive direct evidence of C. burnetii in dairy cows in Greece, including the genetic characterization of strains. The 462 examined dairy farms represented all geographical areas of Greece. One bulk tank milk sample was collected from every farm and tested for the presence of C. burnetii. Molecular genotyping of strains, performed directly on samples, revealed the existence of two separate clades characterized by single nucleotide polymorphism (SNP) genotypes of type 1 and type 2. The two clades were clearly distinguished in multiple locus variable-number tandem repeat analysis (MLVA) by two discriminative loci: MS30 and MS28. Whereas MLVA profiles of SNP-type 2 clade were closely related to strains described in other European cattle populations, the MLVA profile observed within the SNP type 1 clade highlighted a peculiar genetic signature for Greece, related to genotypes found in sheep and goats in Europe. The shedding of C. burnetii bearing this genotype might have yet undefined human epidemiological consequences. Surveillance of the genetic distribution of C. burnetii from different sources is needed to fully understand the epidemiology of Q fever in Greece

Molecular identification of spotted fever group Rickettsia in ticks collected from dogs and small ruminants in Greece

Spotted fever group (SFG) Rickettsia are zoonotic and emerging pathogens with considerable impact in public and animal health. Greece is an endemic country of diseases caused by SFG Rickettsia. This work aims to evaluate the prevalence of SFG Rickettsia in ticks collected from domestic hosts including sheep, goats and dogs. Several genetic markers for bacterial genes, such as 16S rRNA, ompA, ompB, atpA, gltA, recA, dnaA and dnaK, were amplified and sequenced to accurately classified the rickettsial pathogens in the ticks. Taxonomy and species classification of the Rickettsia was achieved by combining phylogenetic and in silico digestion analysis of the gene sequences obtained. A total of 187 ticks were collected and classified at the species level as Ixodes gibosus, Dermacentor marginatus, Haemaphysalis parva, H. sulcata, H. punctata, Hyalomma scavatum, Rhipicephalus sanguineus, R. bursa and Rhipicephalus sp. The results showed that 7.5% of ticks were infected with at least one SFG Rickettsia including R. massiliae (n = 3), R. slovaca (n = 5), R. raoultii (n = 1) and R. hoogstraalii (n = 5), collected from sheep (n = 4), goats (n = 5) and dogs (n = 3). Molecular analysis revealed the presence of novel genetic variants of R. hoogstraalii (in H. sulcata and H. parva from goat and sheep) and R. raoultii (in D. marginatus from goat). These results proof the presence of SFG Rickettsia in domestic hosts in Greece, and support the need for continued monitoring, surveillance and further analyses of other hosts and study areas.This study was financially supported by the State Scholarship Foundation IKY, NSRF 2007–2013. IGFM is funded from the Own Research Program by University of Castilla-La Mancha.Peer reviewe

Coxiella burnetii Shedding in Milk and Molecular Typing of Strains Infecting Dairy Cows in Greece

Ruminants are considered the commonest animal reservoir for human infection of Coxiella burnetii, the Q fever causative agent. Considering the recently described importance of human Q fever in Greece, we aimed at providing the first comprehensive direct evidence of C. burnetii in dairy cows in Greece, including the genetic characterization of strains. The 462 examined dairy farms represented all geographical areas of Greece. One bulk tank milk sample was collected from every farm and tested for the presence of C. burnetii. Molecular genotyping of strains, performed directly on samples, revealed the existence of two separate clades characterized by single nucleotide polymorphism (SNP) genotypes of type 1 and type 2. The two clades were clearly distinguished in multiple locus variable-number tandem repeat analysis (MLVA) by two discriminative loci: MS30 and MS28. Whereas MLVA profiles of SNP-type 2 clade were closely related to strains described in other European cattle populations, the MLVA profile observed within the SNP type 1 clade highlighted a peculiar genetic signature for Greece, related to genotypes found in sheep and goats in Europe. The shedding of C. burnetii bearing this genotype might have yet undefined human epidemiological consequences. Surveillance of the genetic distribution of C. burnetii from different sources is needed to fully understand the epidemiology of Q fever in Greece