Rheumatoid arthritis mediator CD18 expression by Staphylococcus aureus superantigen C in rats.

BACKGROUND AND OBJECTIVES: Microbial superantigens have been reported in the blood and synovial fluid of rheumatoid arthritis patients, raising the question of whether the presence of these superantigens could provoke the induction of inflammatory biomarkers expression or not. The purpose of this study was to examine the Staphylococcus aureus superantigen C on CD18 expression. MATERIALS AND METHODS: The superantigen C was purified by ultrafiltration. Immunoblotting was performed using a specific antibody. Also, 50 micrograms of superantigens (toxin) were injected intraperitoneally and intra-articularly into separate rat groups. Blood was collected and RNA extracted. Then, the cDNA was synthesized. The expression of CD18 marker was evaluated using RT-real-time PCR, and the results were descriptively analyzed. RESULTS: The results of this study revealed that 50 µg of toxin, injected intra-articularly and intraperitoneally, showed the surplus expression of the marker CD18 in the blood of rats after 20 days. By this method, the expression of the marker CD18 was significantly different between rats that received the superantigen intra-articularly and intraperitoneally (2.10; 2.3 and 3.3 folds) and the controls (P≤ 0.05). CONCLUSION: The results indicated that the presence of Staphylococcal of superantigen C in the body of rats has enhanced the expression of the CD18 inflammatory marker more than 3 times. This valuable finding is an introduction to further research and could provide new methods to prevent and control inflammatory diseases, including rheumatoid arthritis.

Diversity of virulence genes in Brucella melitensis and Brucella abortus detected from patients with rheumatoid arthritis.

The presence of Brucella melitensis and Brucella abortus genomes were investigated in the synovial fluid (SF) samples from 90 patients with rheumatoid arthritis (RA). DNA extraction and PCR assay were performed for simultaneous identification and discrimination of B. melitensis and B. abortus from the SF using three specific primers. After gel electrophoresis, the PCR products were confirmed by DNA sequencing. The cbg, omp31, manA, virB, and znuA virulence genes typing were performed by multiplex-PCR. Of the 90 samples, 14 were positive for B. melitensis (n = 9; 10%) and B. abortus (n = 5; 5.5%). The virulotyping of positive samples revealed the presence of all five virulence genes in B. melitensis. The virB, cbg, and om31 were detected in all five samples of B. abortus. In addition, zhuA and manA were detected in three (60%) and four (80%) samples, respectively, of the B. abortus-positive samples. Moreover, a total of 94.2% and 89.2% of the 14 positive samples were also found positive for manA and znuA, respectively. Our findings revealed that the Brucella spp. genomes can be detected in the SF of RA patients by the PCR-based method. We thus suggest that physicians should consider the Brucella spp. as indicators of potential RA for the timely diagnosis and treatment of RA.

Bacteriological Aspects of Hand Washing: A Key for Health Promotion and Infections Control.

The aim of this review is to show the historical aspects of hands washing for healthy life and explains how can reduce the transmission of community-acquired infectious agents by healthcare workers and patients. This review article is prepared based on available database. The key words used were hands washing, risk assessment, hands hygiene, bacterial flora, contamination, infection, nosocomial, tap water, sanitizer, bacterial resistance, hands bacterial flora, washing methods, antiseptics, healthcare workers, healthcare personnel, from PubMed, ScienceDirect, Embase, Scopus, Web of Sciences, and Google Scholar. Data were descriptively analyzed. The insistence on hand washing has a history of 1400 years. The research results indicate that the bacteria released from the female washed hands in wet and dry condition was lower than from the male's hands with a significance level (3 CFU vs. 8 CFU; confidence interval 95%, P ≤ 0.001). The valuable results of the study indicated that released amount of bacterial flora from wet hands is more than 10 times in compared to dry hands. In addition, established monitoring systems for washing hands before and after patient's manipulation as well as after toilet were dominant indices to prevent the transfer of infectious agents to the patients. Increasing awareness and belief of the healthcare workers have shown an important role by about 30% reduction in the transfection. Hand washing could reduce the episodes of transmission of infectious agents in both community and healthcare settings. However, hand washing is an important key factor to prevent transmission of infectious agents to patients. There is no standard method for measuring compliance. Thus, permanent monitoring of hand washing to reduce the transmission of infections is crucial. Finally, the personnel must believe that hand washing is an inevitable approach to infection control.

Carriage rates of Neisseria meningitidis serogroups: determination among freshmen conscripts before vaccination.

BACKGROUND AND OBJECTIVES: Neisseria meningitidis is transmitted from person-to-person. Thus, close contact with a healthy carrier can facilitate the spread of the bacteria and lead to life-threatening meningococcal disease. The aim of this study was to identify oropharyngeal carriers of N. meningitidis in volunteers preparing for military service before vaccination. MATERIALS AND METHODS: In a cross-sectional study, 226 volunteers entering military service were referred to the Shemiranat Health Center for meningococcal vaccination and assayed. Before vaccination, the participants underwent sampling of the throat using separate swabs. Thayer-Martin Agar medium and microbiological standard methods were used for culture and isolation of the organisms. The bacterial isolates were subjected to DNA extraction and polymerase chain reaction. The obtained data were descriptively analyzed. RESULTS: Out of the 226 (100%) young volunteers, only 18 (8%) yielded Gram-negative diplococci. The results showed the presence of N. meningitidis (carriage rate: 8%) in their oropharyngeal regions. The isolated serogroups were C, A, Y, W-135, and X with frequencies of 50, 22.2, 16.6, 5.5, and 5.5, respectively. DISCUSSION: This study showed that the carriage rate in young volunteers for military service is around 8% before vaccination. Although the rates for serogroups A and C were dominant, the existence of serogroups Y and W indicate the necessary revision of the A/C vaccine. More research is needed to determine serogroup diversity and decrease the risk of meningococcal disease in individual groups.

Staphylococcal Enterotoxin A Detection from Rheumatoid Arthritis Patients' Blood and Synovial Fluid.

INTRODUCTION: Direct detection of microbial super antigens in synovial fluid of patients with rheumatoid arthritis may be able to guide to the design of cost-effective therapies. The purpose of this study was to assess the existence of Staphylococcal enterotoxin A (superantigen A) in the synovial fluid of patients with RA by the PCR and ELISA methods. METHODS: This experimental study was conducted on the synovial fluid of 103 RA patients from Baqiyatallah University of Medical Sciences' Rheumatology Clinic in Tehran, Iran in 2011-2014. Bacterial cultures, polymerase chain reaction with specific primer pairs and enzyme-linked immunosorbent assay (ELISA) methods were used. The PCR products were subjected to sequence as a confirmatory molecular method results. The data were descriptively analyzed by SPSS Version 19. RESULTS: The bacteriological study result indicated that, in four cases (3.8%) of the patients, bacterial strains were isolated. The result of PCR molecular method for staphylococcal enterotoxin A gene showed that, 42 of the patients (40.7%) tested positive for the ent A gene. The results of ELISA were positive for staphylococcal enterotoxin A (superantigen A) in 51 cases (49.51%) of the patients' synovial fluids. The results indicated that the possibility of detecting superantigen A in the SF of RA patients, but the origin of the enterotoxin A gene remained unknown. CONCLUSIONS: The findings of this study may be able to alter the actual theory on the pathogenesis, diagnosis, and treatment of RA patients. In addition, the results have shown the probability of an endogenous origin for the involved superantigen A in RA patients' synovial fluids.

Detection of Epstein-Barr virus in synovial fluid of rheumatoid arthritis patients.

INTRODUCTION: Rheumatoid arthritis (RA) is one of the most common chronic inflammatory disorders. Genes and environmental factors contribute to RA. Epstein-Barr Virus (EBV) has been considered as one the RA pathogeneses. The aim of this study was to detect of the EBV genome in patients with RA. METHODS: In this cross-sectional study, 50 samples of synovial fluid were obtained from patients with RA from 2010-2012. Using a standard of the EBV genome and EBNA-1-specific primers, the method of PCR was set up. Then, all of the samples of synovial fluids separately were subjected to DNA extraction and Polymerase Chain Reaction (PCR) amplification. Data were analyzed using SPSS version 18.0. The statistical analysis was performed by the t-test. RESULTS: The demographic and laboratory characteristic assay revealed that the mean age of patients was 49, and the patients were 60% males and 40% females. In addition, in all cases, the mean rheumatoid factor (RF) levels of the patients were below the normal level. The results of this study showed that the PCR was able to detect EBV DNA in > 60% of the cases. CONCLUSION: The results of this study indicated that EBV was frequently detected in the synovial fluid of RA patients. Thus, EBV may be a strong candidate that can act at several levels of the pathophysiology of RA. However, these findings also indicated that EBV may play a role in the pathogenesis of RA. However, the possible relationship between RA and EBV must be determined by further research.

Simultaneous Detection of Mycoplasma pneumoniae, Mycoplasma hominis and Mycoplasma arthritidis in Synovial Fluid of Patients with Rheumatoid Arthritis by Multiplex PCR.

BACKGROUND: It has been recognized that infectious agents, such as different bacteria and viruses, may play a role in the developing of rheumatoid arthritis (RA). Recently, the mycoplasma species has been implicated in the pathogenesis of RA. AIM: The aim of this study was to design a multiplex PCR for rapid and simultaneous detection of Mycoplasma pneumoniae, Mycoplasma hominis, and Mycoplasma arthritidis in the synovial fluid of patients with rheumatoid arthritis (RA). METHODS: A total of 131 synovial fluid (SF) samples from patients with RA were assayed. Mycoplasma pneumoniae (ATCC: 29342), M. hominis (native strain), and the synthetic complete genome of M. arthritidis mitogen (MAM) superantigen were used as controls. All SF samples were subjected to DNA extraction separately and multiplex PCR was performed. The PCR products were confirmed by sequencing. RESULTS: The designed multiplex PCR was able to detect M. pneumoniae, M. hominis, and M. arthritidis in the SF of patients with RA with a frequency of 30 (22.9%), 23 (17.5%) and 13 (9.9%), respectively. CONCLUSION: In this study, the overall detection of the Mycoplasma species in RA patients was 53.4%; thus, we recommend the application of multiplex PCR assays when searching for a specific anti mycoplasma treatment for RA patients.

Bacteriological and molecular assessment of staphylococcal enterotoxin e in the blood of patients with rheumatoid arthritis.

BACKGROUND: Rheumatoid arthritis (RA) is the most common chronic inflammatory disease of unknown etiology. In this regard, the role of bacterial superantigens (as an effective agent) were considered. OBJECTIVES: This study aimed to assess staphylococcal enterotoxin E in the blood of patients with rheumatoid arthritis. PATIENTS AND METHODS: A total of 83 blood samples of patients with RA were studied. All of patient's blood samples have been cultured. Polymerase chain reaction (PCR) and ELISA methods have been used to assess the existence of staphylococcal enterotoxin E (entE). The data were analyzed through descriptive statistics. RESULTS: During this study and after sequential sub cultures, only 5 bacterial strains were isolated. Based on the results of biochemical tests, just one case was detected as Staphylococcus aureus. The result of molecular diagnosis of enterotoxin E gene was 13.25%. The results of ELISA were 40.96% positive for staphylococcal enterotoxin E. CONCLUSIONS: In this study, staphylococcal enterotoxin E (superantigen E) was detected in the blood of patients with RA, but its origin is unknown, because no staphylococcus enterotoxin E producer was isolated. This finding could provide a good model for the diagnosis and treatment of RA. However, the results of this study have shown some evidence regarding endogenous origin of involved superantigens in patients with rheumatoid arthritis.

Assay of Blood and Synovial Fluid of Patients With Rheumatoid Arthritis for Staphylococcus aureus Enterotoxin D: Absence of Bacteria But Presence of Its Toxin.

BACKGROUND: Rheumatoid arthritis (RA) is the most common chronic inflammatory disease. The staphylococcal superantigens are considered as the causative agent of RA disease. OBJECTIVES: This study aimed to assess the presence of staphylococcal enterotoxin D in synovial fluid and blood of patients with RA. PATIENTS AND METHODS: A total of 120 blood and SF samples of patients with RA were studied. Bacterial culture, primer pairs design, polymerase chain reaction (PCR), and enzyme-linked immunosorbent assay (ELISA) methods have been used to assess of the staphylococcal enterotoxin D. The data were analyzed through descriptive statistics. RESULTS: During this study and after sequential subcultures, only 5 bacterial strains were isolated. The results of PCR showed the presence of staphylococcal enterotoxin D gene in almost 50% of SF and also in 48.4% of blood samples of patients with RA. Similarly, the ELISA method detected staphylococcal enterotoxin D in 36.16% of SF and in 33.33% of blood of patients with RA. CONCLUSIONS: The result of this study showed that a high percentage of patients with RA have shown staphylococcal enterotoxin D (superantigen D) or entD gene in SF and in blood. However, the origin of this superantigen was not clarified and no Staphylococcus aureus enterotoxin D producer was isolated. This finding indicates other role of this superantigen besides its intoxication. Therefore, staphylococcal enterotoxin D as a biomarker may provide a good model for the diagnosis and treatment of patients with RA.

Determination of vancomycin minimum inhibitory concentration for ceftazidime resistant Streptococcus pneumoniae in Iran.

BACKGROUND: In the context of growing health concerns over antibiotic resistance, the evaluation of the minimum inhibitory concentration (MIC) of vancomycin for Streptococcus pneumoniae (S. pneumoniae) strains resistant to ceftazidime becomes important for guiding health policy makers. The aim of this study was to determine vancomycin MIC of ceftazidime resistant S. pneumoniae strains. METHODS: Fifty identified serotypes of ceftazidime resistant S. pneumoniae strains were included in the study. The vancomycin MIC of the above mentioned bacteria was determined based on the 0.5 McFarland standards, by using a microdilution broth and the Etest method. RESULTS: The results showed that out of 50 ceftazidime resistant strains of S. pneumoniae, 46 strains (92%) have shown a vancomycin MIC ≤0.19 - 0.1.5 µg/ml and only four strains (8%) have shown a vancomycin MIC equal to 1.5 µg/ml and the related maximum zone of inhibition was of 10 millimeter diameters. CONCLUSIONS: The results of this investigation point out the emergence of S. pneumoniae strains with a vancomycin MIC ≥1.5 µg/ml, which were resistant to ceftazidime. This finding uncovers a major health concern: a vancomycin MIC higher than 1.5 µg/ml and maximum zone of inhibition of only 10 millimeter. These findings represent an important warning for health authorities globally, concerning the treatment of patients, as the occurrence of S. pneumoniae strains with decreased vancomycin susceptibility has been demonstrated.

Staphylococcal enterotoxin C in synovial fluid of patients with rheumatoid arthritis.

BACKGROUND: In the previous studies using the commercial ELISA kit, the existence of staphylococcal superantigens has been reported in synovial fluid of patients with rheumatoid arthritis (RA). OBJECTIVES: This study aimed to design molecular methods to detect staphylococcal enterotoxin C in synovial fluid of patients with rheumatoid arthritis. MATERIALS AND METHODS: In this experimental study, Staphylococcus aureus strain producing enterotoxin C was used as the reference strain. The polymerase chain reaction (PCR) was set up by design a specific pair of primers. Besides bacterial culture, 50 synovial fluid samples of patients with rheumatoid arthritis were subjected to DNA extraction, and then PCR amplification was carried out according to the protocol. All samples were examined by ELISA method for enterotoxin C. The data were descriptively analyzed. RESULTS: The results of bacterial culture were negative for all samples. The results showed that 66% (33 cases) of samples contained entC gene and only 46% (23 cases) have also enterotoxin C. The interesting finding was that the results of ELISA and PCR were the same and have shown only 22 positive cases (44%samples) for staphylococcal enterotoxin C. CONCLUSIONS: Based on the findings of this study, S. aureus enterotoxin C (SEC) has been detected in synovial fluid of patients with rheumatoid arthritis by PCR and ELISA methods. These valuable findings may describe the exact etiology of the RA and as well as change the methods of its diagnosis and treatment. This is the first research, which has shown the staphylococcal entC gene in synovial fluid of RA patients. However, S. aureus strains can produce more than 20 types of enterotoxins. Therefore, its involvement on rheumatoid arthritis pathogenesis makes an important challenge in the future. In this regard, further investigation on the other enterotoxins is necessary.

Design of PCR-based method for detection of a gene-encoding Mycoplasma arthritidis mitogen superantigen in synovial fluid of rheumatoid arthritis patients.

BACKGROUND AND OBJECTIVES: Mycoplasma arthritidis mitogen (MAM) superantigen has been shown to induce chronic arthritis, which resembles human rheumatoid arthritis (RA) in a rodent model. However, its role as a causative agent in human RA is not well understood yet. The aim of this study was to investigate the presence of MAM superantigen gene in the synovial fluid (SF) of RA patients. MATERIALS AND METHODS: The MAM superantigen gene a reference was synthesized based on GenBank Data base (Gene ID: 6418105). Specific primer pairs were designed and PCR amplification was performed for MAM superantigen gene detection. A total of 133 SF samples of RA patients were assayed. The PCR products were subjected to sequencing and were descriptively analyzed. RESULTS: The results of the PCR product sequencing showed the method has objective applicability and accuracy. The sensitivity of the PCR reaction for the reference DNA template was 1ng/ml. The PCR results assay of the 133 SF samples raveled that, 9.7% and 22.5% of them were positive for the MAM superantigen gene and Mycoplasma pneumoniae (M. pneumoniae), respectively. CONCLUSION: In this study, two Mycoplasma genomes were detected with increased frequency in RA SF patients' samples. This finding appears to be a promising instrument in the etiological diagnostic of RA patients and could also lead to improved treatment selection. Further research on the other Mycoplasma species present in the SF of RA patients is essential.

Serotype distribution and antibiotics susceptibility pattern of Streptococcus pneumonia in Iran.

BACKGROUND: The development of antibiotic resistance among Streptococcus pneumoniae strains has caused significant health problems worldwide. OBJECTIVES: The aim of this study was to determine antibiotic resistance pattern and serotypes distribution of Streptococcus pneumoniae strains isolated from clinical specimens. MATERIAL AND METHODS: A total of fifty Streptococcus pneumoniae strains were isolated from Tehran Hospital's laboratory from 2008 to 2012. Antimicrobial susceptibility testing was performed using broth microdilution method and minimum inhibitory concentration (MIC) of each strain was determined. to verify the resistant strains and demonstrate the presence of antibiotic resistant genes, the PCR was performed. RESULTS: The study showed that three strains (6%) and six strains (12%) indicated intermediate resistance and complete resistance to penicillin, respectively, 58% strains were susceptible to ceftazidime, two ones (4%) indicated resistance to ciprofloxacin, one (2%) indicated intermediate resistance to ceftriaxone , two strains (4%) indicated complete resistance and four (8%) strains indicated resistance to vancomycin. CONCLUSIONS: The emergence of Streptococcus pneumoniae strains with multiple resistance needs permanent monitoring of antibiotic susceptibility patterns of clinical isolates. We have found that ceftazidime is not a suitable drug for choosing the treatment of pneumococcal infections.

Bacterial meningitis: a new risk factor.

BACKGROUND: The purpose of this study is to discuss a possible new risk factor for the bacterial meningitis. METHODS: Cerebrospinal fluid collected from 270 patients was assayed. An enzyme immunosorbent assay for the detection of Staphylococcal enterotoxins A to E was used. RESULTS: The results indicated that the frequency of Coagulase Negative Staphylococci (CoNS) was 35 (20.46%). An important finding of this research was that the CoNS isolates produced enterotoxin C and D or enterotoxin C and E. CONCLUSIONS: This is the first report of enterotoxin-producing Coagulase Negative Staphylococci isolated from CSF patients. Therefore, these enterotoxins probably act as risk factors in the bacterial invasion into central nervous system.

Molecular screening of staphylococcal enterotoxin B gene in clinical isolates.

OBJECTIVE: The role of staphylococcal enterotoxin B (SEB) in food poisoning is well known, however its role in other diseases remains to be explored. The aim of this study is the molecular screening and characterization of the SEB gene in clinically isolated strains. MATERIALS AND METHODS: In this experimentally study, 300 Staphylococcus aureus (S. aureus) strains isolated from clinical samples were assayed. The isolated strains were confirmed by conventional bacteriological methods. Polymerase chain reaction (PCR) was used to determine the enterotoxin B (ent B) gene. Assessment of toxin production in all strains that contained the ent B gene was then performed. Finally, using specific antibody against SEB, a Western-blot was applied to confirm detection of enterotoxin B production. RESULTS: RESULTS indicated that only 5% of the 300 clinically isolated S. aureus contained the ent B gene. All strains which contained the ent B gene produced a proteinous enterotoxin B. The results of sequence determination of the PCR product were compared with the gene bank database and 98% similarity was achieved. The results of the Western-blot confirmed that enterotoxin B was produced in strains that contained the ent B gene. CONCLUSION: The results of this study indicate that 5% of clinically isolated S. aureus strains produce enterotoxin B. Considering that the enterotoxin B is an important superantigen, it is possible that a delay in diagnosis and lack of early proper treatment can cause an incidence of late complications, particularly in staphylococcal chronic infections. For this reason, it is suggested that in addition to detecting bacteria, an enterotoxin B detection test should be performed to control its toxigenicity.