Sfa Genes Research Articles

Virulence factors, biofilm-forming ability, and antimicrobial resistance of urinary Escherichia coli strains isolated from hospitalized patients.

Escherichia coli is the most frequent cause of urinary tract infections. We investigated the possible associations between the origin of strains, antimicrobial resistance, the presence of urovirulence factors, and biofilm-forming ability. Antibiotic susceptibility of E. coli strains was tested by disk diffusion method. Hemagglutination assays were performed for phenotypic characterization of the cell surface. Multiplex PCR was used for detection of virulence genes and for determination of phylogenetic relationships. The resistance to ampicillin (55.5%) and tetracycline (39.3%) was significantly more frequent than to other antimicrobial agents. The fim gene was present in 92.5% of strains. The sfa and pap genes were found in 53.8% and 38.7% of strains, respectively. The pap gene was significantly less frequently detected in strains from dialysis patients. The hly gene was present in 18.5% of strains. The aer gene was detected in 52.6% and cnf in 12.1%, while afa was detected in 4.6% of strains. Most strains belonged to the B2 and D phylogenetic groups. The aer gene was significantly associated with strains producing strong biofilms. The E. coli strains causing cystitis in hospitalized patients differed in terms of resistance to antibiotics, virulence genes, and potential for biofilm formation.

Detection of pap, sfa, afa, foc, and fim Adhesin-Encoding Operons in Uropathogenic Escherichia coli Isolates Collected From Patients With Urinary Tract Infection.

Background:Uropathogenic Escherichia coli (UPEC) with its virulence factors is the most prevalent cause of urinary tract infection (UTI).Objectives;This study aimed to determine the occurrence of fim, pap, sfa, and afa genes among 100 UPEC isolates collected from patients diagnosed with UTI.Materials and MethodsA total of 100 UPEC isolates were obtained from urine samples of patients with UTI. The prevalence of 5 virulence genes encoding type 1 fimbriae (fimH), pili associated with pyelonephritis (pap), S and F1C fimbriae (sfa and foc) and afimbrial adhesins (afa) were determined through PCR method. We also investigated the phylogenetic background of all isolates. In addition, the distribution of adhesin-encoding operons between the phylogroups was assessed.Results:The prevalence of genes encoding for fimbrial adhesive systems was 95% for fim, 57% for pap, 16% for foc, and 81% for sfa. The operons encoding for afa afimbrial adhesins were identified in 12% of isolates. The various combinations of detected genes were designated as virulence patterns. The fim gene, which occurred in strains from all phylogenetic groups (A, B1, B2, and D) was evaluated and no significant differences were found among these groups. Conversely, significant differences were observed in relation to pap, afa, foc, and sfa operons.Conclusions:These results indicate that the PCR method is a powerful genotypic assay for the detection of adhesin-encoding operons. Thus, this assay can be recommended for clinical use to detect virulent urinary E. coli strains, as well as epidemiological studies.

Detection of virulence genes in Escherichia coli isolated from patients with cystitis and pyelonephritis

Uropathogenic Escherichia coli (UPEC) is a common cause of ascending urinary tract infections including cystitis and pyelonephritis. The purpose of this study was to investigate virulence genes among Escherichia coli isolated from patients with cystitis and pyelonephritis. Between December 2012 and June 2013, 150 E. coli isolates from hospitalized patients with pyelonephritis (n = 72) and cystitis (n=78) were collected at Shahid Beheshti Hospital in Kashan. A PCR assay was used to evaluate the presence of virulence genes including pap, hly, aer, sfa, cnf, afa, traT, and pathogenicity island (PAI) markers in isolates. Of the total 150 UPEC isolates, 130 (86.7%) were found to carry the virulence genes studied. Nineteen different virulence patterns were identified. The most prevalent virulence pattern was UPEC including traT-PAI operons. The pap, traT, aer, hly, and PAI operons were more prevalent among patients with pyelonephritis than cystitis, and the sfa, afa, and cnf genes were not detected in any of the isolates. Higher virulence gene diversity was found among pyelonephritis UPEC isolates in comparison to cystitis UPEC isolates, showing that UPEC strains that cause pyelonephritis need more virulence factors.

Virulence Factor Diversity Between Imipenem Resistant and Imipenem Susceptible Strains of Escherichia coli Isolated from Hospitalized Patients with Severe Urinary Tract Infections

cultured immediately and those that were E. coli-positive were analyzed for the antimicrobial susceptibility pattern against imipenem using disk diffusion method. Imipenem resistant and imipenem susceptible strains were analyzed for the presence of sfa, afa, pap, hly, cnf1 and fim virulence factors using the PCR technique. Fifty out of 70 (71.42%) children urines and 55 out of 90 (61.11%) seniors urine samples harbored E. coli. The incidence of resistance against imipenem in children and seniors were 6% and 9.09%, respectively. In the other hand, 99.39% of tested strains were susceptible to imipenem. Total prevalence of pap, cnf1, hly, sfa, afa and fim genes in the imipenem resistant strains were 12.5% 25%, 50%, 75%, 62.5% and 25%, respectively. Prevalence of the sfa, afa and hly genes was lower in the imipenem susceptible strains of E. coli, while the pap, cnf1 and fim genes was entirely higher in the imipenem susceptible strains. This finding suggests that imipenem resistance could be directly associated with decreased prevalence of pap, cnf1 and fim virulent genes. However, the findings of the present study are novel and valuable but more courtesy studies are necessary to authorize them.

Biofilm and fluoroquinolone resistance of canine Escherichia coli uropathogenic isolates.

BackgroundEscherichia coli is the most common uropathogen involved in urinary tract infection (UTI). Virulence of strains may differ, and may be enhanced by antimicrobial resistance and biofilm formation, resulting in increased morbidity and recurrent infections. The aim of this study was to evaluate the in vitro biofilm forming capacity of E. coli isolates from dogs with UTI, by using fluorescent in situ hybridization, and its association with virulence genes and antimicrobial resistance.FindingsThe proportion of biofilm-producing isolates significantly increased with the length of incubation time (P < 0.05). Biofilm production was significantly associated with fluoroquinolone resistance at all incubation time points and was independent of the media used (P < 0.05). Biofilm production was not associated with cnf1, hly, pap and sfa genes (P > 0.05), but was significantly associated with afa, aer and the β-lactamase genes (P < 0.05).ConclusionsTo the best of our knowledge, this is the first report showing significant association between biofilm production and fluoroquinolone resistance in E. coli isolates from dogs with UTI. Biofilm formation may contribute to UTI treatment failure in dogs, through the development of bacterial reservoirs inside bladder cells, allowing them to overcome host immune defenses and to establish recurrent infections.

Correlation Between &lt;i&gt;in vitro&lt;/i&gt; Biofilm Formation and Virulence Properties of Extra-Intestinal Pathogenic &lt;i&gt;Escherichia Coli&lt;/i&gt; (Expec)

About 65 Escherichia coli strains isolated from extra-intestinal sites were included in this study to determine their biofilm formation capability and virulence properties. About 87% ExPEC isolates were motile and 12.3% were found to be non-motile. Only 12.30% isolates were strong biofilm former (SBF >1.5) whereas 18.46% isolates were non biofilm former. About 71% isolates were capable to express curli but curli expression is lower in isolates from pus and peritoneal fluid. About 41.53% isolates have moderate capability (CSH value 21-50%) to adhere to hydrocarbons whereas 20% didn’t show any hydrocarbon adherence capability. Most of the isolates showed moderate cell aggregation (60-80%) potential with S. maltophila but 15.4% isolates were found to be non-aggregative. Forty-one (63.07%) strains were resistant to 2 or more of the most commonly clinically used antibiotics. 44.6% isolates were haemolytic, 35.3% showed colicin activity, 47.7% were MRHA positive, 49.2% had SAT value ≤2.0 M and 9.2% were protease producer. About 77% (50 out of 65) of the isolates harbor type 1 gene. csgA gene was found in 61.5% (40) isolate, papC was found in 64.6% (42), afa and sfa genes were found in 67.7% (44) each. Biofilm formation may contribute to the pathogenic potential of ExPEC.

Prevalence of adhesive genes among uropathogenic Escherichia coli strains isolated from patients with urinary tract infection in Mangalore

The study was carried out to detect the adhesive genes pap (pyelonephritis associated pili), sfa (S fimbrial adhesin) and afa (afimbrial adhesin) from Escherichia coli strains isolated in patients diagnosed with urinary tract infection (UTI). A total of 23% of the isolates were positive for pap, sfa and afa genes with a prevalence of 60.87% (14/23), 39.1% (9/23) and 39.1% (9/23), respectively. Prevalence of multiple adhesive genes was 8.7% (2/23) for pap and afa, 30.43% (7/23) for pap and sfa. Significant numbers of isolates were positive for Congo red binding (80%) and haemolysin production 60%. The prevalence of multiple adhesive genes indicate the potential to adhere and subsequently cause a systemic infection among UTI patients.

Avian ExtraintestinalEscherichia coliExhibits Enterotoxigenic-Like Activity in theIn VivoRabbit Ligated Ileal Loop Assay

Avian pathogenic Escherichia coli (APEC) strains harbor a number of virulence genes and cause extraintestinal diseases, such as septicemia, swollen-head syndrome, salpingitis, and omphalitis in poultry. APEC strains are not known to cause intestinal diseases. Herein, for the first time, it is reported that APEC strains were able to induce an enterotoxigenic-like effect in rabbit ligated ileal loops. Strain SEPT362 caused cell detachment of the intestinal villi, which also showed a flattened and wilted appearance, but the integrity of the tight junctions was maintained. Additionally, this strain did not adhere to enterocytes in vivo, although adhesin encoding genes ( fimH, csgA, lpfA2-3, and ECP) were present while other lpfA types, sfa, afa, papC, and ral genes were not. This enterotoxigenic-like activity was conserved after thermal treatment of the supernatant at 65°C but not at 100°C. Moreover, experiments based on filtering with different molecular weight cut-off (MWCO) pore sizes demonstrated that the component associated with the observed biological effect has a molecular weight >100 kDa. Blast search and polymerase chain reaction assays for known E. coli virulence factors showed that strain SEPT362 harbors the gene encoding for the toxin EAST-1 and the serine protease autotransporter (SPATE) Tsh, but is negative for genes encoding for the toxins LT-I, STh, STp, Stx1, Stx2, CNF-1, CNF-2, CDT and the SPATEs Sat, Pic, Vat, SigA, SepA, EatA, EspP, or EspC. A cloned copy of the tsh gene in E. coli K-12 was also tested and was shown to have an enterotoxic effect. These results suggest that APEC might induce fluid accumulation in the rabbit gut. The Tsh autotransporter seems to be one of the factors associated with this phenotype.

Association of some virulence genes with antibiotic resistance among uropathogenic Escherichia coli isolated from urinary tract infection patients in Alexandria, Egypt: A hospital-based study.

Uropathogenic Escherichia coli (UPEC) is the infecting agent most frequently involved in urinary tract infections (UTIs) worldwide. UPEC resistance to commonly used antibiotics represents a major health problem all over the world. Several factors have been associated with UPEC resistance to antibiotics. The present study deployed a molecular approach to explore the association between some UPEC virulence genes and antibiotic resistance among patients with UTI in Alexandria, Egypt. The study revealed a significant association between presence of the pap gene and resistance to gentamicin; however, it was not significantly associated with resistance to β-lactam antibiotics, quinolones, aminoglycosides, nitrofurantoin and trimethoprim/sulfamethoxazole. The genes sfa, aer and cnf1 were not significantly associated with UPEC resistance to any of the tested antibiotics. In conclusion, resistance of UPEC isolates in the present study could be attributed to other virulence factors.

Virulence Factors and O-Serogroups Profiles of Uropathogenic Escherichia Coli Isolated from Iranian Pediatric Patients

Background:Uropathogenic Escherichia coli O- Serogroups with their virulence factors are the most prevalent causes of UTIs.Objectives:The present investigation was performed to study the virulence factors and O-Serogroups profiles of UPEC isolated from Iranian pediatric patients.Patients and Methods:This cross sectional investigation was performed on 100 urine samples collected from hospitalized pediatrics of Baqiyatallah Hospital, Tehran, Iran. Midstream urine was collected to decrease potential bacterial, cellular and artifactual contamination. All samples were cultured and those with positive results were subjected to polymerase chain reactions to detect pap, cnf1, afa, sfa and hlyA genes and various O- Serogroups.Results:We found that 37.5% of boys and 75% of girls had positive results for Escherichia coli. We also found that O1 (19.33%), O2 (13.33%), O6 (13.33%), O4 (11.66%), and O18 (11.66 %) were the most commonly detected Serogroups. Totally, the serogroup of 5% of all strains were not detected. In addition, all of these O- Serogroups were pap+, cnf1+, hlyA+, and afa+. Totally, pap (70 %), cnf1 (56.66 %), and hlyA (43.33 %) were the most commonly detected virulence genes in the both studied groups of children. The sfa (30 %) and afa (26.66 %) genes had the lowest incidence rates.Conclusions:Special health care should be performed on UTIs management in Iranian pediatric patients. Extended researches should be performed to evaluate relation between other O-Serogroups and virulent genes.

DISTRIBUTION OF VIRULENCE ASSOCIATED GENES IN ISOLATED ESCHERICHIA COLI FROM AVIAN COLIBACILLOSIS

BACKGROUND: Colibacillosis is one of the most prevalentdiseases in the world that causes multimillion-dollar annuallosses. OBJECTIVES: In order to evaluate molecular epidemiologyof some virulence associated factors in Escherichia coli,isolated from poultry, the presence of iut A, iss, hly F, omp T, iroN, afa, sfa (S)and pap G (II) were investigated by multiplex PCRassay. METHODS: Two hundred thirty four Escherichia coliisolated from avian colibacillosis (APEC) and fifty four fecal E.coli isolates from the feces of apparently healthy birds (AFEC)were investigated for presence of some virulence associatedgenes by two panel of multiplex PCR. Statistical analysis wasperformed using |c2 test. the p-value was |£|0.05. RESULTS:Among 234 E. coli strains associated with colibacillosis and 54AFEC strains, 85% of isolates were positive for at least one of thevirulence gene. The three most prevalent genes in E. coli isolatedfrom colibacillosis were hly F (77.3%), omp T(73%) and iss(68.2%). Iut A, iro Nand pap G (II) were detected in 157 (67.4%),152 (65.2%) and 41(17.6%) respectively. None of isolatesharbored sfa (s) and afa genes. Several combination patterns ofvirulence genes were detected. Combination of hly F, omp T(70.8%) was the most prevalent pattern. CONCLUSIONS: theprevalence of iss, hly F, omp T, iro N genes in APEC isolates wassignificantly more than AFEC strains and probably these genesplay an important role in the pathogenesis of APEC strains.

Prevalence of antimicrobial resistance in relation to virulence genes and phylogenetic origins among urogenital Escherichia coli isolates from dogs and cats in Japan

To assess the status of antimicrobial resistance (AMR), identify extraintestinal virulence factors (VFs) and phylogenetic origins, and analyze relationships among these traits in extraintestinal pathogenic Escherichia coli (ExPEC) isolates from companion animals. 104 E coli isolates obtained from urine or genital swab samples collected between 2003 and 2010 from 85 dogs and 19 cats with urogenital infections in Japan. Antimicrobial susceptibility of isolates was determined by use of the agar dilution method; a multiplex PCR assay was used for VF gene detection and phylogenetic group assessment. Genetic diversity was evaluated via randomly amplified polymorphic DNA analysis. Of the 104 isolates, 45 (43.3%) were resistant to > 2 antimicrobials. Phylogenetically, 64 (61.5%), 22 (21.2%), 13 (12.5%), and 5 (4.8%) isolates belonged to groups B2, D, B1, and A, respectively. Compared with other groups, group B2 isolates were less resistant to all tested antimicrobials and carried the pap, hly, and cnf genes with higher frequency and the aer gene with lower frequency. The aer gene was directly associated and the pap, sfa, hly, and cnf genes were inversely associated with AMR. Randomly amplified polymorphic DNA analysis revealed 3 major clusters, comprised mainly of group B1, B2, and D isolates; 2 subclusters of group B2 isolates had different VF and AMR status. CONCLUSIONS AND CLINICAL RELEVANCE; Prevalences of multidrug resistance and human-like phylogenetic origins among ExPEC isolates from companion animals in Japan were high. It is suggested that VFs, phylogenetic origins, and genetic diversity are significantly associated with AMR in ExPEC.

Antimicrobial resistance, virulence profiles, and phylogenetic groups of fecal Escherichia coli isolates: A comparative analysis between dogs and their owners in Japan

In this study, fecal Escherichia coli isolates (n=188) from 34 dog-owner pairs and 26 healthy control humans (2 isolates per individual) were tested for susceptibility to 6 antimicrobials and screened for virulence genes. Genetic diversity between canine and owner isolates was evaluated by pulsed-field gel electrophoresis (PFGE). Canine isolates exhibited significantly different rates of resistance to four and two antimicrobials, compared to control and owner isolates, respectively. Of the genes examined, the prevalence of sfa, hly, and cnf genes in canine isolates were higher than in control isolates, but not than in owner isolates. These results suggest that characteristics of owner isolates are somewhat similar to canine isolates, compared to isolates from non-dog owners. In addition, PFGE analysis revealed that transfer of E. coli between owners and their dogs had occurred within 3/34 (8.8%) households. Considering the effects of dog ownership on the population of E. coli isolates from owners, further epidemiological studies are required.

Phenotypical characterization and adhesin identification in Escherichia coli strains isolated from dogs with urinary tract infections.

Pathogenic strains of Escherichia coli are the most common bacteria associated with urinary tract infections in both humans and companion animals. Standard biochemical tests may be useful in demonstrating detailed phenotypical characteristics of these strains. Thirteen strains of E. coli isolated from dogs with UTIs were submitted to biochemical tests, serotyping for O and H antigens and antimicrobial resistance testing. Furthermore, the presence of papC, sfa, and afa genes was evaluated by PCR, and genetic relationships were established using enterobacterial repetitive intergenic consensus PCR (ERIC-PCR). The antimicrobial that showed the highest resistance rate among the isolates was nalidixic acid (76.9%), followed by cephalotin (69.2%), sulfamethoxazole + trimethoprim (61.5%), tetracycline (61.5%), streptomycin (53.8%), ciprofloxacin (53.8%), ampicillin (46.2%), gentamicin (30.8%) and chloramphenicol (23.1%). No isolate was resistant either to meropenem or nitrofurantoin. Among the five clusters that were identified using ERIC-PCR, one cluster (A) had only one strain, which belonged to a serotype with zoonotic potential (O6:H31) and showed the genes papC+, sfa+, afa-. Strains with the genes papC-, sfa+, afa- were found in two other clusters (C and D), whereas all strains in clusters B and E possessed papC-, sfa-, afa- genes. Sucrose and raffinose phenotypic tests showed some ability in discriminating clusters A, B and C from clusters D and E.

Comparative genomics of Escherichia coli isolated from patients with inflammatory bowel disease

BackgroundInflammatory bowel disease (IBD) is used to describe a state of idiopathic, chronic inflammation of the gastrointestinal tract. The two main phenotypes of IBD are Crohn's disease (CD) and ulcerative colitis (UC). The major cause of IBD-associated mortality is colorectal cancer. Although both host-genetic and exogenous factors have been found to be involved, the aetiology of IBD is still not well understood. In this study we characterized thirteen Escherichia coli strains from patients with IBD by comparative genomic hybridization employing a microarray based on 31 sequenced E. coli genomes from a wide range of commensal and pathogenic isolates.ResultsThe IBD isolates, obtained from patients with UC and CD, displayed remarkably heterogeneous genomic profiles with little or no evidence of group-specific determinants. No IBD-specific genes were evident when compared with the prototypic CD isolate, LF82, suggesting that the IBD-inducing effect of the strains is multifactorial. Several of the IBD isolates carried a number of extraintestinal pathogenic E. coli (ExPEC)-related virulence determinants such as the pap, sfa, cdt and hly genes. The isolates were also found to carry genes of ExPEC-associated genomic islands.ConclusionsCombined, these data suggest that E. coli isolates obtained from UC and CD patients represents a heterogeneous population of strains, with genomic profiles that are indistinguishable to those of ExPEC isolates. Our findings indicate that IBD-induction from E. coli strains is multifactorial and that a range of gene products may be involved in triggering the disease.

The DNA Sequence of the Escherichia coli O22 O-Antigen Gene Cluster and Detection of Pathogenic Strains Belonging to E. coli Serogroups O22 and O91 by Multiplex PCR Assays Targeting Virulence Genes and Genes in the Respective O-Antigen Gene Clusters

Multiplex polymerase chain reaction (PCR) assays were developed for detection of pathogenic strains belonging to Escherichia coli serogroups O22 and O91. The O-antigen gene cluster of E. coli O22 was sequenced to identify genes that could be employed as targets for serogroup-specific PCR assays. The wzx and wzy genes in the O-antigen gene clusters of E. coli O22 and E. coli O91 were selected as target genes. The assays were serogroup-specific when tested against 72 E. coli O22 strains and 57 E. coli O91 strains isolated from food, humans, and animals, representative strains belonging to 168 E. coli O serogroups and non-E. coli bacteria. Furthermore, 72 E. coli O22 strains and 57 E. coli O91 strains isolated from food, water, animals, and humans were tested by the PCR for the presence of six and 19 virulence genes, respectively, associated with pathogenic E. coli strains. Based on the PCR screening results, multiplex PCR assays targeting the O22 wzy gene and the cnf-1 and sfa genes in E. coli O22 and the O91 wzy gene, conserved sequences of stx1 and stx2 genes, and the astA and cdt-III genes in E. coli O91 were developed to detect and identify pathogenic strains belonging to serogroups O22 and O91. Furthermore, E. coli O22 and O91 were detected by multiplex PCR assays targeting the wzx or wzy genes and conserved sequences of the stx1 and stx2 genes in ground beef samples inoculated with approximately two colony-forming units (CFU)/25 g after 18-h enrichment. The results demonstrate that the E. coli O22 and O91 wzx and wzy gene sequences were specific for the respective serogroups and can be used as diagnostic markers for rapid identification of these serogroups as an alternative to serotyping. The multiplex PCR assays targeting the O22 and O91 wzx and wzy genes and virulence genes can be used to identify and to detect pathogenic strains of these serogroups in food and fecal samples.

Some adhesins of avian pathogenic Escherichia coli (APEC) isolated from septicemic poultry in Brazil

Three hundred and fifty strains of E. coli isolated from septicemic poultry from seven states of Brazil were examined for presence of nine adhesion-encoding genes, hemagglutination and adherence to chicken tracheal cells (in vitro). Analysis of the strains by colony hybridization tests demonstrated that 93.7% of the isolates were fim +, 17% pap+ and 5.7% were sfa+. The mannose sensitive fimbriae occur with similar frequency in APEC isolated from all Brazilians states, while significant differences among pap and sfa genes distributions were observed. The results showed that 0.85% and 0.28% of APEC were positive for genes that encoded enteroaggregative adhesins and EPEC adherence factor, respectively. None of APEC was positive for DA, afa, Bfp and Eae probes. The adherence to chicken tracheal cells showed 96% positive strains, while hemagglutination assays showed 26.5% of the isolates were mannose sensitive and 21.7% were mannose resistant.

P.425 Non-alcoholic fatty liver disease in the Philippines: comparable with other nations?

Uropathogenic Escherichia coli (UPEC) is a common cause of ascending urinary tract infections including cystitis and pyelonephritis. The purpose of this study was to investigate virulence genes among Escherichia coli isolated from patients with cystitis and pyelonephritis.Between December 2012 and June 2013, 150 E. coli isolates from hospitalized patients with pyelonephritis (n = 72) and cystitis (n = 78) were collected at Shahid Beheshti Hospital in Kashan. A PCR assay was used to evaluate the presence of virulence genes including pap, hly, aer, sfa, cnf, afa, traT, and pathogenicity island (PAI) markers in isolates.Of the total 150 UPEC isolates, 130 (86.7%) were found to carry the virulence genes studied. Nineteen different virulence patterns were identified. The most prevalent virulence pattern was UPEC including traT–PAI operons. The pap, traT, aer, hly, and PAI operons were more prevalent among patients with pyelonephritis than cystitis, and the sfa, afa, and cnf genes were not detected in any of the isolates.Higher virulence gene diversity was found among pyelonephritis UPEC isolates in comparison to cystitis UPEC isolates, showing that UPEC strains that cause pyelonephritis need more virulence factors.

Transcriptional analysis of the sfa determinant revealing mmRNA processing events in the biogenesis of S fimbriae in pathogenic Escherichia coli.

Among the virulence factors present in pathogenic extraintestinal Escherichia coli strains, expression of fimbrial adhesins is necessary for attachment to the host tissues and subsequent colonization. Occurrence of the sfa determinant coding for the S fimbriae is widespread among the uropathogens and meningitis isolates. The sfa operon consists of nine genes. In the biogenesis of S fimbriae, the proteins encoded by the sfa genes are presumably required in a specific stoichiometry. In the present work we studied how differential expression of the sfa operon genes occurs. Our findings indicate that a number of endoribonucleolytic cleavages occur in the mRNA from the sfa operon, and we detected the presence of different distinct transcriptional products, including sfaBA, sfaA, sfaADE, and sfaGSH. The sfaGSH transcript represents the three distal genes of the sfa operon, which code for the minor subunits of the S fimbriae. Analysis of the proteins in S fimbriae suggested that expression of the sfaGSH transcript provides equimolar amounts of the minor subunits. Furthermore, we showed that in the generation of the major sfaA transcript, the processing included RNase E endoribonuceolytic cleavage of the precursor sfaBA transcript. We suggest that posttranscriptional mRNA processing events result in differential gene expression important to achieve the stoichiometry necessary for fimbrial adhesin biogenesis.

Detection of urovirulence factors in Escherichia coli by multiplex polymerase chain reaction

Primers to amplify the genes encoding the virulence factors of uropathogenic Escherichia coli, such as pilus associated with pyelonephritis ( pap), haemolysin ( hly), aerobactin ( aer) and cytotoxic necrotizing factor 1 ( cnf1) genes, were designed. The above primers along with previously reported primers for S fimbriae ( sfa) and afimbrial adhesin I ( afaI) genes were combined to develop a multiplex polymerase chain reaction (PCR) for detection of the respective virulence factors and for the identification of uropathogenic E. coli. The multiplex PCR to detect pap, sfa, afaI, hly, aer and cnf1 genes was highly specific and the sensitivity was found to be about 5 × 10 3 colony forming units of E. coli per ml. A total of 194 E. coli strains isolated from patients with simple acute cystitis were examined by the multiplex PCR and the results were in complete agreement with that obtained by DNA colony hybridization test. The multiplex PCR developed was, therefore, concluded to be a useful, sensitive and rapid assay system to identify uropathogenic E. coli.