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Introduction: Staphylococcus aureus, coagulase-negative staphylococci, Enterococcus spp., Enterobacteriaceae, Pseudomonas aeruginosa, and Acinetobacter baumanii have been found to be the most prevalent bacteremia-causing bacteria in patients with septicemia. Early detection of bloodstream infection (BSI) is crucial in the clinical setting. A multiplex PCR method for identification of these agents in clinical samples has been developed in parallel by conventional microbiological methods., Methodology: The target genes selected for each of the organisms were very specific for designing primers. Design of primers was done using Mega4, Allel ID6, Oligo6, and Oligo analyzer software. The test comprises a universal PCR from the 16S rDNA gene and multiplex PCR from the rpoB, gyrA, sss, and chromosome X (as an internal control)., Results: The sensitivity and specificity for universal PCR and multiplex PCR in comparison with BC were 83.87% and 91.58%, and 74.19% and 91.58%, respectively. The positive predictive value (PPV) and the negative predictive value (NPV) for these two PCRs were 76.47% and 94.57%, and 74.19% and 91.58%, respectively. PCR failed to identify bacteria which were found conventionally in only 3.96% and 6.34% of the cases by universal and multiplex PCR (mostly bacteria not included in the PCR cassette). In 6.34% of the cases, multiplex PCR afforded identification of bacteria, but BC showed no bacteria in the sample., Conclusions: The multiplex PCR approach facilitates the detection of bacteremia in blood samples within a few hours. Rapid detection of bacteria by multiplex PCR appears to be a valuable tool, allowing earlier pathogen-adopted antimicrobial therapy in critically ill patients.

Aim: The aims of the present study were to determine the antibiotic susceptibility profils with particular emphasis on susceptible or resistant strains to macrolides and lincosamids antibiotics and to determine possible antibiotic resistance mechanisms occurring in group B streptococci (GBS) strains using PCR assay and disk diffusion method.Methods: A total of 62 clinical GBS strains were investigated. Antibacterial susceptibility testing was performed using the disk diffusion method and inducible resistance test for clindamycin by standard double disk diffusion or D-zone test for all isolates to differentiate macrolide resistance phenotype (M), constitutive macrolide-lincosamide-streptogramin B phenotype (cMLS) and induced macrolide-lincosamide-streptogramin B phenotype (iMLS). In addition, minimum inhibitory concentrations (MIC) of penicillin were determined for all isolates. Finally, possible existence of antibiotic resistance genes for erythromycin , and ) and for clindamycin ) were examined among isolates using PCR assay.Results: All 62 isolates were susceptible to penicillin, ampicillin, linezolid, cefazoline and vancomycin. However, 93.5% (n=58) of isolates showed an increased MIC to penicillin. The overall rate of erythromycin resistance was 35.5% (n=22). All erythromycin-resistant isolates displayed the M phenotype (100%, n=22). All three erythromycin resistance genes (i.e. , and ) were found in erythromycin-resistant isolates.Conclusion: It was concluded that prescribing antibiotic without antibacterial susceptibility tests should be prevented because of the high prevalence of erythromycin-resistant GBS strains and the fact that erythromycin-resistant GBS strains has shown an increased MIC to penicillin, as the drug of choice for treating GBS infections.

Background: Escherichia coli has appeared as an important opportunistic pathogen responsible for nosocomial infections in patients with immunodeficiency, particularly in leukemia patients. New Delhi metallo-beta-lactamase is an enzyme originally found in Enterobacteriaceae.Case Presentation: In this study, 80 isolates of Escherichia coli and Klebsiella pneumoniae were collected over the course of 2 years from two medical centers in Tehran, Iran. Production of carbapenemase was detected in the isolates using modified Hodge test. New Delhi metallo-beta-lactamase-1 genes were detected by polymerase chain reaction amplification with specific primers. Two New Delhi metallo-beta-lactamase-1-producing Escherichia coli strains were isolated from two Iranian patients with leukemia. These two patients were 6 and 15 years old, one female and the other male, from two oncology centers in Iran. The isolates were resistant to carbapenems (imipenem, meropenem), and two isolates were positive for carbapenemase production by modified Hodge test.Conclusions: The emergence of New Delhi metallo-beta-lactamase-1-producing Escherichia coli is a threat for leukemia patients in oncology and hematology departments. We conclude that the incidence of multidrug resistant pathogens has increased among patients with leukemia and is life threatening. [ABSTRACT FROM AUTHOR]

In recent years, the prevalence of carbapenem-resistant Pseudomonas aeruginosa isolates has become a worldwide concern. Rapid and accurate detection of carbapenemase-producing P. aeruginosa isolates is so important. The aim of this study was to evaluate the performance of the phenotypic methods such as Modified Hodge test (MHT), CarbaNP (CNPt), combined double-disk synergy test (CDDT), and carbapenem inactivation method (CIM) for rapid and accurate detection of clinical carbapenemase production of P. aeruginosa isolates. This study was performed on 97 P. aeruginosa strains, which were isolated from clinical samples in Hamadan hospitals, western Iran in 2017-2018. Antibiotic susceptibility testing was performed using disk diffusion and minimum inhibitory concentration (MIC) by E-test method. We evaluated the performance of MHT, CarbaNP, CDDT, and CIM tests in comparison to polymerase chain reaction (PCR) for the detection of carbapenemase-producing isolates. Additionally, the presence of carbapenem-resistant genes was investigated using the PCR method. Our findings showed that the highest resistance was to cefoxitin (94.8%). Moreover, among the carbapenem antibiotics, the highest resistance was to imipenem (49.4%). Among the 49 carbapenem-resistant isolates, 42 (85.7%) isolates were MIC positive. The results of phenotypic tests showed that CarbaNP, CIM, CDDT, and MHT tests were positive in (48/49, 97.95%), (46/49, 93.87%), (27/49, 57.44%), and (25/49, 53.19%) of isolates, respectively. CarbaNP and CIM tests showed high sensitivity, specificity, positive predictive values (PPV), and negative predictive values (NPV) compared to PCR in P. aeruginosa isolates. CarbaNP and CIM tests are highly sensitive and specific tests for identifying carbapenemase-producing P. aeruginosa isolates. [ABSTRACT FROM AUTHOR]

Aim: This study determined the genes encoding the binding and receiving factors of iron and microbial biofilm in E. coli strains isolated from mucosal samples of patients with colorectal cancer and inflammation of the colorectal compared to healthy people. Background: Colorectal cancer is one of the most important malignancies in recent years. Escherichia coli is the most important infectious agents associated with colorectal cancer that has numerous virulence factors such as iron uptake and adhesion factors included in the process of inflammation and colorectal cancer. Methods: Of the three healthy, inflammatory bowel diseases and colorectal cancer groups, 40 Escherichia coli strains isolated after confirmation by biochemical and molecular methods. After determining the isolates phylogroups, the frequency of genes was measured by PCR method. The biofilm formation of isolates was performed using Crystal Violet method. Results: In the determination of the bacteria phylogroups, the colorectal cancer isolates had a maximum incidence of phylogroups B2 and A. In the analysis of fimH, papA, papC, iutA, ireA and fyuA genes, the highest frequency was observed in these two phylogroups. The presence of ireA gene in bacterial isolates from three groups showed a significant difference (P value: 0.004). There was also no significant difference in biofilm productions in Escherichia coli strains isolated from the three groups. Conclusion: Unlike previous studies focusing solely on Escherichia coli toxins, we found that iron absorption and adhesion factors could be effective in developing inflammatory bowel diseases and colorectal cancer. It was also determined that biofilm formation is a specific characteristic of Escherichia coli isolated from the healthy colon. [ABSTRACT FROM AUTHOR]

Objectives: The purpose of this study was to determine the presence of IMP and OXA genes in clinical strains of Pseudomonas aeruginosa (P. aeruginosa) that are carriers of the ampC gene. Methods: In this study, 105 clinical isolates of P. aeruginosa were collected. Antibiotic resistance patterns were determined using the disk diffusion method. The strains carrying AmpC enzymes were characterized by a combination disk method. Multiplex-PCR was used to identify resistance and virulence genes, chi-square test was used to determine the relationship between variables. Results: Among 105 isolates of P. aeruginosa, the highest antibiotic resistance was to cefotaxime and aztreonam, and the least resistance was to colictin and ceftazidime. There were 49 isolates (46.66%) that showed an AmpC phenotype. In addition, the frequencies of the resistance genes were; OXA48 gene 85.2%, OXA199, 139 3.8%, OXA23 3.8%, OXA2 66.6%, OXA10 3.8%, OXA51 85.2% and OXA58 3.8%. The IMP27 gene was detected in 9 isolates (8.57%) and the IMP3.34 was detected in 11 isolates (10.47%). Other genes detected included; lasR (17.1%), lasB (18%) and lasA (26.6%). There was a significant relationship between virulence factors and the OX and IMP genes (p ≤ 0.05). Conclusion: The relationship between antibiotic resistance and virulence factors observed in this study could play an important role in outbreaks associated with P. aeruginosa infections. [ABSTRACT FROM AUTHOR]

The rapid increase of drug resistance and failure of available antibiotics to treat biofilm-associated infections is of great health concern. Accordingly, our study aimed to evaluate the synergistic antibacterial, biofilm inhibitory, and biofilm removal activities of melittin in combination with colistin, imipenem, and ciprofloxacin against multidrug-resistant (MDR) strong biofilm producer Acinetobacter baumannii isolates. The kinetics of biofilm formation were evaluated for the isolates for 144 h. Minimum inhibitory concentrations (MICs), minimum bactericidal concentrations (MBCs), minimum biofilm inhibitory concentrations (MBICs), and biofilm removal activities for melittin and combinations with antibiotics were determined. Inhibition of biofilm-associated protein (bap) expression by melittin was evaluated with real-time polymerase chain reaction (PCR). Field emission scanning electron microscopy (FE-SEM) was used to visualize the effect of synergism on the inhibition of biofilm production. The geometric means of the fractional inhibitory concentration index (FICi) for melittin–colistin, melittin–imipenem, and melittin–ciprofloxacin combinations were calculated as 0.31, 0.24, and 0.94, respectively. Comparing the geometric means of the removal activity for melittin, colistin, imipenem, and combinations of them in both 6 and 24 h showed a significant difference between the groups (p-value < 0.05). Exposure to melittin induced a statistically significant downregulation of bap mRNA levels in all isolates at sub-MIC doses. Analysis of the FE-SEM results demonstrated that the synergism of melittin–colistin at 0.125–0.25 μg inhibited biofilm formation completely. In conclusion, our findings indicate that melittin possesses considerable potential for use in combination with colistin and imipenem to treat infections caused by MDR strong biofilm producer A. baumannii isolates. [ABSTRACT FROM AUTHOR]

Objectives: We sought to evaluate the relationship between the prevalence of antibiotics resistance and virulence factors genes in methicillin-resistant Staphylococcal aureus (MRSA) and methicillin-sensitive S. aureus (MSSA) strains from clinical samples taken in west Iran. Methods: We performed a cross-sectional study using 100 MRSA and 100 MSSA samples isolated from clinical specimens. We used biochemical methods to identify the isolates, which were confirmed by the polymerase chain reaction (PCR) assay. Antibiotic susceptibility testing was performed using disk diffusion. PCR detected the presence of virulence factors, including enterotoxin genes, toxic shock syndrome toxin-1 (TSST-1), and exfoliative toxin. Results: The majority of MRSA isolates exhibited a high level of resistance to common antibiotics and susceptible to vancomycin, while most MSSA isolates were also resistant to erythromycin and ciprofloxacin. The prevalence of Staphylococcal enterotoxins (SEs) were reported 147 (73.5%). Among 100 MRSA samples, 92 (92.0%) harbored SAggenes. The most frequent toxin gene was sea (45.0%) followed by sec (39.0%). Among 100 MSSA isolates, 89 (89.0%) harbored SAg genes and the most prevalent genes were sea (42.0%), sek (38.0%), sec (35.0%,), and TSST-1 (10.0%). The prevalence of TSST-1 and exfoliative toxin genes in MRSA samples were 12 (12.0%). The association of SAg genes with MRSA and MSSA isolates showed a high prevalence of enterotoxin seq,seg, and sei in MRSA than MSSA with a statistically significant difference (p < 0.050). Conclusions: The prevalence of MRSA and the association of pathogenic agents with antibiotics resistance genes can lead to the emergence of strains with higher pathogenicity and less susceptibility. [ABSTRACT FROM AUTHOR]

Acinetobacter baumannii potential to form biofilm and exhibit multiple antibiotic resistances may be responsible in its survival in hospital environment. Accordingly, our study was aimed to determine the correlation between ability of biofilm formation and the frequency of biofilm related genes with antibiotic resistance phenotypes, and also the categorization of their patterns in clinical and environmental isolates. A total of 75 clinical and 32 environmental strains of the A. baumannii were collected and identified via API 20NE. Antibiotic susceptibility was evaluated by disk diffusion and microdilution broth methods. Biofilm formation assay was performed by microtiter plate method. OXA types and biofilm related genes including Bla OXA-51 , Bla OXA-23 , Bla OXA-24 , Bla OXA-58 , bap, bla PER-1 , and ompA were amplified by PCR. The rate of MDR A. baumannii in clinical isolates (100%) was higher than environmental (81.2%) isolates (p < 0.05). Among 10 antibiotypes, the predominant resistance pattern in clinical and environmental isolates was antibiotypes I (85.3 and 78.1%, respectively). Analysis of the frequency of bla OXA -23 gene revealed a statistically significant difference between clinical (85.3%) and environmental (68.7%) isolates (p < 0.05). The prevalence of strong biofilm producers in clinical and environmental isolates were 31.2%–58.7%, respectively. In the clinical and environmental isolates, the frequencies of ompA, bla RER-1 and bap genes were 100%, 53.3%, 82.7% and 100%, 37.5%, 84.4% respectively. Statistical analysis revealed a significant correlation between the frequency of MDR isolates and biofilm formation ability (p = 0.008). The high frequency of antibiotype I would be indicated that an outbreak has been happened earlier and an endemic strain is currently being settled in the hospital environment. It would be suggested that if there was no difference in the frequency of pattern I and biofilm formation ability between clinical and environmental isolates, it is a critical point representing the higher risk of bacterial transmission from environment to the patients. The resulting data would be assisted in the improvement of disinfection strategies to better control of nosocomial infections. One dominant resistance pattern has shown among clinical and environmental isolates. The frequency of blaOXA-23 had significant difference between clinical and environmental isolates. The presence of bap gene in the A. baumannii isolates was associated with biofilm formation. There was a significant correlation between multiple drug resistance and biofilm formation. The clinical isolates had a higher ability to form strong biofilms compared to the environmental samples. [ABSTRACT FROM AUTHOR]

Background and Aims: Staphylococcus aureus adhesion factors can reinforce the pathogenicity of the bacteria. The aim of this study was to identify adhesion factors among clinical isolates of methicillin-resistant S. aureus and determine the association between these factors and antibiotic resistance patterns. Materials and Methods: In an analytical study from October 2016 to April 2017, 302 clinical isolates of S. aureus were confirmed by biochemical tests. Methicillin-resistant strains were determined by phenotypic methods. Multiplex PCR method was used to identify adhesion factors. In this way, bbp, cna, eno and ebpS genes were identified among different isolates. Results: A total of 302 clinical isolates of S. aureus were isolated from different clinical samples including wound, blood, urine, trachea, catheter, swabs. Of 302 isolates, 123 were methicillin resistant and 73.53% and 75.7% of the isolates were resistant to erythromycin and penicillin, respectively. The incidence of resistance genes among among methicillin resistant S. aureus isolates were as follows: bbp (10 isolates: 6.89%), cna (6 isolates: 13.4%), eno (28 isolates: 19.31%) and ebpS (19 isolates: 13.1%),. There was a significant correlation between the antibiotic resistance patterns and the frequency of adhesion factors (P≤0.05). Conclusions: According to the results, there was a significant correlation between adhesion factors and antibiotic resistance among methicillin-resistant isolates of S. aureus. [ABSTRACT FROM AUTHOR]

Streptococcus agalactiae is acommensalorganism, but it may cause infection in susceptible hosts. The aim of this study was to evaluate PCR assay compared with conventional culture method for direct detection of Streptococcus agalactiae. Total of 203 paired low vaginal swabs were collected from women at 35-37 weeks of pregnancy from June 2013 through February 2014 for detection of Streptococcus agalactiae using PCR assay targeting 16S rRNA, cfb, scpB, and atr genes and culture method following broth enrichment. The results were recorded and evaluated for determining of sensitivity, specificity, positive and negative predictive values of PCR assaycompared with culture method. Prevalence of Streptococcus agalactiae was determined as 7.39% (n=15) using culture method; 19.70% (n=40) by PCR targeting 16S rRNA gene; 18.23% (n=37) by targeting atr gene; 17.24% (n=35) by cfb gene; and 8.87% (n=18) by scpB gene. Generally, a total of 49 specimens were considered true positive (27 samples by PCR assay using the four genes in sum, 4 samples only by atr gene PCR, 3 samples only by cfb gene PCR, 2 samples only by culture method, and 13 samples by PCR assay and culture method in common) and prevalence of Streptococcus agalactiae determined 24.14% in Hamadan. The current data demonstrated that performing only culture method for detecting GBS from pregnant women leads to missed false negative carrier individuals. Thus, it is recommended that both the PCR assay and conventional culture method to be performed in order to detect Streptococcus agalactiae. [ABSTRACT FROM AUTHOR]

Background: Staphylococcus aureus is the most important cause of nosocomial infections acquired in the community. Protein A is a major component of Staphylococcus aureus cell wall. In analysis of the nucleotide sequence Protein A encoding spa, locus x consists of 24 base pairs which repeat with high polymorphism. In this study, the spa gene of Staphylococcus aureus isolated from clinical specimens were obtained from patients admitted to the hospital and healthy carriers. Methods: In a cross-sectional study, a total of 200 samples were collected. One hundred fifty samples were obtained from hospitalized patients and 50 samples obtained from staff nasal swabs in Hamadan University Hospitals from October 2013 to August 2014. Disk diffusion antibiotic susceptibility tests performed. The antibiotics studied were Vancomycin (30 μg), Cefoxitin (15 μg) Gentamicin (10 μg), Tetracycline (30 μg), Trimethoprim/sulfamethoxazole (25 μg), Ciprofloxacin (5 μg), Erythromycin (15 μg), Clindamycin (2 μg), Rifampin (5 μg). The tests performed according to the guidelines of clinical and laboratory standards institute (CLSI). It also detect the mecA gene of Methicillin-resistant Staphylococcus aureus strains (MRSA) and genes spa which encodes the protein A by polymerase chain reaction (PCR). The PCR products using a polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method with enzyme Rsa I (Afa I) were prepared. Results: This methicillin-resistant Staphylococcus aureus strain (MRSA) had the highest sensitivity and resistance to ciprofloxacin and clindamycin. Totally, 8 amplicon with different sizes for the spa gene were identified. A total of 9 patterns polymerase chain reaction- restriction fragment length polymorphism (PCR-RFLP) were found. Some of these patterns between Staphylococcus aureus isolated from clinical specimens and nasal carriers were common. Conclusion: There is a similar pattern of spa gene among patients admitted to the hospital and staff, according to our findings. Analysis of the patterns can reduced transmission of infection in both hospital staff and patients. Also it can help the physicians for correct management of infections. [ABSTRACT FROM AUTHOR]

BACKGROUND: Pseudomonas aeruginosa is one of the most common nosocomial pathogens with high mortality rates. Organisms such as Pseudomonas aeruginosa have the ability to develop high level MDR (Multi drug resistance). The MexAB-OprM system is one of the largest multi-drug resistant efflux pumps with high levels of expression and the first finding of the RND (Resistance-nodulation-division) family in P. aeruginosa. AIM: For better understanding of the antibiotic resistance mechanism in P. aeruginosa, this study was conducted to determine the expression of the genes encoding these efflux pumps in 100 strains of P. aeruginosa isolated from patients admitted to various hospitals in Hamadan using quantitative Real-Time PCR (qRT-PCR). METHODS: This study examined 100 strains of P. aeruginosa isolated from patients admitted to various hospitals in Hamadan. Then, 31 samples were selected based on collected specimen type and their antibiotic susceptibility pattern; i.e., the samples with reduced susceptibility to antibiotics, particularly carbapenems. Antibiotic disk diffusion method was performed for aminoglycoside, quinolone and carbapenem. Furthermore, MIC method was performed for ciprofloxacin, gentamicin and imipenem. Finally, qRT-PCR was used for determining the efflux pump genes expression. RESULTS: Among eight selected antibiotics, the greatest resistance was to levofloxacin (61.2%, n = 19) and the lowest one to imipenem (9.6%, n = 3). All isolates (100%, n = 31) exhibited efflux pump MexAB-OprM genes but different expression was observed in different strains. The result of gene expression indicated that significant differences in expression of MexR (P value = 0.003), OprD (P value < 0.001), and MexB (P value = 0.026) genes. In addition, there was high level of MexR gene expression in bacteria that leads to reduced expression of MexA, MexB, and OprM. The OprD gene was presented in all strains but different expression has been observed. CONCLUSION: Identifying the bacterial resistance mechanisms is very complicated. Although efflux pump MexAB-OprM plays an important role in antibiotic resistance in P. aeruginosa, because of acting the efflux pumps on antibiotics in a non-specific manner, it is elusive to consider or describe an antibiotic resistance based on the presence or absence of an efflux pump. [ABSTRACT FROM AUTHOR]

Objective: Sepsis is a systemic inflammatory response associated with high mortality rates in the clinical setting. A multiplex endpoint polymerase chain reaction (PCR) based assay for rapid detection of enterobacteriaceae involved in septicemia, which included Internal Control (IC) and 16S rDNA, is presented here. To develop a panel of primers for DNA fragments of 16S rDNA, enterobacteriaceae, IC, and evaluate analytical sensitivity and specificity of the test Materials and Methods: Primers for amplification of enterobacteriaceae, IC, and16S rDNA were designed, and then PCR was performed. Minimal analytical sensitivity was determined by cloning and colony PCR, and specificity was tested on the basis of their respective standard strains. This study is a cross-sectional Model. Results: Our results showed the rpoB gene as the most promising target for detection of enterobacteriaceae by PCR amplification. Specificity and sensitivity of endpoint PCR were 100%, 100%, and 100%, and 10, 1, and 100 copies/reaction for enterobacteriaceae, IC, and 16S rDNA, respectively. Conclusion: The molecular panel presented offers the advantage of an easy, reliable, and cost-effective system when compared to other molecular detection methods. However, further evaluation is needed. Our assay holds promising for more rapid pathogens related in clinical sepsis. [ABSTRACT FROM AUTHOR]

Backgrand: Pseudomonas aeruginosa is a common cause of nosocomial infections. It exhibits innate resistance to a wide range of antibiotics. This study was performed to determine clonal characteristic of P. aeruginosa isolated from clinical specimens, hospital means, and hospital personnel by PCR- ribotyping patterns. Methods: a total of 104 P. aeruginosa were isolated from clinical and environmental samples (59 clinical, 45 hospital means and hospital personnel). P. aeruginosa was identified by standard bacteriological methods, mucoid colony morphotypes, and antibiotic resistance rate. The genomes of isolates were extracted and all considered species were confirmed by 16S rDNA- based PCR assay. Then all isolates were genotyped by the 16S - 23SrDNA and Hinf1 restriction enzyme technique. Results: antibacterial sensitivity pattern of isolates showed clinical and environmental specimens were approximately identical (high antibiotic resistance to Ceftazidime and low antibiotic resistance to Amikacin). Colony morphotypes of specimens revealed that mucoid type of clinical isolates were more than that of environmental isolates. Among clinical and environmental strains P1; (570 bp) was the most prevalence pattern. Conclusions: antibiotic resistance, phenotypic characterization, and PCR- ribotyping pattern showed there is clonal relatedness between clinical and environmental isolates and environment could be a main reservoir for P. aeruginosa infections in hospital. [ABSTRACT FROM AUTHOR]

Brucellosis is a widespread zoonotic disease causing considerable economic and public health problems. Despite animal vaccination, brucellosis remains endemic in some areas such as Iran, especially in the western Iranian province of Hamadan. We sought to detect some of the most common virulence-associated genes in Brucella isolated from human blood cultures to determine the prevalence of some virulence genes among Brucella isolates. Fifty-seven isolates were studied from patients with a clinical diagnosis of brucellosis who referred to the Infectious Diseases Ward of Sina Hospital in Hamadan Province, Iran, between April 2013 and July 2014. Blood samples were collected for the diagnosis of brucellosis using the BACTEC blood culture system. All of these isolates were confirmed by the bcsp31 Brucella-specific gene. We detected 11 virulence-associated genes of Brucella, namely cβg, virB, znuA, ure, bvfA, omp25, omp31, wbkA, mviN, manA, and manB, which are important for the pathogenesis of this bacterium in the intracellular environment by multiplex PCR. Totally, 149 patients with a clinical diagnosis of brucellosis were enrolled in this study. Fifty-seven (38.3%) patients had positive blood cultures. On biochemical and molecular testing, all of the isolates were Brucella melitensis. Ten of the virulence genes were detected among all of the 57 isolates, but the bvf gene was detected in 53 (93%) isolates. The high prevalence of virulence-associated genes among the Brucella isolates detected in Hamadan Province, Iran, underscores the pathogenicity of this bacterium in this region. [ABSTRACT FROM AUTHOR]