Characterization of group A streptococci causing invasive diseases in Sri Lanka.

Group A ß haemolytic streptococcus (GAS) or Streptococcus pyogenes is a human pathogen that causes an array of infections, including pharyngitis, cellulitis, impetigo, scarlet fever, toxic shock syndrome, and necrotizing fasciitis. The present study characterizes 51 GAS isolates from invasive infections in Sri Lanka, focusing on resistance profiles, genetic determinants of resistance, and virulence markers. Isolates were tested for sensitivity to penicillin, erythromycin, clindamycin, and tetracycline. The presence of erm(A), erm(B), and mef(A) was detected in erythromycin-resistant isolates, while tet(M) was detected in the tetracycline-resistant isolates. PCR was used to identify SpeA, SpeB, SpeC, SpeF, SpeG, smez, and ssa as virulence markers. Selected GAS isolates were emm-typed using the updated CDC protocol. All 51 isolates were susceptible to penicillin. The number of isolates non-susceptible to erythromycin was 16. The commonest resistance determinant identified was erm(B) (11/16). Tetracycline non-susceptibility was found in 36 (70.6 %) isolates and 26 of them contained the tet(M) gene. Thirteen (25.5 %) isolates were resistant to both tetracycline and erythromycin, while 12 (23.5 %) isolates were sensitive to both antibiotics. The commonest virulence markers detected among the isolates were SpeB (44, 86.3 %), SpeG (36, 70.6 %), and SpeF (35, 68.6 %), while SpeJ (15, 29.4 %), SpeA (10, 19.6 %), and ssa (5,9.8 %) were less common. The emm types were diverse. In conclusion, the GAS isolates studied showed resistance to erythromycin and tetracycline, while retaining universal susceptibility to penicillin. Additionally, these isolates exhibited diverse genetic backgrounds, displaying varying patterns of virulence genes and emm types.

Colonization with selected antibiotic resistant bacteria among a cohort of Sri Lankan university students.

BACKGROUND: Antibiotic Resistance is an imminent global public health threat. Antibiotic resistance emerged in healthcare settings and has now moved on to the community settings. This study was conducted to identify the rates of asymptomatic colonization with selected antibiotic resistant organisms, (Methicillin Resistant Staphylococcus aureus (MRSA), Extended Spectrum Beta Lactamase (ESBL) producing Escherichia coli and Klebsiella spp and carbapenem resistant E.coli and Klebsiella spp) - among a group of university students in Sri Lanka. Identification of genetic determinants of MRSA and ESBL was an additional objective of the study. METHODS: A self - collected nasal swab and a peri-rectal swab collected after passing stools were obtained. Routine microbiological methods were used for the isolation S.aureus from the nasal swab and E.coli and Klebsiella species from the peri-rectal swab. Antibiotic sensitivity testing was performed as recommended by clinical and laboratory standard institute (CLSI). Three (3) genes that are responsible for ESBL production; blaCTX-M, blaSHV, and blaTEM were tested using previously described primers and PCR procedures. Identification of MecA and PVL genes attributed to MRSA was also done with PCR. RESULTS: A total of 322 participants between 21 and 28 years were recruited representing 5 different faculties of study. Seventy one (22.0%) were colonized with S.aureus and 14 among them with MRSA, making the MRSA colonization rate of 4.3%. Forty five (15%) of the participants were colonized with an ESBL producing E.coli or Klebsiella spp. No one was colonized with carbapenem resistant E.coli or Klebsiella species. Of the 45 ESBL producers the commonest genetic determinant identified was blaCTX-M (n = 36), while 16 isolates had blaTEM and 7 had blaSHV. Similarly, of the 14 isolates identified as MRSA, 3 (21.4%) were found to be PVL positive while 11 (78.6%) were MecA positive. CONCLUSIONS: A high rate of colonization with ESBL producing E.coli and Klebsiella species was noted in our study group.

Pneumococcal colonization among healthy and hospitalized vaccine-naive Sri Lankan children.

BACKGROUND: Streptococcus pneumoniae continues to cause mortality and morbidity despite availability of effective vaccines. Pneumococcal colonization is considered a pre-requisite for disease. Identifying the serotypes circulating in a given locale is important for surveillance purposes as well as for assessing the need for vaccination. Aim of the present study was to identify nasopharyngeal pneumococcal colonization rates in healthy children and children with respiratory tract infections in central Sri Lanka. METHOD: A total of 450 nasopharyngeal swabs (NPS) of children aged between 2 months and 2 years were collected from two groups; healthy children and children hospitalized with respiratory symptoms. NPS samples were processed using conventional laboratory techniques to isolate S. pneumoniae. Antibiotic susceptibility patterns of pneumococcal isolates were identified using CLSI disc diffusion method and minimum inhibitory concentration (MIC) was determined by micro-broth dilution method. RESULTS: Pneumococcal colonization rate among healthy children was 31.8% (143/450) it was 39.8% (179/450) in children hospitalized with respiratory symptoms. MIC for penicillin and cefotaxime ranged between 0.015 to 4 µg/ml and <0.015 to 16 µg/ml respectively. All isolates were susceptible to levofloxacin, vancomycin, linezolid and rifampicin. Erythromycin and tetracycline non-susceptibility rates were >50% in both groups. The predominant serotypes identified were 19F (n = 66, 20.5%), 6B (n = 43, 13.4%), 6A (n = 30, 9.3%), 23F (n = 28, 8.7%) and 14 (n = 20, 6.2%). Among healthy children, presence of school going children at home and the number of household members were significantly associated with pneumococcal colonization while in hospitalized children, pneumococcal colonization was significantly associated with presence of school going children at home. CONCLUSION: Pneumococcal colonization rates were considerably higher in both study cohorts and the commonest serotypes were 19F, 6B, 6A, 23F and 14. Antibiotic resistance rates were also relatively higher among the pneumococcal isolates.

Synergistic activity between Triphala and selected antibiotics against drug resistant clinical isolates.

BACKGROUND: Triphala is an indigenous medical product used for a variety of diseases. This study was conducted to determine the effect of Triphala on antibiotic properties of gentamicin and oxacillin against multi-drug resistant organisms. METHODS: The checkerboard method was used to determine the synergy of Triphala with gentamicin and oxacillin against multi-drug resistant (MDR) Gram negative bacilli and methicillin-resistant Staphylococcus aureus (MRSA) using 2,3,5-triphenyltetrazolium chloride (TTC) assay. Fractional inhibitory concentration (FIC) index was calculated. RESULTS: When tested alone, the minimum inhibitory concentration (MIC) values of gentamicin for Gram negative isolates ranged from 8 to > 64 µg/ml. The MIC values of gentamicin for the Gram negative isolates ranged from 1 to 32 µg/ml when tested with Triphala. The FIC index was < 1 indicating a synergistic interaction in 10 of the 11 isolates and it was 1 indicating an additive effect in one isolate. The MIC values of oxacillin for MRSA isolates ranged from 4 to > 16 µg/ml with all MICs being equal to or higher than the resistance cut-off level. The MIC level with the addition of Triphala ranged from 0.25 to 4 µg/ml. FIC index was < 1 for all tested isolates indicating a synergistic interaction. CONCLUSIONS: Triphala has synergistic activity with gentamicin against the selected MDR Gram negative bacilli and with oxacillin against MRSA isolates warranting further studies on the possibility of clinical use.

Terminalia bellirica fruit extracts: in-vitro antibacterial activity against selected multidrug-resistant bacteria, radical scavenging activity and cytotoxicity study on BHK-21 cells.

BACKGROUND: Identification of novel sources for developing new antibiotics is imperative with the emergence of antibiotic resistant bacteria. The fruits of Terminalia bellirica (Gaertn) Roxb., widely used in traditional medicine, were evaluated for antibacterial activity against multidrug-resistant (MDR) bacteria, antioxidant activity and cytotoxicity. METHODS: Twelve solvent extracts of T. bellirica fruits were prepared by direct aqueous extraction and sequential extraction with dichloromethane, methanol and water using Soxhlet, bottle-shaker and ultrasound sonicator methods. Antibacterial activity of the extracts was tested against 16 strains MDR bacteria-methicillin-resistant Staphylococcus aureus (MRSA), extended spectrum ß-lactamase (ESBL) producing Escherichia coli and MDR Acinetobacter spp., Klebsiella pneumoniae and Pseudomonas aeruginosa-and 4 control organisms, using the cut-well diffusion method. The minimum inhibitory concentration (MIC) was determined using an agar dilution method. The radical scavenging activity of six antibacterial extracts was screened against 2,2'-diphenyl-2-picrylhydrazyl (DPPH) and correlation was established between EC50 (50% effective concentration) values and the total phenolic content (TPC). Cytotoxicity was determined for the most potent antibacterial extract on baby hamster kidney (BHK-21) cells by Tryphan Blue exclusion method. Statistical analysis was carried out by one-way analysis of variance at significant level p < 0.05 using "SigmaPlot 10" and "R 3.2.0" software. RESULTS: All aqueous and methanol extracts displayed antibacterial activity (MIC 0.25-4 mg/mL) against all strains of MRSA, MDR Acinetobacter spp. and MDR P. aeruginosa. The sequential aqueous extracts (MIC, 4 mg/mL) inhibited ESBL producing-E. coli. None of the extracts exhibited activity against MDR K. pneumoniae (MIC > 5 mg/mL). The sequential methanol extract (Soxhlet) recorded high antibacterial activity and the highest DPPH radical scavenging activity (EC50, 6.99 ± 0.15 ppm) and TPC content (188.71 ± 2.12 GAE mg/g). The IC50 (50% inhibition concentration) values of the most potent antibacterial extract-the direct aqueous extract from reflux method-on BHK-21 cells were 2.62 ± 0.06 and 1.45 ± 0.08 mg/ml with 24 and 48 h exposure, respectively. CONCLUSIONS: Results indicate that T. bellirica fruit is a potential source for developing broad-spectrum antibacterial drugs against MDR bacteria, which are non-toxic to mammalian cells and impart health benefits by high antioxidant activity.

Maternal vaginal colonization with selected potential pathogens of neonatal sepsis in the era of antimicrobial resistance, a single center experience from Sri Lanka.

BACKGROUND: Maternal vaginal colonization with antibiotic resistant organisms is a growing concern in countries with high antibiotic resistance rates. METHODS: A low vaginal swab was collected from mothers on admission, on discharge and a peri-rectal swab was collected from the neonates born to these mothers on discharge. Routine microbiological methods were used to identify the colonization rates for Escherichia coli, Klebsiella spp. and Streptococcus agalactiae. RESULTS: The pre-delivery colonization rate among the 250 participants for total Enterobacteriaceae was 18.8%. The colonization rates for Klebsiella spp., E. coli and S. agalactiae were, 12.4, 5.6 and 14.8% respectively. Two Klebsiella spp. and two E. coli isolates were confirmed to be exentend spectrum ß lactamase (ESBL) producers with the commonest resistant determinant being blaCTX-M. Post-delivery swabs were collected from 130 participants and the colonization rates were 41.5% for Enterobacteriaceae, 25.4% for Klebsiella spp., 10.8% for E. coli, and 10.8% for S. agalacteiae. Three Klebsiella isolates and one E. coli isolate were confirmed to be ESBL producers with the commonest resistant determinant being blaCTX-M. Considering the 130 participants with both samples, there was a significant increase in the colonization with any Enterobacteriaceae and Klebsiella spp. (p < 0.05). Peri-rectal swabs were collected from neonates in 159 instances. The isolation rates for Enterobacteriaceae was 34%. The genus specific isolation rate for Klebsiella was 21.4% while the rates for E. coli and S.agalactiae were 10.1 and 5.7% respectively. Two of the E. coli were confirmed to be ESBL producers while none of the klebsiellae were identified to be so. Considering these 159 instances where both the mother and baby were sampled, random amplification of polymorphic DNA (RAPD) analysis revealed that Enterbacteriaceae with same strain type was present in 6.9% of the instances, indicating possible transfer between the mother and neonate. The transfer rate for ESBL producers were 0.6%. CONCLUSIONS: The lower level of antimicrobial resistance among these potentially community acquired isolates is encouraging. However, in view of the increasing level of resistance reported elsewhere in the region, regular monitoring is warranted.

Spread of resistant gram negatives in a Sri Lankan intensive care unit.

BACKGROUND: Infections with multi drug resistant (MDR) organisms are a major problem in intensive care units (ICUs). Proper infection control procedures are mandatory to combat the spread of resistant organisms within ICUs. Well stablished surveillance programmes will enhance the adherence of the staff to infection control protocols. The study was conducted to assess the feasibility of using basic molecular typing methods and routine hospital data for laboratory surveillance of resistance organisms in resource limited settings. METHODS: A retrospective study was conducted using consecutive Gram negative isolates obtained from an ICU over a six month period. Antibiotic sensitivity patterns and random amplified polymorphic DNA (RAPD) based typing was performed on the given isolates. RESULTS: Of the seventy isolates included in the study, seven were E.coli. All E.coli were MDRs and Extended Spectrum ß lactamse (ESBL) producers carrying bla CTX-M. Fourteen isolates were K.pneumoniae, and all were MDRs and ESBL producers. All K.pneumoniae harboured bla SHV while 13 harboured bla CTX-M. The MDR rate among P.aeruginosa was 13% (n=15) while all acinetobacters (n=30) were MDRs. Predominant clusters were identified within all four types of Gram negatives using RAPD and the ICU stay of patients overlapped temporally. CONCLUSION: We propose that simple surveillance methods like RAPD based typing and basic hospital data can be used to convince hospital staff to adhere to infection control protocols more effectively, in low and middle income countries.