pilot study on microbial flora of water contained in shisha water pipes and their identification by MALDI-TOF MS

Objective: The water containing jar of the shisha water pipe is constantly moist and is a potential site for bacterial growth and biofilm formation. This study aims to examine the microbial flora and bacterial load of the water in the shisha water jar so as to determine possible risks to users

Materials and methods: Three shisha cafes participated voluntarily in the study. Seventeen de-identified water samples were collected from Shisha pipes being used by various customers at the participating Shisha Cafes. Samples were collected using sterile precautions. Ten micro L from the water samples were inoculated on a blood agar plate and incubated at 37[degree sign]C for 24 hours. The bacterial CFUs/mL were counted manually and organisms isolated were identified by Proteomic fingerprinting of ribosomal proteins using a Bruker Biotyper [Trade Mark] working on the principle of Matrix Assisted Laser Desorption Ionization- Time of Flight Mass Spectrometry, [MALDI-TOF MS]

Results: Seventeen samples were collected. Eleven samples showed microbial growth of 100 to 3000 CFUs/ml with a median value of 400 CFUs/ml. Six samples were sterile. Of the eleven samples, six samples had one organism each, two samples had two organisms each, one sample had three organisms and the last sample had five organisms. The 18 organisms isolated were distributed among nine species: Acidovorax temperans [3 isolates], Burkholderia vietnamiensis [3 isolates], Candida_pelliculosa [1 isolate], Delftia acidovorans [1 isolate], Enterobacter cloacae [3 isolates], Escherichia hermannii [1 isolate], Klebsiella pneumonia [2 isolates], Pseudomonas aeruginosa [1 isolate], and Raoultella ornithinolytica [1 isolate]. Two isolates could not be identified

Conclusions: The organisms isolated have been referenced in literature as pathogens or emerging pathogens capable of causing infections. Presence of significant amounts of pathogens in Shisha water suggests the strong possibility of inhalation of aerosols containing microbes by shisha smokers. The poly microbial nature of the isolates especially suggests the formation of bacterial biofilms. Interestingly almost all the isolates were gram negative bacteria, which contain lipopolysaccharide [LPS] in their cell walls. LPS is a microbial protein that stimulates innate immunity. LPS has been shown to be an etiological agent of acute and chronic airway obstruction and disease. Thus LPS from bacterial aerosols can contribute to the causation of COPD in chronic users, simultaneously exposing them to the risk of infection by newer emerging pathogens

Prevalence and antimicrobial susceptibility pattern of Methicillin-resistant Staphylococcus aureus among clinical samples in a tertiary care hospital

To determine the prevalence and antimicrobial susceptibility pattern of MRSA isolates among clinical samples in a tertiary care hospital. This cross sectional study was carried out among patients between the age group one and seventy years, admitted to a tertiary care hospital in India. A total of 2850 clinical samples from patients attending various clinical departments were collected over a period of two years. staphylococcus aureus was identified by culture characteristics and confirmed by tube coagulase test. Methicillin resistance was determined by oxacillin [1 micro g] disc diffusion method using MHA with 5% NaCl supplementation. Antimicrobial susceptibility testing was done by Kirby-Bauer's method for all the isolated MRSA strains. The prevalence for age, gender and clinical samples were calculated. Of the total 2850 clinical samples studied, 1262 [44%] samples were from pyogenic materials. 490 [17%] from aspirates and 1098 [39%] from wound secretions of a patients of various clinical departments. 586 samples were positive for S. aureus. Out of the 586 S. aureus isolated, 236 [40.2%] isolates were positive for MRSA and 350 [59.7%] isolates were positive for MSSA. The yield of MRSA was the highest among 20-40 year age group [47.4%], predominantly males and were mostly from surgical post-operative patients [39.8%]. All the 236 MRSA isolates were 100% susceptible to Vancomycin and 100% resistant to Penicillin. The Prevalence of MRSA in our hospital was 40.2%. We conclude that knowledge of prevalence and antibiotic sensitivity pattern of MRSA help the clinician in providing the first line treatment in the health care settings. Also we recommend a universal rapid MRSA screening strategy for patients admitted for surgery in order to control and minimize the spread of MRSA in patients

Staphylococcus aureus: prevalence among normal healthy individuals

To determine the prevalence of Staphylococcus aureus among normal healthy individuals in relation to age, gender and site of isolation. This cross sectional study was carried out among normal healthy individuals in the age group 15-65 years, at Gulf Medical University and Gulf Medical College Hospital and Research Center, Ajman. The study included a detailed proforma of all the volunteers. The nasal and throat swabs were collected with strict aseptic precautions and were subjected to direct microscopic examination and culture on appropriate media. Staphylococcal isolates were subjected for Tube Coagulase test and growth on Mannitol Salt Agar [MSA] and were grouped as Staphylococcus aureus and Coagulase Negative Staphylococci [CoNS]. Of the 127 human volunteers screened, 67 were from GMU [22 were staff and 45 M.B.B.S students] and 60 volunteers from GMCH and RC, Ajman [doctors, staff nurses and ward boys]. Staph, aureus isolation was the highest among the 21-30 year age group [37%] with male predominance [59.6%] and mostly from the nasal swabs [56.45%]. The yield of Staph. aureus isolates from the nasal mucosa and throat were only from 49 normal individuals, hence the prevalence of Staph. aureus in our study being 38.5%. Among the total 124 Stapylococcal isolates, 62 [50%] were Staph. aureus and 62 [50%] were coagulase negative staphylococci. Of the 62 Staph aureus isolates, 35 [56.45%] were from nasal swabs and 27 [43.54%] from throat swabs. Of the 62 coagulase negative Staphylococcus isolates, 44 [70%] were from nasal swab, four [6%] from throat swab and 14 [22%] were from both nasal and throat swabs. The study indicates an alarming prevalence rate of Staph. aureus. Hence we conclude that the data observed reflected the need for a complete revision for all the infection control programs at GMU and GMCH and RC which includes education programs, aseptic techniques, hospital hygiene and periodic microbiological surveillance in order to control the spread of infection

Betalactamase production of Staphylococcus aureus: a comparison study of different iodometric methods

This study was undertaken to detect Beta-lactamase producing Staphylococcus aureus among healthy individuals by using different iodometric methods [filter paper, agar plate and tube]. The study was carried out among 127 normal healthy volunteers of Gulf Medical University and Gulf Medical College Hospital and Research Center, Ajman. Nasal and throat swabs were collected with strict aseptic precautions. All the swabs were examined by direct microscopy and inoculated into appropriate culture media. Staphylococcus aureus was identified by tube coagulase test and growth on MSA. Beta-lactamase enzyme activity of Staph. aureus was determined by the filter paper, agar plate and tube lodometric methods. Antimicrobial susceptibility testing by Kirby-Bauer's method was performed for all Beta-lactamase producing Staph. aureus. Out of 127 healthy volunteers screened, Staph. aureus were isolated from the nasal and throat swabs from 62 subjects. Of the 62 Staph. aureus isolates, 40 [64.5%] were positive by all the three iodometric methods. Six isolates [9.6%] were positive by two methods [4 isolates by filter paper and tube methods, one isolate by filter paper and agar plate methods and one isolate by agar plate and tube methods]. Four isolates [6.4%] were positive by one of these methods. Twelve isolates [19.4%] were totally negative by all the three methods. Overall susceptibility of filter paper and tube methods were found to have similar results 45 [72.5%]. Whereas the overall susceptibility of agar plate method was 42 [67.7%]. Antimicrobial susceptibility was 100% to Vancomycin and Rifampicin, and 100% resistance to Penicillin and Ampicillin. The study indicates that the filter paper and tube iodometric methods were accurate and superior to the agar plate method in the detection of Staphylococcal Beta-lactamase. We conclude that the alarming prevalence rate of Beta-lactamase among normal healthy individuals indicates the need for treatment by Beta-lactamase-susceptible antibiotics

Detection of methicillin-resistant strains of staphylococci by multiplex polymerase chain reaction

A rapid and accurate detection procedure for Methicillin resistance among Staphylococci isolates through the amplification of specific gene determinants by Multiplex PCR was done in the present study. A total 586 Staphylococcal clinical isolates were studied. Clinical samples including pyogenic materials, aspirates and wound secretions were collected under aseptic conditions from patients attending various departments of a tertiary care hospital. staphylococcus aureus isolates were identified by culture characteristics and confirmed by tube coagulase test. Methicillin resistance was determined by Oxacillin disc diffusion method [1 micro] using MHA with 5% NaCl supplementation. The multiplex PCR was used as the gold standard for the detection of mec A genes. Thirty five MRSA isolates were subjected for mec A gene detection by multiplex PCR at Sir Dorabji Tata center for Research in Tropical Diseases, IISc Campus, Bangalore, India. The multiplex PCR was performed according to the procedure of Oliveira et al. by using the lysates of Staphylococcal strains. [MRSA] as templates and Oligonucleotide sequences as primers, a 162 and 530-bp region of mec A, the structural gene of a low-affinity penicillin-binding protein [PBP2'] was amplified and detected by agarose gel electrophoresis. Of the total 586 Staphylococcal aureus isolates studied, 236 [40.2%] samples were positive for MRSA and 350 [59.7%] samples were MSSA by Oxacillin disc diffusion method [1 micro] using MHA with 5% NaCl supplementation. Thirty five isolates were subjected for mec A gene detection by multiplex PCR. Thirty three MRSA isolates [94%] were found to be positive at 162 and 530 base pairs and 2 MRSA strains were mec A negative. We conclude that the PCR assay isw a rapid and accurate procedure for the diagnosis of MRSA infection