The emergence of carbapenem-resistance and New Delhi metallo-ß-lactamase-1 (blaNDM-1) among Salmonella spp. in Kerman, Iran.

Background and Objectives: Salmonella species (spp) are the most prevalent zoonotic pathogens that cause outbreaks of gastroenteritis worldwide. Therefore evaluation of the profile of antibiotic resistance, virulence factors, and plasmid replicon types in these bacteria is necessary to control and prevent the spread of potentially pathogenic and drug-resistant strains. Materials and Methods: This study was performed on 39 Salmonella spp. The antibacterial susceptibility of isolates to various antibiotic agents was determined using disk diffusion test. ß-lactamases (bla) including ESBLs, AmpC, MBLs, and virulence genes were detected by PCR methods. Plasmid incompatibility groups among the isolates were identified using PCR-based replicon typing (PBRT). Results: The most prevalent virulent gene was phoP/Q (84.6%). slyA, sopB, and stn were identified in 79.4% (n=31), 69.2% (n=27), and 2.5% (n=1) of the isolates, respectively. The antibiotic susceptibility testing showed that 30.7% of the isolates were ESBL-producing. blaTEM (41%; n=16) was the most frequent ß-lactamase gene among the isolates followed by blaNDM-1 (15.4%; n=6), blaDHA (7.7%; n=3), and blaCTX-M (1.5%; n=1). Six different plasmid replicon types, including IncP (n=9; 23%), IncFIC (n=3; 7.70%), IncY (n=3; 7.70%), IncI1-Iγ (n=2; 5.12%), IncFIIAs (n=1; 2.56%), and IncN (n=1; 2.56%) were observed among the isolates. Conclusion: Our study showed the emergence of carbapenem-resistant and blaNDM-1 among Salmonella spp. for the first time in Kerman, Iran. Since Salmonella spp. plays an important role in the transmission of resistance genes in livestock and humans in the food chains, so more stringent control policies are recommended to prevent the circulation of drug-resistant and potentially pathogenic strains from animals to humans.

Antibiotic resistance profiling and phylotyping of human-diarrheagenic Escherichia coli pathotypes detected from diarrheic and non-diarrheic calves in Iran.

BACKGROUND: Escherichia coli (E. coli) serves as a common indicator of gut microbiota and is utilized for monitoring antimicrobial resistance determinants in food-producing animals. This study aimed to investigate antimicrobial resistance patterns in virulence gene-positive E. coli isolates obtained from 340 healthy and diarrheic calves. METHODS AND RESULTS: A total of 340 fecal swab samples were obtained from diarrheic (n = 170) and healthy (n = 170) calves for 12 months from different farms in Kerman, Iran. The samples were phenotypically analyzed to detect E. coli isolates and antibiotic resistance. Also, antimicrobial resistance genes, diarrheagenic E. coli pathotypes, and phylogenetic background were screened by PCR. Fifteen percent (51/340) of E. coli isolates were positive for at least one of the examined virulence genes (VGs); the prevalence of VGs in E. coli isolates from healthy calves (36/170; 21.17%) was higher than that in diarrheic cases (15/170; 8.82%). Out of the 51 VG-positive isolates, six pathotypes including Shiga toxin-producing E. coli (STEC; 27.45%), enterotoxigenic E. coli (ETEC; 23.52%), enterohemorrhagic E. coli (EHEC; 19.6%), necrotoxigenic E. coli (NTEC; 19.6%), enteropathogenic E. coli (EPEC; 15.68%), enteroinvasive E. coli (EIEC; 1.96%) and three hybrid pathotypes including ETEC/STEC, ETEC/EHEC, and STEC/EIEC were detected among the strains. Antimicrobial resistance (AR) was observed in 98.03% of the VG-positive isolates, which was the same for both healthy and diarrheic calves. The maximum prevalence rate of AR was found against trimethoprim/sulfamethoxazole (49.01%; 3/51), while the minimum prevalence rate was against gentamycin (5.88%; 25/51). Among the VG-positives, phylotype A was found to be the most prevalent followed by B1 and D phylotypes. CONCLUSIONS: The prevalence of VG-positive E. coli isolates was higher in healthy calves compared to diarrheic cases. AR was widespread among VG-positive isolates. These findings suggest that calves may serve as potential reservoirs of antimicrobial-resistant hybrid pathotypes of E. coli.

High prevalence of ß-lactam and fluoroquinolone resistance in various phylotypes of Escherichia coli isolates from urinary tract infections in Jiroft city, Iran.

BACKGROUND: Urinary tract infection (UTI) is one of the most prevalent infectious diseases with worldwide health threatening. Antimicrobial resistant strains of Escherichia coli (E. coli) are a common cause of UTI which were identified as a treatment challenge. This study aimed to assay the prevalence of common ß-lactam resistance genes including blaTEM, blaSHV, blaCTX-M and blaCMY and phenotypic resistance to commonly used ß-lactam and fluoroquinolone antibiotics in UTIs. These factors were evaluated in various phylogenetic groups (phylotypes) of E. coli isolates. Real-time PCR was applied to detect ß-lactam resistance genes and conventional PCR was used to determine the phylotypes. Phenotypic resistance against ß-lactams (ceftazidime, cefotaxime, aztreonam and ceftriaxone) and fluoroquinolones (ciprofloxacin) were identified by the disc diffusion technique. The ability of extended spectrum ß-lactamases (ESBLs) production in E. coli isolates was detected using the combined disc diffusion method. RESULTS: The prevalence of resistance genes were 89.6% for blaTEM, 44.3% for blaCTX-M, 6.6% for blaSHV and 0.9% for blaCMY. The two high prevalent phylotypes were B2 (29.2%) and D (17.9%) followed by E (14.1%), F (9.4%), C (6.6%) and 10.3% of isolates were unknown in phylotyping. Disc diffusion results showed high prevalence of antibiotic resistance to cefotaxime (88.6%), aztreonam (83%), ceftireaxon (77.3%), ceftazidime (76.4%) and ciprofloxacin (55.6%). Totally, 52.8% of isolates were found as phenotypical ESBL-producers. CONCLUSIONS: This study's results confirmed an explosion of antibiotic resistance amongst E. coli isolates from UTI against ß-lactams and fluoroquinolones. Findings explain the necessity of deep changes in quantity and quality of drug resistance diagnosis and antibiotic therapy strategies. More studies are suggested to better and confident evaluations.

Molecular characterization and antimicrobial resistance of potentially human-pathogenic Escherichia coli strains isolated from riding horses.

BACKGROUND: Transmission of antimicrobial resistant and virulent Escherichia coli (E. coli) from animal to human has been considered as a public health concern. This study aimed to determine the phylogenetic background and prevalence of diarrheagenic E. coli and antimicrobial resistance in healthy riding-horses in Iran. In this research, the genes related to six main pathotypes of E. coli were screened. Also, genotypic and phenotypic antimicrobial resistance against commonly used antibiotics were studied, then phylo-grouping was performed on all the isolates. RESULTS: Out of 65 analyzed isolates, 29.23 % (n = 19) were determined as STEC and 6.15 % (n = 4) as potential EPEC. The most prevalent antimicrobial resistance phenotypes were against amoxicillin/clavulanic acid (46.2 %) and ceftriaxone (38.5 %). blaTEM was the most detected resistance gene (98.4 %) among the isolates and 26.15 % of the E. coli isolates were determined as multi-drug resistant (MDR). Three phylo-types including B1 (76.92 %), A (13.85 %) and D (3.08 %) were detected among the isolates. CONCLUSIONS: Due to the close interaction of horses and humans, these findings would place emphasis on the pathogenic and zoonotic potential of the equine strains and may help to design antimicrobial resistance stewardship programs to control the dissemination of virulent and multi-drug resistant E. coli strains in the community.

Public health aspects of Shiga toxin-producing Escherichia coli (STEC) strains in sheep and goats of Bakhtiari pastoral tribe, Iran.

Nomadic populations do not have permanent settlements as they move their livestock between grazing areas in different seasons; such movements may have great impact on dissemination of food-borne pathogens in various regions. The aim of this study was to characterize Shiga toxin-producing Escherichia coli (STEC) strains as a food-borne pathogen in sheep and goats of Bakhtiari pastoral tribe in Iran. In the present study, 72 fecal samples were obtained from 26 sheep and 46 goats. First, all recovered E. coli isolates were screened for stx gene. After detection of stx-positive isolates, the virulence genes including stx1, stx2, eae, ehly, saa, astA, subAB, terD, and the genetic markers of O Island 57 (Z2098 and Z2099) were investigated. Also fifteen important STEC O-serogroups were determined using PCR assays. Results showed that 27 animals (27/72; 37.5%) carried STEC strains including 16/26 (61.6%) sheep and 11/46 (23.9%) goats. All STECs were eae-negative but 81.4% (22/27) were positive for saa. The most prevalent virulence profile was stx1/stx2/ehly/saa/subAB (37%; 10/27). Most STECs (24/27) were positive for at least one of the selected OI-57 markers. The O91 (n = 6), O5 (n = 3), O113 (n = 1), O128 (n = 1), and O104 (n = 1) were the detected O-serogroups in this study. It is concluded that such moving animal populations could have public health concerns which have to be addressed in the future.

Domestic and game pigeons as reservoirs for Escherichia coli harbouring antimicrobial resistance genes.

OBJECTIVES: In Iran, pigeons are kept and bred on the roofs of houses, which indicates the potential significance of pigeons in the dispersal of antimicrobial resistant Escherichia coli strains. Here we characterized antimicrobial resistance genotypes in relation to phenotypic presentations and phylogenetic backgrounds of the E. coli isolates from household pigeons in Kerman in southeast Iran. METHODS: Totally, 152 faecal E. coli isolates from domestic and household pigeons were screened for 13 antimicrobial resistance genes, blaTEM, blaSHV, blaCTX-M, sulI, sulII, dhfrI, dhfrV, aadA, aac(3)-I, tetA, tetB, floR and qnrA, by conventional polymerase chain reaction (PCR) technique. Clermont phylogenetic background of E. coli strains was studied and antibiotic resistance of all strains was assessed for seven antibiotics. RESULTS: The antimicrobial resistance genes blaTEM, tetA, tetB and aadA were detected in 52.6%, 6.5%, 6.5% and 5.9% of the isolates, respectively. PCR phylotyping revealed that a significant number of isolates within A0 (54%), A1 (70%), B1 (57.6%), B22 (75%) and D1 (87.5%) phylogroups were positive for the studied resistance genes. One phenotypic resistance pattern (trimethoprim-sulfamethoxazole) was associated with the presence of the corresponding gene sul2. CONCLUSIONS: An alarming rate of phenotypic resistance was observed in this study. Many isolates were positive for the screened resistance genes. According to the phylogenetic background, most resistant isolates belonged to the commensal phylotypes, representing significant role of commensal strains as a source of resistance genes. These findings highlight the role of the pigeon as disseminator of resistant E. coli strains.

Detection of zoonotic diarrheagenic pathotypes of Escherichia coli in healthy household dogs.

BACKGROUND AND OBJECTIVES: Intestinal pathotypes of Escherichia coli belong to the companion animals may poses potential risk to public health following zoonotic transmission. Therefore, this study was proposed to determine the virulence genes associated to diarrheagenic E. coli strains isolated from healthy pet dogs and their owners in the southeast of Iran, Kerman province. MATERIALS AND METHODS: Totally 168 E. coli isolates were collected from 49 healthy household dogs and their owners. Seventy isolates were obtained from non-pet owners as control group. Presence or absence of the virulence genes including eae, stx1, stx2, st1, lt1, ipaH, cnf1 and cnf2 were screened by conventional polymerase chain reaction (PCR) and dissemination pattern of the genes were studied among the various hosts. RESULTS: PCR examinations showed that the most frequent virulence gene was ipaH (6.1%) in dogs followed by eae in dog owners (6.1%) and in controls (8.6%). The most frequent pathotypes in dogs, their owners and controls were EIEC (6.1%), EHEC (4.08%) and EPEC (8.5%), respectively. In one of studied houses, both of dog and its owner harbored E. coli strains with same virulence profile (stx1/eae) and pathotype (EHEC). CONCLUSION: These results collectively indicate that healthy household dogs probably are the mild reservoir of potential virulent E. coli strains with possible active transmission to their contact owner. However, even non-pet owners seemed to be a notable source of intestinal pathotypes, especially EPEC, for their environment. Transmission of E. coli pathotypes may occurs by direct contact with the reservoirs or ingestion of contaminated food. These pathotypes are potentially virulent and creates public health hazards. Further studies are needed for better understanding of dissemination mechanisms of E. coli pathotypes among humans and their pets.

An overview of diarrheagenic Escherichia coli in Iran: A systematic review and meta-analysis.

BACKGROUND: Diarrheagenic Escherichia coli (DEC) is a common enteric pathogen that causes a wide spectrum of gastrointestinal infections, particularly in developing countries. This is a systematic review and meta-analysis to determine the prevalence of DEC in various geographical regions in Iran. MATERIALS AND METHODS: English (PubMed, Web of Science, Scopus, Embase, Cochrane Library, and Google Scholar) and Persian (IranMedex, SID, Magiran, and Iran Doc) databases were comprehensively searched from January 1990 to April 2017. Study selection and data extraction were performed by two independent reviewers. After assessing heterogeneity among studies, a random effects model was applied to estimate pooled prevalence. Data analyses were done with the Stata software (version 12.0). This meta-analysis was registered with PROSPERO, number CRD42017070411. RESULTS: A total of 73 studies with 18068 isolates were eligible for inclusion within the meta-analysis. The results of random effects model showed that the most prevalent DEC pathotypes were enterotoxigenic E. coli (ETEC) (16%; 95% confidence interval [CI]: 11%-23%), enteroaggregative E. coli (11%; 95% CI: 8%-15%), atypical enteropathogenic E. coli (EPEC) (11%; 95% CI: 8%-14%), Shiga toxin-producing E. coli (9%; 95% CI: 6%-13%), diffuse adherent E. coli (6%; 95% CI: 6%-12%), enteroinvasive E. coli (4%; 95% CI: 2%-6%), and typical EPEC (3%; 95% CI: 1%-5%). CONCLUSION: This study showed that DEC infections in the Iranian population have low frequency. Our data suggest that the ETEC pathotype can be regarded as one of the most important etiological agents of diarrhea in this country. However, the prevalence of DEC pathotypes is diverse in different regions of Iran.

Molecular detection of Shiga toxin-producing and antibiotic-resistant Escherichia coli isolates from buffaloes in southwest of Iran.

Three hundred fifteen bacteriological samples were obtained from feces and both external and visceral cavity surfaces of carcasses of 105 healthy buffalo slaughtered in southwest of Iran. Confirmed Escherichia coli isolates were examined for antimicrobial resistance phenotypically and were screened for stx1, stx2, and eae genes and their subtypes and assessment of antimicrobial resistance genes by regular PCR and RFLP techniques. One hundred forty-five E. coli were isolated from feces (96 isolates) and external (37) and internal (12) surfaces of carcasses. Results showed that the prevalence of STEC, EPEC, and EHEC pathotypes was 2.8%, 0.7%, and 0.7% respectively. Among 6 (4.13%) positive isolates for examined genes, 4 (2.8%) isolates were positive for stx1, 3 (2.1%) for stx2, and 2 (1.4%) for eae gene. The detected genes were classified into stx1a (4 isolates), stx2a, stx2b, stx2c, eae-ß, and unknown subtypes. The most prevalent antibiotic resistance gene was sulII (11.03%). The tetB, qnrB, floR, blaTEM, blaSHV, and aadA genes were found to a lesser extent, and all isolates were negative for blaCTX-15, blaOXA, aac(3)-I, tetA, cat1, qnrA, sulI, dhfrI, and dhfrV genes. Twelve combination patterns of antibiotic-resistant genes were observed. Maximum phenotypically resistance rate was against doxycycline (91.83%), and the minimum was against ceftazidime and florfenicol (2.75%). E. coli isolates from feces and carcasses of slaughtered buffalo can be considered a mild reservoir for stx and eae genes. However, healthy buffaloes could be considered a potential reservoir of multiple antibiotic resistance genes in E. coli isolates.

Molecular investigation of the colicinogenic Escherichia coli strains that are capable of inhibiting E. coli O157:H7 in vitro.

BACKGROUND: Escherichia coli O157:H7 is a highly virulent human pathogen with severe consequences following infection, which claims many lives worldwide. A suggested method for controlling this bacterium is the competitive elimination through using probiotic bacteria that prevent its colonization. Some nonpathogenic E. coli strains that produce antibacterial colicins are among these probiotic bacteria. We aimed to isolate and characterize the colicinogenic E. coli strains from diarrheic and healthy sheep that inhibit E. coli O157:H7, which could be used as possible probiotic sources. A total of 292 E. coli isolates (146 from each diarrheic and healthy sheep) were screened for the presence of colicin and virulence genes. The phylogenetic group/subgroup determination was performed by PCR. In vitro evaluation of inhibitory effect of colicinogenic isolates on E. coli O157:H7 was done phenotypically. RESULTS: The frequency of diarrhea associated colicinogenic E. coli isolates was significantly higher than those isolated from healthy sheep. An association between ETEC and the genes coding for colicin-V & colicin-Iab in diarrheic E. coli isolates was observed. Moreover, there was an association between ipaH and Colicin-V encoding genes. Furthermore, E. coli isolates showing bacteriocinogeny while possessing no virulence genes had a frequency of 97.67 and 11.94% in healthy and diarrheic isolates, respectively. Of these strains, five isolates (3.42%) from diarrheic and twenty-five isolates (17.12%) from healthy sheep inhibited O157:H7 strain. Additionally, colicin E1 and colicin Iab genes were more prevalent in B1 phylogroup. CONCLUSIONS: These results signified that healthy sheep could be considered as a potential source for anti-O175:H7 bacterial isolates.

ECOR phylotyping and determination of virulence genes in Escherichia coli isolates from pathological conditions of broiler chickens in poultry slaughter-houses of southeast of Iran.

Avian pathogenic Escherichia coli (APEC) are responsible for wide ranges of extra-intestinal diseases in poultry including colibacillosis, cellulitis, coligranuloma and yolk sac infection. Numbers of virulence are considered important in the pathogenicity of these diseases. The aims of the present study were phylogenetic typing and virulence genes detection in Escherichia coli isolates from colibacillosis and cellulitis of broiler chickens in poultry slaughterhouses of Shahrbabak region, Kerman, Iran. A total number of eighty three E. coli isolates were taken from broiler chickens with colibacillosis and thirty four isolates were taken from carcasses with cellulitis in the industrial slaughterhouses. Biochemically confirmed E. coli isolates were subjected to polymerase chain reaction assay to determine phylogenetic groups and presence of pap C, sfa/focDE, iucD, afaIB-C, hlyA, fimH and crl virulence genes. Colibacillosis isolates were belonged to A (54.21%), B1 (7.22%), B2 (6.03%) and D (32.53%) phylogroups. Whereas, the isolates from cellulitis cases were belonged to three main phylogroups; A (55.88%), B1 (5.88%) and D (38.24%). Statistical analysis showed a specific association between the presence of crl virulence gene and phylogroups of A and D in colibacillosis isolates. The results showed that the isolates from both diseases in broiler chickens could be assigned to various phylogenetic groups (mainly A(. Also, the virulence genes profile of cellulitis E. coli is completely different from that of colibacillosis in this region.

Serogroups, subtypes and virulence factors of shiga toxin-producing Escherichia coli isolated from human, calves and goats in Kerman, Iran.

AIM: The present study was conducted to detect the occurrence, serogroups, virulence genes and phylogenetic relationship of shiga toxin-producing Escherichia coli (STEC) in human, clave and goat in Kerman (southeast of Iran). BACKGROUND: STEC have emerged as the important foodborne zoonotic pathogens causing human gastrointestinal disease and confirming the risk to public health. METHODS: A total of 671 fecal samples were collected from diarrheic patients (n=395) and healthy calves (n=156) and goats (n=120) and screened for the presence of stx gene. Furthermore, the prevalence of stx1 and stx2 variants, serotypes (O157, O145, O103, O26, O111, O91, O128, and O45), phylogenetic groups and the presence of ehxA, eae, hylA, iha and saa virulence genes were studied. RESULTS: Prevalence of STEC in human diarrheic isolates was 1.3% (5 isolates), in claves was 26.3% (41 isolates) and in goats was 27.5% (33 isolates). stx1 gene was the most prevalent variant and detected in 75 isolates. Furthermore, stx1c was the most predominant stx subtype, found in 56 isolates. The ehxA identified in 36 (45.6%) isolates, followed by iha 5 (6.3%), eaeA 4 (5.1%), hlyA 2 (2.5%) and saa 2 (2.5%). Most of the isolates belonged to phylogroup B1. Only two O26 and one O91 isolates were detected in our study. CONCLUSION: Our results show that STEC strains were widespread among healthy domestic animals in the southeast of Iran.

Pathogenic potential of Shiga toxin-producing Escherichia coli strains of caprine origin: virulence genes, Shiga toxin subtypes, phylogenetic background and clonal relatedness.

BACKGROUND: All over the world, Shiga toxin-producing Escherichia coli (STEC) are considered as important zoonotic pathogens. Eight serogroups have the greatest role in the outbreaks and diseases caused by STEC which include O26, O45, O103, O111, O113, O121, O145 and O157. Ruminants, especially cattle are the main reservoirs but the role of small ruminants in the epidemiology of human infections has not been thoroughly assessed in many countries. The objective of this research was to investigate the pathogenic potential of the STEC strains isolated from slaughtered goats. In this study, a total of 57 STEC strains were recovered from 450 goats and characterized by subtyping of stx genes, O-serogrouping, phylo-typing and DNA fingerprinting. RESULTS: Amongst 57 STEC strains isolated from goats, the prevalence of stx1 was significantly more than stx2 (98.2% vs. 24.5%; P ≤ 0.05), and 22.8% of strains harbored both stx1 and stx2 genes. Three (5.2%) isolates were characterized as EHEC, which carried both eae and stx genes. A total of five stx-subtypes were recognized namely: stx1c (94.7%), stx1a (53.7%), stx2d (21%), stx2c (17.5%), and stx2a (15.7%). In some parts of the world, these subtypes have been reported in relation with severe human infections. The stx subtypes predominantly occurred in four combinations, including stx1a/stx1c (35%), stx1c (31.5%), stx1c/stx2a/stx2c/stx2d (5.2%) and stx1c/stx2c/stx2d (%5.2%). In serogrouping, the majority of STECs from goats did not belong to the top 8 serogroups but two strains belonged to O113, which has been recognized as an important pathogenic STEC in Australia. Interestingly, none of stx + eae + isolates belonged to the tested serogroups. In phylo-typing the isolates mostly belonged to phylo-group B1 (82.4%), followed by phylo-group A (12.3%). STEC strains showed a substantial diversity in DNA fingerprinting; there were 24 unique ERIC-types (with a ≥95% similarity) among the isolates. CONCLUSIONS: Despite the fact that the top 8 STEC serogroups were uncommon in caprine strains, the presence of highly pathogenic stx subtypes indicates that small ruminants and their products can be considered as an overlooked public health risk for humans, especially in developing countries which consume traditional products.

Recombinant M2e-HA2 fusion protein induced immunity responses against intranasally administered H9N2 influenza virus.

Influenza is a highly contagious respiratory tract disease and is considered a serious community health problem. Influenza viruses possess multiple conserved epitopes which are used for designing universal vaccines. To this aim, the gene coding for N-terminal part of M2e (SLLTEVET) and HA2 (GLFGAIAGF), was synthesized, linked by a (Gly4Ser)4 peptide linker, and cloned into pGS-21a vector. Afterwards, the construct was transferred into E. coli BL21 (DE3) cells to produce the designed antigenic protein called M2e-HA2. Immunization of mice with these peptides significantly induced humoral immune responses against the influenza virus. Three weeks after the last booster, mice were inoculated intranasally with 1 × 106 EID50 of H9N2 virus. The results indicated that the recombinant M2e-HA2 fusion protein could protect mice against H9N2 virus.

Role of antigen-43 on biofilm formation and horizontal antibiotic resistance gene transfer in non-O157 Shiga toxin producing Escherichia coli strains.

BACKGROUND AND OBJECTIVES: The objectives of this study were to evaluate the antibiotic resistance profiles, biofilm formation, presence of antigen 43 (Ag43) gene, and transfer of antibiotic resistance phenotype among non-O157 Shiga toxin producing Escherichia coli (STEC). MATERIALS AND METHODS: From October 2014 to November 2015 a total of 276 stool samples were collected from healthy calves, goats and 395 patients with the sign of nonbloody diarrhea and screened for presence of stx and serotype O157 genes by polymerase chain reaction (PCR) technique. Susceptibility to 14 antibiotics was determined as per CLSI guideline. Presence of Ag43 and intimin (eaeA) genes were detected by PCR. Biofilm formation was measured by microtiter plate method. Conjugation was carried out by membrane filter technique. RESULTS: We isolated 74 (93.6%) non-O157 STEC strains from 41 calves, 33 goats and 5 (6.3%) patients' stools, however, no O157 serotype was detected in our study. Resistance was observed most commonly to tobramycin (66.2%), kanamycin (48.6%), and amikacin (29.7%) and less frequently to ciprofloxacin (4.1%), amoxicillin-clavulanic acid (5.4%), and ceftriaxone (9.5%) in isolates recovered from calves and goats fecal samples, whereas, all human isolates were sensitive to ceftazidime, ciprofloxacin, tobramycin and imipenem, respectively. Furthermore, Ag43 was detected in 60 STEC isolated from animals and 5 human origins (no eaeA gene was found in this study). Biofilm formation from Ag43+ and Ag43- colonies showed 20 isolates with strong biofilm activities. Cefotaxime resistance phenotype was transferred to E. coli ATCC 25922.1 (Nalr) by conjugation at a frequency of 1.6×10-4. CONCLUSION: From the above results we concluded that, human infections with non-O157 STEC were significantly low in Kerman. Ag43 was insignificant with biofilm quantity in most cases.

Phylogenetic typing and molecular detection of virulence factors of avian pathogenic Escherichia coli isolated from colibacillosis cases in Japanese quail.

Colibacillosis caused by avian pathogenic Escherichia coli (APEC) is an economic threat to the poultry industry throughout the world. Some of the virulence genes may enhance the ability of E. coli isolates to grow in the tissues of broilers. The APEC strains are assigned to a few distinct phylogenetic groups. The purpose of the present study was to detect the virulence genes and phylogenetic groups of E. coli isolates from colibacillosis cases in Japanese quail in 2014 in Kerman, Iran. In the present study, one hundred and two E. coli isolates were obtained from dead Japanese quails with colibacillosis. E. coli isolates were confirmed by standard biochemical and bacteriological methods. DNA of E. coli isolates was extracted by boiling method. The confirmed E. coli isolates were investigated to detect the phylogenetic groups and virulence genes including sfa/focDE, afaIBC, papEF by PCR methods. E. coli isolates were classified into A (62 isolates), B1 (24 isolates), B2 (12 isolates) and D (four isolates) phylogenetic groups. Among examined isolates nine isolates (8.82%) were positive for papE-F, five isolates (4.90%) for afaIB-C and two isolates (1.96%) for sfa/focD-E genes. Based on our findings, E. coli isolates from colibacillosis of Japanese quail could be assigned to various phylogenetic groups (mostly A and D), and they may contain the adhesion genes in a low prevalence.

Genotypic analysis of virulence genes and antimicrobial profile of diarrheagenic Escherichia coli isolated from diseased lambs in Iran.

The aim of the present study was to determine the analysis of virulence genes and antimicrobial profile of diarrheagenic Escherichia coli isolated from diseased lambs. Two hundred ninety E. coli isolates were recovered from 300 rectal swabs of diarrheic lambs and were confirmed by biochemical tests. The pathotype determination was done according to the presence of genes including f5, f41, LTI, STI, bfp, ipaH, stx 1 , stx 2 , eae, ehlyA, cnf 1 , cnf 2 , cdIII, cdIV, and f17 by PCR method. Sixty-six isolates (23.72%) possessed the STI gene and categorized into entrotoxigenic E. coli (ETEC). Nine isolates (3.1%) and five isolates (1.72%) were positive for the cnf1 and cnf2 genes which categorized into necrotoxic E. coli (NTEC). Hundred and seventeen isolates (40.34%) harbored stx 1 and/or stx 2 and classified as Shiga toxin-producing E. coli (STEC). Thirteen isolates (4.48%) were assigned to atypical entropathogenic E. coli (aEPEC) and possessed eae gene. Two isolates (0.68%) were positive for ipaH gene and were assigned to entroinvasive E. coli (EIEC). Statistical analysis showed a specific association between eae gene and STEC pathotype (P < 0.0001). The most prevalent resistance was observed against lincomycin (96.5%) and the lowest resistance was against kanamycine (56.02%), respectively. The high prevalence of STEC and ETEC indicates that diarrheic lambs represent an important reservoir for humans. ETEC may play an important role for frequent occurrence of diarrhea in lambs observed in this region. Due to high antibiotic resistance, appropriate control should be implemented in veterinary medicine to curb the development of novel resistant isolates.

High Frequency of Diarrheagenic Escherichia coli in HIV-Infected Patients and Patients with Thalassemia in Kerman, Iran.

This study was conducted on patients with thalassemia and HIV-infected patients to determine the frequency of diarrheagenic Escherichia coli in Kerman, Iran. We analyzed 68 and 49 E coli isolates isolated from healthy fecal samples of patients with thalassemia and HIV-infected patients, respectively. The E coli isolates were studied using a multiplex polymerase chain reaction to identify the enterotoxigenic E coli (ETEC), enterohemorrhagic E coli (EHEC), and enteropathogenic E coli (EPEC) groups. Statistical analysis was carried out to determine the correlation of diarrheagenic E coli between HIV-infected patients and patients with thalassemia using Stata 11.2 software. The frequency of having at least 1 diarrheagenic E coli was more common in patients with thalassemia (67.64%) than in HIV-infected patients (57.14%; P = .25), including ETEC (67.64% versus 57.14%), EHEC (33.82% versus 26.53%), and EPEC (19.11% versus 16.32%). The results of this study indicate that ETEC, EHEC, and EPEC pathotypes are widespread among diarrheagenic E coli isolates in patients with thalassemia and HIV-infected patients.

Phylogenetic background of enterotoxigenic and enteroinvasive Escherichia coli from patients with diarrhea in Sirjan, Iran.

BACKGROUND AND OBJECTIVES: Diarrheagenic Escherichia coli (DEC) strains are a major cause of intestinal syndromes in the developing countries. The aim of this study was to determine the prevalence of enterotoxigenic E. coli (ETEC) and enteroinvasive E. coli (EIEC) in relation to phylogenetic background from patients with diarrhea. MATERIALS AND METHODS: A total of 110 E. coli isolates were obtained from diarrhea patients in Sirjan, southeast of Iran. The E. coli isolates were confirmed using biochemical and bacteriological tests. DNA of E. coli isolates was extracted by boiling method and checked for existence of ETEC (LT and ST genes) and EIEC (ipaH gene) pathotypes and also characterize the phylogenetic groups on the basis of presence or absence of the chuA, yjaA genes and an anonymous DNA fragment, TspE4. C2 by multiplex PCR. RESULTS: Out of 110 E. coli isolates, 32 (29.09%) were positive for ETEC (LT and ST genes) and 6 (5.45%) possessed EIEC (ipaH gene) pathotypes. Isolates fall into four phylogenetic groups: A (39.09%), B1 (20%), B2 (15.45%) and D (25.45%). Phylotyping of isolates of DEC indicated they were distributed in four phylogenetic groups including A (12 isolates), B1 (7), B2 (9) and D (10). CONCLUSION: In this study, the DEC isolates were segregated into different phylogenetic groups. The majority of isolates belonged to phylo-groups A and D.

Phylogenetic typing and detection of extended-spectrum ß-lactamases in Escherichia coli isolates from broiler chickens in Ahvaz, Iran.

This study was conducted to reveal the phylogenetic background, to detect the genes encoding TEM, SHV and CTX-M-15 extended-spectrum ß-lactamases (ESBL), and to analyze their distribution in phylo-groups of 150 Escherichia coli isolates from broiler chickens in Ahvaz (Southwest of Iran). Seventy- five cloacal swabs from healthy birds (fecal isolates), and 75 heart blood samples from birds with colibacillosis (septicemic isolates) were obtained. All isolates were phylotyped and screened for ESBL genes by polymerase chain reaction (PCR). The fecal isolates belonged to four main phylo-groups, including 41 isolates (54.67%) to A, 9 (12.00%) to B1, 5 (6.67%) to B2, and 20 (26.67%) to D. Of septicemic isolates, 37 isolates (49.33%) were classified as phylotype A, 5 (6.67%) as B1, 10 (13.33%) as B2, and 23 (30.67%) as D. In molecular analysis, a total of 72 isolates (35 fecal and 37 septicemic) were identified to harbor ESBL genes, which were distributed in phylo-groups A, B1, B2, and D. Regardless of the type of isolate, blaCTX-M-15 gene was the most common genotype, followed by blaTEM and blaSHV genes. This study suggests that broiler chickens in Iran are infected to ESBL genes- harboring Escherichia coli strains which may be spread to the food chain through fecal contamination of carcasses during slaughtering.