Elucidating the bacterial inactivation mechanism by argon cold atmospheric pressure plasma jet through spectroscopic and imaging techniques.

AIMS: This study aims to assess the potential bacterial inactivation pathway triggered by argon (Ar) cold atmospheric pressure plasma jet (CAPJ) discharge using spectroscopic and imaging techniques. METHODS AND RESULTS: Electrical and reactive species of the Ar CAPJ discharge was characterized. The chemical composition and morphology of bacteria pre- and post-CAPJ exposure were assessed using Fourier transform infrared (FTIR), Raman micro-spectroscopy, and transmission electron microscopy (TEM). A greater than 6 log reduction of Escherichia coli and Staphylococcus aureus was achieved within 60 and 120 s of CAPJ exposure, respectively. Extremely low D-values (<20 s) were recorded for both the isolates. The alterations in the FTIR spectra and Raman micro-spectra signals of post-CAPJ exposed bacteria revealed the degree of destruction at the molecular level, such as lipid peroxidation, protein oxidation, bond breakages, etc. Further, TEM images of exposed bacteria indicated the incurred damages on cell morphology by CAPJ reactive species. Also, the inactivation process varied for both isolates, as evidenced by the correlation between the inactivation curve and FTIR spectra. It was observed that the identified gas-phase reactive species, such as Ar I, O I, OH•, NO+, OH+, NO2-, NO3-, etc. played a significant role in bacterial inactivation. CONCLUSIONS: This study clearly demonstrated the effect of CAPJ exposure on bacterial cell morphology and molecular composition, illuminating potential bacterial inactivation mechanisms.

Antiseptic efficacy and plasma chlorhexidine levels following two different methods of application of 1% aqueous chlorhexidine gluconate for skin disinfection in preterm newborns: a randomized controlled trial.

OBJECTIVE: Many neonatal units have started using Chlorhexidine gluconate for neonatal skin antisepsis. However, there is in-vitro evidence of inhibition of neurite growth. The current study aimed to compare two methods of its local application, for the extent of systemic absorption and antiseptic efficacy. STUDY DESIGN: Parallel group, blinded, randomised trial, at a Level III, neonatal intensive care unit. Between December 2020 to July 2022, neonates from 28 to 34 weeks gestation, were randomized to local skin antisepsis by either- (a) 1% aqueous chlorhexidine (CHG aq) followed by cleansing off the residual agent with sterile water swab (Cleansing group) or (b)1% CHG aq followed by air drying (No cleansing group). The outcome measures were the proportion of post antisepsis skin swabs with no/insignificant growth, and the plasma chlorhexidine levels. RESULTS: Of the total of 457 enrollments (Cleansing: n = 230; No Cleansing: n = 227), 216 (93.91%) in "Cleansing" vs. 221 (97.36%) in "No cleansing" (risk difference -3.45%, 95% CI -7.2 to 0.28%; p = 0.072) had no/insignificant growth post-antisepsis. The lower bound of the confidence interval crossed the pre-specified non-inferiority limit of 5%. The median (IQR) plasma chlorhexidine levels were not significantly different between the two groups (7.9 (5.6, 17.9)) ng/mL in Cleansing vs. 6.5 (4.6, 17.7) in No cleansing groups (p = 0.437). CONCLUSION: Cleansing with sterile water after application of chlorhexidine in preterm neonates was not shown to be non-inferior compared to no cleansing, for skin antisepsis efficacy. Systemic absorption occurred to a similar extent despite cleansing off the residual agent. TRIAL REGISTRATION NUMBER: CTRI/2020/10/028719.

Early organism identification by Matrix Assisted Laser Desorption Ionization Time of Flight Mass Spectrometry (MALDI-TOF-MS) decreases the time to appropriate antibiotic modifications for common bacterial infections.

Objective: MALDI-TOF-MS facilitates the identification of microorganisms from positive cultures in a timely and accurate manner. It eliminates the necessity for the application of biochemicals and operates on the principle of proteomics. It decreases the time required to report culture results. Prompt detection and notification of the pathogen, prior to the disclosure of antimicrobial susceptibilities, could potentially shorten the duration until the initial antibiotic adjustment is necessary, thereby influencing patients' clinical prognoses. Methodology: Fifty patients in the conventional arm and one hundred patients in the interventional arm were compared in a pre and post quasi-experimental study conducted at a tertiary care centre in North India. Patients with positive cultures from medical wards and intensive care units were included. Comparing the time to first antibiotic modification after culture positivity, MALDI-TOF-MS-based identification, and clinical outcomes in both arms was the primary objective. Antibiotic modifications, escalation, and de-escalation were all recorded. Results: The intervention arm exhibited a substantially shorter median time to first antibiotic modification (2010 mins vs 2905 mins, p=0.002) than the conventional arm. In the interventional group, a total of 44 out of 100 antibiotic modifications were implemented. Of these, 19 (43.3%) were determined solely by the MALDI report, without the anticipation of susceptibility assessments. De-escalation of antibiotics constituted the pre-dominant form of modification (47.4%). The difference between the 27% and 32% mortality rates in the intervention arm and the conventional arm was not statistically significant (p=0.52). Conclusion: MALDI-TOF-MS facilitates the modification of antibiotics early on. The primary benefit lies in the reduction of superfluous antibiotic usage. Early organism identification and reporting prior to the availability of susceptibility results did not result in any mortality benefit. This strategy, when combined with a strong antimicrobial stewardship programme, can aid in the reduction of antibiotic use.

Metagenome Analysis of Cholesteatoma-associated Bacteria: A Pilot Study.

Introduction: Cholesteatoma usually harbors a poly-microbial infection. As the diversity of bacterial pathogens in the Indian COM is unknown, we set out to identify the bacteria associated with cholesteatoma disease in different patients of North India using targeted metagenomic analysis of the 16 S rRNA gene. Methods: We recruited 15 patients of cholesteatomatous chronic otitis media (COM), who underwent surgical disease clearance. We divided these patients into four groups based on the four clinic-radiological stages categorized as per the EAONO/JOS joint consensus statement classification. Representative samples were extracted during the surgery and sent for bacterial culture and sensitivity and 16 S rRNA gene metagenomic analysis. Results: While 12 (80%) of the patients belonged to clinical Stage I/II; one patient had an extracranial complication (stage III) and two patients had an intracranial complication (stage IV). Our detailed bacterial metagenomics analyses showed that while phylum Proteobacteria was most abundant (reads up to ∼ 95%) in specimens from nine patients, phylum Firmicutes was most abundant (up to ∼ 80%) in specimens from four patients. Gamma (γ) Proteobacteria and Epsilon (ε) Proteobacteria were the most abundant class amongst Proteobacteria. Class Tissierellia stood out as the most abundant Firmicutes (40-60%), followed by Clostridia (20%) and Bacilli (10%). There was negligible difference in the bacterial profiles across all four clinical stages. Conclusion: Cholesteatoma is primarily associated with Proteobacteria and Firmicutes phyla, even in complicated disease. Further studies with a larger sample size are required to validate our findings. Supplementary Information: The online version contains supplementary material available at 10.1007/s12070-024-04678-9.

Retrospective Analysis of Hospital Acquired Infection and Antibiotic Resistance in Coronary Care Unit (CCU), Adult, and Pediatric Cardiology Wards.

BACKGROUND: Antibiotics resistance is an paramount threat affecting the whole world but nowhere situation is as gloomy as in India. No study till date regarding epidemiology of hospital acquired infections in coronary care units(CCU) and cardiology wards from India. From Indian perspective it is the first observational study to analyse microbiological profile and antibiotic resistance in CCU. The purpose of this observational study is to explore the epidemiology and importance of infections in CCU patients. METHODOLOGY: After ethics committee approval, the records of all patients who were admitted in coronary care units, adult and pediatric cardiology wards surgery between January 2020 and December 2021 were reviewed retrospectively. The type of organism,source of infection ,age wise distribution and seasonal variability among patients who developed hospital acquired infection (HAI) were determined. RESULTS: 271 patients developed microbiologically documented HAI during from January 2020 to December 2021. Maximum number of organisms(78/271 28.78%) are isolated from urinary samples ,followed by blood stream(60/271 22.14%) and Endotracheal tube (54/271 19.92%). Acinetobacter baumanii (53/271, 19.5%) being the most common isolate among all the samples taken . Acinetobacter was the most frequent pathogens isolated in patients with LRTI and blood stream infection while E. coli was from urinary tract infection . In the adult population, infection with E. coli(24.6%) is the most common followed by Klebsiella pneumoniae (12.8%) and Acinetobacter baumanii (10.1%). In the pediatric population Acinetobacter baumanii (38.6%%) is the most common followed by Klebsiella pneumoniae (20.5%) and Methicillin Resistant Staphylococcus aureus, MRSA (6.8%). Commonly used antibiotics eg ciprofloxacin,ceftazidime and amikacin were found to be resistant against the top three isolates. CONCLUSION: Urinary tract was the most common site of infection and Gram-negative bacilli, the most common pathogens in adult as well as pediatric population. Antibiotic resistance was maximum with commonly isolated microorganisms.

Regional variations in antimicrobial susceptibility of community-acquired uropathogenic Escherichia coli in India: Findings of a multicentric study highlighting the importance of local antibiograms.

Objectives: Evidence-based prescribing is essential to optimize patient outcomes in cystitis. This requires knowledge of local antibiotic resistance rates. Diagnostic and Antimicrobial Stewardship (DASH) to Protect Antibiotics (https://dashuti.com/) is a multicentric mentorship program guiding centers in preparing, analyzing and disseminating local antibiograms to promote antimicrobial stewardship in community urinary tract infection. Here, we mapped the susceptibility profile of Escherichia coli from 22 Indian centers. Methods: These centers spanned 10 Indian states and three union territories. Antibiograms for urinary E. coli from the outpatient departments were collated. Standardization was achieved by regional online training; anomalies were resolved via consultation with study experts. Data were collated and analyzed. Results: Nationally, fosfomycin, with 94% susceptibility (inter-center range 83-97%), and nitrofurantoin, with 85% susceptibility (61-97%), retained the widest activity. The susceptibility rates were lower for co-trimoxazole (49%), fluoroquinolones (31%), and oral cephalosporins (26%). The rates for third- and fourth-generation cephalosporins were 46% and 52%, respectively, with 54% (33-58%) extended-spectrum ß-lactamase prevalence. Piperacillin-tazobactam (81%), amikacin (88%), and meropenem (88%) retained better activity; however, one center in Delhi recorded only 42% meropenem susceptibility. Susceptibility rates were mostly higher in South, West, and Northeast India; centers in the heavily populated Gangetic plains, across north and northwest India, had greater resistance. These findings highlight the importance of local antibiograms in guiding appropriate antimicrobial choices. Conclusions: Fosfomycin and nitrofurantoin are the preferred oral empirical choices for uncomplicated E. coli cystitis in India, although elevated resistance in some areas is concerning. Empiric use of fluoroquinolones and third-generation cephalosporins is discouraged, whereas piperacillin/tazobactam and aminoglycosides remain carbapenem-sparing parenteral agents.

Changes in antimicrobial resistance of Escherichia coli isolated from community-associated urinary tract infection before and during the COVID-19 pandemic in India.

PURPOSE: The impact of the COVID-19 pandemic on antimicrobial resistance (AMR) is largely studied in healthcare settings. There is a need to understand the fluctuations in AMR during pandemic at the community level. With urinary tract infection (UTI) being one of the most common infections in the community, the AMR profile of community-acquired UTI (CA-UTI) is considered representative AMR at the community level. METHODS: The study was taken in a cohort of patients with a clinical diagnosis of CA-UTI. The four study sites represented different community health centres in India. Escherichia coli isolates were analysed phenotypically and genotypically for AMR pre-COVID (October 2019-February 2020) and in the first (March 2020-February 2021) and second waves of COVID-19 (March 2021-December 2021). RESULTS: E. coli was the predominant uropathogen (229, 82%). Increased susceptibility to nitrofurantoin was observed during the pandemic. Reduced susceptibility to first-line oral antibiotics and carbapenems was seen during the second wave, and an increased minimum inhibitory concentration (MIC50) to beta-lactams and fluoroquinolones was seen during the pandemic. Genomic analysis of E. coli isolates showed some AMR genes (aacC1, aacC4, SHV, QepA) only during the second wave. CONCLUSION: One good outcome of the pandemic was increased susceptibility to nitrofurantoin, while drawback was a significant decrease in susceptibility to oral antibiotics during the second wave and increased MIC50 of some antibiotics. Decreased susceptibility to last-resort carbapenems and the occurrence of various AMR genes during the second wave of the pandemic are of great concern.

Antimicrobial efficacy of eravacycline against emerging extensively drug-resistant (XDR) Acinetobacter baumannii isolates.

PURPOSE: Drug-resistant Acinetobacter baumannii is an emerging threat. This study has been conducted to observe the efficacy of eravacycline along with the RND-efflux pump system. METHODS: A cross-sectional study was done collecting 48 clinical isolates of Acinetobacter baumannii. MICs of 15 antibiotics were detected along with BMD of tigecycline and eravacycline. PCR products of drug-resistant regulatory genes were sequenced and analyzed. RESULTS: Of the total 48 Isolates, 35 (72.91%) were XDR and 13 (27.08%) were MDR. Out of all, 60.41% of isolates were found to be susceptible to eravacycline by BMD according to both FDA and EUCAST guidelines. A 2-fold decline of MIC50/90 was observed with the use of eravacycline compared to tigecycline. RND-efflux genes like AdeC in 30 (62.5%) isolates and Regulatory gene AdeS in 29 (60.41%) isolates were detected, explaining the existing resistance mechanism. CONCLUSIONS: XDR Acinetobacter poses an escalating threat due to its resistance to multiple antibiotics, raising serious concerns in healthcare settings. Eravacycline is an encouraging new drug for empirical use in severe infection caused due to the same. Molecular investigation and strict antimicrobial stewardship should be followed to control the emergence, and a better understanding of mechanisms of resistance to prevent the spread of drug-resistant isolates.

Detection of Aminoglycoside Modifying Enzyme (AME) genes in Acinetobacter baumannii isolates and the inhibitory effect of efflux pump activity on drug susceptibility pattern.

INTRODUCTION: The development of aminoglycoside modifying enzymes (AMEs) and increased efflux activity are considered important aminoglycosides resistance mechanisms. AIM: This study is focused on the detection of the AMEs gene and assessing the effect of efflux pump inhibitor on the reversal of A. baumannii drug susceptibility. METHODOLOGY: Bacterial DNA was amplified using AMEs gene-specific primers. Isolates were also investigated for efflux pump activity using efflux pump inhibitor (EPI) i.e. Carbonyl cyanide m-chlorophenyl hydrazone (CCCP) and the impact of both mechanisms was analyzed. RESULTS: Among A. baumannii isolates, 55% isolates (n â€‹= â€‹22/40) were identified to have aminoglycoside modifying enzymes genes; ant(3')-I gene (50%, 11/22), aac(6')-Ib gene (45.4%, 10/22), aph(3')-I gene (18.1%, 4/22) and aac(3)-I (9.1%, 2/22). Total 70% isolates have shown MIC alteration in different classes of drugs in response to EPI-CCCP. Such alteration was found in 100% amikacin sensitive and 58.6% amikacin resistant, 93.7% and 57.1% gentamicin sensitive and resistant isolates respectively. CONCLUSION: The presence of aminoglycosides modifying enzymes was frequent among aminoglycosides resistant A. baumannii isolates and the coexistence of efflux pumps activity also plays an important role to increase drug resistance. REPOSITORIES: Genbank and their accession numbers are MT903331[aac(3)-I], MT903332 MT903333 [ant(3')-I], MT903334, MT903335 [aph(3')-I)] and MT903336, MT940242 [ aac(6')-Ib].

A Quality Improvement (QI) initiative in diagnostic stewardship of urine culture in an era of Laboratory Information Systems (LIS).

Standard urine culture is the gold standard for diagnosing urinary tract infections (UTIs) but fails to differentiate true UTI from asymptomatic bacteriuria, which is important to prevent the overuse of antibiotics. Correlation with the presence or absence of pyuria can be helpful in giving a hint of the true situation. With the help of Laboratory Information System (LIS), patients' urinalysis reports can be conveniently accessed and compared simultaneously with appropriate reports. In our study, a quality improvement initiative was planned for appropriate reporting of urine culture and antimicrobial susceptibility testing using information obtained through LIS.

Distinct epigenetic signatures of classical and hypervirulent Klebsiella pneumoniae.

Emergence and spread of the hypervirulent pathotype of Klebsiella pneumoniae have significantly increased infection rates in community as well as healthcare settings. There is an increasing interest to identify discriminating features between classical K. pneumoniae (cKp) and hypervirulent K. pneumoniae (hvKp) to facilitate our understanding of the rapid emergence and dissemination of the hypervirulent pathotype. Here, we sought to identify unique epigenetic signatures of hvKp pathotype that differ from its classical counterpart using single-base resolution methylome analysis of native DNA sequencing on the Oxford Nanopore Technologies platform. The overall global adenine methylation in GATC motifs (i.e., Dam methylation motif) and cytosine methylation in CCWGG motifs (i.e., Dcm methylation motif) were significantly higher in hvKp isolates compared to that in cKp isolates, irrespective of their position in chromosomes or putative extra-chromosomal genetic elements. Notably, we observed significant enrichment of hypermethylated GATC and CCWGG motifs in the virulome of hvKp compared to hvKp genes not directly associated with virulence. We also observed increased methylation of GATC and CCWGG motifs in the capsule synthesis locus of hvKp isolates compared to cKp isolates. Furthermore, we identified several differentially methylated genes (DMGs) between the two pathotypes; interestingly, these DMGs include metal ion transporters, multidrug efflux pumps, transcriptional regulators of stress response, and genes associated with biofilm formation. Our results highlight hypermethylation of GATC and CCWGG motifs as unique epigenetic signatures of hvKp isolates.IMPORTANCEHypervirulent Klebsiella pneumoniae (hvKp) is a more virulent and rapidly evolving hypermucoviscous pathotype of classical K. pneumoniae (cKp). The hypervirulent pathotype is a major public health concern and is associated with high infection rates in community as well as hospital settings. With the recent emergence of multidrug-resistant hvKp, it has become imperative to investigate non-classical mechanisms such as epigenetics in addition to canonical biochemical and genetic mechanisms that delineate and differentiate the hypervirulent pathotype from its classical counterpart. Here, we identify genome-wide differences in adenine and cytosine methylation marks at well-characterized motifs between the two pathotypes. Overall, significantly higher levels of methylation were observed across chromosomal DNA and extrachromosomal elements in hvKp compared to cKp. Among hvKp isolates, the genes associated with virulence are particularly enriched for methylation marks. Our findings shed light on how epigenetic signatures may help distinguish the pathogenic potential of bacteria.

Umbilical Cord Blood IgA Levels and Bacterial Profile in Preterm Neonates Born with Maternal Risk Factors for Early-Onset Neonatal Sepsis.

OBJECTIVES: To investigate the IgA levels and bacterial profile in umbilical cord blood (UCB) samples of mothers with risk factors compared to those without risk factors; and to understand the link between UCB culture positivity and neonatal outcomes [early-onset sepsis (EOS) or death within 7 d of life]. METHODS: This is a pilot prospective case-control study. Mothers with preterm deliveries (gestational age <34 wk) were enrolled in two groups- Cases: Those with antenatal risk factors (prolonged duration of rupture of membranes of ≥24 h or chorioamnionitis) and controls: Those without these two risk factors. Serum IgA levels was assayed and microbiological culture was tested in UCB samples. 16S sequencing to determine the UCB microbiome was performed in a subset of samples (n = 15). Neonates were followed-up for the occurrence of EOS or death until 7 d of life. RESULTS: Forty-nine mothers as cases and 50 mothers as controls were consecutively enrolled. No significant difference was observed in the IgA levels (60.5 vs. 58.1 mg/L; p = 0.71), neonatal blood culture positivity (4.1% vs. 8.0%; p = 0.41) and UCB culture positivity (30.6% vs. 26.0%; p = 0.61) in the two groups. No difference was observed between the groups in occurrence of EOS or death within 7 d of life. Proteobacteria, Firmicutes and Actinobacteria were the most abundant phyla. Serratia, Bifidobacterium, Collinsella, Meganomas and Blautia being the most common genera. CONCLUSIONS: Cord blood IgA concentration could not differentiate the neonates at-risk of infection due to its presence in both the groups.

Procalcitonin for Detecting Culture-Positive Sepsis in Neonates: A Prospective, Multicenter Study.

INTRODUCTION: It is unclear if serum procalcitonin (PCT) estimated at sepsis suspicion can help detect culture-positive sepsis in neonates. We evaluated the diagnostic performance of PCT in culture-positive sepsis in neonates. METHODS: This was a prospective study (February 2016 to September 2020) conducted in four level-3 units in India. We enrolled neonates suspected of sepsis in the first 28 days of life. Neonates with birth weight <750 g, asphyxia, shock, and major malformations were excluded. Blood for PCT assay was drawn along with the blood culture at the time of suspicion of sepsis and before antibiotic initiation. The investigators labeled the neonates as having culture-positive sepsis or "no sepsis" based on the culture reports and clinical course. PCT assay was performed by electrochemiluminescence immunoassay, and the clinicians were masked to the PCT levels while assigning the label of sepsis. Primary outcomes were the sensitivity, specificity, and likelihood ratios to identify culture-positive sepsis. RESULTS: The mean birth weight (SD) and median gestation (IQR) were 2,113 (727) g and 36 (32-38) weeks, respectively. Of the 1,204 neonates with eligible cultures, 155 (12.9%) had culture-positive sepsis. Most (79.4%) were culture-positive within 72 h of birth. The sensitivity, specificity, and positive and negative likelihood ratios at 2 ng/mL PCT threshold were 52.3% (95% confidence interval: 44.1-60.3), 64.5% (60.7-68.1), 1.47 (1.23-1.76), and 0.74 (0.62-0.88), respectively. Adding PCT to assessing neonates with 12.9% pretest probability of sepsis generated posttest probabilities of 18% and 10% for positive and negative test results, respectively. CONCLUSION: Serum PCT did not reliably identify culture-positive sepsis in neonates.

A Comparative Review of ICMR, WHO, and EMA Guidelines for Good Clinical Laboratory Practices.

With the advancement of clinical research and the increased burden on laboratory services, there is an unmet need for guidelines regarding proper laboratory functioning and reliable data generation. Several organizations from all over the world have published guidelines for these clinical and research laboratories. Good Clinical Laboratory Practices (GCLP) are stepwise procedures aimed at strengthening the quality of test results produced by all clinical laboratories engaged in human sample analysis. In this article, we attempt a comparison of the GCLP guidelines recently issued by the Indian Council of Medical Research with the guidelines released by the World Health Organization and the European Medicines Agency. Also, we have included and discussed several suggestions that, if included, will lead to the strengthening of the laboratory practices used for both research and patient care for overall improvement in the Indian healthcare system.

Tick-tock, beat the clock: comparative analysis of disc diffusion testing with 6-, 10-, and 24-h growth for accelerated antimicrobial susceptibility testing and antimicrobial stewardship.

Disc diffusion testing by Kirby-Bauer technique is the most used method for determining antimicrobial susceptibility in microbiological laboratories. The current guidelines by The Clinical and Laboratory Standards Institute (CLSI) 2022 specify using an 18- to 24-h growth for testing by disc diffusion. We aim to determine if using an early growth (6 h and 10 h) would produce comparable results, thus ultimately leading to reduced turnaround time. Six-hour, 10-h, and 24-h growths of 20 quality control strains and 6-h and 24-h growths of 48 clinical samples were used to perform disc diffusion testing using a panel of appropriate antimicrobial agents. Disc diffusion zone sizes were interpreted for all and comparative analyses were performed to determine categorical agreement, minor errors (mE), major errors (ME), and very major errors (VME) according to CLSI guidelines. On comparing with the standard 24 h of incubation, disc diffusion from 6-h and 10-h growths of quality control strains showed 94.38% categorical agreement, 5.10% mE, 0.69% MEs, and no VMEs. Disc diffusion testing for the additional 40 clinical samples yielded a similarly high level of categorical agreement (98.15%) and mE, ME, and VME of 1.29%, 1.22%, and 0% respectively. Disc diffusion testing using early growth is a simple and accurate method for susceptibility testing that can reduce turnaround time and may prove to be critical for timely patient management.

Microbiological Characterization and Clinical Facets of Elizabethkingia Bloodstream Infections in a Tertiary Care Hospital of Eastern India.

Purpose: Elizabethkingia is an emerging non-fermenting Gram-negative bacillus (NFGNB) causing bloodstream infections (BSI) associated with high mortality. It demonstrates a unique antimicrobial profile in showing susceptibility to antimicrobials effective against Gram-positive bacteria. This study was undertaken to determine the overall frequency of Elizabethkingia BSI, associated risk factors, microbiological susceptibility, and clonal relationship of Elizabethkingia isolates using Enterobacterial Repetitive Intergenic Consensus Polymerase Chain Reaction (ERIC-PCR). Patients and Methods: Elizabethkingia isolates obtained from the blood culture of admitted patients (August 2020-December 2021) were identified by the VITEK 2 system and subjected to an antimicrobial susceptibility test by standard procedures. Demographics, co-morbidities, risk factors for survival, and outcome were summarized and analyzed by Chi-square test, Kaplan-Meier curve, and Cox regression. Clonal relatedness between Elizabethkingia isolates was analyzed using ERIC­PCR fingerprinting with the "PAST: Paleontological statistics software package". Results: Of 13,747 blood samples received during the study period, 13.59% were culture positive, and 14.60% were NFGNBs. The frequency of Elizabethkingia spp. among all NFGNBs in BSI was 29.30%, and the overall prevalence in BSI was 4.21%. In patients with Elizabethkingia BSI, Foley's catheter was present in 81.25% of the cases. 100% susceptibility was observed to linezolid, followed by vancomycin (98.75%) and chloramphenicol (89.5%). The 30-day mortality rate in the patients of Elizabethkingia BSI was 26.25%. The Presence of COVID-19, pneumonia, diabetes mellitus (DM), mechanical ventilation (MV), and prior antibiotics were significantly different (p<0.05) between the survival and death groups. ERIC-PCR profile dendrogram of Elizabethkingia isolates showed ten major clusters indicating high genetic diversity. Conclusion: Elizabethkingia was responsible for one-third of NFGNB BSI in a single-center study, with approximately 26% of 30-day all-cause mortality. Most isolates were susceptible to linezolid, vancomycin, and chloramphenicol. COVID-19 was the most significant risk factor associated with mortality. ERIC-PCR of Elizabethkingia isolates exhibited high genetic diversity.

Genome profiling of uropathogenic E. coli from strictly defined community-acquired UTI in paediatric patients: a multicentric study.

BACKGROUND: Urinary tract infection (UTI) in children is a common bacterial infection. The emergence of extended-spectrum beta-lactamases (ESBLs) poses a major challenge against the treatment of uropathogens. We aimed to characterize the E. coli isolates recovered from children with UTI for their resistance profile and circulating sequence types (ST). METHODS: Children (> 1.5-18 years of age) from different community health centres of India with symptoms of UTI were enrolled. Isolates causing significant bacteriuria were identified by Matrix-Assisted Laser Desorption Ionization Time of Flight Mass Spectrometry (MALDI-TOF MS) and tested for antimicrobial susceptibility by the automated system, VITEK-2 (Biomeriux, Durhum, US). Nineteen E. coli isolates (15 ESBL positive and 4 ESBL negative) were sequenced in Oxford Nanopore platform followed by core-genome phylogeny, accessory genome cluster analysis, identification of sequence types, mobile genetic elements, genetic antimicrobial resistance markers. The correlation between detection of antimicrobial resistance genes with phenotypic resistance profiles was also investigated. RESULTS: Eleven percent of children had significant bacteriuria [male:female-1:1, > 50% were 11-18 years of age group]. E. coli was predominant (86%) followed by K. pneumoniae (11%). Susceptibility of E. coli was highest against fosfomycin (100%) followed by carbapenems (90.7%) and nitrofurantoin (88.8%). ST131 (15.8%) and ST167 (10.5%) found as high-risk clones with the presence of plasmid [IncFIB (63.1%), IncFIA (52.6%)], and composite transposon [Tn2680 (46.6%)] in many isolates. Few isolates coharboured multiple beta-lactamases including blaNDM-5 (33.3%), blaOXA-1 (53.3%), blaCTX-M-15 (60%) and blaTEM-4 (60%). CONCLUSIONS: This study highlights horizontal transmission of resistance genes and plasmids in paediatric patients at community centers across the nation harbouring multidrug-resistant genes such as blaNDM-5 and blaCTX-M-15 associated with high-risk clones ST131 and ST167. The data is alarming and emphasizes the need for rapid identification of resistance markers to reduce the spread in community. To our knowledge, this is the first multicentric study targeting paediatric UTI patients from the community setting of India.

The Gram stain: Implication in blood culture reporting.

Bloodstream infections are life-threatening. They are responsible for prolonged hospital stays and high healthcare costs. Clinical microbiology has an essential role to play in its management. Direct Gram-stain reporting from positive blood culture bottles has been found helpful. We aimed to evaluate the use of the automated blood culture system in adjunct to the conventional Gram-stain technique, with a direct antimicrobial susceptibility test on the second day. Concordance was highly satisfactory.