Background & Objectives: Although isoniazid is the most efficient in killing the tuberculosis bacilli, resistance to this drug also develops most readily. Mutations in katG, inhA and ahpC are responsible for isoniazid resistance in a large proportion of tuberculosis cases. The frequency of these mutations varies with population samples, however. This study provided the first molecular characterization of Isoniazid-resistance in M. Tuberculosis strains that is widely applicable for rapid drug resistance detection. Materials and Methods: The study was a descriptive and analytical and the presence of mutations in specific regions of the katG, inhA and ahpC genes was analyzed in 32 M. tuberculosis Isoniazid-resistant strains in Isfahan and Tehran. To determine the mutations in codon 315 of katG, PCR-RFLP technique was performed. In this way, 355 bp PCR products were digested by MspI and MspA1I. Mutations in inhA and ahpC genes were detected by sequencing. Results : The frequency of mutations in the katG 315(Ser→Thr), inhA and ahpC were detected in 71.9%, 18.9% and 6.2% of the 32 Isoniazid resistant isolates, respectively. Mutation was not found in one of the isolates. Conclusion : The PCR-RFLP with MspI, being able to detect katG Ser315Thr substitution, can identify more Isoniazid-resistant strains with mutations at codon 315 in the katG. Elucidation of the molecular basis of Isoniazid resistance in M. tuberculosis has led to the development of different genotypic approaches for the rapid detection of Isoniazid resistance in clinical isolates. The results also suggest that the detection of the Ser315Thr in the katG gene may be used as a rapid screening method for identifying Isoniazid-resistant clinical M.tuberculosis isolates recovered from Isfahan and Tehran tuberculosis centers.
Antibiotic resistant Acinetobacter baumannii has emerged as one of the most problematic hospital acquired pathogens around the world. This study was designed to investigate the presence of antibiotic resistant A. baumannii in various hospital environments.Air, water and inanimate surface samples were taken in different wards of four hospitals and analyzed for the presence of A. baumannii. Confirmed A. baumannii isolates were analyzed for antimicrobial susceptibility and also screened for the presence of three most common OXA- type carbapenemase-encoding genes.A. baumannii was detected in 11% (7/64) of air samples with the highest recovery in intensive care units (ICUs). A. baumannii was also detected in 17% (7/42) and 2% (1/42) of surface and water samples, respectively. A total of 40 A. baumannii isolates were recovered and analysis of antimicrobial susceptibility showed the highest resistance towards ceftazidime (92.5%, 37/40). 85% (34/40) and 80% (32/40) of the isolates were also resistant to imipenem and gentamicin, respectively. Resistance genes analysis showed that 77.5% (31/40) strains contained OXA-23 and 5% (2/40) strains contained OXA-24, but OXA-58 was not detected in any of the strains.Detection of antibiotic resistant A. baumannii in various samples revealed that hospital environments could act as a potential source for transmission of A. baumannii infections especially in ICUs. These results emphasize the importance of early detection and implementation of control measures to prevent the spread of A. baumannii in hospital environments.
Mycobacteria are a heterogeneous group of bacteria in terms of their genotypic features and disease association. The accurate and rapid detection of genus Mycobacterium andMycobacterium tuberculosis complex (MTC) isolates from primary culture is important due to timely and appropriate antibiotic therapy. In this study, we developed a novel multiplex real time PCR for rapid identification of Mycobacteria from other genus of bacteria and Mycobacterium tuberculosis complex from non-tuberculosis Mycobacteria(NTMs). Genetic targets for primer designing and identification of genus Mycobacteriumand Mycobacterium tuberculosis complex (MTC) included the 16SrDNA and 16S-23S internal transcribed spacer (ITS). Multiplex real time PCR was setup with reference toMycobacterium strains. Results of real time PCR were analyzed with melting curves and determined melting temperature of genus Mycobacterium and Mycobacterium tuberculosis complex. For the assessment of the sensitivity and specificity of technique, fifty three clinical isolates of Mycobacteria were collected from different Tuberculousis Centers in Iran. Multiplex real time PCR results were compared by PCR-based sequencing method with amplification and sequencing of 16S–23S ribosomal RNA internal transcribed spacer (ITS) for identification of Mycobacterium species. Of 53 isolates, 52 (98%) isolates identified as genus Mycobacterium and 34 isolates as MTC (65%). Sensitivity and specificity of multiplex real time PCR were 96%, 100% for genusMycobacterium and 100%, 100% for Mycobacterium tuberculosis complex (MTC), respectively. The multiplex real time PCR with melting curve analysis in the present study is a reliable and suitable method for rapid and accurate identification of genusMycobacterium and Mycobacterium tuberculosis complex.
Key words: Mycobacterium tuberculosis complex, multiplex real time PCR, rapid identification
Background: Pseudomonas aeruginosa, is the most common pathogen in patients with cystic fibrosis (CF) that shows various resistance to antibiotics, acquires mucoidity and multiple genotypes. This survey was performed to study phenotypic and genotypic variations among P. aeruginosa isolates in CF patients at Alzahra Hospital in Isfahan, Iran. Materials and Methods: The isolates of Pseudomonas aeruginosa from CF patients at Alzahra Hospital was identified by appropriate biochemical and microscopic tests, then performed antibiotic resistance tests and mucoid colony morphotyping. The genum of isolates extracted and confirmed on 16S rDNA-based PCR assay and typed on 16S rDNA-23SrDNA spacer, restricted with Hinf1 restriction enzyme. Results: P. aeruginosa was isolated from 21 of the 59 CF patients (35.5%), Out of 21 isolates 9 (42.8%) strains were revealed mucoid morphotype. 81.8% isolates of mucoid strains were resistance to at least one of four antibiotics (GM, AN, PIP and CP). Most of the isolates (86%) showed resistance to ceftazidime. Ribotyping revealed two patterns (P1, P5).Conclusion: The isolates of P. aeruginosa showed meaningful difference between drug resistance to antibiotics. The majority of P. aeruginosa isolated from CF patients showed pattern1 of PCR-Ribotyping.
Proteus mirabilis (P. mirabilis) is a frequent cause of catheter-associated urinary tract infections. This study aims to investigate the anti-infective effect of Alhagi maurorum extract (AME), the traditional medicinal plant in the middle east, on the biofilm-forming P. mirabilis isolates. Hydroalcoholic extract and oil of A. maurorum were characterized by HPLC and GC-MS. The antiproliferative, anti-biofilm, and bactericidal activity of AME at various concentrations were assessed by turbidity, crystal violet binding, and agar well diffusion assays, respectively. The AME's effect on adhesion and quorum sensing (QS) were investigated by in vitro adhesion assay on cell culture and agar overlay assay using Janthinobacterium lividum (ATCC 12472) as a biosensor strain. In addition, the expression level of selected genes involved in QS and biofilm regulation were determined by quantitative Real-Time PCR. Furthermore, the bladder phantom model was created to evaluate the assays and investigate the catheter's calcium deposition. The most effective chemical compounds found in AME were tamarixetin, quercetin, and trans-anethole. Although AME did not inhibit swarming motility, it reduced biofilm production and exerted a concentration-dependent anti-adhesive and anti-QS activity against P. mirabilis. AME also downregulated the expression level of selected genes involved in biofilm formation and QS. This study showed that AME as a natural compound reduced biofilm formation of P. mirabilis by targeting virulence factor genes, quorum sensing, and other strategies that include preventing the adhesion of P. mirabilis to the cells. The results suggest that A. maurorum extract might have the potential to be considered for preventing UTIs caused by P. mirabilis.
Introduction. Viral infections are increasingly an important cause of central nervous system (CNS) complications.Hypothesis/Gap Statement. There is no comprehensive insight about CNS infections due to viral agents among Iranian children.Aim. This study aimed to investigate the viral aetiology, clinical and epidemiological profile of children with acute infections of the CNS.Methodology. A prospective study was conducted on children at the referral hospital in Isfahan, Iran, from June 2019 to June 2020. A multiplex PCR assay was used to detect the viral causative agent in cerebrospinal fluid and throat/rectal swab samples.Results. Among 103 patients with eligible criteria, a confirmed or probable viral aetiology was detected in 41 (39.8 %) patients, including enteroviruses - 56.1 %, herpes simplex virus 1/2 (HSV-1/2) - 31.7 %, Epstein-Barr virus - 17.1 %, varicella-zoster virus (VZV) - 9.7 %, influenza A virus (H1N1) -4.9 % and mumps - 2.4 %. There was a higher proportion of PCR-positive samples in infants than in other age groups. Encephalitis and meningoencephalitis were diagnosed in 68.3 % (28/41) and 22 % (9/41) PCR-positive cases, respectively.Conclusion. The findings of this research provide insights into the clinical and viral aetiological patterns of acute CNS infections in Iran, and the importance of molecular methods to identify CNS viruses. HSV and VZV were identified as important causes of encephalitis in young children.
Carriage of S. aureus in the anterior nares seems to play a significant role in the pathogenesis of infection. This study aimed to determine the molecular characteristics and antibiotic susceptibility pattern of S. aureus isolates obtained from the nasal carriage of health care workers (HCWs). This study was performed during July 2014 to July 2015 at three tertiary care hospitals. Nasal samples were collected from the nasal cavity of HCWs. Standard microbiological methods were used for identification of S. aureus isolates. Antibiotic susceptibility pattern was determined by the disc diffusion method. Determination of SCCmec typing and virulence genes was performed by the PCR method. From the isolates of 340 nasal swab samples of HCWs, 65 S. aureus strains (19%) including 22 (33.8%) MRSA were isolated. The highest sensitivity for MRSA isolates was towards vancomycin and rifampicin, each with 90.9%. Overall, 17% (11/65) and 92.3% (60/65) of S. aureus isolates were positive for pvl and hla genes, respectively. The rates of SCCmec types II, III, IV, V and I among MRSA isolates were 36.4 %, 22.7 %, 22.7 %, 9.1% and 4.5% respectively. The results of the present study indicate that S. aureus nasal carriage with potential virulence ability still remains a significant healthcare problem, especially in hospital environments.
Background: Ethambutol (EMB) is the first-line drug used for the treatment of tuberculosis. Recent reports on the EMB-resistant isolates of Mycobacterium tuberculosis in different geographical regions of the world have revealed the EMB resistance patterns of M. tuberculosis and mutations in the embB gene. Objectives: In this study, we determined the emb locus in sensitive and resistant Iranian M. tuberculosis isolates using two effective methods for the detection of point mutation, i.e., single-strand conformational polymorphism (SSCP) and direct sequencing. Methods: Thirty-two M. tuberculosis isolates from the Isfahan tuberculosis center were characterized by conventional methods and specific amplification of the regions of difference (rd) gene and internal transcribed spacer (its) gene. Observing standard operational procedures, EMB susceptibility tests were performed on the LJ medium using the proportion method with 2, 5, and 10 μg/mL concentrations. PCR-SSCP and direct sequencing were used to detect different kinds of mutation in the embB gene with precision. Results: In a total of 32 isolates, two isolates (6.25%) were found to be resistant to EMB in 2, 5, and 10 μg/mL concentrations. Single-strand conformational polymorphism showed altered mobility with triple bands in the resistant isolates and double bands in the sensitive isolates. In the two EMB-resistant cases, mutation was found to occur codons 309and 299. Conclusions: We concluded from the results that the frequency of EMB-resistant M. tuberculosis cases in Iran is lower than that of many other regions. The PCR-SSCP technique can separate resistant isolates from sensitive isolates. The sequencing results of this study showed mutation in codons 309 and 299 of the embB gene. In none of the resistant isolates, mutation was observed in codon 306. Further studies are required to determine other point mutations and analyze other genetic loci associated with EMB resistance in M. tuberculosis isolates in Iran.
'%3e%3cg id='e'%3e%3cg id='c' fill='none' stroke='%23fff' stroke-width='2'%3e%3cpath id='b' d='M0 1h26M1 10V5h8v4h8V5h-5M4 9h2m20 0h-5V5h8m0-5v9h8V0m-4 0v9' transform='scale(1.4)'/%3e%3cpath id='a' d='M0 7h9m1 0h9' transform='scale(2.8)'/%3e%3cuse xlink:href='%23a' y='120'/%3e%3cuse xlink:href='%23b' y='145.2'/%3e%3c/g%3e%3cg id='d'%3e%3cuse xlink:href='%23c' x='56'/%3e%3cuse xlink:href='%23c' x='112'/%3e%3cuse xlink:href='%23c' x='168'/%3e%3c/g%3e%3c/g%3e%3cuse xlink:href='%23d' x='168'/%3e%3cuse xlink:href='%23e' x='392'/%3e%3c/g%3e%3cg fill='%23da0000' transform='matrix(45 0 0 45 315 180)'%3e%3cg id='f'%3e%3cpath d='M-.548.836A.912.912 0 0 0 .329-.722 1 1 0 0 1-.548.836'/%3e%3cpath d='M.618.661A.764.764 0 0 0 .422-.74 1 1 0 0 1 .618.661M0 1l-.05-1L0-.787a.31.31 0 0 0 .118.099V-.1l-.04.993zM-.02-.85 0-.831a.144.144 0 0 0 .252-.137A.136.136 0 0 1 0-.925'/%3e%3c/g%3e%3cuse xlink:href='%23f' transform='scale(-1 1)'/%3e%3c/g%3e%3c/svg%3e)