Antibiotic Susceptibility and Biofilm Production among Coagulase Negative Staphylococci Isolated from Clinical Samples at Tertiary Care Hospital.

BACKGROUND: Coagulase Negative Staphylococci have been widely associated with medical device implant treatment and immune-compromised patients. Despite having increasing interest in Coagulase Negative Staphylococci, few studies from Nepal have reported the association of these organisms with urinary tract infections, conjunctivitis, high vaginal swabs, and cerebrospinal fluid. This study was carried out to determine antibiotic susceptibility pattern and biofilm production among Coagulase Negative Staphylococci isolated from clinical samples at tertiary care hospital. METHODS: This study was a hospital based cross-sectional study in which 3690 clinical samples were included. Isolation and identification of isolates was done following standard microbiological protocol. Coagulase Negative Staphylococci were identified phenotypically on the basis of gram staining, slide and tube coagulase test and by various sugar fermentation tests. Antibiotic susceptibility test was done following Kirby Bauer disk diffusion method (Clinical and Laboratory Standards Institute 2020). Biofilm production was determined by Tissue Culture Plate technique. RESULTS: A total of 113 isolates of Coagulase Negative Staphylococci were detected. Among them S. epidermidis (45.1%), S. saprophyticus (23.9%), S. haemolyticus (16.8%), S. hominis (5.3%), S. capitis (2.7%), -----S. cohini (1.8%), S. lugdunensis (1.8%) and S. sciuri (2.7%) were identified phenotypically. All isolates were found to be resistant against Ampicillin and 111 (98.2%) were sensitive against Linezolid.23.9% of CoNS were strong biofilm producers, 19.5% moderate and 56.6 % were non/weak biofilm producers. CONCLUSIONS: It requires susceptibility test for prescribing antibiotics against Coagulase Negative Staphylococci in hospital and the misuse of antibiotics should be prevented.

Higher Rate of Extreme Drug-resistant Klebsiella pneumoniae Infections among Cardiac Patients.

BACKGROUND: The accelerating rate of carbapenems resistance in Klebseilla pneumoniae isolates has put the treatment option worrisome. The effective strategy to ameliorate this alarming situation is possible through enhancing the combination therapy and appropriate laboratory diagnosis. Hence, the study was focused on identifying carbapenemase-producing K. pneumoniae and their antibiogram pattern. METHODS: A total of 944 clinical samples from patients attending Sahid Gangalal National Heart Center were processed from September 2019 to March 2020 to identify the possible bacterial pathogens following the standard microbiological procedures. K. pneumonaie isolates were further subjected to antibiotic susceptibility testing by the modified Kirby Bauer disc diffusion technique. Phenotypic confirmation of carbapenemase production was done by the modified carbapenemase inactivation method. The minimum inhibitory concentration of colistin was determined by the broth microdilution method. RESULTS: Of the total 944 samples, 15.47% (146) samples showed bacterial growth, among which 23.97% (35) were K. pneumoniae. Out of 35 K. pneumoniae isolates, 45.71% (16) were multidrug-resistant followed by 42.86% (15) extensively drug-resistant. Fourteen isolates of K. pneumoniae were carbapenemase producers among which 20% (7) were serine carbapenemase while 20% (7) showed metallo-?-lactamase production. All the carbapenemase-producing K. pneumoniae were susceptible to colistin with <0.125µg/ml. Carbapenemase activity showed statistically significant with multidrug resistance (p=<0.05). CONCLUSIONS: An increasing resistance to the carbapenem drugs showed a great problem in the management of K. pneumoniae infections among immunocompromised patients especially cardiac patients however, colistin can be still an ultimate choice of drug for disease management.

gyrA ser83 mutation among fluoroquinolone-resistant Salmonella enterica serovars from enteric fever patients in tertiary care hospital, Kathmandu.

BACKGROUND: The management of enteric fever through antibiotics is difficult these days due to the emerging resistance of Salmonella to various antimicrobial agents. The development of antimicrobial resistance is associated with multiple factors including mutations in the specific genes. To know the current status of mutation-mediated fluoroquinolone-resistance among Salmonella enterica serovars; Typhi, Paratyphi A, B and C, this study was focused on detecting gyrA ser83 mutation by restriction digestion analysis of gyrA gene using HinfI endonuclease. RESULTS: A total of 948 blood samples were processed for isolation of Salmonella spp. and 3.4% of them were found to be positive for Salmonella growth. Out of the 32 Salmonella isolates, 2.2% were S. Typhi and 1.2% were S. Paratyphi A. More interestingly, we observed less than 5% of isolates were resistant to first-line drugs including chloramphenicol, cotrimoxazole and ampicillin. More than 80% of isolates were resistant to fluoroquinolones accounting for 84.4% to levofloxacin followed by 87.5% to ofloxacin and 100% to ciprofloxacin by disc diffusion methods. However, the minimum inhibitory concentration method using agar dilution showed only 50% of isolates were resistant to ciprofloxacin. A total of 3.1% of isolates were multidrug-resistant. Similarly, 90.6% of the Salmonella isolates showed gyrA ser83 mutation with resistance to nalidixic acid. CONCLUSIONS: The increased resistance to fluoroquinolones and nalidixic acid in Salmonella isolates in our study suggests the use of alternative drugs as empirical treatment. Rather, the treatment should focus on prescribing first-line antibiotics since we observed less than 5% of Salmonella isolates were resistant to these drugs.

Comparison of Nasal Colonization of Methicillin-Resistant Staphylococcus aureus in HIV-Infected and Non-HIV Patients Attending the National Public Health Laboratory of Central Nepal.

BACKGROUND: Staphylococcus aureus is a cardinal source of community- and hospital-acquired infection. HIV infection is a well-recognized risk factor for methicillin-resistant S. aureus (MRSA) carriage and infection. Intrinsically developed antibiotic resistance has sharply increased the burden of MRSA which is often associated with morbidity and mortality of the patients. Moreover, nasal carriage of S. aureus plays a significant role in spread of community-associated (CA) S. aureus infections. METHODS: This study was conducted from June 2016 to December 2016 at National Public Health Laboratory (NPHL), Kathmandu, with an aim to assess the rate of S. aureus nasal carriage and MRSA carriage among HIV-infected and non-HIV patients. A total of 600 nonrepeated nasal swabs were analyzed following standard microbiological procedures, where 300 swabs were from HIV-infected patients while remaining 300 were from non-HIV patients. The isolates were identified on the basis of colony characteristics and a series of biochemical tests. The antibiotic susceptibility test (AST) was performed by the modified Kirby-Bauer disc diffusion method. Inducible clindamycin resistance in isolates was confirmed by the D-test method. RESULTS: Overall, out of 600 nasal swabs of patients tested, 125 (20.8%) were S. aureus nasal carriers which included 80 out of 300 (26.66%) among HIV-infected patients and 45 (15%) out of 300 among non-HIV patients, and the result was statistically significant (p=0.0043). Among the isolated S. aureus, 11 (13.8%) MRSA were confirmed in HIV-infected while 3 (6.7%) MRSA were detected from non-HIV patients. A higher number of S. aureus carriers was detected among HIV-infected males 40 (26.49%), whereas MRSA carriage was more prevalent among HIV-infected females 7 (5.1%). Among the HIV-infected, patients of age group 31-40 years were the ones with highest carriage rate 36 (45%), while in non-HIV patients, the highest rate 13 (28.9%) of carriage was detected among the patients of age group 21-30 years. Statistically significant difference was found between S. aureus carriage and HIV infection in patients (p < 0.05). Higher rate 2/3 (66.7%) of inducible clindamycin resistance in MRSA was detected from non-HIV patients in comparison to HIV-infected patients 7/11 (63.63%) while the result was statistically insignificant (p > 0.05). All the MRSA isolates (100%) were resistant against co-trimoxazole while ciprofloxacin showed high rate of sensitivity towards both MSSA and MRSA. None of the isolates were detected as VRSA. The major factors associated with nasal colonization of S. aureus were close personal contact, current smoking habit, and working or living in a farm (p < 0.05). CONCLUSION: Regular surveillance and monitoring of MRSA nasal carriage and antibiotic susceptibility pattern are of prime importance in controlling S. aureus infections especially in high risk groups like HIV-infected patients.

Surgical Site Infections and Antimicrobial Resistance Pattern.

BACKGROUND: Post-operative surgical site infections (SSIs) are among the leading cause of morbidity and increased medical expense. The aim of this study is to isolate identify and study antimicrobial susceptibility pattern of microorganism from surgical wound of admitted patients. METHODS: This retrospective study was carried at the Microbiology Laboratory of Bharatpur hospital, Nepal, from May 2015 to October 2015. The pus samples were cultured and antibiotic susceptibility determined in vitro by Kirby Bauer's disc diffusion method following clinical and Laboratory Standards Institute (CLSI) 2014 recommendation. RESULTS: Of the total 250 samples, 194 (77.6%) showed bacterial growth. Staphylococcus aureus was 47.4% and Escherichia coli 20.60 %. Of 194 isolates 39.2% were multi drug resistant. Amikacin was sensitive in 93.1% of Gram positive isolates and 81.8% of gram negative isolates. CONCLUSIONS: Bacterial growth is common in surgical site. Staphylococcus aureus and Escherichia coli were multidrug resistant. Grampositive and gram negative isolates were commonly sensitive to Amikacin.

Bacteriological Study of Otitis Media and Its Antibiotic Susceptibility Pattern.

BACKGROUND: Otitis Media is a prevailing and notorious infection in developing countries causing serious local damage and threating complication. Mainly in developing countries like Nepal, Otitis Media results because of illiteracy, poverty and poor hygiene. The aim of this study was to determine the profile of Otitis Media, its causative agents and their antibiotic susceptibility pattern. METHODS: The study included 263 pus samples from 240 patients attending ENT department of Bharatpur hospital from May 2015 to January 2016. Samples were processed in microbiology department for bacteria using standard operating protocol. Antibiotic susceptibility testing was performed for all bacterial isolates by Kirby Bauer disc diffusion method and the results were interpreted according to clinical and laboratory standard institute (CLSI) guideline. RESULTS: Out of 240 patients, 121 were female and 119 were male. Highest incidence of Otitis Media was observed in 1-10 year age group. Out of 263 samples taken from 240 patients, 216 showed bacterial growth. Gram negative bacteria predominated and the most common bacteria isolated were Staphylococcus aureus 36.11% followed by Pseudomonas aeruginosa 33.33% and Coagulase Negative Staphylococci 8.08%. All bacterial isolates were sensitive to gentamycin. Staphylococcus aureus was sensitive to Amikacin and gentamycin. All gram negative bacterial isolates were sensitive to Imipenem and gentamycin. 100% of Pseudomonas aeruginosa was sensitive to Imipenem. CONCLUSIONS: Staphylococcus aureus was the most predominant organism isolated from the pus swab followed by Pseudomonas aeruginosa and all the isolated organisms were sensitive to Gentamycin.

Co-existence of beta-lactamases in clinical isolates of Escherichia coli from Kathmandu, Nepal.

BACKGROUND: The trend of extended-spectrum beta-lactamases producing Escherichia coli (ESBL-EC) is increasing in Nepal. Limited studies have been reported investigating ESBL types and carbapenemases in E. coli. METHODS: A cross sectional study was conducted between June 2012 to January 2013 in Kathmandu Medical College and Teaching Hospital, Nepal. Non-repetitive clinical samples from out-patient department (OPD) and Intensive Care Units (ICU) were processed for bacteriological culture and identification of E. coli. Antibiotic susceptibility test, screening and phenotypic confirmation for ESBLs and carbapenemases and PCR (blaCTX-M, blaSHV and blaTEM-type ESBLs, blaVIM, blaIMP and blaNDM-1-type carbapenemases, and class 1 integron element integrase gene) were performed. Clones were resolved by PCR-Randomly Amplified Polymorphic DNA. RESULTS: Out of 332 non-repetitive clinical specimens processed for culture and identification 160 (48.2%) were culture positive. Of which, 93 (58.1%) were E. coli. Of these, 24 (25.8%) were phenotypically confirmed as ESBL-EC and 3 (12.50%) of 24 ESBL-EC were carbapenemase producers. blaCTX-M-type ESBL was most common (23, 95.8%) followed by blaTEM (7, 29.2%) and blaSHV (3, 12.5%). blaVIM, blaIMP and blaNDM-1 were present in 3, 2 and 2 ESBL-EC, respectively. Class 1 integron element was present in 18 (75.0%) ESBL-EC. Nine isolates possessed more than one type of beta-lactamases. Interestingly, all carbapenemase producers were isolated form ICU and co-existence of blaCTX-M, blaSHV, blaTEM, blaIMP, blaVIM and blaNDM-1 beta-lactamases was documented in one ESBL-EC (EC104). All most all isolates had different RAPD patterns. CONCLUSIONS: For the first time in Nepal, high prevalence of blaCTX-M-type ESBL and co-existence of ESBLs and carbapenemases has been described. Continuous monitoring and surveillance and proper infection control and prevention practices will limit the further spread of these super-bugs within this hospital and beyond.