In vitro Salmonella typhi biofilm formation on gallstones and its disruption by Manuka honey

Biofilm is a complex community of single or different types of microorganisms [bacteria, viruses, fungi, protozoa] attached to a surface and stick to each other through production of extracellular matrix. Salmonella typhi forms biofilm on cholesterol gallstones resulting in carrier state. Once formed, biofilm is difficult to treat. To date cholecystectomy is the only cure for this condition. Manuka honey is known to have tremendous antibiofilm activity against various organisms. S. typhi biofilm was grown in vitro on clinical samples of human cholesterol gallstones by Gallstone tube assay method for 12 days. Biofilm mass was quantified on day 1, 5, 7, 9 and 12 by crystal violet assay and was also examined by scanning electron microscope. Three concentrations w/v of Manuka honey [40%, 60% and 80%] were used, each one at 24, 48 and 72 hours. The most effective concentration [80% w/v] was repeated on two sets of gallstones. Biofilm mass was re quantified by crystal violet assay and was examined by scanning electron microscope. S. typhi formed uniform biofilm on cholesterol gallstone surface. The optical density measurements exhibited a rising pattern with time thereby indicating an increase in biofilm mass. It was 0.2 on day 1 and 0.9 on day 12. With 80% w/v Manuka honey, biofilm mass decreased most effectively with 0.5 OD after 72 hours. Biofilm formation by S, typhi on gallstones is surface specific and bile dependant. Either increasing the duration [beyond 72 hours] of the effective concentration [80% w/v] of honey or increasing the concentration [above 80%] of honey for a specific duration [72 hour] may cause complete disruption of the S. typhi biofilm on gallstone. S. typhi forms biofilm on cholesterol gallstones surface in vitro and it can be visualized by scanning electron microscopy. Biofilm mass can be quantified using crystal violet assay. Among various concentrations 80% Manuka honey for 72 hours is most effective in disrupting S. typhi biofilm on gallstones in vitro as evident from crystal violet assay

Carbapenemase intrudes K. pneumoniae in local clinical isolates

To determine the sensitivity pattern of the isolates under study against commonly used antimicrobials. The study was carried out at the Department of Microbiology, University of Health Sciences Lahore. Two hundred Clinical isolates [n=200] of Enterobacteriaceae were collected from various tertiary care hospitals of Lahore. The isolates were identified by their morphology and cultural characteristics. API 20E was used for their biochemical profile. Kirby-Bauer disk diffusion method was employed for their susceptibility according to CLSI 2009 guidelines. Modified Hodge Test was used for testing carbapenemase production. Further confirmation was done using EDTA disk potentiation method. Susceptibility pattern of two hundred isolates showed multidrug resistance pattern. All the isolates showed least resistance to Imipenem and Meropenem [0.5%] followed by Amikacin [24%] and Tetracycline [61%]. Only one strain of K. pneumoniae was found to be resistant to carbapenems and it was confirmed to be carbapenemase producer by Modified Hodge Test, and metallo [3-lactamase producer by EDTA disk potentiation method. Carbapenemase has intruded local isolates of K. pnuemoniae. It can result in outbreak of carbapenem resistant strains as it can spread through vertical as well as horizontal transmission. The carbapenem resistant isolates must be contained in order to prevent its further spread among other members of Enterobacteriaceae and other bacteria

Bacteriological analysis of drinking water from 100 families of Lahore by membrane filtration technique and chromagar

Water borne diseases are caused by pathogenic microorganisms. In Pakistan, the availability of safe water is only 40% to 60%. Therefore it becomes imperative to determine the bacteriological status of drinking water. A few laboratories perform such an evaluation and, that too, by the old method technique i.e Most Probable Number [MPN]. We evaluated 100 samples of drinking water from some areas of Lahore by the Metnbrane Filtration Technique [MFT] using CHRO-Magar. Using this technique in one step a much large volume of water can be evaluated quantitatively in a short time and with ease. Use of CHROMagar straightaway confirms the presence of Escherichia coli which is accepted universally as the indicator of fecal contamination. It was a cross sectional study. A volume of 100 ml water was filtered under the vacuum pressure through Millipore membrane filters. After filtration, membrane filters were placed on CHRO Magar and incubated at 35°C for 24 hr. Escherichia coli appeared as blue coloured colonies while coliforms yielded colonies of pink colour. Escherichia coli were further identified by API 2OE and confirmed by Eijkman test. Escherichia coli was grown from 42% samples [all Eijkman positive]. Coliform organisms were grown from 54% specimens. It was alarming that 59% of drinking water was unsatisfactory for human consumption

'Gastro' of mirpur khas [Sindh] is resolved

Diarrhoeal disease is a common cause of major public health concern in many parts of the world including Pakistan. Eighteen stool samples were received from Civil Hospital Mirpur Khas [Sindh]. All the specimens were processed for culture and antimicrobial susceptibility according to Clinical Laboratory Standards Institute [CLSI] guidelines. Eight out of eighteen [44.4%] samples were positive for Vibrio cholerae. The isolates were gram negative rods, showed darting motility and were Oxidase positive. Contact with distilled water immobilized all these strains [Mdw]. API 20NE was used for the biochemical identification and serological confirmation was done with 'difco' antisera. Kirby-Bauer disc diffusion method was performed for their respective susceptibility to various antibiotics. All these isolates were confirmed to be Vibrio cholerae O1 Biotype El Tor Serotype Ogawa. The isolates were generally sensitive to majority of the antibiotics but resistant to nalidixic acid except one strain. Six out of eight isolates were resistant to co-trimoxazole. Cholera must be suspected in outbreaks of such kind of 'gastro'

Anti bacterial activity of Nigella sativa against clinical isolates of methicillin resistant Staphylococcus aureus

Methicillin resistant Staphylococcus aureus [MRSA] continues to be one of the commonest pathogens encountered in clinical as well as laboratory practice. It has become a major health problem worldwide. Newer antimicrobials/agents are urgently needed to combat this problem MRSA resistance to various anti-staphylococcal agents. In the back-drop of this difficult situation Nigella sativa commonly known as black seed [ethanolic extract] was aimed at to evaluate if it had any anti-staphylococcal activity. The extract was prepared by reflux extraction method. Disc diffusion and in agar dilution methods were performed to assess the antibacterial activity. Staphylococcus aureus ATCC 25923 was used as the standard reference strain. All tested strains of MRSA were sensitive to N. sativa extract at a concentration of 4 mg/disc while the extract had an MIC range of 0.2-0.5 mg/ml. The results indicated that N. sativa has inhibitory effect on MRSA. This finding warrants necessity of further investigation of this product of folk medicine

Rapid isolation of mycobacteria-need of the hour in our settings

To compare Bactec MGIT 960 with LJ media in terms of time taken for the initial isolation of mycobacteria. A total of 100 AFB [acid fast bacillus] positive sputum samples were processed and inoculated in both the Lowenstein Jensen [LJ] media and mycobacterium growth indicator tube [MGIT] tubes. Of the 100 samples, positive growth was obtained from all the samples on both the MGIT and LJ media. In MGIT 53% samples grew in 4 days, 30% in 5 days and 17% in 6 days [Mean=4.6 days] while on LJ media, 44% grew in 30 days, 20% in 35 days and 36% in 44 days [Mean=37 days]. Significant difference was observed between two systems with a p-value of less than 0.05. Bactec MGIT 960 is a much faster and efficient system for the initial isolation of mycobacteria