315 Veterinaria Italiana 2022, 58 (3), 315-323. doi: 10.12834/VetIt.2555.16997.2 Accepted: 07.03.2022 | Available on line: 31.12.2022 1University Manouba, Institution de la Recherche et de l'Enseignement Supérieur Agricoles, Service de Microbiologie et d’Immunologie, Ecole Nationale de Médecine Vétérinaire, 2020 Sidi Thabet, Tunisie. 2University Manouba, Institution de la Recherche et de l'Enseignement Supérieur Agricoles, Laboratoire de Parasitologie, École Nationale de Médecine Vétérinaire, 2020 Sidi Thabet, Tunisie. *Corresponding author at: Service de Microbiologie et d’Immunologie, Ecole Nationale de Médecine Vétérinaire, 2020 Sidi Thabet, Ariana, Tunisie. Tel.: +216 71 552 200, Mobile: +216 97003060 , Fax: +216 71 552 441, E‑mail: lilia_messadi@yahoo.fr. Ghassan Tayh1, Asma Ben Haj Yahia1, Rachid Selmi1, Sarrah Landolsi1, Faten Ben Chehida1, Aymen Mamlouk1, Mohamed Habib Jemli2, Monia Dâaloul-Jedidi1 and Lilia Messadi1* Keywords E. coli O157:H7, Camels, Shiga-like toxin genes, Enterohemorrhagic Escherichia coli, Enterohaemolysin (ehxA), Antimicrobial resistance. Summary Shiga-toxin-producing E. coli (STEC) is a foodborne pathogen associated with outbreaks worldwide that can be identified in the feces and in the meat of food-producing animals. Our study aimed to evaluate the incidence of E. coli O157:H7 in the feces of diarrheic camels (Camelus dromedarius) in Tunisia. From January 2018 to April 2019, 120 unduplicated fecal samples were obtained from diarrheic camels located in southern Tunisia. Non-sorbitol-fermenting colonies were confirmed as E. coli O157 via latex agglutination test and were screened for the presence of rfbEO157, fliCH7, stx1, stx2, eaeA, and ehxA genes by PCR. All isolates were examined for their susceptibility to 21 antibiotics. Of the 70 E. coli isolates that were recovered from 120 diarrheic camels, 4 (5.7%) were identified as STEC O157:H7. All isolates harbored ehxA and eae genes. Shiga toxin genes stx2 and stx1 were present in 50% and 25% of isolates, respectively. All E. coli O157:H7 isolates were sensitive to amoxicillin/clavulanic acid, cefotaxime, cefepime, aztreonam, colistin, and sulfamethoxazole-trimethoprim. All isolates belonged to the phylogroup E. This is the first report of E. coli O157:H7 isolates from diarrheic camels in Tunisia with a prevalence of 4 isolates (3.3%) amongst 120 fecal samples. This study supports the necessity for a platform purposed for regular screening and surveillance programs in food-producing animals and meat products, to perform early and rapid identification of food-borne pathogens. Prevalence of Escherichia coli O157:H7 isolated from fecal samples of diarrheic camels in Tunisia (HUS), and/or hospitalization (Al-Ajmi et  al. 2019, Falup-Pecurariu et al. 2019). These bacteria represent a significant public health concern and have the efficiency to produce Shiga toxin type 1 (stx1) and Shiga toxin type 2 (stx2), which are very potent toxins and are the main virulence determinants of this pathogen. Importantly, stx2-producing strains cause more severe infections than stx1-producing strains (Ogura et  al. 2015). In fact, purified stx2 is 1,000 times more toxic to human renal endothelial cells than stx1. The other main virulence factors produced by these serotypes are enterohaemolysin (ehxA) and intimin (eae) (Sperandio and Nguyen 2012). STEC O157 is the most clinically important serogroup but some serogroups non-O157 are also clinically significant foodborne pathogens, including STEC  O26, O45, O103, O111, O121, and O145 (Al-Ajmi et al. 2019, Hegde et al. 2012). Introduction Escherichia coli is a Gram-negative, rod-shaped bacterium, that normally colonizes the intestine of human and most animals and is considered an opportunistic pathogen (Tayh et  al. 2016). Some strains of E.  coli are capable to cause severe diseases in the human gut. They are recognized as enterohemorrhagic E.  coli (EHEC) belonging to intestinal pathogenic (diarrheagenic) strains that cause gastroenteritis. These strains cause illness in humans by producing an effective toxin known as Shiga toxin (Meng et al. 2012). Shiga-toxin-producing E.  coli (STEC) still known as VTEC (verotoxin-producing Escherichia coli) brings together several serotypes like EHEC pathogenic to humans, as food-borne bacteria linked with outbreaks worldwide. They can be associated with severe bloody diarrhea, hemolytic uremic syndrome 316 Veterinaria Italiana 2022, 58 (3), 315-323. doi: 10.12834/VetIt.2555.16997.2 E. coli O157:H7 in diarrheic camels Tayh et al. Bacterial isolation and identification Fecal samples were enriched in buffered peptone water overnight at 37  °C, and then 10 µl were plated on MacConkey agar for 18-24 hrs at 37  °C. The E.  coli isolates were identified by conventional biochemical tests. E.  coli isolates were stored at -  20  °C in brain heart infusion broth supplemented with 20% glycerol. Identification of E. coli O157:H7 One isolated strain was streaked on a plate of Sorbitol MacConkey with Cefixime Tellurite (SMAC-CT) agar. The bacterial plate was incubated overnight at 37 °C. After incubation, all colonies which are not able to ferment sorbitol (colorless/white colonies) were selected as probably E. coli O157. All E. coli colonies of non-sorbitol-fermenting on SMAC-CT medium were tested for the presence of the O157 antigen by agglutination test (DrySpot™ E.  coli O157 Latex Agglutination Test, Thermo Fischer Scientific). DNA extraction of bacteria genome The extraction of the E.  coli genome was carried out by boiling method. Bacterial colonies were suspended in 1 ml of sterile distilled water. After centrifugation at 13,000 rpm for 5 min, the supernatant was removed, replaced by 100 ml of sterile distilled water, heated at 95 °C for 10 minutes, and then kept at - 20 °C to be used for amplification by polymerase chain reaction (PCR). The PCR was carried out in this study including positive (E.  coli O157:H7 isolated from cattle in our laboratory) and negative (DNA-free water) controls. Detection of O157 and fliCh7 genes by PCR PCR amplification was used to detect the rfbE and fliCh7 genes encoding for O157 and H7 antigens of E.  coli O157:H7 strains, respectively, using oligonucleotide primers listed in Table I. DNA amplification reactions were carried out using a DNA thermal cycler (2720 thermal cycler, Applied Biosystem by Life Technologies, Singapore) with the following program: one cycle of denaturation at 94 °C for 5 min; 35 cycles of denaturation at 94 °C for 45  sec, annealing for 45 sec at 52  °C and 60  °C for O157 for detecting rfbE and extension at 72  °C for 45  sec; and a final extension at 72  °C for 10 min for detecting fliCh7. The PCR amplification products were separated by gel electrophoresis with 1.5% agarose and visualized under ultraviolet (UV) light using ethidium bromide staining. According to the Centers for Disease Control and Prevention (CDC), the dissemination of the STEC to humans might take place through contaminated food such as beef meat, fruits and vegetables, contaminated water, or via contact with contaminated animals or persons (CDC 2019). It is found in the intestines of healthy cattle, goats, and sheep which are considered natural reservoirs and feed may be contaminated with livestock manure (Ferens and Hovde 2011). According to El-Gallas and colleagues (El-Gallas et  al. 2006), in Tunisia, 3.4% of isolates from human stool samples were E. coli O157:H7. However, the incidence reports of E.  coli O157:H7 in camels (Camelus dromedaries) are rare. Studies from the United Arab Emirates (UAE) (Moore et  al. 2002, Al-Ajmi et al. 2019), Iran (Rahimi 2012), Kenya (Baschera et al. 2019) and Iraq (Mohammed Hamzah et  al. 2013) failed to isolate these bacteria among camel fecal samples. Furthermore, the same failure to detect this pathogen was reported in African countries (Egypt, Somalia, Djibouti, Kenya, and Sudan) (El-Sayed et  al. 2008). However, in some countries, the prevalence of STEC O157:H7 isolates among healthy camel feces, has been reported, as in UAE was 4.3 (6/140) (Al-Ajmi et  al. 2020), Saudi Arabia (2.4% and 11.5%) (Al Humam 2016, Bosilevac et  al. 2015) and Iran (2%) (Sami and Adeli 2013), as well as from diarrheic camel feces in Egypt (17.9%) (El-Hewairy et al. 2009). No data are available on the prevalence of E.  coli O157:H7 among camels in Tunisia. We selected diarrheic camels to determine whether E.  coli O157 is an agent of diarrhea as this has not been studied in Tunisia and can potentially pose a risk to people in contact with diarrheic animals. Therefore, this study aimed to determine the prevalence, virulence factors, and antimicrobial resistance profiles of STEC O157:H7 among fecal samples of diarrheic camels. To the best of our knowledge, this is the first report of STEC O157:H7 in camels in Tunisia. Materials and methods Samples collection One hundred and twenty fecal samples were collected from diarrheic camels of southern Tunisian regions (Douz, Tozeur, Gabes and Ben Guerden) from January 2018 to April 2019. Only one specimen per animal was included. These samples were transported appropriately to the Microbiology Laboratory at the National School of Veterinary Medicine of Sidi Thabet for bacterial isolation and further investigations. 317Veterinaria Italiana 2022, 58 (3), 315-323. doi: 10.12834/VetIt.2555.16997.2 Tayh et al. E. coli O157:H7 in diarrheic camels Antimicrobial sensitivity test The antimicrobial sensitivity was studied by the disk-diffusion method on Mueller-Hinton agar plates according to the guidelines and clinical breakpoints of the Antibiogram Committee of the French Society (CASFM-Vétérinaire 2018) using twenty-one antibiotics discs belonging to eight classes comprising μg/disk (Bio Rad, France): twelve β-lactams [(amoxicillin  (25), amoxicillin/ clavulanic acid (20/10), ticarcillin/clavulanic acid (75/10), cefotaxime  (30), ceftazidime (30), cefepime (30), cefoxitin (30), aztreonam (30) ertapenem (10), piperacillin (30), cefalotine (30), cefuroxime (30)], and nine non-β-lactams antibiotics [chloramphenicol (30), gentamicin (15), nalidixic acid (30), enrofloxacin Detection of virulence factors The presence of stx1, stx2, and ehxA virulence genes with uidA gene was screened in E.  coli O157:H7 isolates by the multiplex PCR method and eae by the simplex method using primers listed in Table  I. The PCR thermocycler apparatus was used to perform PCR reactions. The reaction consists of 25  cycles of DNA denaturation for 1 min at 95  °C, primer annealing for 1 min at 56 °C, and first extension for 1 min at 72  °C. The last step is the final extension for 5 min at 72 °C. The separation of PCR bands was performed by electrophoresis using 2% agarose gel, the PCR products were watched under UV light by ethidium bromide stain. Table I. Primers for PCR amplification of E. coli O157:H7. PCR reaction Gene Primer sequence (5’-3’) Size of PCR product (bp) Annealing temperature (°C) Reference Phylogenetic genes Quadruplex chuA chuA.1b: ATGGTACCGGACGAACCAAC 288 60 Clermont et al. 2013 chuA.2: TGCCGCCAGTACCAAAGACA yjaA yjaA.1b: CAAACGTGAAGTGTCAGGAG 211 60 Clermont et al. 2013 yjaA.2b: AATGCGTTCCTCAACCTGTG TspE4C2 TspE4C2.1b: CACTATTCGTAAGGTCATCC 152 60 Clermont et al. 2013 TspE4C2.2b: AGTTTATCGCTGCGGGTCGC arpA AceK.f: AACGCTATTCGCCAGCTTGC 400 60 Clermont et al. 2013 AceK.r: TCTCCCCATACCGTACGCTA Group E arpA ArpAgpE.f: GATTCCATCTTGTCAAAATATGCC 301 57 Clermont et al. 2013 ArpAgpE.r: GAAAAGAAAAAGAATTCCCAAGAG Group C trpA trpAgpC.1: AGTTTTATGCCCAGTGCGAG 219 59 Clermont et al. 2013 trpAgpC.2: TCTGCGCCGGTCACGCCC Internal control trpA trpBA.f: CGGCGATAAAGACATCTTCAC 489 57 Clermont et al. 2013 trpBA.r: GCAACGCGGCCTGGCGGAAG Virulence factors Shiga toxin type 1 stx1 F: CAGTTAATGTGGTGGCGAAGG 348 bp 56 Sjöling et al. 2015 R: CACCAGACAATGTAACCGCTG Shiga toxin type 2 stx2 F: ATCCTATTCCCGGGAGTTTACG 584 bp 56 Sjöling et al. 2015 R: GCGTCATCGTATACACAGGAGC Enterohaemolysin ehxA F: GCATCATCAAGCGTACGTTCC 534 bp 56 Grispoldi et al. 2017 R: AATGAGCCAAGCTGGTTAAGCT Enteropathogenical attachment and effacement eae F: TGCGGCACAACAGGCGGCGA 629 pb 58 Ranjbar et al. 2017 R: CGGTCGCCGCACCAGGATTC Others Part of O-antigen 157 O157 F: CGGACATCCATGTGATATGG 259 bp 52 Paton and Paton 1998 R: TTGCCTATGTACAGCTAATCC Encoding H7 flagellar antigens fliCH7 F: GCGCTGTCGAGTTCTATCGAGC 625 bp 60 Al-Ajmi et al. 2020 R: CAACGGTGACTTTATCGCCATTCC Beta-glucuronidase uidA F: ATCACCGTGGTGACGCATGTCGC 486 bp 56 Heininger et al. 1999 R: CACCACGATGCCATGTTCATCTGC 318 Veterinaria Italiana 2022, 58 (3), 315-323. doi: 10.12834/VetIt.2555.16997.2 E. coli O157:H7 in diarrheic camels Tayh et al. biochemical tests as E. coli. Testing of E. coli isolates on SMAC-CT and by agglutination test, revealed four isolates (5.71%) considered as presumptive E.  coli O157:H7 isolates. Thus, 4 E. coli O157 were identified out of 120 camels (3.33%). The frequency of isolation in this study was different between males and females with statistical significance (P = 0.045) (Table  II). The difference between the rate of isolation according to the age of the animals was not statistically significant (P = 0.657) (Table  II). Furthermore, there was no statistically significant difference between the isolation frequency of O157 according to animal’s origin (P = 0.231) (Table II). The confirmation of E.  coli O157 by specific genes showed that uidA, O157 and fliCh7 were present in all isolates. The virulence genes eae and ehxA were present in the four isolates; stx1 and stx2 were present in one and two isolates, respectively (Table III). All isolates belong to the phylogroup E. All isolates were susceptible to cefoxitin, enrofloxacin, florfenicol, colistin, and sulfamethoxazole-trimethoprim. Furthermore, 75% of the isolates were susceptible to piperacillin, cephalotin, cefotaxime, nalidixic acid and tetracycline. However, 50% of the isolates were (5), tetracycline (30), sulfamethoxazole/trimethoprim (1.25/23.75), streptomycin (10), florfenicol (30) and colispot]. Detection of phylogenetic groups The phylogenetic groups (A, B1, B2 and D) were identified in O157 isolates by quadruplex PCR using chuA, yjaA genes and the TspE4‑C2 fragment. The phylogroups C and E were detected by PCR using arpA and trpA genes. The primers and the method were previously described by Clermont and colleagues (Clermont et al. 2013). Data analysis The data of the camels; age, gender and origin were analyzed with the frequency of E.  coli O157 by the Statistical Package for the Social Sciences (SPSS) version 26 software (IBM Corporation, Somers, NY). The comparison of data was performed using Pearson's Chi-square with a P  <  0.05 value of statistical significance. Results A total of 70 non-duplicate isolates isolated from 120 diarrheic camel samples from four major cities in southern Tunisia were identified by conventional Table II. Prevalence of E. coli O157:H7 in Tunisian camels according to gender, age groups and region. Factors No. (%) samples No. E. coli O157 Rate Gender Male 61 (50.8) 4 6.56% Female 59 (49.2) 0 0 Total 120 (100) 4 3.33% P-value = 0.045 Age (years) < 1 70 (58.3) 2 1.67% 1-5 28 (23.3) 2 1.67% 6-10 6 (5.0) 0 0 ≥ 10 16 (13.4) 0 0 Total 120 (100) 4 3.34% P = 0.657 Origin (city) Gabes 51 (42.5) 4 7.84% Douz 34 (28.3) 0 0 Tozeur 19 (15.8) 0 0 Ben Guerden 16 (13.4) 0 0 Total 120 (100) 4 3.34% P = 0.231 Table III. Characteristics of the 4 E. coli O157:H7 isolated in Tunisian camels. Bacterial code Specific genes Virulence genes O157 fliCH7 uidA stx-1 stx-2 eae ehxA D5 + + + - + + + D10 + + + - + + + D25 + + + - - + + D42 + + + + - + + Figure 1. PCR for detection of virulence genes. Lane M = Ladder 100 bp; lane 1, 2, 3, 4 = Positive samples; Agarose gel concentration: 2%. 600 bp M 1 2 3 4 500 bp 400 bp stx2; 584 bp ehxA; 534 bp iudA; 486 bp stx1; 348 bp 319Veterinaria Italiana 2022, 58 (3), 315-323. doi: 10.12834/VetIt.2555.16997.2 Tayh et al. E. coli O157:H7 in diarrheic camels Interestingly, in Iran, Salehi and colleagues (Salehi et  al. 2011) detected the serotype O157:H7 in two dromedaries with haemorrhagic diarrhea. The occurrence of E.  coli O157 was investigated among 70 E. coli isolated from feces of 120 diarrheic animals in the main southern Tunisian cities. This is the first report of E. coli O157 isolation from camels in Tunisia and also the first report in animals in Tunisia. The incidence of E.  coli O157:H7 in camel’s fecal samples was 5.71%, with four isolates amongst 70 strains. The occurrence studies of this bacterium in camels are limited; a study conducted in the UAE was unsuccessful in detecting E.  coli O157:H7 in fresh feces of racing camels (Moore et  al. 2002). The same failure to identify E. coli O157 was among 400 camel fecal samples obtained from African countries (Egypt, Somalia, Djibouti, Kenya, and Sudan) (El-Sayed et al. 2008) and also from 40 camel samples in Iran (Rahimi 2012). More recently, in a study in Kenya, none of 163 fecal samples of grazing dromedaries contained STEC O157:H7 (Baschera et al. 2019). An Iraqi study was performed on a zoo animals in Baghdad; they detected 24 E.  coli O157:H7 out of 174  fecal samples from bear, deer, pony, horses, zebra, ostrich, hyena, llama, goat, and jaguar. However, no E.  coli O157:H7 was identified among camels (Mohammed Hamzah et al. 2013). Our methods used for the detection and isolation of O157:H7 are considered appropriate but non-selective, so the results might be an underestimate of the actual prevalence of O157:H7 among our samples. The ISO 16654:2001 method is the standard for detection of E.  coli O157 but it requires separation and concentration using immunomagnetic beads coated (Tozzoli et al. 2019). Our findings were compatible with the results of a study in the UAE (Al-Ajmi et  al. 2020) in which the prevalence of E.  coli O157 was 4.3%. They used an isolation method like the ISO 16654 method. The demonstration of E.  coli O157 in camels was also reported in Saudi Arabia among 200 camel fecal samples in six isolates (11.5%) from 52 E. coli isolates (Al Humam 2016). In Iran, they reported 2% of 150 healthy camel feces were E.  coli O157 (Sami and Adeli 2013). Another report from Iran demonstrated that one (1.3%) strain was E.  coli O157 out of 75 camel meat samples using the protocol of selection of sorbitol negative colonies and PCR amplification of the O-antigen encoding region of the O157 gene and flagellar H7 gene (fliC) (Hajian et al. 2011). A high prevalence (17.9%) was recorded among 60 Egyptian camel calves; half of them suffered from diarrhea (El-Hewairy et  al. 2009). In Iraq, Al-Gburi (Al-Gburi 2016) detected E. coli O157:H7 with 19% in healthy camels by the same protocol as our study. resistant to cefuroxime, ticarcillin-clavulanic acid and streptomycin (Table IV). Discussion Escherichia coli O157:H7 infections in humans originally came from food of animal or vegetable origin via food contamination with feces containing this pathogen. This particularly concerns ruminants such as cattle, sheep and goats which are considered the major natural reservoirs for this serovar and mainly implicated in human infections (Jo et  al. 2004). Due to the consumption of camel meat in Tunisia, specifically in the southern part, and given the absence of studies on this subject, it was an established decision to study the prevalence of E.  coli O157:H7 in camels. The choice of diarrheic dromedaries is related to the lack of data on this pathology in Tunisia and on the zoonotic risk of serotype O157:H7, in order to raise awareness of this risk among individuals responsible for the maintenance and care of dromedaries (breeders, veterinarians, butchers, slaughterhouse employees etc). To our knowledge, in Tunisia, only one study has focused on E.  coli isolated from diarrhoeic camels but has not targeted STEC (Bessalah et  al. 2016). Table IV. Antimicrobial susceptibility of E. coli O157:H7 Tunisian camel isolates. Antimicrobial Susceptible Intermediate Resistant Ampicllin 25% 75% 0 Piperacillin 75% 25% 0 Cephalotin 75% 25% 0 Cefuroxime 50% 0 50% Cefoxitin 100% 0 0 Cefotaxime 75% 25% 0 Ceftazidime 25% 50% 25% Cefepime 50% 50% 0 Aztreonam 25% 75% 0 Ertapenem 50% 50% 0 Amoxicillin/ clavulanic acid 25% 75% 0 Ticarcillin/ clavulanic acid 50% 0 50% Gentamicin 50% 50% 0 Streptomycin 25% 25% 50% Nalidixic acid 75% 25% 0 Enrofloxacin 100% 0 0 Chloramphenicol 50% 50% 0 Florfenicol 100% 0 0 Colistin 100% 0 0 Sulfamethoxazole/ trimethoprim 100% 0% 0% Tetracycline 75% 0 25% 320 Veterinaria Italiana 2022, 58 (3), 315-323. doi: 10.12834/VetIt.2555.16997.2 E. coli O157:H7 in diarrheic camels Tayh et al. important epidemiological marker for the rapid detection of STEC strains (Schwidder et  al. 2019). Another important virulence factor is intimin protein (eae) which plays an important role in the attaching of host intestinal mucosa within the colonization and causes severe infections (Sperandio and Nguyen 2012). In the present study, the STEC O157:H7 was confirmed by the identification of specific genes rfb‑O157 and fliCH‑7 using PCR as well as assessment of Shiga toxins genes (stx1, stx2) and enterohaemolysin (ehxA). Our finding revealed that eae and ehxA genes were present in all isolates; stx2 and stx1 were present in two and one isolates, respectively. In a recent study, out of the 12 E.  coli O157 isolates from fecal samples of healthy camels, stx2 was found in all isolates, eae in 11 (91.7%), hlyA in 11 samples (75%) and stx1 was absent in all isolates (Al-Ajmi et al. 2020). In a study conducted in Saudi Arabia, they found one isolate carrying both Shiga toxins genes (stx1 and stx2), and like to our results, all 6  isolates of E. coli O157:H7 possessed the eae gene (Bosilevac et  al. 2015). Rahimi and colleagues (Rahimi et  al. 2012) confirmed 2.0% of 50 camel samples as E. coli O157:H7 but they failed to identify any virulence gene (stx1, stx2, hlyA and eae). Antimicrobials have saved millions of human lives and their use has made a significant contribution to the improvement of the health of animals and humans. Antimicrobial resistance development is known as a global health threat. Antimicrobial agents are used in animals intended for human consumption to make animals healthier, reducing diseases and mortality (Oliver et  al. 2011). The increased quantity of antibiotic residues in food-producing animals participates in the increase of antimicrobial resistance problems. Misuse and overuse of antimicrobials constitute a serious health enigma in many countries, such as using them for growth progression and infection prevention; furthermore, in several regions, the antibiotic quantities used in animals are four times larger than the quantities used in humans (Hudson et al. 2017). The results of this study revealed that all isolates were susceptible to most tested antibiotics except cefuroxime, ticarcillin/clavulanic acid and streptomycin with resistance detected in 50% of the isolates and some isolates were intermediate to antibiotics which might progress to resistance. The results of an Iraqi study showed resistance of E.  coli O157:H7 isolates to all the antibiotics tested in the study (doxycycline, cephalexin, erythromycin, clarithromycin, ceftriaxone, ampicillin, cloxacillin, rifampin and carbenicillin) except to trimethoprim (Al-Gburi 2016). A report in Iran revealed the resistance of E. coli O157:H7 to most tested antibiotics (ampicillin, erythromycin, gentamycin, nalidixic acid, Many reports confirmed that the prevalence of E.  coli O157 isolated from camel feces was lower than that of those reported in cattle (Bosilevac et al. 2015, Sami and Adeli 2013) suggesting that camels cannot be considered the major natural reservoir for this pathogen in comparison with cattle. Considering the low number of E.  coli O157:H7 in Tunisia and few positive reports on camels in other studies, it can be concluded that the camel is not a significant source of STEC O157:H7 infection; practically, the studied camels share environments or pastures or pens with cattle, goats, and sheep. These ruminants can therefore be the source of STEC contamination of dromedaries. Surveillance and screening programs on camel products remain necessary for the prevention of outbreaks, particularly for food-borne infections, especially if those camels share spaces with ruminants. The frequency of E.  coli O157 was greater in males than in females with statistical significance, while there was no significant difference according to age or region. Our results correlate with another study in Saudi Arabia which reported that the prevalence of E.  coli O157:H7 was significantly greater in young males than among females (Al-Ajmi et  al. 2020). On the contrary, Baschera and colleagues (Baschera et al. 2019) reported from a Kenyan study a prevalence of 14.5% (17/117) of female camels and 6.7% (3/45) of male camels. Another study has found no statistically significant difference according to gender and age among camel samples (Al-Ajmi et al. 2020, Al-Gburi 2016). In the present study, no statistically significant relationship was found between E.  coli O157:H7 prevalence and the camel’s origin. All E. coli O157:H7 were isolated from animals sampled in Gabes and these results highlight the high risk of the spread of this pathogen among animals from that region. Moreover, this finding strongly encourages us to carry out investigations on a larger scale of camels from Gabes and to try to find possible particularities of the camel farms of this region, and further investigations are necessary to study E. coli O157:H7 portage in cattle, sheep and goats from Gabes. Serotype O157:H7 might carry stx1 and stx2 genes encoding Shiga toxins. Strains that possess stx2 are associated with human illness more frequently than strains carrying stx1 (Bosilevac et  al. 2015). Both factors play important roles in the progress of hemolytic uremic syndrome. Epidemiological evidence of STEC O157 has revealed that strains carrying stx2 are associated with more severe human infections than strains carrying stx1 (Manning et  al. 2008). Furthermore, STEC possesses hemolysin encoded by plasmid-carried enterohaemolysin gene (ehxA) which is associated with diarrheal disease and HUS (Fu et  al. 2018). This factor is an 321Veterinaria Italiana 2022, 58 (3), 315-323. doi: 10.12834/VetIt.2555.16997.2 Tayh et al. E. coli O157:H7 in diarrheic camels the region suggest that camels might be considered as the natural reservoir and play an important role in the infection of humans with this pathogen. Our results supply some baseline data concerning the occurrence of E.  coli O157:H7 in camels and their antimicrobial resistance that could be used in future studies. This work leads to highly recommended surveillance and screening programs on camel products to prevent outbreaks. Furthermore, it supports the necessity for a platform purposed to regular screening and surveillance programs in food-producing animals and meat products, to allow early and rapid identification of food-borne pathogens. More studies should be performed to characterize the prevalence of STEC O157 and other non-O157 pathogens in animals and meat food products with a large number of isolates. Funding This work was supported by the research project PEER 7-349 funded by the USAID “Monitoring of antimicrobial resistance of bacteria for a better health of animals in Tunisia”. doxycycline, streptomycin, kanamycin, tetracycline, chloramphenicol and amoxicillin) and susceptible to cefuroxime (Tanzifi et al. 2015). In a UAE study, E. coli O157 isolated from the fecal samples of camels were 100% susceptible to cefotaxime, chloramphenicol, ciprofloxacin, norfloxacin, and polymyxin B (Al-Ajmi et al. 2020). The results of our study showed that the E.  coli O157 isolates belonged to the phylogroup  E, which is in agreement with other studies (Coura et al. 2015, Tenaillon et al. 2010). Antibiotic resistance in these isolates can be used as an indicator of antibiotic use in the animals or their environment. These resistances can be useful for studying the isolates, but O157:H7 infections in humans are not treated with antibiotics as this can induce expression of Shiga toxins and worsen the disease. Conclusions This is the first report of E. coli O157:H7 isolated from diarrheic camels in Tunisia. The presence of this pathogen in our study and some positive reports in 322 Veterinaria Italiana 2022, 58 (3), 315-323. doi: 10.12834/VetIt.2555.16997.2 E. coli O157:H7 in diarrheic camels Tayh et al. Al-Ajmi D., Rahman S. & Banu S. 2019. Complete genome sequence of Escherichia coli O157: H7 strain Al Ain, isolated from camel feces in the United Arab Emirates. Microbiol Res Announc, 8, e01171-19. 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