High Minimum Inhibitory Concentration Research Articles

Prevalence of T. rubrum and T. interdigitale Exhibiting High MICs to Terbinafine in Clinical Samples Analyzed in the Portuguese Mycology Reference Laboratory

Cutaneous fungal infections represent a significant burden worldwide with a high impact on public health. Accurate identification of dermatophyte species causing these infections is vital for an appropriate treatment. Terbinafine is the primary agent against Trichophyton species due to its clinical efficacy; however, cases of elevated minimum inhibitory concentration (MIC) have been reported, raising clinical and epidemiological concerns. Herein, we aimed to detect Trichophyton rubrum and Trichophyton interdigitale isolates collected from clinical samples with terbinafine-high MICs (TRB-hMIC). A total of 168 isolates, recovered from 2017 to 2023, were identified as T. rubrum complex (140/83.4%) or T. interdigitale (28/16.7%) and further screened regarding their terbinafine susceptibility. Four isolates with capacity to grow in terbinafine media were detected by screening, and these and a further sixteen random isolates were submitted to the broth microdilution method. This methodology confirmed the four (2.4%) isolates as TRB-hMIC. One T. rubrum and three T. interdigitale showed a minimum inhibitory concentration (MIC) higher than 1 mg/L. Partial sequencing of the SQLE gene identified point mutations in T. rubrum (Phe397Iso) and in one T. interdigitale (Phe397Leu) isolate. Notably, in the other two T. interdigitale isolates with TRB-hMIC, no point mutations in the SQLE gene were identified. In conclusion, TRB-hMIC isolates (T. rubrum and T. interdigitale) were identified in clinical samples analyzed in Portugal, as antifungal susceptibility testing is a crucial routine for identifying treatment failures and also for epidemiological purposes aiming to monitor the dynamics of terbinafine resistance.

Dissemination of arr-2 and arr-3 is associated with class 1 integrons in Klebsiella pneumoniae clinical isolates from Portugal.

\nKlebsiella pneumoniae is a common pathogen of healthcare-associated infections expressing a plethora of antimicrobial resistance loci, including ADP-ribosyltransferase coding genes (arr), able to mediate rifampicin resistance. The latter has activity against a broad range of microorganisms by inhibiting DNA-dependent RNA polymerases. This study aims to characterise the arr distribution and genetic context in 138 clinical isolates of K. pneumoniae and correlate these with rifampicin resistance. All isolates were subjected to whole-genome sequencing for species identification, typing and AMR genes identification, along with the determination of the minimum inhibitory concentration (MIC) of rifampicin. Molecular detection of arr genes and class 1 integrons was performed for rifampicin-resistant isolates. Efflux activity was investigated as a possible determinant of rifampicin resistance in isolates devoid of known genetic determinants. Twelve isolates exhibited high rifampicin MICs (≥ 64mg/L), 124 showed intermediate MICs (16-32mg/L) and two displayed low (8mg/L) MICs. Two arr allelic variants, arr-2 and arr-3, were found across one and nine K. pneumoniae isolates, respectively, all within class 1 integrons, including a newly described integron, and all associated with high rifampicin MICs (≥ 64mg/L). Elevated resistance levels were additionally linked to increased arr-2/3 expression and closer proximity to the promoter. No arr gene or rpoB mutations were found across the remaining two isolates and no correlation between efflux activity and high-level rifampicin resistance was found for both isolates. In conclusion, this study demonstrates that arr genes confer high levels of rifampicin resistance in K. pneumoniae highlighting its widespread dissemination within class 1 integrons.

Antifungal susceptibility profile of Candida species and uncommon yeasts from drug abusers with oral candidiasis

In Iran, there is limited information regarding the species distribution and antifungal susceptibility profiles of yeast isolates from drug addicts suffering from oral candidiasis (OC). In this study, 104 yeast isolates, including 98 Candida species and 6 uncommon yeasts, were collected from 71 drug abusers with OC. The susceptibility profiles of Candida spp. and uncommon yeasts to amphotericin B (AMB), itraconazole (ITC), nystatin (NYC), fluconazole (FLC), and caspofungin (CAS) were evaluated using the CLSI broth microdilution method. The prevalence of OC in the sampled population was found to be 29%. The susceptibility profile of Candida spp. revealed remarkable sensitivity, with 100% and 99% of isolates susceptible to NYC and AMB, respectively. However, concerning levels of resistance or non-wild-type minimum inhibitory concentrations (MICs) were observed, with 13.2% of Candida isolates showing resistance to FLC, 13.2% to ITC, and 16.3% to CAS. Notably, 35.2% of patients showed mixed yeast species, while 5.1% of Candida isolates exhibited multidrug resistance. The analysis of the uncommon yeast species showed that the overall frequencies of the highest MICs were observed for CAS. Furthermore, within the six non-Candida species identified, Hanseniaspora opuntiae and one isolate of Pichia kluyveri exhibited resistance to FLC and ITC, respectively, while all non-Candida species were susceptible to AMB and NYC. Additionally, one isolate of Pichia kluyveri exhibited simultaneously high MICs to two drugs ITC and CAS. Furthermore, the Hanseniaspora opuntiae isolate showed high MICs to CAS and FLC. The findings from the present study suggest that AMB and NYC can be suitable choices for empiric treatment of both common Candida species and uncommon yeast infections in substance abuse patients.

Whole genome analysis revealed the role of bla OXA-23 and bla OXA-66 genes in carbapenem resistance of Acinetobacter baumannii strains

ABSTRACT Acinetobacter baumannii is a multidrug-resistant bacterium that has emerged as a significant nosocomial pathogen globally and renowned for its ability to acquire antimicrobial resistance (AMR) genes. However, understanding of its resistance mechanisms to certain drug classes remains limited. This study focused on four bacterial strains (AB863, AB889, AB930, and AB960) exhibiting carbapenem resistance. They demonstrated high minimum inhibitory concentration (MIC) (128 mg/L) to meropenem and were categorized as extensively drug-resistant strains. Subsequently, they were identified as A. baumannii through 16S rRNA gene sequence analysis and species-specific PCR targeting the bla OXA51-like gene. Three strains were sequenced for their genomes to study the genetic determinants and functional relevance of carbapenem resistance. The draft genome length of the strains ranged from 3.8 to 4.0 Mbp. A total of 16 antibiotic resistance genes including the genes bla OXA-23 and bla OXA-66 which mediate carbapenem resistance were identified in the genomes. A comprehensive multilocus sequence typing analysis involving 95 A. baumannii strains from different Asian countries assigned the four strains to sequence type 2 (ST2), the most predominant ST circulating in Asia. Comparative genome analysis also revealed bla OXA-66 as the most dominant variant of bla OXA-51-like gene and also a widespread distribution of bla OXA-23 gene. In addition, various mobile genetic elements associated with AMR genes and three efflux pumps families were detected in the genomes of the strains. Transformation of bla OXA-23 and bla OXA-66 genes resulted in meropenem resistance in the transformant which exhibited a MIC of 2 mg/L, thus confirming direct involvement of both genes in carbapenem resistance.

Analysis of Efflux Pump Contributions and Plasmid-Mediated Genetic Determinants in Ciprofloxacin-Resistant Salmonella.

This study aimed to explore the interactions among genetic determinants influencing ciprofloxacin resistance in Salmonella. Treatment with PAβN, an efflux pump inhibitor, resulted in a 4-32-fold reduction in the minimum inhibitory concentration (MIC) across all 18 ciprofloxacin-resistant Salmonella isolates. Notably, isolates without point mutations reverted from resistance to sensitivity. The efflux pump played a crucial role in resistance development, particularly in serovar Enteritidis, where PAβN treatment caused a more significant MIC reduction (16-32-fold) in five strains carrying the GyrA (Asp87Tyr) mutation, which initially exhibited high MICs (8 μg/mL). Several resistance genes were identified on transferable plasmids: oqxAB and aac(6')-Ib-cr were associated with IncF plasmids in S. Enteritidis, IncA/C plasmids in S. Typhimurium, and IncHI2 plasmids in S. Virchow. Additionally, qnrS1 and/or qepA were carried by IncA/C plasmids in S. Thompson. Whole-genome sequencing revealed the presence of an oqxAB module integrated into the chromosomal DNA of S. Derby. Although the MICs of ciprofloxacin in transconjugants and transformants remained low (1-4 μg/mL), they exceeded the clinical breakpoint for susceptibility. These findings highlight the synergistic impact of efflux pumps and plasmid-mediated resistance mechanisms, contributing to the increasing prevalence of ciprofloxacin resistance and posing a significant threat to food safety.

In Vitro Dynamic Model Evaluation of Meropenem Alone and in Combination with Avibactam Against Carbapenemase-Producing Klebsiella pneumoniae.

Background: A potential strategy to maintain the efficacy of carbapenems against carbapenemase-producing Klebsiella pneumoniae (CPKP) is their combination with carbapenemase inhibitors. To address these issues, the effectiveness of a novel combination of meropenem with avibactam against CPKP was studied. Additionally, the applicability of a pharmacokinetically-based approach to antibiotic/inhibitor minimum inhibitory concentration (MIC) determinations to better predict efficacy was examined. Methods: CPKP strains were exposed to meropenem alone or in combination with avibactam in an in vitro hollow-fiber infection model. Treatment effects were correlated with simulated antibiotic and antibiotic/inhibitor combination ratios of the area under the concentration-time curve (AUC) to the MIC (AUC/MIC). All MICs were determined at standard and at high inocula; combination MICs were determined using the conventional approach with fixed avibactam concentration or using the pharmacokinetic (PK)-based approach with a fixed meropenem-to-avibactam concentration ratio, equal to the respective drug therapeutic AUC ratios. Results: Meropenem alone was not effective even against a "susceptible" CPKP strain. The addition of avibactam significantly improved both meropenem MICs and its effectiveness. The effects of meropenem alone and in combination with avibactam (merged data) correlated well with AUC/MIC ratios only when MICs were determined at high inocula and using the PK-based approach (r2 0.97); the correlation was worse with the conventional approach (r2 0.73). Conclusions: The effectiveness of meropenem/avibactam against CPKP is promising. A single "effect-AUC/MIC" relationship useful for predicting meropenem efficacy (alone or in combination with avibactam) was obtained using MICs at high inocula and combination MICs determined using a PK-based approach.

Characterization of an Enterococcus sp. SMC-9 strain isolated from bile of a patient with cholangitis.

The genus Enterococcus is increasingly recognized for its involvement in various human infections, with several species known to be pathogenic. This study characterized Enterococcus sp. SMC-9, isolated from bile of a patient with cholangitis, and compared its characteristics with those of Enterococcus montenegrensis CoE-012-22T, recently isolated from dried beef sausage. A comprehensive analysis, encompassing phylogenetic, genomic, and phenotypic studies, confirmed that strain SMC-9 belongs to the same species as E. montenegrensis CoE-012-22T. However, comparative genomic analysis revealed key differences in virulence and antibiotic resistance gene profiles between the two strains. Notably, genes related to exopolysaccharide biosynthesis and the L-rhamnose biosynthesis pathway were found exclusively in strain SMC-9, suggesting their role in the strain's colonization of the biliary tract and its involvement in cholangitis. Additionally, the tetracycline resistance gene tet(M), which was absent in E. montenegrensis CoE-012-22T, was identified in strain SMC-9, explaining its high tetracycline minimum inhibitory concentration (>16 μg/mL). These findings highlight the unique pathogenic traits of strain SMC-9 compared to E. montenegrensis CoE-012-22T. Our study underscores the significant genetic and phenotypic variations that can exist among strains within the same species, highlighting the critical need for strain typing to assess their potential impact on patient outcomes and public health.

Multilocus sequence typing, antimicrobial susceptibility, and phage discovery of motile Aeromonas species from the pond-cultured goldfish Carassius auratus on two farms in Japan

Motile Aeromonas spp., including Aeromonas hydrophila, A. caviae, and A. veronii biovar sobria can cause motile aeromonad disease and pose serious threats to the aquaculture industry. Motile aeromonads showing high minimum inhibitory concentrations (MICs) to widely used various antibiotics in global aquaculture production have been reported. Several studies have shown that A. hydrophila phages are effective in treating infected fish. However, in Japan, epidemiological reports and phage studies of motile Aeromonas spp. isolated from fish are scarce. In this work, we visited two goldfish farms in Japan between 2020 and 2021, and obtained 33 motile Aeromonas isolates from Carassius auratus exhibiting symptoms of motile aeromonad disease. Multilocus sequence typing analysis revealed that the isolated motile aeromonads belonged to a wide variety of sequence types (STs) including new STs. The ST2007 accounted for 50% (6/12) of A. hydrophila isolates, and each was isolated at different timepoints and farms. The isolated bacteria exhibited high MICs for oxolinic acid, nitrofural, sulfamerazine, and oxytetracycline; 13 isolates (39%) which presented with MICs above commercially used concentrations of oxolinic acid, nitrofural, and sulfamerazine in Japan and were classified as non-wild types for oxytetracycline. The subsequently isolated phages showed high specificity for the motile aeromonads isolates.

Phenotypic and Genotypic Analysis of Antimicrobial Resistance in Mycoplasma hyopneumoniae Isolated from Pigs with Enzootic Pneumonia in Australia.

Mycoplasma hyopneumoniae, an important cause of enzootic pneumonia in pigs in many countries, has recently been shown to exhibit reduced susceptibility to several antimicrobial classes. In the present study, a total of 185 pig lung tissue samples were collected from abattoirs in Australia, from which 21 isolates of M. hyopneumoniae were obtained. The antimicrobial resistance profile of the isolates was determined for 12 antimicrobials using minimum inhibitory concentration (MIC) testing, and a subset (n = 14) underwent whole-genome sequence analysis. MIC testing revealed uniformly low values for enrofloxacin (≤1 μg/mL), florfenicol (≤8 μg/mL), lincomycin (≤4 μg/mL), spectinomycin (≤4 μg/mL), tetracycline (≤0.5 μg/mL), tiamulin (≤2 μg/mL), tildipirosin (≤4 μg/mL), tilmicosin (≤16 μg/mL) tulathromycin (≤2 μg/mL), and tylosin (≤2 μg/mL). Higher MICs were observed for erythromycin (MIC range: 16-32 μg/mL), gamithromycin, and tilmicosin (MIC range of both: 32-64 μg/mL). Whole-genome sequencing of the isolates and additional screening using mismatch amplification mutation assay PCR did not identify any known genetic resistance markers within 23S rRNA (macrolides), DNA gyrase A, and topoisomerase IV genes (fluoroquinolones). The WGS data also indicated that the Australian M. hyopneumoniae isolates exhibited limited genetic diversity and formed a distinct monophylectic clade when compared to isolates from other countries. These findings indicate that Australian M. hyopneumoniae likely remains susceptible to the major antimicrobials used to treat enzootic pneumonia in pigs and have evolved in isolation from strains identified in other pig-producing countries.

Antibacterial Activity of Clindamycin/BPO in Combination With Adapalene

Introduction: Acne guidelines recommend the addition of benzoyl peroxide (BPO) to antibiotics to reduce resistance in Cutibacterium acnes (C. acnes). Pairing the antibiotic/BPO combination with a retinoid, such as adapalene, may further increase treatment efficacy, although research on adapalene’s antibacterial activity is limited. To determine if adapalene improves antimicrobial activity, this in vitro study evaluated minimum inhibitory concentration (MIC) of clindamycin, adapalene, and BPO alone or in combination against both susceptible and resistant C. acnes isolates. Methods: Part 1a: C. acnes sensitivity to clindamycin, adapalene, or BPO was assessed via MIC values obtained by the broth microdilution method, with lower MIC indicating higher susceptibility. Part 1b: The effect (synergistic, additive, antagonistic, or indifferent [no interaction]) of combining adapalene with clindamycin or BPO on C. acnes inhibition was evaluated using a checkerboard assay, wherein two test compounds are combined in varying concentrations. Part 2: Susceptibility to single formulations of clindamycin, adapalene, and BPO, and fixed-dose combination formulations of an antibiotic (clindamycin or erythromycin) with BPO was determined by measuring antibacterial zone of inhibition using agar diffusion method, with larger diameter indicating increased bacterial inhibition. Results: Part 1a: Clindamycin demonstrated strain-dependent activity (MIC range: <0.125-64 μg/ml), while BPO and adapalene had no/low activity as indicated by high MIC (>512 and >64, respectively) against the 6 acne-associated strains tested. Part 1b: The combination of adapalene with clindamycin resulted in an additive effect for 3 and no interaction for 1 strain, whereas adapalene with BPO did not result in any interaction against the 4 acne-associated strains tested. Part 2: Activity of single formulations varied, with adapalene 0.1% having no activity (zone of inhibition: 0 cm) against any of the 8 acne-associated C. acnes strains tested. The fixed-dose combination formulations had generally similar activity against all strains tested (range: 0.9-5 cm). Conclusions: In these in vitro analyses, adapalene and BPO had no/low activity against C. acnes, whereas clindamycin alone or in combination formulations demonstrated strain-dependent activity. The relatively high MIC of BPO is not unexpected and aligns with previous studies in which the in vitro activity of BPO was low compared to its high in vivo antimicrobial activity. Further, addition of the retinoid adapalene had an additive effect on the antimicrobial activity of clindamycin against 3 out of 4 C. acnes strains tested, but no effect on the activity of BPO in vitro. Taken together, these data suggest that when combined with clindamycin/BPO, adapalene may enhance antimicrobial activity, while also bringing its own, unique retinoid mechanism of action to the triple combination, adding to clindamycin's bacteriostatic and anti-inflammatory properties. Funding: Ortho Dermatologics

Resistance tofluoroquinolones among Klebsiella pneumoniae isolates collected from patients referred to the teaching hospitals of Ardabil university of medical sciences in 2020

Background. Klebsiella pneumoniae (KP) is one of the most important causes of hospital-acquired infection and one of the most important causes of ventilator-associated pneumonia. During the present study, we investigated the resistance to fluoroquinolones among KP isolates collected from patients referred to hospitals affiliated with Ardabil University of Medical Sciences in 2020. Methods. In this descriptive, cross-sectional study, the minimum inhibitory concentration (MIC) of ciprofloxacin (CLX) was determined by the agar dilution method, and mutations in the gyrA and parC genes and the presence of resistance genes, such as accA, qnr, and qepA, were evaluated using the polymerase chain reaction method. Results. Out of 100 isolates, 43% (n = 43) were resistant to CLX. The MIC range of CLX was 0.5 µg/mL to 1024 µg/mL. Isolates with a high MIC of CLX were investigated for mutations in the gyrA and parC genes. In the investigation of mutations in gyrA, a substitution from serine to isoleucine was observed at position 83, while no mutation was found in parC. In 10 isolates (10%), the presence of the aac (6')-Ib-cr gene was detected, while qepA and qnr resistance genes were not observed in any of the isolates. Conclusion. Based on the findings, a high frequency of the aac (6')-Ib-cr gene was reported in this study. Since this gene is located on a plasmid, it can be easily transferred between KP strains in a hospital setting and leads to the spread of resistance to fluoroquinolones. Practical Implications. The emergence of bacterial resistant strains is on the rise and has become a serious public health problem worldwide. Infection with these strains leads to complicated diseases with worse outcomes, prolonged hospitalization, increased healthcare costs, an increased need to use second-line drugs with costs, and treatment failures.

Fusarium spp. causing invasive disease in humans: A case series from north India.

Owing to their inherent resistance to different classes of antifungals, early identification of Fusarium spp. is crucial. In this study, 10 clinical isolates were included from patients with invasive fusariosis involving lungs, sinuses, or both. Clinico-radiological data were collected. Samples were processed by standard laboratory procedures. Three gene regions (ITS, TEF1, and RPB2) were amplified by PCR for multilocus sequencing. Fusarium MLST, FUSARIUM-ID, and FUSARIOID-ID databases were used for final identification. Antifungal susceptibility testing was performed by broth microdilution following CLSI M38-A3 and Sensititre™ YeastOne™ YO9 plate. Pulmonary involvement was seen in all patients, and sino-nasal involvement was present in six. Radiologically, consolidations and cavitations were present in eight and six cases, respectively. Halo sign was present in six; reverse halo sign was also found in three of them. Direct microscopy showed septate hyphae that were morphologically different from those found in aspergillosis. Results of the molecular identification were as follows: two Fusarium irregulare, one Fusarium pernambucanum, one Fusarium incarnatum, one Fusarium sp. FIESC 30, two Fusarium keratoplasticum, one Fusarium falciforme, one Fusarium pseudonygamai, and one Fusarium delphinoides. For both Fusarium solani (FSSC) and Fusarium incarnatum-equiseti (FIESC) species complexes, amphotericin B had the lowest minimum inhibitory concentrations (MICs). Importantly, for terbinafine, all FIESC isolates had low MICs, while FSSC isolates had high MICs. In some cases, early identification of Fusarium spp. is possible by means of morphology of hyphae on direct microscopy and findings on radiology. Molecular identification, at least to the species complex level, is crucial for the choice of antifungals.

Rosmarinic Acid Exhibits Antifungal and Antibiofilm Activities Against Candida albicans: Insights into Gene Expression and Morphological Changes.

Candida species, opportunistic pathogens that cause various infections, pose a significant threat due to their ability to form biofilms that resist antifungal treatments and immune responses. The increasing resistance of Candida spp. and the limited availability of effective treatments have prompted the research of natural compounds as alternative therapies. This study assessed the antifungal properties of RA against Candida species, focusing on its impact on C. albicans biofilms and the underlying mechanisms. The antifungal efficacy of RA was evaluated using the CLSI M27-A3 microdilution method on both fluconazole-susceptible and -resistant strains. Biofilm formation by C. albicans was assessed through a crystal violet assay, while its antibiofilm activity was analyzed using an MTT assay and field emission scanning electron microscopy (FESEM). Gene expression related to biofilm formation was studied using quantitative real-time PCR (qRT-PCR), and statistical analysis was performed with an ANOVA. Among the 28 Candida strains tested, RA exhibited minimum inhibitory concentration (MIC) values ranging from 160 to 1280 μg/mL. At a 640 μg/mL concentration, it significantly reduced the expression of genes associated with adhesion (ALS3, HWP1, and ECE1), hyphal development (UME6 and HGC1), and hyphal cAMP-dependent protein kinase regulators (CYR1, RAS1, and EFG1) in RAS1-cAMP-EFG1 pathway (p < 0.05). FESEM analysis revealed a reduction in hyphal networks and disruptions on the cell surface. Our study is the first to demonstrate the effects of RA on C. albicans adhesion, hyphae development, and biofilm formation through gene expression analysis with findings supported by FESEM. This approach distinguishes our study from previous studies on the effect of RA on Candida. However, the high MIC values of RA limit its antifungal potential. Therefore, more extensive research using innovative methods is required to increase the antifungal effect of RA.

Phenotypic heterogeneity in bacteria: the rise of antibiotic persistence, clinical implications, and therapeutic opportunities.

The rising incidence of antimicrobial resistance (AMR) and the diminishing antibiotics discovery pipeline have created an unprecedented scenario where minor infections could become untreatable. AMR phenomenon is genetically encoded, and various genetic determinants have been implicated in their emergence and spread. Nevertheless, several non-genetic phenomena are also involved in antibiotic treatment failure which requires a systematic investigation. It has been observed that in an isogenic population of bacteria, not all cells behave or respond the same way to an antibiotic, because of the inherent heterogeneity among them. This heterogeneity is not always heritable but rather phenotypic. Three distinct types of phenotypic heterogeneity, namely tolerance, persistence, and heteroresistance have been observed in bacteria having significant clinical implications influencing the treatment outcome. While tolerance is when a population can survive high doses of antibiotics without changing the minimum inhibitory concentration (MIC) of the drug, persistence occurs in a subpopulation of bacteria that can survive exposure to high antibiotic doses. In contrast, when a subpopulation shows a very high MIC in comparison to the rest of the population, the phenomenon is called heteroresistance. In this article, we have highlighted bacterial persistence with a focus on their emergence and the underlying molecular mechanisms. Moreover, we have tried to associate the genome-wide methylation status with that of the heterogeneity at a single-cell level that may explain the role of epigenetic mechanisms in the development of persistence.

Colistin Insusceptibility in Carbapenem-Resistant Enterobacteriaceae Isolates From a Tertiary Referral Centre.

Background Antimicrobial resistance has developed significant importance as a worldwide health concern more so in the 21st century. This is more specifically observed among the Enterobacteriaceae family, the major group of Gram-negative bacteria. Resistance to carbapenems and colistin antimicrobials from the reserve group, in critical infection treatment poses a substantial therapeutic challenge. This study aims to detect resistance to both carbapenems and colistin in Enterobacteriaceae isolates, emphasizing the importance of monitoring. Materials and methods A laboratory-based study investigated 82 Carbapenem-Resistant Enterobacteriaceae (CRE) isolates from hospitalized patients at Krishna Hospital & Medical Research Centre, Karad, India, for two years (November 2021-November 2023). The Kirby-Bauer disc diffusion method was used to determine the antimicrobial susceptibility following Clinical Laboratory Standards Institute 2022 recommendations. To validate Carbapenem-Resistant Enterobacteriaceae isolates, the Modified Carbapenem Inactivation Method (mCIM) was used, and Colistin resistance was ascertained using Broth Microdilution Susceptibility Testing (BMD). Results Among 309 Enterobacteriaceae isolates, 82 (26.5%) exhibited carbapenem resistance, with 75 confirmed by Modified Carbapenem Inactivation Method (mCIM) testing. For patients in the age group 51-60 years (20%), with men being the most prevalent, the predominance of CRE isolates was from the Intensive Care Unit (44%), mostly from urine samples (34.6%). The most dominant CRE was Klebsiella pneumoniae (62.7%), followed by Escherichia coli (32%). Colistin resistance was found in 14 (18.7%) of CRE isolates, with the highest minimum inhibitory concentration (MIC) at 4 µg/mL (n=10) and 16 µg/mL (n=4). Colistin-resistant isolates (n=10) were sensitive to amikacin (71.4%) and resistant to many antibiotics. Conclusion The extreme rise of colistin-resistant cases among CRE in healthcare settings is a solemn alarm. Among the most effective treatments, aminoglycosides amikacin and netilmicin show the greatest sensitivity against these colistin-resistant CRE infections. The findings stress the difficulties in treating such infections and the dire need for solid antimicrobial stewardship and novel therapeutic strategies.

SPECIES IDENTIFICATION AND IN VITRO ANTIFUNGAL SUSCEPTIBILITY TESTING OF ASPERGILLUS ISOLATED FROM VARIOUS CLINICAL SAMPLES

Objectives: Aspergillus is ubiquitous mold species present in environment in the form of spores. Infection due to Aspergillus is uncommon in immunocompetent individuals unless they possess any abnormality or have undergone any treatment with corticosteroids. In immunocompromised individuals, infection by this fungus is common among people with prolonged neutropenia, transplants, and human immunodeficiency virus infection. The symptoms are diverse, ranging from allergic reactions to invasive aspergillosis and life-threatening complications. The aim of this study is to isolate and identify Aspergillus species obtained from various clinical specimens and perform antifungal susceptibility testing (AFST). Methods: A total of 200 isolates in which positive direct microscopic findings correlated well with culture growth for Aspergillus were included. Antifungal susceptibility was performed by micro broth dilution technique according to clinical laboratory standard institute M38-A2 guidelines. Results: Aspergillus flavus was found to be predominant species followed by Aspergillus fumigatus. AFST was performed, where all Aspergillus strains exhibited low minimum inhibitory concentration (MIC) and minimum effective concentration (MEC) values for most of antifungal drugs tested except for one strain of A. flavus, which exhibited high MIC for voriconazole and high MEC for caspofungin. Conclusion: Polyenes, azoles, and echinocandin resistance are emerging in many parts of the world. Therefore, continued application of AFST combined with morphological characterization for species identification is critical in detecting the emergence of resistance among various Aspergillus species to reduce mortality, morbidity, and overcome treatment failure.

Prevalence and factors associated with human colonisation and vancomycin minimum inhibitory concentration of &lt;em&gt;Staphylococcus aureus&lt;/em&gt; in Colombo, Sri Lanka

Introduction: Staphylococcus aureus is known to cause a wide variety of diseases of varying severity in humans. The objectives of this study were to determine the prevalence of S. aureus and methicillin resistant S. aureus (MRSA) colonisation, assess factors associated with MRSA colonisation, identify antibiotic susceptibility patterns of MRSA isolates, determine distribution of vancomycin minimum inhibitory concentrations (MIC) of S. aureus and factors associated with high vancomycin MIC values (≥2 µg/ml). Methods: A descriptive cross sectional study was conducted among 341 participants aged 18 to 84 years selected randomly from the Colombo district from 11th December 2018 to 10th April 2019. Samples were collected from the nose, axilla, groin of participants and identification and sensitivity testing of S. aureus were performed based on standard operation procedures given in the Laboratory manual in microbiology published by the Sri Lanka College of Microbiologists (2011). Results: Prevalence of S. aureus and MRSA colonisation in the sample population was 34.6% and 13.78% respectively. All MRSA isolates were susceptible to linezolid. MRSA susceptibility to gentamicin (86%), co-trimoxazole (88 %), and chloramphenicol (80%) was high. Erythromycin had the lowest susceptibility (20%). Inducible clindamycin resistance was demonstrated in 31% of isolates. Meat consumption and consumption of antibiotics within the previous six months showed a significant association with MRSA colonisation. Vancomycin MIC of S. aureus ranged from ≤ 0.5 to 2 µg/ml. Conclusion: As prevalence of MRSA colonisation is 13.78%, more attention should be paid when managing patients with suspected S. aureus infections presenting from the community. Further research on associated factors and environmental triggers are warranted with a larger sample size and different study design to formulate national regulations and guidelines.

Phenotypic and Genetic Characteristics of Carbapenemase-Producing Enterobacterales Isolates at Okayama University Hospital.

Spread of carbapenemase-producing Enterobacterales (CPE) is an ongoing public health issue worldwide, including in Japan. In this study, we investigated the phenotypic and genetic characteristics of CPE isolates at Okayama University Hospital over the 5 years (2013-2018) prior to the outbreak of the 2019 coronavirus pandemic. Of 24 carbapenem-resistant Enterobacterales isolated during the study period, we identified 8 CPE isolates harboring blaIMP-1 (5 isolates) and blaIMP-6 genes (3 isolates). Bacterial species and carbapenem susceptibility patterns exhibited diversity. Minimum inhibitory concentrations (MICs) of meropenem were generally higher than those of imipenem and biapenem. Results of pulsed-field gel electrophoresis demonstrated that neither clonal nor plasmid-mediated outbreaks of blaIMP-harboring CPE isolates have developed at our hospital. One Klebsiella oxytoca isolate showed a high MIC (128 μg/mL) of meropenem, which could be explained by the high plasmid copy number. Subsequent analysis of this isolate may elucidate the intricacies of carbapenem resistance profiles among CPE isolates. Collectively, our findings underscore the necessity for ongoing genetic surveillance of CPE, complemented by tailored approaches for infection prevention and control.

First Confirmed Description of Acremonium egyptiacum from Greece and Molecular Identification of Acremonium and Acremonium-like Clinical Isolates.

Acremonium and the recently separated acremonium-like genera, such as Sarocladium, are emerging causes of opportunistic disease in humans, mainly post-traumatic infections in immunocompetent hosts, but also invasive infections in immunocompromised patients, such as those undergoing transplantation. Acremonium egyptiacum has emerged as the major pathogenic Acremonium species in humans, implicated mainly in nail but also in disseminated and organ specific infections. In this first study of acremonium-like clinical isolates in Greece, 34 isolates were identified and typed by sequencing the internal transcribed spacer, and their antifungal susceptibility was determined by a modified CLSI standard M38 3rd Edition method for filamentous fungi. A. egyptiacum was the primary species (18 isolates) followed by Sarocladium kiliense (8), Acremonium charticola, Gliomastix polychroma, Proxiovicillium blochii, Sarocladium terricola, Sarocladium zeae, and Stanjemonium dichromosporum (all with one isolate). Two isolates, each with a novel ITS sequence, possibly represent undescribed species with an affinity to Emericellopsis. All three A. egyptiacum ITS barcode types described to date were identified, with 3 being the major type. Flutrimazole, lanoconazole, and luliconazole presented the lower minimum inhibitory concentration (MIC) values against A. egyptiacum, with a geometric mean (GM) MIC of 2.50, 1.92, and 1.57 μg/mL, respectively. Amphotericin B, itraconazole, posaconazole, voriconazole, terbinafine, amorolfine, and griseofulvin MICs were overall high (GM 12.79-29.49 μg/mL). An analysis of variance performed on absolute values showed that flutrimazole, lanoconazole, and luliconazole were equivalent and notably lower than those of all the other drugs tested against A. egyptiacum. Antifungal susceptibility of the three different A. egyptiacum genotypes was homogeneous. Overall, the high MICs recorded for all systemically administered drugs, and for some topical antifungals against the tested A. egyptiacum and other acremonium-like clinical isolates, justify the routine susceptibility testing of clinical isolates.

Intraperitoneal daptomycin dosing for peritonitis may be inadequate: a Monte Carlo simulation approach to optimize dosing and outcomes

A two-compartmental mathematical pharmacokinetic model with first-order elimination of patients receiving CAPD of 4 exchanges for 6 h with 2 L of dialysate used in each cycle was developed to predict daptomycin disposition in 120 h of therapy. The pharmacodynamic target was plasma AUC/MIC equal to or greater than 666. The dose that achieved at least 90% of the probability of target attainment was defined as an optimal dose. Administering intraperitoneal 300 mg daily for 1 exchange daily regimen would be sufficient to treat peritonitis with S. aureus infection with MICs of 0.25 mg/L in patients undergoing CAPD. A higher dosage may be required for infections with a higher minimum inhibitory concentration. Pharmacodynamic targets and MICs significantly contributed to daptomycin doses in this setting. Clinical validation of our recommendations is recommended.